10 1021@acsami 0c12410

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Review
Functional polymer nanofibers: from spinning
fabrication techniques to recent biomedical applications
Danilo Martins dos Santos, Daniel S. Corrêa, Eliton S
Medeiros, Juliano Oliveira, and LUIZ Henrique C. MATTOSO
ACS Appl. Mater. Interfaces, Just Accepted Manuscript • DOI: 10.1021/acsami.0c12410 • Publication Date (Web): 16 Sep 2020
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Page 1 of 102 ACS Applied Materials & Interfaces
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2 Functional polymer nanofibers: from spinning fabrication
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techniques to recent biomedical applications
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Danilo Martins dos Santos 1, Daniel S. Correa 1, Eliton S. Medeiros 2, Juliano E.
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13 Oliveira 3, Luiz H. C. Mattoso 1
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17 1 Nanotechnology National Laboratory for Agriculture (LNNA), Embrapa
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20 Instrumentação, 13560-970, São Carlos, SP, Brazil.
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22 2 Materials and Biosystems Laboratory (LAMAB), Department of Materials
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24 Engineering (DEMAT), Federal University of Paraíba (UFPB), Cidade Universitária,
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27 58.051-900, João Pessoa, PB, Brazil.
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3 Department of Engineering, Federal University of Lavras (UFLA), 37200-900, Lavras,
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31 MG, Brazil.
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ACS Applied Materials & Interfaces Page 2 of 102
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2 Abstract
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7 Functional polymeric micro/nanofibers have emerged as promising materials for the
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9 construction of structures potentially useful in biomedical fields. Among all kinds of
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11 technologies to produce polymer fibers, spinning methods have gained considerable
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14 attention. Herein, we provide a recent review on advances in the design of micro- and
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16 nanofibrous platforms via spinning techniques for biomedical applications. Specifically,
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18 we give emphasizes on electrospinning, solution blow spinning, centrifugal spinning,
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and microfluidic spinning approaches. We first introduce the fundamentals of these
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23 spinning methods and then highlight the potential biomedical applications of such
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25 micro- and nanostructured fibers for drug delivery, tissue engineering, regenerative
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27 medicine, disease modeling and sensing/biosensing. Finally, we outline the current
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30 challenges and future perspective of spinning techniques for the practical applications of
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32 polymer fibers in the biomedical field.
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37 Keywords: functional nanofibers, electrospinning, solution blow spinning, centrifugal
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39 spinning, microfluidic spinning, biomedical applications.
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2 1. Introduction
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4 In the past decades, considerable effort has been devoted to create novel
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7 biomaterials with appropriate biophysical and biochemical properties capable to
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9 regenerate damaged tissues and restore their functions1. Among engineered
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11 biomaterials, polymer nanofibers have appeared as potential candidate for varied
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14 biomedical applications including drug delivery2,3, biosensing4,5, tissue engineering6,7
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16 and regenerative medicine8,9. The appealing features of polymer nanofibers include the
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18 high surface area/volume ratio, mechanical flexibility, ease of fabrication, the
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possibility of surface or even bulk modification to produce functional nanofibers as well
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23 as their capability to mimic many aspects of physiological microenvironment in
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25 vivo3,10,11.
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27 Indeed, the importance of polymer nanofibers can be perceived by the increasing
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30 number of articles published in the last years in indexed journals. A quick survey
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32 performed in the Web of Science (Thomson Reuters), using the keywords "polymer"
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34 and "nanofiber" yielded the results presented in Fig. 1, where (a) refers to published
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37 items in each year and (b) refers to Citations in each year. The results refer to the period
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39 2010 2019, and clearly evidence the importance of this technological hot topic.
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31 year. Data were retrieved from Web of Science (Thomson Reuters) by searching articles
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using the combined words "Polymer" AND "nanofibers" in the period 2010-2019 period
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Current nanofiber fabrication techniques include top-down and bottom-up
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43 approaches. In top-down approaches, larger bulk material is converted into nanofibers
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45 by using physical12,13, biological14,15 or chemical16,17 treatments. On the other hand,
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47 bottom-up approaches relies on assembling the matter at the molecular level to obtain
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50 nanofibers, including phase separation18, interfacial polymerization19,20, electrochemical
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52 assembly20, and especially spinning techniques11,21. The latter has gained increasing
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54 popularity within the last decade by the widespread use of electrospinning22–25, solution
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blow spinning26–28, centrifugal spinning29,30, and microfluidic spinning31–33 techniques,
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59 which allow the preparation of polymer nanofibers presenting a broad range of complex
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architectures suitable for biomedical application34,35.
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2 To date, many excellent review articles have provided an overview of specific
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4 spinning technique for biomedical applications21,31,33,36–39. However, to the best of our
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7 knowledge, a review comparing spinning methods and exclusively devoted to their
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9 biomedical applications was still missing. In this context, here we critically examine
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11 recent advances and compare the main fiber spinning techniques, namely
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14 electrospinning, solution blow spinning, centrifugal spinning and microfluidic spinning
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16 focusing on their unique features to produce nanofibers for biomedical applications,
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18 including drug delivery, tissue engineering, regenerative medicine, disease modeling
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and sensing/biosensing. We also discuss on challenges, limitations and perspectives
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23 related to the techniques and how they can impact their biomedical applications. We
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25 envisioned that this review article will provide important guide to the rational and
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27 practical design of nanofibrous structures intended for biomedical applications.
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32 2. Spinning methods for producing polymer nanofibers
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34 Current spinning methods generally involve the extrusion of a polymer through
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37 a spinneret, which is then subjected to external driving forces (e.g. electrical, gas
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39 pressurized and centrifugal forces) and/or mechanical drawing to form jets that are
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41 stretched generating fibers upon solidification as a result of precipitation or drying
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process40. According to spinning method, materials choice and processing parameters,
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46 fiber-based materials presenting different architectures and properties can be obtained.
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48 Various spinning methods have been developed for producing micro/nanofibers from a
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50 broad range of synthetic and natural polymers as well as polymer blends36,41. In this
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53 section, we provide a description of the most used spinning methods including
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55 electrospinning, solution blow spinning, centrifugal spinning, and microfluidic spinning
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57 pointing out their advantages and disadvantages in terms of fiber productivity and
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60 versatility.
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4 2.1. Electrospinning
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7 Electrospinning is the most common fiber-forming technique and it is based on
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9 the application of electrostatic forces for generating polymer nanofibers of distinct
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11 morphology and diameters from solutions, suspensions, or melts. This method,
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14 originally reported in 189942, regained considerable attention in 1990’s mainly from
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16 Darrell Reneker and Jayesh Doshi studies43, which proposed the term “electrospinning”
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18 and demonstrated the viability of this technique for producing long and continuous
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fibers with diameters down to the nanometer scale by using different polymers.
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23 Remarkable progress has been made in the last two decades with regard to the
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25 development of electrospinning techniques in order to control the morphology of the
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27 deposited nanofibers as well as to increase fiber productivity and versatility. These
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30 techniques include solution electrospinning, melt electrospinning, coaxial
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32 electrospinning, near-field electrospinning, multi-jet electrospinning and needless
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34 electrospinning36,40. By properly selecting the electrospinning method and controlling
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37 the process parameters as well as material composition, nanofibers with distinct
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39 morphologies can be obtained, as displayed in microscopy images in Fig. 2: (a) porous
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41 nanofibers44; (b) aligned nanofiber45; (c) patterned nanofiber46; (d) core-shell
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nanofibers47; (e) hollow nanofiber48; (f) helicoidal nanofibers49; (g) nanoribbons50; (h)
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46 nanofiber yarns51 (i) nanoneedles52. Next, we describe and compare the major features
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48 of selected electrospinning methods including near-field electrospinning, magnetic-
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50 assisted electrospinning, coaxial electrospinning, multi-nozzle electrospinning,
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53 nozzleless-free electrospinning.
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34 Fig. 2. SEM images of electrospun nanofibers of varied morphology: (a) porous
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36 nanofibers; (b) aligned nanofiber; (c) patterned nanofiber; (d) core-shell nanofibers; (e)
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38 hollow nanofiber; (f) helicoidal nanofibers; (g) nanoribbons; (h) nanofiber yarns (i)
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nanoneedles. Panel (a) reprinted with permission from ref (44). Copyright 2016 Elsevier.
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43 Panel (b) reprinted with permission from ref (45). Copyright 2016 Elsevier. Panel (c)
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45 reprinted with permission from ref (46). Copyright 2019 Elsevier. Panel (d) reprinted
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with permission from ref (47). Copyright 2020 Elsevier. Panel (e) reprinted with
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50 permission from ref (48). Copyright 2017 Elsevier. Panel (f) reprinted with permission
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52 from ref (49). Copyright 2015 American Chemical Society. Panel (g) reprinted with
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54 permission from ref (50). Copyright 2016 Elsevier. Panel (h) reprinted with permission
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57 from ref (51). Copyright 2019 Springer Nature. Panel (i) reprinted with permission from
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59 ref (52). Copyright 2015 American Chemical Society.
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2 2.1.1. Solution electrospinning
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4 A typical setup for solution electrospinning consists of a high-voltage power
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7 supply, a syringe pump, a spinneret and a conductive collector, as illustrated in Figure
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9 3a. Electrospinning is governed by an electrohydrodynamic process, during which a
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11 polymer solution is injected through the spinneret to produce a pendant liquid droplet
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14 that is then subject to an electric field applied by the voltage source. As a consequence,
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16 charge accumulates on the droplet surface, elongating the droplet into a jet, followed by
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18 stretching, elongation and solidification through solvent evaporation, ultimately
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yielding to polymer nanofibers. Such fibers are usually deposited onto a collector
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23 forming a non-woven web 53.
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56 Fig. 3. Schematic illustration of distinct electrospinning set-ups: (a) solution
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electrospinning; (b) melt electrospinning; (c) near-field electrospinning; (d) magnetic-
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assisted electrospinning; (e) coaxial electrospinning; (f) rotating wire electrospinning.
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2 Panel (a) adapted with permission from ref (54). Copyright 2016 Elsevier. Panel (b)
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4 reprinted with permission from ref (54). Copyright 2016 Elsevier. Panel (c) reprinted
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7 with permission from ref (55). Copyright 2020 American Chemical Society. Panel (d)
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9 reprinted with permission from ref (56). Copyright 2019 Elsevier. Panel (f) reprinted
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11 with permission from ref (57). Copyright 2017 Taylor & Francis.
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16 The formation, morphology, and diameter of electrospun nanofibers are largely
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18 influenced by the polymer and solvent properties, such as conductivity, viscosity, and
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molar mass. Processing parameters also play an important role, including the strength of
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23 applied electric field, the distance between spinneret and the collector, and the flow rate
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25 of the liquid), as well as ambient parameters (humidity and temperature of the
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27 surroundings)23,36. Another key parameter is the collector design, which affects the
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30 productivity, diameter, and morphology of fibers as well as the final architecture of the
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32 fibrous construct. A great variety of collectors have been used including aluminium foil,
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34 liquid bath, rotating drum, rotating disc, wire mesh, rotary metal funnel among others23.
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37 The proper choice of solvent and polymer is considered an important criteria on
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39 the solution electrospinning process. The solvent should be able to dissolve polymers in
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41 appropriate concentrations and present suitable volatility to allow the fiber formation
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and avoid clogging the needle36. Organic solvents, including formic acid, acetone,
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46 chloroform, dichloromethane, alcohols, dimethylformamide (DMF), dimethyl sulfoxide
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48 (DMSO), hexafluoroisopropanol (HFIP), tetrahydrofuran (THF), and trifluoroethanol,
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50 among others, are commonly used for electrospinning23,58,59. Some polymers require a
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53 mixture of different solvents in order to obtain suitable solutions for electrospinning60.
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55 A broad range of synthetic and natural polymers as well as polymer blends have
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57 been successfully applied in solution electrospinning. Synthetic polymers including
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60 poly(lactic acid), poly(lactic-co-glycolic acid), poly(vinyl alcohol),
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2 poly(vinylpyrrolidone), polycaprolactone, and poly(3-hydroxybutyrate) are classic
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4 materials that have been electrospun into nonwovens for diverse biomedical
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7 applications. In the last decade, more attention has been given to the use of biopolymers
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9 such as chitosan, chitin, dextran and alginate23,61,62 However, the poor
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11 electrospinnability of pure biopolymers imposes limits to their practical use. In this
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14 case, a successful strategy employed is blending them with an easily electrospinnable
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16 polymer to obtain an appropriate solution. Such strategy can result in a superior material
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18 that combines the benefits of both polymers with suitable properties for biomedical
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applications60–62.
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23 Although solution electrospinning has been in the spotlight of different research
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25 fields due to its great versatility, simplicity and low cost of the processing system, some
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27 challenges for its wide-spread industrial application still remains. First, the inherent use
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30 of organic solvents on electrospinning process involves economic and environmental
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32 concerns54. In addition, some residual solvents in electrospun materials can be
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34 cytotoxic, thereby limiting their use in biomedical applications. Another disadvantage
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37 of conventional solution electrospinning process relies on is its low production rate and
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39 high cost per gram of produced nanofibers36,63
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2.1.2. Melt electrospinning
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46 Melt electrospinning is typically employed to produce fibers from polymers that
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48 are difficult to dissolve in solvents before electrospinning (e.g. Polypropylene (PP),
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50 Polyethylene (PE) and Polyphenylene sulfide (PPS)). Conventional melt electrospinning
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53 apparatus is formed by a supply zone, electrical heating components, a high-voltage
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55 power supply, a spinneret and a conductive collector, as shown in Figure 3b. During
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57 melt electrospinning, the melted polymer is extruded through the spinneret and a high
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60 voltage potential is applied between the collector and spinneret, from which a charged
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2 jet is ejected. The polymer jet is then stretched towards the grounded collector where
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4 solidifies quickly forming ultrafine solid fibers54,64.
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7 The melt electrospinning process is mainly governed by the flow rate of
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9 polymer, drawing temperature, spinneret-to-collector distance, structural characteristics
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11 of polymer and applied voltage65. Fibers possessing diameters down to the micrometer
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14 scale can be obtained through optimization and controll of melt electrospinning
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16 processing parameters. In comparison to solution electrospinning, melt electrospinning
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18 offers some advantages for being a solvent-free process. The absence of solvent is
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environmentally friendly and improves the safety and productivity of the process 54. In
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23 addition, melt spinning has a lower manufacturing cost and allows the precise control of
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25 fibers deposition. Another important advantage compared to its solution counterpart, is
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27 that melt electrospinning can be coupled with fused deposition modeling (FDM)
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30 principle, which allows the preparation of complex structures with high resolution and
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32 precision, thus increasing its potential in numerous applications, particularly in
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34 advanced biomedical field such as tissue engineering and regenerative medicine, where
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37 cell alignment, guidance or protection is important 66–69.
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39 On the other hand, the number of polymers appropriate for melt electrospinning
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41 is much lower than that for solution electrospinning. Few polymers commercially
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available have been explored on melt electrospinning, including poly(ɛ-caprolactone)
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46 (PCL), polyamide-6, polyethylene, poly(methyl methacrylate) (PMMA), polyurethane,
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48 poly(ethylene terephthalate) (PET), poly(l-lactide) (PLLA) and poly(lactic-co-glycolic
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50 acid) (PLGA)36,54,70. The high heating temperature may result in polymer degradation
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53 and limits the processing of thermal sensitives polymers such as natural polymers as
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55 well as compounds such as drugs or bioactive molecules, which poses a great challenge
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57 for applications of melt electrospinning in biomedical field71. In order to improve the
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60 mechanical and tissue compatibility properties of melt electrospun fibers, some
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2 strategies including post-electrospinning technologies and the use of additives
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4 compounds in the polymer melt have been adopted72,73. The scalability of melt
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7 electrospinning process is also a great challenge and some strategies including multi-
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9 needle and needleless configurations, which will be described in the next sections, hold
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11 great potential to upscale fiber production63,74,75
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16 2.1.3. Near-field electrospinning
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18 Near-field electrospinning (NF-ES) offers a straightforward route to precisely
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control the positions of the deposited fibers in applications that require arranged or
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23 patterned micro/nanoscale fibrous structures. In the NF-ES approach, a sharp probe tip
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25 is utilized to dip liquid polymer solution or melted polymer and it is usually positioned
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27 close to the collector55,76,77. The shorter tip-to-collector distance (usually 500 μm – 5
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30 cm) drastically reduces the applied voltage (0.6 – 3 KV) and the bending instability in
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32 spinning, thus favoring the deposition of fibers in high spatial definition in a 3D motion
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34 platform (Figure 3c) 55. It has been demonstrated that two-dimensional (2D) as well as
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37 three-dimensional (3D) structures, can be fabricated by NF-ES, which expands the
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39 potential application of fibers in diverse fields such as tissue engineering76, energy
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41 harvesting78 and flexible sensors79.
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Indeed, NF-ES presents a number of advantages compared to conventional
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46 electrospinning, including reduction in applied voltage and material consumption as
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48 well as position-controlled deposition of the fibers along X, Y, and Z directions. Despite
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50 these advantages, the fibers prepared by NF-ES are usually thicker than those prepared
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53 by typical electrospinning process 55. Furthermore, the large-scale preparation of fibers
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55 is limited by the complexity of NF-ES setup and by the low feed rates of polymer
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2 2.1.4. Magnetic-assisted electrospinning
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4 Magnetic-assisted electrospinning is an approach for producing nanofibers that
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7 enables a better control over the fiber deposition, alignment and fiber diameter. As
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9 illustrated in Figure 3d, the setup is similar to the conventional configuration except for
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11 the application of a magnetic field close to the collector56. In this modified
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14 electrospinning method, the branching and splitting of spinning jets is reduced by
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16 applying an adjustable magnectic field that balances off the forces that creates
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18 instability, thus yielding to more uniform nanofibers and controlled deposition. In
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addition, the magnetic field also increases the velocity of the jet reaching the substrate
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23 and the internal alignment of polymeric chains, leading the reduction of fiber
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25 diameter56,80. Such are desirable for fabricating well-ordered nanofibers in a parallel
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27 fashion along the magnetic field lines for the development of biomedical devices and
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30 implants such as scaffolds.
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34 2.1.5. Coaxial electrospinning
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37 In coaxial electrospinning, the setup is essentially the same as the conventional
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39 configuration except for the use of a coaxial needle consisting of two concentric aligned
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41 hollow needles. In this case, two polymeric solutions or melt polymers are injected
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separately through the outer and inner needles by using two syringe pumps, as shown in
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46 Figure 3e81,82. Coaxial electrospinning has allowed the preparation of nanofibers with
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48 controlled compositions for the core and sheath layer83–86. This technique has also been
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50 explored to prepare nanofibers from unspinnable liquids, which are spun in fiber form
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53 as the core or sheath solution under the guidance of an easily electrospinnable polymer
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55 solution47,87. A variety of materials, including proteins88, oils89, and drugs47 have been
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57 electrospun as a core structure in core-sheath nanofibers. Coaxial electrospinning also
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60 enables the production hollow nanofibers (Figure 1e). During this process, a post-
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2 electrospinning step is required for selectively removing the sacrificial core from the as-
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4 spun core–sheath nanofibers90,91.
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7 Another important approach to prepare core-sheath nanofibers consists in
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9 emulsion electrospinning, where a water-in-oil (W/O) or an oil-in-water (O/W)
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11 emulsion are electrospun by using a single nozzle. During the process organization of
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14 the emulsified droplets into two distinct phases is carried as the solvent evaporates from
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16 the spinning jet92. This strategy holds great potential on the incorporation of sensitive
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18 bioactive molecules such as peptides, enzymes, hydrophobic and hydrophilic drugs93.
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23 2.1.6. Multi-nozzle electrospinning
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25 Despite the development of new methods to fabricate polymer nanofibers, the
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27 large-scale preparation of fibers is still somewhat limited. In order to increase fiber
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30 productivity, multi-noozles electrospinning employing a similar setup to conventional
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32 electrospinning process has been proposed, but using now of an array of needles, as
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34 ilustrated in Figure 4a, where multiple jets are simultaneously ejected from the
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37 noozles63,74. Although this technique offers great potential to increase the mass
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39 production, it still remains a challenging task because of the following factors: (i) inter-
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41 jet perturbations, which greatly affects the morphology and fiber formation resulting in
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inhomogeneous fibers, fiber–fiber bonding, and/or poor fiber distribution in the
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46 collected mat ; (ii) low feed rates through needles are required to prepare continuous
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48 nanoscale nanofibers, thus limiting the throughput of process; (iii) the clogging and
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50 cleaning of the needles94–96
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53 Although there are challenges and difficulties, some companies (e.g. Yflow®97,
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55 Inovenso Inc.98, Uepartner99) have been commercializing machines based on multi-
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57 needle electrospinning technology to produce nanofibers in industrial and semi-
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60 industrial scale. An innovative example is an industrial machine developed by Yflow®
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2 (Fig. 4b), which system is formed by a great number of needles (Fig 4c) able to produce
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4 large quantities of nanofiber-based products (Fig 4d)97. For comparison, conventional
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7 single-nozzle electrospinning reaches productivity lower than 100 mg per hour, while
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9 the production capacity of some industrial machines is superior to 200 g/hour63.
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42 Fig. 4. Multi-needle arrangement (a) Industrial multi-nozzle electrospinning machine
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44 developed by Yflow® (b) photograph of multi-nozzle device based injection (c)
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47 photograph of collected electrospun nonwoven (d). Reprinted from ref (97).
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49
50
51 2.1.7. Nozzleless electrospinning
52
53
54 Nozzleless-electrospinning, also known as needleless electrospinning and
55
56 spinneret-free electrospinning, functions similarly to electrospinning, but in this case, it
57
58 allows the spontaneous charged jet initiation on a free liquid surface without the use of a
59
60
nozzle (Figure 5a and b)40,100. In this case, the spinning jets are self-assembled and the
16
1
2 distance between each spinline self-stabilizes, thus affording the improvement on
3
4 productivity. Compared with multi-needle electrospinning, the nozzleless-free
5
6
7 electrospinning present some advantages that include simple design, robustness against
8
9 clogging as well as high troughput101–104. However, issues associated to this method
10
11 should be considered such as solvent evaporation from polymer solution bath, water
12
13
14 absorption on the reservoir and solvent ignition due to corona discharge in extreme
15
16 cases. Furthermore, the liquid flow rate and the emanation of uniform jets is not easy to
17
18 be precisely controlled. To address these issues, the ambient parameters (temperature
19
20
and relative humidity) can be controlled by using an enclosed system. In addition, the
21
22
23 modification of the setup by immerging rotatory solid spinnerets including metal
24
25 cylinder, disc, wire (Figure 3f), spiral wire coil as well as a metallic ball on the spinning
26
27 solution bath may improve the uniformity and the number of the jets formed36,40.
28
29
30 Elmarco Inc.105 produces and commercializes an industrial machine based on
31
32 nozzleless electrospinning named Nanospider™ NS 8S1600U (Figure 5c). In this
33
34 approach, a thin layer of a liquid polymer is continuously deposited on wire electrodes,
35
36
37 yielding to multiple jets when a high electric field is applied. This device can produce
38
39 over 20 000 000 m2/year of PA6-based nonwoven sheet presenting a layer width of 1,6
40
41 m (base weight: 0.03 g/m2) formed by fibers with a diameter around 150 nm105.
42
43
Although high throughput nanofibers production can be reached by using a needless
44
45
46 electrospinning machine, the preparation of fiber with diverse morphologies such as
47
48 core-sheath remains a challenge for this process.
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27 Fig. 5 - (a) schematic illustration of a nozzleless electrospinning setup, where the shape
28
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30 of spinneret is represented at the top right corner; (b) photograph of the linear flume
31
32 spinneret spinning process, insert image: a magnified image of multiple jet formation;
33
34 (c) photograph of the Nanospider NS 8S1600U industrial machine developed by
35
36
37 Elmarco Inc. Panel (a) and (b) reprinted with permission from ref (100). Copyright 2020
38
39 MDPI. Panel (c) reprinted from ref (105).
40
41
42
43
44
The nanofiber production rate can be greatly improved by multi-nozzle
45
46 electrospinning or Nozzleless electrospinning; however, the scale-up production is still
47
48 a great challenge, once it requires more complex and expensive equipment63. In this
49
50 sense, alternatives to the above-mentioned electrospinning methods have been
51
52
53 developed, including solution blow spinning26–28,106 and centrifugal spinning29,30. These
54
55 methods have gained great attention due to its capability and feasibility in generating
56
57 large quantities of nanofibers with well-defined structures at high speed and low cost.
58
59
60 Other important issues that remain challenging in electrospinning, as well as in solution
18
1
2 blow spinning and centrifugal spinning techniques, are the accumulation of the
3
4 evaporated solvent, which poses environmental and health concerns, and the absence of
5
6
7 spatiotemporal control of the composition, size, and morphology of the fibers26–33,54,106.
8
9 In order to address these issues, microfluidic spinning31–33 has emerged as a promising
10
11 alternative spinning method. In the next sections are presented the fiber formation
12
13
14 process, spinning systems, material types, and processing-structure relationships of the
15
16 above-mentioned alternative spinning methods.
17
18
19
20
2.2. Solution blow spinning
21
22
23 Solution blow spinning (SBS) is a gas-assisted fiber production method that has
24
25 gained considerable interest in the last years due to its simplicity, high productivity, and
26
27 versatility. Such technique is especially useful for spinning fibers from poorly
28
29
30 electrospinnable materials with low dielectric constant and/or electrical
31
32 conductivity26,27. SBS or slight variations of this technique have also been named in the
33
34 literature as air spinning or air-jet spinning107,108, solution blowing109,110, gas jet
35
36
37 spinning111, jet spraying112, air brushing112, solution spraying113, and pressure-driven
38
39 spinning114 . The conventional set up for solution blow spinning consists of concentric
40
41 nozzles, a source of compressed gas such as air, nitrogen, synthetic air, oxygen or
42
43
argon, a fiber collecting system, and an infusion pump to control the polymer ejection
44
45
46 rate, as schematically illustrated in Figure 6.
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Fig. 6. Schematic depiction of the Solution Blow Spinning (SBS) apparatus used for
26
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28 fiber production. (a) Infusion pump; (b) pressurized gas; (c) pressure gauge; (d) nozzle;
29
30 (e) polymer nanofibers leaving the nozzle and being directed to the collector (f); (g)
31
32 nanofibers mat deposited onto the collector, (h) nanofibers mat deposited in situ. In (d)
33
34
35 is also displayed a zoomed cross-sectional view of the nozzle, displaying the inner (by
36
37 which the polymer solution flows through) and outer (by which the pressurized air
38
39 flows through) nozzles, as well the solution cone and the fibers. Panel (g) and (h)
40
41
42 reprinted with permission from ref (27). Copyright 2009 Wiley.
43
44
45
46 During the SBS, the polymer solution is forced through the inner nozzle at a
47
48
constant feed rate, resulting in the formation of a droplet at the tip of the inner nozzle,
49
50
51 which is stretched by the high-pressure stream of compressed gas flowing through the
52
53 outer nozzle. The compressed air exiting the nozzle causes the surface of the drop to
54
55 shape into a cone similar to Taylor’s cone in electrospinning. When a critical air
56
57
58 pressure is exceeded, the aerodynamic forces overcome the surface tension forces, and a
59
60 jet erupting from the top of this cone is accelerated towards the collection target. As the
20
1
2 jets travel through the surrounding environment (across the working distance), the
3
4 polymer solution is stretched, and the solvent rapidly evaporates forming a web of
5
6
7 micro- and nanofibers that can be collected on virtually any target, including living
8
9 tissue, sheets, membranes, liquid surfaces and rotating drums27,115,116.
10
11 Process and material variables are injection rate, gas pressure, polymer
12
13
14 concentration and molecular weight, working distance, and protrusion distance of the
15
16 inner nozzle with respect to the outer nozzle, nozzle diameter and solvent used27,107–
17
18 110,115. In general, solution properties influence on fiber morphology similarly to
19
20
electrospinning and most spinning techniques, in which high concentrations and high
21
22
23 molecular weight polymers – high viscous solutions – give rise to thicker fibers. Solvent
24
25 volatility also imparts on fiber morphology, once low volatile solvents tend to produce
26
27 films rather than fibers, especially if the working distance is short; however, heating can
28
29
30 also be applied in order to increase the rate of solvent evaporation and then produce
31
32 fibers rather than films27,117,118. Higher pressures produce narrower fiber diameter
33
34 distribution, while low pressures produce broad ones. However, none of these
35
36
37 parameters should be considered individually, once the average fiber diameter and
38
39 distribution are a combination of all variables, including surrounding atmosphere itself
40
41 that influences on the rate of solvent evaporation27,115.
42
43
Solution Blow Spinning has been successfully employed to produce fibers of
44
45
46 synthetic and natural polymers from aqueous and non-aqueous solvents as well as
47
48 solvent mixtures with diameters ranging from a few nanometers to several micrometers
49
50 (Table 1). Recent developments include multi-nozzle systems for industrial-scale
51
52
53 production119, aqueous (heated) spinning118, and coaxial nozzles for core-shell fiber
54
55 production120,121.
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60
21
1
2
3
4 Table 1. Some polymeric micro- and nanofibers produced by Solution Blow Spinning.
5
6
7 Material Concentration Fiber diameter References
8
9 PLA, PS, PMMA 5–15% 80 nm–7.8 µm 27
10
11 122
PVDF/ MWCNT 10 % 300 nm –1.9 µm
12
13
Cellulose diacetate 12 % 950 ± 410 nm 123
14
15
16 6 wt. (PMMA)
17 PMMA-PAN 400 – 600 nm 109
18 8% (PAN)
19
20 115
21 PDLA 4–8% 70–2000 nm
22
Soy Protein/PA6 (core/shell) 14–20% 582–2051 nm 28
23
24
25 Cellulose/PEO (core/shell) 2–10% 0.26–1.9 µm 124
26
27 111
28
PEO, PVP 2–6% 100 – 3600 nm
29
Polyvinyl butyral (PVB) 7–13% 0.69 – 2.37 µm 125
30
31
32 PAN 8 – 20% 100 – 800 nm 126
33
34 127
PLA/PEO (blends), 5% 180 – 1200 nm
35
36
PLA, PCL, PEO 6% 100 – 520 nm 128
37
38
39 PEO 6 – 10% 400 – 1000 nm 129
40
41 PVA 12 – 20 % 300 – 3040 nm 118
42
43 130
44 Chitosan/PVA 8% 400 – 600 nm
45
Soy protein, PVA, PEO 8 – 17 % 500 – 600 nm 119
46
47
48 PA6 14 – 22 % 150 – 750 nm 131
49
50 132
51
PLA/PVP 5 – 20 % 400 – 1800 nm
52
Polystyrene 6 – 14 % 200 – 1000 nm 133
53
54
55 PCL 6–8% 200 – 1200 nm 134
56
57 135
Cellulose 5% 1 – 2.5 µm
58
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60
22
1
2 Since its first report in 200927, there has been an increasing number of papers
3
4 concerned with several aspects (materials and process) of Solution Blow Spinning.
5
6
7 Compared with electrospinning, SBS present some advantages including no need of
8
9 applying an electrical voltage, low cost; ease of fabrication and scaling 3D structures
10
11 with larger mesh size. In addition, SBS allows for portability and in principle any
12
13
14 surface, even biological substrates, can be coated by SB-spun fibers, which greatly
15
16 expands its potential in numerous applications, especially in biomedical field.
17
18 Furthermore, the scale-up capacity and the ability to produce sufficient fiber quantities
19
20
within reasonable time frames of SBS is higher than electrospinning26,63. Recently, a
21
22
23 modified SBS apparatus containing 400-600 needles was developed. This system has a
24
25 production capacity of 400 g/h with a density of 0.1–20 g/m2 and holds great potential
26
27 to scale-up fiber productivity63. However, as a major challenge of SBS, it is the
28
29
30 accumulation of the evaporated solvent during the process, which requires solvent
31
32 removal system and pose environmental, health and recycling concerns.
33
34
35
36
37 2.3. Centrifugal spinning
38
39 Centrifugal spinning is a simple and controllable process that uses centrifugal
40
41 forces to create fibers136. This technique has also been denoted as rotary jet
42
43
spinning30,137 and forcespinning138. As illustrated in Figure 7a, the basic centrifugal
44
45
46 spinning setup is comprised of an electric motor driven rotating spinneret coupled to a
47
48 polymer reservoir and a collector136. In this approach, the surface tension and viscosity
49
50 of melted polymer or polymer solution injected through the nozzle is overcomed by
51
52
53 centrifugal forces, resulting in the ejection of a jet, which is then elongated and
54
55 stretched by the air vortices (Figure 7bi and 7bii). During this phase, the solvent
56
57 evaporates and the solidification of the filament takes place, with the consequent
58
59
60 formation of fibers that are deposited on the collector (Figure 7biii)137.
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17 Fig. 7 – (a) schematic illustration of rotary jet spinning setup; (b) schematic
18
19 representation of fiber mechanism formation via RJS comprising three main stages: (i)
20
21
22
jet-initiation, (ii) jet-extension, and (iii) solvent evaporation. Panel (a) reprinted with
23
24 permission from ref (136). Copyright 2015 American Chemical Society. Panel (b)
25
26 reprinted with permission from ref (137). Copyright 2010 American Chemical Society.
27
28
29
30
31 The diameter and morphology of centrifugal spun fibers can be adjusted by
32
33 properly controlling the fluid viscoelasticity, rotation speed, diameter, length, and
34
35 structure of the nozzle as well as the distance between the nozzle and the collector. As
36
37
38 in electrospinning, centrifugal spinning has a wide versatility in terms of polymers and
39
40 solvents. In contrast to electrospinning, centrifugal spinning does not require a high
41
42 voltage supply and consists in a low cost process with high potential for industrial
43
44
45
scaling136,137,139. In this regard, FibeRio® company developed a centrifugal spinning
46
47 industrial equipment called FibeRio’s Cyclone™ Fibre Engine FX able to spin
48
49 nanofibers with diameters as low as 500 nm and with productivity up to 12 kg/h with
50
51 line speeds superior to 200 m/min, which is higher than electrospinning-based industrial
52
53
54 setups39. One another important advantage of centrifugal spinning is the easy
55
56 preparation of aligned nanofibers, which are potentially useful in tissue engineering
57
58 applications such as musculoskeletal and neural engineering140,141. Although centrifugal
59
60
spinning and SBS are able to achieve superior fiber productivity, the preparation of
24
1
2 nanofibers with diversified morphologies such as core-sheath or patterned nanofibers by
3
4 using these techniques remains yet as challenge.
5
6
7
8
9 2.4. Microfluidic spinning
10
11 Over the last decade, microfluidic spinning has emerged as a simple and cost-
12
13
14 effective strategy to produce fibers at micro- and nano-scale exhibiting diverse
15
16 structures and compositions31–33. In this method, two different fluids are simultaneously
17
18 injected within microscale channels through separate input ports forming a 3D coaxial
19
20
flow at the channel intersection, as illustrated in Fig 8a, where the core fluid consists in
21
22
23 polymer precursor solution and the sheath fluid acts as a lubricant to facilitate fiber
24
25 extrusion and in some cases also as a crosslinking solution33. Fiber formation occurs via
26
27 solidification of the core fluid by using strategies such as photopolymerization,
28
29
30 chemical or ionic crosslinking reactions, nonsolvent-induced phase separation, and
31
32 solvent evaporation32,142, as shown in Fig. 8b. By adjusting the design and dimensions
33
34 of the microchannel as well as the composition, viscosity, surface tension, and flow
35
36
37 rates of fluids, micro/nanoscale fibers with controllable shapes including cylindrical,
38
39 flat, core-shell, hollow, Janus, triple, and helical can be prepared31,143,144.
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Fig. 8 – (a) Schematic illustration of a microfluidic spinning chip setup; (b) schematic
26
27 representation of fiber solidification methods via (i) photopolymerization; (ii) ionic
28
29 crosslinking; (iii) chemical crosslinking and (iv) solvent exchange. Panel (a) reprinted
30
31
with permission from ref (145). Copyright 2019 Elsevier. Panel (b) adapted with
32
33
34 permission from ref (31). Copyright 2019 Elsevier.
35
36
37
38 Diverse biocompatible and biodegradable synthetic polymers including
39
40
41 Poly(ethylene glycol) diacrylate (PEG-DA)146, 4-hydroxybutyl acrylate (4-HBA)147,
42
43 polylactide (PLA)148, polyvinyl alcohol (PVA)149, polycaprolactone (PCL)150, and
44
45 poly(N-isopropyl acrylamide) (poly(NIPAAM))151 have been successfully spun by
46
47
48
using microfluidic spinning. In addition, biomacromolecules such as alginate152,
49
50 collagen153, silk fibroin154,155, and chitosan156 have been widely spun either alone or in
51
52 combination with synthetic or natural polymers to fabricate micro/nanofibrous.
53
54
Depending on the fluid composition, an appropriate solidification method should
55
56
57 be selected during the fiber formation process. For example, the preparation of
58
59 alginate-based fibers generally involves the use of sheath fluid solutions that acts as a
60
lubricant and ionic crosslinking solution (CaCl2) to promote fiber solidification via
26
1
2 gelation of the polymer157. In some cases, the fiber solidification method relies on the
3
4 Schiff base reaction156, nucleophilic crosslinking reactions158 or diffusion-based solvent
5
6
7 exchange150 between the core and sheath fluid. Another commonly used fiber
8
9 solidification approach is the photopolymerization, where the core fluid is composed by
10
11 photopolymerizable monomers incorporated with a photoinitiator, while the sheath fluid
12
13
14 is a lubricant solution, and the fiber formation occurs via UV light irradiation onto the
15
16 flow stream146,151. Despite UV-curing consists of a simple, fast, and stable solidification
17
18 method, its use on tissue engineering can be hindered due to the potential damage of
19
20
cells by UV radiation31.
21
22
23 The precise and easy spatiotemporal control of the composition, size and
24
25 morphology of the fibers is one of the greatest advantages of microfluidic spinning over
26
27 conventional spinning methods32. In addition, the use of non-harmful solvents, the non-
28
29
30 exposition of fluids to high voltage, and temperature allow the incorporation of sensitive
31
32 materials such as enzymes, genes, growth factors, and cells into the fibers without loss
33
34 of their activity, which is highly useful for the design of materials for regenerative
35
36
37 medicine and tissue engineering applications33. Another important feature of this
38
39 spinning technique is the possibility to construct 3D fibrous structures by reeling,
40
41 weaving, and direct writing, which is highly useful in the design of scaffolds143,159.
42
43
Although microfluid spinning exhibits several advantages, the scalability of this
44
45
46 technique still poses a significant challenge due to the difficulties in designing
47
48 nanoscale microfluidic chips and injecting fluids into the channels33. Besides, the
49
50 relatively short time required to the fiber solidification is a great challenge and efforts
51
52
53 should be devoted to development of solidification strategies regarding applications in
54
55 biomedical field31. Moreover, to date, the number of materials for biomedical
56
57 applications that can be spun by using microfluidic spinning is quite lower compared to
58
59
60 those employed in conventional spinning methods such as electrospinning31–33. Despite
27
1
2 this s microfluidic spinning is still in its infancy when compared to other spinning
3
4 methods, some companies (e.g. MicroFIT160; Micronit161) have been commercializing
5
6
7 microfluidic devices for spinning, which is highly beneficial for the practical
8
9 applications of microfluidic-spun fibers. Aiming to facilitate the comparison between
10
11 each spinning technique described in this review, Table 2 lists the major features of
12
13
14 each method.
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
28
1
2 Table 2. Comparison of fiber spinning processes and their advantages and disadvantages.
3
4 Spinning Technique
5 Feature Solution Blow Centrifugal
Electrospinning Microfluidic spinning
6 spinning spinning
7 Micro-scale fluid
Driving force Electrostatic Pressurized gas Centrifugal
8 dynamics
9 Process parameters: Process parameters: - Process - Process
10 applied voltage (5 - 35 gas pressure (1 - 80 parameters: parameters:
11 kV); working distance (5 psi); working distance rotation speed microchanel design,
12 - 20 cm); feeding rate (10 - 50 cm) and (2,000 - 16,000 feeding rate,
13 (0.01 - 1 mL min-1), protrusion distance; rpm), working solidification method.
14 nozzle structure. nozzle structure; distance (10 - 30 - Solution
15 Solution/melted feeding rate (0.02 - 1 cm); nozzle parameters:
16 polymer parameters: mL min-1). diameter. Composition,
17 composition, Solution parameter: Solution/melted viscosity, and surface
18 conductivity, viscosity, composition,viscosity, polymer tension.
19 Parameters solvent volatilility solvent volatilility, parameters:
20 (*solution surface tension. composition,
21 electrospinning), surface Enviromental viscosity, surface
22 tension. parameters: Relative tension; solvent
23 Enviromental humidity and volatilility
24 parameters: Relative temperature (*solutions)
25 humidity and Enviromental
26 temperature parameters:
27 humidity and
28 temperature
29
Typical Fiber 50 nm - 500 µm 100 nm - 3 µm 50 nm - 3 µm 400 nm - 500 µm
30
diameter
31
range
32
Diverse
33 Diverse Diverse Diverse
(e.g. Patterned,
34 (e.g. Patterned, Random, (e.g. Random, (e.g. Random,
Random, aligned,
35 Morphology aligned, janus, core- aligned, core-shell, aligned, core-
janus, core-shell,
36 shell, hollow ) hollow) shell, hollow)
hollow)
37
38
39 - Spatiotemporal
- Suitable for diverse
40 control of the
polymers; - Suitable for diverse - Suitable for
41 composition, size and
- Sub-micron diameters polymers; diverse polymers;
42 morphology of the
easily attained - Low cost; - Low cost;
43 fibers
Advantages - Allows Portability; - Simplicity; - Simplicity;
44 - Mild process
- Production of 3D - Portability; - Scale-up is
45 conditions.
fibrous structure. - Scale-up is possible possible
46 -Production of 3D
- Scale-up is possible
47 fibrous structure.
48
49 - Solvent removal
50 required;
- Require high voltage;
51 - Polymers
- Solvent removal - Time consuming;
52 - Solvent removal require some
required (*solution - Poor mechanical
53 required; thermal stability
electrospinning); properties of fibers;
54 Disadvantages - Low reproducibility (*melt spinning)
- Polymers require some - Scale-up is still
55 of the fiber diameter; - Low
thermal stability (*melt a challenge
56 reproducibility of
electrospinning)
57 the fiber
58 diameter.
59
60
29
1
2 3. Biomedical applications of polymer fibers
3
4 3.1. Drug delivery
5
6
7 Controlled delivery of therapeutic agents to specific target in a sustained manner
8
9 has been considered a highly promising approach for overcoming the limitations of the
10
11 systemic administration, since that approach improves the therapeutic index and reduces
12
13
14 the adverse side effects162. The efficiency of a drug delivery system greatly depends on
15
16 the choice of drug carrier, which ideally should possess biocompatibility, high drug
17
18 loading efficiency and effective cellular uptake as well as be able to maintain drug
19
20
concentrations in an appropriate therapeutic window163. Recently, polymeric micro- and
21
22
23 nanofibers have gained widespread interest for such platforms due to their great
24
25 versatility in terms of material choice, easy preparation and drug encapsulation as well
26
27 as desirable drug release profile3,164–167. Indeed, a variety of therapeutic agents including
28
29
30 antibiotics168, anticancer drugs169, RNA170 and bioactive proteins171 have been
31
32 successfully incorporated in polymeric nanofibers of diverse architectures that afford
33
34 different drug release profiles including zero-order release172, sequential release173,
35
36
37 spatiotemporal release174, biphasic release175 and stimuli-triggered release176. The
38
39 modulation of drug release kinetic on polymeric nanofibers have been achieved by
40
41 properly choosing the spinning preparation method and material composition.
42
43
Among spinning techniques that have been used for the fabrication of
44
45
46 polymeric micro- and nanofibers, solution electrospinning has been the most studied
47
48 method in the field of drug delivery3,164,177. The reasons for this great interest include
49
50 the versatility in terms of material composition combined to the possibility of creating
51
52
53 fibers with different morphologies and structures, such as core-sheath47 and janus178,
54
55 which allows the control over the drug release kinetics. Core-sheath structured fibers is
56
57 one the most promising platforms for controlled drug delivery, since the release of
58
59
60 incorporated drug can be modulated by varying the composition of inner and outer
30
1
2 layers, besides providing a more sustained release of the encapsulated agents compared
3
4 to their monolithic counterparts, due to the existence of outer barrier layer86,93.
5
6
7 Additionally, core-sheath nanofiber offers the opportunity to one-step co-incorporation
8
9 of multiple therapeutic agents with different solubility characteristics179,180.
10
11 Although solution blow spinning and centrifugal spinning methods have been
12
13
14 considered promising techniques in the development of drug delivery platforms, there
15
16 are some bottle necks related to the processing parameters control and fabrication of
17
18 customized nozzles to be achieved. Microfluidic spinning holds great potential for the
19
20
design of sustained delivery systems of sensitive active compounds31,32,157,181,182.
21
22
23 However, its use is still in infancy as compared to other spinning techniques. Singh et
24
25 al.121 developed core-sheath nanofibers by using a modified solution blow spinning
26
27 apparatus (Figure 9a). A mixture of poly-D,L-lactic acid (PDLLA) and model proteins
28
29
30 composed the nanofibers core, while the sheath was made of polycaprolactone (PCL)-
31
32 polyethylene oxide (PEO) blend. The fibers exhibited a defined core-sheath structure
33
34 (Figure 9b, c), sustained release of bovine serum albumin (BSA) (Figure 8 d) and basic
35
36
37 fibroblast growth factor (bFGF) (Figure 8 e) over 14 days. In vitro tests were carried out
38
39 using human bone marrow-derived mesenchymal stem cells and confirmed that the
40
41 core-sheath nanofiber mat demonstrated cytocompatibility as well as stimulate their
42
43
proliferation and differentiation. The authors also highlighted the feasibility of the
44
45
46 developed system for the direct deposition of core-sheath fibers scaffold containing
47
48 bioactive molecules at the patient bedside, as illustrated in Figure 8 a.
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50
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53
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60
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48 Fig. 9 – Schematic illustration of co-axial solution blow spinning (a); SEM (b) and
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51
TEM (c) images of core–shell nanofibers; release profile of BSA protein loaded in the
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53 variable diameters of core in core–shell nanofibers (d); bFGF release profile from the
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55 core–shell nanofiber mat, where the right Y axis represents bFGF release and left Y axis
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57
corresponds to the cumulative release of bFGF over the period of 14 days of the cell
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32
1
2 culture (e). Reprinted with permission from ref (121). Copyright 2018 American Chemical
3
4 Society.
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7
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9 Mamidi et al.29 prepared pH-responsive polycaprolactone
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11 (PCL)/mercaptophenyl methacrylate functionalized carbon nano-onions (f-CNOs)
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13 composite nanofibers by centrifugal spinning to deliver doxorubicin (DOX). The
14
15
16 authors found that the π-π stacking interactions between DOX and f-CNOs have led to
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18 long-term sustained release of DOX over 15 days under simulated physiological
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20 conditions. Additionally, in vitro tests demonstrated the composite nanofiber not to be
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23 cytotoxic toward human fibroblast cells, indicating its potential for biomedical
24
25 application.
26
27 As previously mentioned, the composition of electrospun fibers plays a key
28
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role in regulating the drug release kinetics. A great variety of synthetic and natural
31
32 polymers as well as polymer blends have widely been explored in the development of
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34 nanofiber-based drug delivery systems. Currently, stimuli-responsive polymer-based
35
36 nanofiber delivery systems has attracted significant attention in the field of drug
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39 delivery. These systems have been designed in order to trigger drug release in a
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41 spatiotemporal manner through physical and/or chemical stimuli. These stimuli can be
42
43 endogenous, including pH variation, redox gradient, enzymes-sensitive, or exogenous
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46 such as light, magnetic field (MF), ultrasound, electric field, temperature, among
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48 others169,176,183–186. Particularly, stimuli-responsive polymer-based nanofiber delivery
49
50 systems sensitive to noninvasive exogenous stimuli has gained wide interest. Such
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53
strategy afford a targeted and reliable tuning of drug release by an operator, resulting in
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55 regulation of the dosage of the therapeutic agent according to specific needs of patient,
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57 improving therapeutic efficacy and reducing side effects169,184,187,188.
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33
1
2 Altinbasak et al.168 developed a photothermally activated platform based on
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4 reduced graphene oxide (rGO)-embedded poly(acrylic acid) (PAA) electrospun
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7 nanofiber mats for on-demand release of antibiotic ampicillin upon irradiation in the
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9 near-infrared. The authors found the antibiotic-loaded nanofiber mats suitable for
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11 treating superficial skin infections. Puiggalí-Jou et al.189 successfully prepared poly(ε-
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14 caprolactone) (PCL) microfibers loaded with poly(3,4-ethylenedioxythiophene)
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16 nanoparticles (PEDOT NPs) and curcumin using solution electrospinning. The PEDOT
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18 NPs behaved as electroactuators upon application of well-defined potential pulses,
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while the release of curcumin from the PCL microfibers was boosted by an external
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23 electrical stimulus. Ballesteros et al recently reported an approach to produce a smart
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25 nanomaterial based on polycaprolactone (PCL) electrospun nanofibers which were
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27 surface-modified by a photoresponsive nanogel containing silver nanoparticles
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30 (AgNps)186. Upon light irradiation at a specific wavelength, (405 nm) AgNps were
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32 slowly released from the nanogel, yielding the system a strong antibacterial activity
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34 against S. aureus and E. coli, indicating the potential of this smart nanomaterial for
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37 application in light-responsive wound dressing. Perera et al.188 utilized centrifugal
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39 spinning to produce poly(vinyl alcohol) ultrathin fibers incorporated with citric acid-
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41 coated Fe3O4 magnetic nanoparticles (MNPs) to be used as an on-demand drug carrier
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43
of acetaminophen. In this study, the drug release was triggered by a moving external
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46 magnetic field, which demonstrated the developed platform as an efficient remote-
47
48 controlled method for drug delivery.
49
50 An electrically responsive drug release core–shell hybrid microfibers hydrogel
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53 system was fabricated via microfluidic spinning followed by dip-coating, where the
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55 core was composed of spun bacterial cellulose loaded with diclofenac sodium while the
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57 shell layer consisted of a dip-coated poly(3,4-ethylenedioxythiophene) (PEDOT)
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60 conductive polymer190. The release behavior of the diclofenac sodium from microfibers
34
1
2 was properly modulated by the application of external electrical stimulation. In addition,
3
4 the hybrid fiber platform demonstrated biocompatibility and electroactivity, therefore
5
6
7 indicating its potential use for nerve regeneration.
8
9 Despite the considerable advances in on-demand externally triggered
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11 nanofiber-based drug release systems, most studies have been limited to in vitro proofs-
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13
14 of-concept and clinical translation of such technologies remains yet to be further
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16 developed. The main challenges are associated with the chemical complexity, biological
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18 responses, and predictability of externally triggered release vehicles when used in vivo
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20
physiological environment. Additionally, the design of these drug delivery systems
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23 involves nondegradable materials, which may hinder their applications in biomedical
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25 field. In this perspective, further studies are still necessary to address these issues.
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27
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30 3.2. Regenerative medicine and tissue engineering
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32 Lesions in tissues or organs are one of the most frequent problems faced by
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34 medicine. Currently therapeutic options (allograft, xenograft, autograft and implant
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37 biomedical devices) have increased the quality of life of society191,192 However, these
38
39 approaches are usually associated with several limitations, including feasibility of
40
41 donors, infection, poor integration with the host and the potential rejection of the
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43
implant. Thus, regenerative medicine and tissue engineering has been developed in
44
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46 order to overcome these limitations and to find revolutionary therapies able to heal the
47
48 tissue damage through the repair or creation of tissues or organs aiming at the
49
50 restoration of tissue functions193–195.
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53 Regenerative medicine and tissue engineering involve concepts of diverse areas
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55 including engineering, chemistry, biology and materials science. The development of
56
57 functional tissues helpful in repairing damaged organs is a complex process and
58
59
60 requires scaffolding materials that resemble the cellular microenvironment and possess
35
1
2 physical and biochemical properties to support cell attachment, migration, growth and
3
4 tissue maturation194,196. Over the last years, fibrous scaffolds prepared by
5
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7 electrospinning, solution blow spinning and centrifugal spinning have become a
8
9 promising multimodal platform for tissue engineering and regenerative medicine. These
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11 constructs closely resemble the architecture of the extracellular matrices (ECM) of the
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14 native tissues and exhibits high porosity and surface-area-to-volume ratios.
15
16 Additionally, they can be tuned in terms of material composition, surface
17
18 characteristics, mechanical properties, degradability as well as distribution pattern,
19
20
which increases the potential of a close interaction with the cells to trigger essential
21
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23 physiological processes, leading to cell assembly into functioning units34,197,198.
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25 In the past two decades, a large number of articles have focused on evaluating
26
27 the potential of fibrous scaffold in regenerative medicine and tissue engineering for
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29
30 various applications including bone, cartilage, neural and cardiac tissue regeneration,
31
32 comprehensively reviewed in this section.
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34
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37 3.2.1. Bone regeneration
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39 Bone is a highly dynamic tissue formed by organic (mainly collagen nanofibers)
40
41 and inorganic (nanocrystalline hydroxyapatite) components disposed in a hierarchical
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43
structure ranging from the nano- to macroscale199. When injured, bone tissues have self-
44
45
46 healing capability depending on the size and extent of the damage, but in case of severe
47
48 loss, therapeutic approaches are required to repair or regenerate the damaged tissue.
49
50 Despite recent advances in tissue engineering and regenerative medicine, reconstruction
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53 of critical-sized bone defects is still challenging200.
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55 Advanced bone healing approaches included a wide range of biomaterials that
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57 mainly mimic the composition, structure, and properties of bone extracellular matrix
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60 with osteogenic activity. Nanofibrous composites formed by synthetic and natural
36
1
2 polymers combined with mineral components such as hydroxyapatite201, calcium
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4 phosphate202, β-tricalcium phosphate203 and glass ceramics204 have been considered as a
5
6
7 promising approach toward functional bone scaffolds. Despite composites scaffolds
8
9 show some intrinsic osteogenic capacities, they often are not sufficient to promote
10
11 complete bone healing. Additionally, integration of implanted scaffolds with the host
12
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14 tissue is still a great challenge in bone regeneration because they can elicit exacerbated
15
16 immune responses205. In order to modulate inflammatory responses and stimulate bone
17
18 regeneration, many studies have focused on the development of spun fiber-based
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20
scaffolds to sustained release of bioactive molecules including proteins206, peptides207,
21
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23 growth factors208,209 and drugs210,211.
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25 Although conventional spinning techniques, particularly electrospinning, have
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27 been widely explored for producing biomimetic scaffolds in bone tissue engineering,
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30 the inherent small pore diameter of spun two-dimensional (2D) constructs are smaller
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32 than that recommended for the cellular growth into and bone vascularization. In recent
33
34 years, strategies including increasing spinning time, post-processing of the as-spun
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37 fibers, the use of 3D collectors or liquid collectors as well as by combining
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39 electrospinning and additive manufacturing principles have been adopted for developing
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41 3D nanofibrous structures212. For example, Medeiros et al.213 combined solution blow
42
43
spinning with thermally induced phase separation (TIPS) to produce highly porous 3D
44
45
46 structures, as displayed in Fig. 10. This combined SBS-TIPS technique has proven to
47
48 have a great potential in regenerative medicine mimicking the extracellular matrix.
49
50 Addition of Nano-bioactive glass (nBG) provided scaffolds with therapeutic ion
51
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53 releasing properties, pointing its potential applications in non-load bearing bone tissue
54
55 engineering.
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33 Fig. 10. Schematic of the cryogenic SBS process with in situ spraying of ice
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35 microspheres, and subsequent compression into scaffolds with interconnected pores
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37 (A); cotton-wool-like structure produced without compression (B); scaffolds with large
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39
40
interconnected pores (C) (scaffold diameter is 16 mm, sectioned and opened, right
41
42 image), and scaffold produced using small ice microspheres (D). Reprinted with
43
44 permission from ref (213). Copyright American Chemical Society (2016)
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46
Melt electrospinning writing (MEW) combines electrospinning and additive
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49 manufacturing principles and have been emerged as a promising technique to design
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51 ultrathin fibrous scaffolds with controlled architecture. By using a computer-controlled
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53 collector, biocompatible and biodegradable electrospun polymer-based fibers can be
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56 deposited accurately, which allows the control over pore size and interconnectivity of
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58 the resulting scaffolds, whose characteristics are highly desirable for bone tissue
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60 engineering application214. For example, Abdal-hay et al.215 prepared polycaprolactone
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2 (m-PCL)-hydroxyapatite (HAp) composite 3D fibrous structures with high porosity
3
4 (96–98%) and fully interconnected pore architectures by using MEW. Results revealed
5
6
7 that designed structures provided a favorable platform for the infiltration and growth of
8
9 human osteoblasts, indicating great potential for use in mineralized tissue
10
11 reconstruction/regeneration applications.
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14 Despite great advances, the preparation of 3D fibrous scaffolds still presents a
15
16 major challenge because the developed approaches are time-consuming and the process
17
18 scalability is limited. In addition, the mechanical properties of some 3D fibrous
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20
structures such as those prepared by SBS-TIPS are not appropriate to withstand stress
21
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23 and strain in bone tissue, and thus, more efforts must be made to overcome these
24
25 bottlenecks.
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27
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30 3.2.2. Cartilage regeneration
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32 Cartilage regeneration remains challenging due to the low intrinsic regenerative
33
34 capacity of this tissue216. Currently, numerous clinical treatments for cartilage
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36
37 regeneration including microfracture and tissue transplants (decellularized structures,
38
39 xenograft, allograft, autograft) have been investigated217,218. Nonetheless, these
40
41 treatments have a limited capacity to promote full cartilage tissue repair. Tissue
42
43
engineering approach by utilizing spun polymeric fibrous scaffolds have appeared as a
44
45
46 promising strategy to regenerate complex tissues like cartilage21,219.
47
48 Among several techniques used to fabricate fiber-based scaffolds for cartilage
49
50 regeneration, electrospinning is the most widely used strategy, although SBS and
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52
53 centrifugal spinning have gained more importance for practical application . An
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55 interesting feature of these spining techniques relies on their versatility to combine
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57 various types of materials to prepare both aligned and random oriented fibers with
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59
60 mechanical properties similar to that of the native tissue. For instance, skeletally mature
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1
2 articular cartilage possess a hierarchical structure formed by aligned and random fibril
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4 zones220. Christakiran et al.221 prepared a bilayer cartilage scaffold by electrospinning
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7 poly(vinyl alcohol)/Silk random nanofibers onto previously electrospun poly(vinyl
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9 butyrate)PVB-bioactive glass aligned nanofibers. The biphasic mats exhibited spatial
10
11 confinement for the growth and maturation of both osteoblasts and chondrocytes, thus
12
13
14 indicating great potential for osteochondral tissue repair.
15
16 A number of bioactive molecules, including drugs, corticosteroids, and growth
17
18 factors, have been incorporated into fiber scaffolds to control inflammation responses or
19
20
to promote new cartilage tissue formation222–224. In a related work225, growth factor
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23 TGF-β3-containing polymer blend fibers of poly-ε-caprolactone (PCL) and polylactide-
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25 co-glycolide (PLGA) were prepared by electrospinning. The nanofibers meshes
26
27 provided sustained growth factor delivery and demonstrated potential for guiding
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30 synovium-derived stem cell (SDSC)-mediated fibrocartilage regeneration.
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32 Despite the promising prospects provided by spun-fibrous scaffolds, challenges
33
34 and obstacles still exist due to the current limitation of spinning techniques to prepare
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37 3D structures with appropriate porosity and pore size for cartilage tissue engineering. In
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39 this regard, Chen et al.226 fabricated a 3D hybrid construct by freeze-drying suspensions
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41 of electrospun gelatin/PLA nanofibers cross-linked with hyaluronic acid (HA). The
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43
resulting 3D scaffold exhibited porous structure (Fig. 11a), water superabsorbent
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46 capacity (1300 %) (Figure 11 b,c), and elastic property in the wet state. The
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48 chondrocyte cell viability was evaluated in vitro by live and dead assay for 3 days and 7
49
50 days and the results revealed that the scaffold was noncytotoxic (Fig. 11 d and 11 e),
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52
53 and allowed the adhesion and proliferation of cells (Fig. 11 f and 11 g). An in vivo study
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55 indicated the developed scaffold was able to enhance cartilage defects repair in rabbit.
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57 After 12 weeks of implantation, the defect in non-treated group was not filled overall
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60 (Fig. 11h and 11i) and covered with loose fibrous tissue (Fig. 11j), while the treated
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1
2 defect was filled with uniform cartilage-like tissue with a flat surface and the defect
3
4 (Fig. 11k and 11l) covered with regenerated cartilage tissue and chondrocytes (Fig.
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7 11m).
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Fig. 11. SEM image of scaffold (a); photographs showing the scaffold shape before (b)
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46 and after water absorption (c); fluorescence micrographs of chondrocytes seeded on
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48 scaffold for 3 days (d) and 7 days (e), where live and dead cells are identified by green
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50
and red color, respectively; SEM images of chondrocytes cultured for 3 days (f) and 7
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53 days (g); photographs of the cartilage joints non-treated (h) and treated (k) at 12 weeks
54
55 after surgery; histological analysis of defect site, which are indicated by arrows and
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57 dotted lines (OC: original cartilage tissue; RC: repaired cartilage tissue); stained with
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60 Safranin O-fast green (non-treated (i) and treated (l)) and hematoxylin and eosin (non-
41
1
2 treated (j) and treated (m)). Reprinted with permission from ref (226). Copyright 2016
3
4 American Chemical Society.
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6
7
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9 Significant improvements in cartilage regeneration can be achieved through the
10
11 development of stimuli-responsive hierarchical 3D fibrous scaffolds to a spatiotemporal
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14 delivery of bioactive molecules aiming to modulate cell behavior. These scaffolds also
15
16 should present an appropriate pore size to allow infiltration and migration of cells
17
18 involved in the tissue regeneration process.
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23 3.2.3. Peripheral nerve regeneration
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25 Peripheral nerve injuries are clinically common and frequently lead to
26
27 debilitating neurological deficits and neuropathic pain227. Limited regenerative capacity
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29
30 of the nervous system makes treating nerve injuries a challenging task. Current clinical
31
32 golden standard for repairing nerve injuries involves the use of nerve autograft, but this
33
34 therapeutic strategy presents some drawbacks including low availability of donor nerves
35
36
37 and donor-site morbidities228. As promising alternatives for nerve autograft, polymeric
38
39 nanofibrous nerve conduits integrated with biochemical and physical cues have been
40
41 proposed229,230.
42
43
Aligned spun polymeric fibers scaffolds incorporated with bioactive substances
44
45
46 including cell growth factors, peptides and extracellular matrix (ECM) components
47
48 have demonstrated great potential for regeneration of injured nerve229–231. As strategies
49
50 to prepare 3D tubular nerve conduits, polymeric nanofibers can be rolled, deposited on
51
52
53 cylindrical supports or embedded in hydrogel matrices. In a related work231, bone
54
55 marrow stromal cells (BMSCs) were harvested and seeded on a never growth factor
56
57 (NGF)-loaded rolled poly(ɛ-caprolactone) (PCL)/BSA/PEG-diamine nanofibrous
58
59
60 scaffold and cultured with a rotary cell culture system before implantation in rat sciatic
42
1
2 nerve defect model, as show in Fig. 12. In vitro studies showed that bioactivity of
3
4 growth factor was preserved over 28 days. Furthermore, an in vivo study revealed that
5
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7 the prepared nanofibrous scaffolds allows functional peripheral nerve regeneration.
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44 Fig. 12. Schematic representation of nanofibrous nerve conduit preparation and
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46 aplication on in vitro and in vivo studies (a); SEM images of the cross-section (b) as
47
48 well as outer (c) and inner (d) surface of the nanofibrous scaffold. Reprinted with
49
50
51
permission from ref (231) . Copyright 2020 American Chemical Society.
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55 In a recent study232, the potential for neural stem cell (NSCs) proliferation was
56
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examined on aligned centrifugal-spun polycaprolactone (PCL)/Bovin serum albumin
58
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60 (BSA) nanofibers and electrospun PCL/BSA nanofibers with similar diameter and
43
1
2 alignment. Through in vitro studies, it was found that NSCs grew on the aligned
3
4 centrifugal-spun PCL fibers exhibited a bipolar elongation along the direction of the
5
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7 fiber, while NSCs cultured on the aligned electrospun PCL fibers expressed a multipolar
8
9 elongation, thus indicating the higher potential of aligned centrifugal-spun nanofibrous
10
11 scaffolds for nerve injury repair.
12
13
14 Recent trends on nanofibers scaffolds design have focused on the use of
15
16 conductive materials to promote the nerve regeneration through electrical stimulation
17
18 and restore the proper signal-transmission functionality of healed nerves233. Conductive
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polymers (e.g. polypyrrole (Ppy)234, polyaniline (PANI)235), carbon nanomaterials (e.g.
21
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23 reduced graphene236, carbon nanotubes237) and metal nanomaterials (e.g. gold
24
25 nanoparticles192) have been widely investigated for nerve tissue regeneration. Polymeric
26
27 composites and blends nanofibrous scaffolds based on conductive materials and
28
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30 biocompatible and biodegradable polymers (natural or synthetic) have been prepared
31
32 aiming to produce materials with superior electrical conductivity and mechanical
33
34 properties for nerve tissue regeneration application234–237. Another strategy that has been
35
36
37 adopted to produce conductive nerve conduits consists of the post-spinning
38
39 functionalization of nanofibrous scaffolds surface by using carbon-based nanomaterials
40
41 such as reduced graphene oxide and carbon. For example, Wang et al.236 developed
42
43
conductive nanofiber scaffolds via electrospinning PCL/silk fibroin and subsequent
44
45
46 layer deposition of reduced graphene oxide (RGO) onto nanofibrous scaffolds surface
47
48 through in situ redox reaction of the graphene oxide (GO). The conductive nanofibrous
49
50 scaffold was biocompatible and showed the capability to modulate cell functions in
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53 vitro as well as to promote nerve regeneration in vivo into rat sciatic nerve defects
54
55 model at a similar level to the gold standard autograft.
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57
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59
60 3.2.4 Cardiovascular regeneration
44
1
2 Cardiovascular diseases are the leading cause of death around the world and
3
4 their incidence has been increasing drastically in the last decades238. Among the
5
6
7 conditions that can results in heart failure, myocardial infarction, coronary artery
8
9 disease, valve disorders, high blood pressure, and cardiomyopathy are the most
10
11 prevalent239. Over the past decade, cardiac tissue engineering approaches involving 3D
12
13
14 fiber scaffolds have become a potential strategy for regeneration of damaged heart
15
16 tissue240.
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18 To date, numerous spinning techniques including electrospinning241, and
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solution blow spinning242, centrifugal spinning243, and microfluidic spinning32 have
21
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23 been applied in the construction of scaffolds for regeneration of myocardial, valve, and
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25 vascular tissues. Considerable attention has been paid to those spinning techniques that
26
27 allow the controlled deposition of fibers to create hierarchical structures that resemble
28
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30 the native extracellular matrix and provide appropriate mechanical, biochemical, and
31
32 topographical cues to stimulate tissue regeneration. In this regard, a microfiber PCL-
33
34 based 3D scaffold with controlled hexagonal microstructures was prepared via melt
35
36
37 electrowriting (MEW) (Figure 13 a, b and c) to be employed in myocardial
38
39 regeneration73. Aiming to create functional cardiac patches, human induced pluripotent
40
41 stem cell-derived cardiomyocytes (iPSC˗CM) were previously encapsulated in a
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43
collagen-based hydrogel and seeded on the prepared scaffolds (Figure 13 d, e, f). In
44
45
46 vitro studies revealed that engineered structures promoted cell alignment, sarcomere
47
48 content and organization as well as an increase in cardiac maturation-related marker
49
50 expression. Furthermore, the designed scaffold exhibited remarkable biaxial
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53 deformation and supported high tensile strains when implanted via a minimally invasive
54
55 approach on a contracting porcine heart (Figure 13 g) besides to preserve iPSC˗CM
56
57 viability. Despite in vivo data are only a proof-of-principle in a porcine model, this
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60 study indicates the great potential of developed fibrous scaffolds for cardiac repair.
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Fig. 13. Schematic illustration of melt electrowriting (a); photographs of the fabricated
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39 scaffolds acquired from top and lateral perspective (b); Stereoscopic (left) and scanning
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41 electron microscopy (SEM) (right) image of a hexagonal cell with a side length of 400
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44 µm and 20 stacked layers (c); In vitro culture of iPSC‐CM on nanofibrous scaffold (d);
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47 In vitro injectability test of scaffold (e) and its image after injection (f); implantation of
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49 cardiac patch through a minimally invasive procedure on beating porcine heart (g).
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51
Reprinted with permission from ref (73). Copyright 2018 Wiley.
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53
54 One promising approach that has been widely explored is the preparation of
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56 polymeric 3D structures formed by aligned conductive fibers loaded with bioactive
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58 molecules in order to mimic the structural organization of native myocardium tissue,
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60
facilitates the electrical signal transmission as well as to stimulate cell adhesion,
46
1
2 differentiation, and proliferation241. Feiner et al.244 developed a cardiac patch that
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4 integrates cardiac cells with flexible, freestanding electronics and a 3D nanofibrous
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7 electroactive scaffold. This complex structure enabled the on-demand delivery of
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9 lysozyme and the monitoring and control of the engineered-tissue function through
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11 remote external electrical stimulation, thus consisting of an innovative strategy that
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14 holds great potential for repair damaged myocardial tissue.
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16 Recently, MacQueen et al.245 fabricated a PCL/collagen nanofibrous ventricle-
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18 shaped scaffold via pull spinning, a modified centrifugal spinning method246. In this
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study, aligned fibers were deposited on a ventricle-shaped (ellipsoidal) mandrel with
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23 dimensions of a rat ventricle. It was found that designed scaffolds exhibited superior
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25 mechanical properties compared to the extracellular native myocardial tissue. The
26
27 authors also demonstrated through in vitro studies that the designed scaffolds promoted
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30 native-like anisotropic myocardial tissue genesis and presented chamber-level
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32 contractile function, thus showing great potential for engineering 3D cardiac tissues.
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34 These structures were also used in an instrumented bioreactor revealing a suitable
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37 strategy for in vitro cardiology studies.
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39 Polymeric nanofibrous scaffolds have been investigated as an alternative to the
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41 current bioprosthetic heart valve247. Capulli et al.243 developed an automated fabrication
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method of fibrous heart valve scaffolds via deposition of poly-4-
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46 hydroxybutyrate/gelatin centrifugal spun fibers onto a modified collector. The authors
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48 demonstrated the feasibility of the method to design in few minutes tunable scaffolds
49
50 replacements with fibrosal-like fiber alignment and global semilunar valve structure.
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53 Furthermore, the designed scaffolds were easily incorporated into stents and implanted
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55 via a minimally-invasively procedure through the transapical access to the pulmonary
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57 valve position in an ovine model. In vivo results revealed that nanofibrous scaffolds
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1
2 were biocompatible and functional, highlighting the potentiality of this approach to
3
4 translate to a human model.
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7 One of the key challenges associated with valve tissue engineering is the
8
9 development of a scaffold that mimic the leaflet, which comprises the primary
10
11 component of a heart valve and is formed by three layers with three different
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14 orientations: circumferential, random and radial, respectively. Jana and Lerman247
15
16 fabricated via solution electrospinning a trilayered PCL-based nanofibrous scaffold with
17
18 architecture that mimics the three layers of a native heart valve leaflet. The authors
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20
observed that such electrospun structures provided appropriate mechanical properties to
21
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23 support the growth and orientation of seeded porcine valvular interstitial cells (PVICs)
24
25 and their deposited collagen fibrils, allowing the in vitro formation of a viable tissue
26
27 construct after one month of cell incubation. Although further investigation is still
28
29
30 required, these scaffolds hold great potential for cardiac valve tissue engineering
31
32 applications.
33
34 3.2.5. Wound dressings
35
36
37 Skin is the largest organ of human body and protects the inner organs from the
38
39 external environment as well as actuates in metabolic, immunologic, thermoregulatory
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41 and sensory functions. Disruption of skin’s anatomic structure due to trauma, surgery,
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burn or chronic diseases frequently requires the use of wound dressing to promote
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46 regeneration of skin tissue in the damaged area248. A great variety of wound dressings in
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48 the form of hydrogels, films, sponges, and foams are currently available in the market.
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50 In recent years, micro/nanofibers nonwovens have emerged as promising class of
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53 wound dressings60,249. This renewed interest relies on the inherent high porosity and
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55 surface area to volume ratio exhibited by fiber matrices, which affords a proper control
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57 of moisture balance at the wound site and facilitates the gas permeation, transport of
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60 nutrient as well as the metabolic waste elimination. Additionally, these matrices mimic
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2 the native extracellular matrix (ECM) structure, thus serving as a template for
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4 attachment, growth, and migration for skin cells and consequently stimulate tissue
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7 regeneration10,250–252.
8
9 Numerous studies have reported the application of micro/nanofibrous scaffolds
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11 as skin substitutes and wound dressings10,250–252. Chantre et al.253 investigated the
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14 performance of centrifugal spun nanofiber scaffolds composed of hyaluronic acid (HA)
15
16 on cutaneous tissue repair. By centrifugal spinning the researchers were able to prepare
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18 highly porous HA scaffold in large scale. In vitro testing showed that the nanofiber
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20
scaffold afforded rapid and in-depth permeation of seeded dermal fibroblasts and
21
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23 demonstrated biocompatibility and bioactivity up to 7 days in culture. Further in vivo
24
25 studies were performed in an excisional splinted wound healing model in mice
26
27 demonstrating that nanofibrous scaffold accelerated granulation tissue formation,
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30 neovascularization, and reepithelialization, promoting faster wound closure and tissue
31
32 repair within the first week of treatment.
33
34 A fibrous wound dressing was prepared by using a microfluidic spinning device
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37 for creating a micrometer-scale core-sheath hydrogel fibers comprised of fibroblast
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39 growth factor (FGF-2)-loaded fibrous alginate as core material and a self-assembly
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41 peptide hydrogel loaded with ampicillin as a shell material182. The hydrogels fibers
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43
exhibited appropriate mechanical properties and a dual release profile of active
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46 components was achieved, where antibiotic was rapidly released from shell layers,
47
48 while the growth factor was released in a sustained manner from the core layer within 7
49
50 days. Moreover, the hydrogel showed remarkable in vitro antibacterial properties
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53 against both S. aureus and E. coli. as well as wound healing ability on in vivo infected
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55 wound model.
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57 Recently, the use of portable devices for in situ deposition of nanofibers directly
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60 onto wounds sites have been proposed to improve conformability of the matrices to the
49
1
2 wound site according to patient needs254. Portable devices based on solution
3
4 electrospinning principles are largely investigated while the solution blow spinning
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7 portable apparatus is still on the initial stage of development125,254,255. Liu et al.
8
9 examined the performance of a portable handheld electrospinning prototype device (Fig
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11 14 a). Electrospun zein/poly(ethylene oxide)(PEO) nanofibers incorporated with thyme
12
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14 essential oil (TEO) were deposited directly onto partial thickness wounds on mice
15
16 dermal tissue model (Fig. 14b). The authors highlighted the convenience of application
17
18 and demonstrated that deposited nanofiber dressing displayed appropriate gas-
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20
permeability, surface hydrophilicity, high conformability to the wound site as well as
21
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23 antimicrobial activity towards S. aureus and E. Coli (Figure 14 c). Animal based in
24
25 vivo testing were performed at days 3, 7, and 11 (Fig. 14 d) and revealed that developed
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27 electrospun nanofibers greatly improves the wound healing process within 11 days. The
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30 authors also concluded that in situ application of nanofibrous dressings have the
31
32 potential to reduce infection and cross-contamination rates.
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37 Fig. 14. (a) Image of a handheld electrospinning device (b) in situ electrospinning of
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39 fibrous dressing on wound site (c) antibacterial activity of fibrous dressing towards S.
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41 aureus and E. coli. and (d) wounds healing at 0, 3, 7, and 11 days after injury for no
42
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44 treated wound and after treatment with zein/PEO and zein/PEO/TEO nanofiber
45
46 dressings. Reprinted with permission from ref (255). Copyright 2020 American Chemical
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48 Society.
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50
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A new generation of interactive dressings capable of preventing undesired side
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53 effects such as infection and promoting on-demand therapeutic agents release has been
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55 pursued. Toward this end, Tamayol et al.256 incorporated thermo-responsive particles of
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PEGylated-chitosan loaded with antibiotics into poly(glycerol sebacate)-
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60 poly(caprolactone) (PGS-PCL) electrospun nanofibrous. A flexible heater was
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1
2 integrated into the nanofibrous platform to allow on-demand release of commercial
3
4 antibiotics including cefazolin and ceftriaxone. Delivery of drugs was electronically
5
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7 controlled by adjusting the applied voltage and consequently the scaffold temperature,
8
9 which increase resulted in a higher release rate. In vitro studies showed that the smart
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11 platform was biocompatible, biodegradable and effective against S. aureus and E. coli
12
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14 bacterial strains, thus indicating its potential as a wound dressing.
15
16 Despite promising outcomes exhibited by spun fiber wound dressings, there is
17
18 still much left to be done to translate these materials into clinical applications. In
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addition, studies should focus on the development of multifunctional fibrous wound
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23 dressings that simultaneously stimulates wound healing, avoids infection occurrence,
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25 allows to monitor of the wound healing status as well as provides drug delivery in a
26
27 controlled manner.
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30
31
32 3.3. Nanofibers-based structures for disease modeling and drug screening
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34 In recent years, 3D nanofibers structures have been applied as a scaffold to the
35
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37 development of in vitro models that mimics the physiological and pathological
38
39 microenvironment of tissues and organs for disease modeling, drug screening, and
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41 toxicity testing257,258. This approach offers an alternative for predicting the toxicity and
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43
efficacy of therapies without the necessity of in vivo animal models as well as holds the
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46 potential to reduces the time and cost of development of new drugs259–262
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48 To date, several studies have been devoted to the application of three-
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50 dimensional (3D) spun fibrous materials as scaffold for generating in vitro tumor
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53 models for breast cancer159,263,264, pancreatic cancer265,266, colorectal cancer267, prostate
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55 cancer268, glioblastoma269, bladder cancer270, and melanoma271. For example, Yang et
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57 al.272 proposed an innovative approach to developed a lung-on-a-chip structure that
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60 mimics the alveolar microenvironment by using poly(lactic-co-glycolic acid) (PLGA)
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1
2 electrospun nanofiber membrane as the chip substrate and cell scaffold. The authors
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4 performed cell culture and co-culture of human non-small cell lung cancer cells (A549)
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7 and human fetal lung fibroblasts (HFL1) on the chip, and evaluated the efficacy of the
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9 anti-tumor drug gefitinib. The study revealed that A549 cell drug resistance might be
10
11 attributed to the IGF-1 secreted by HFL1cells, which results in activation of the
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14 PI3K/Akt signal pathway after inhibition of the EGFR-related signal pathway by the
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16 drug, thus leading in decreased sensitivity of the tumor cells to chemotherapeutic drugs.
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18 In addition, the occurrence of tumor invasion was investigated by co-culturing A549,
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HFL1, and human umbilical vein endothelial cells (HUVECs), and the authors found
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23 that A549 cells could trigger endothelial cell apoptosis or death followed by the
24
25 occurrence of tumor invasion. Overall, the proposed lung-on-a-chip system presents
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27 great potential to be used in personalized treatment of lung tumors.
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30 In another study, a three-dimensional (3D) bone tumor model was engineered by
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32 culturing human mesenchymal stem cells (MSCs) in osteogenic conditions on
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34 electrospun poly(ε-caprolactone) (PCL) scaffolds followed by decellularization of these
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37 constructs, which were subsequently used to study Ewing’s sarcoma (ES) cells
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39 behavior273. The resistance of ES cells toward chemotherapeutic agents including
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41 doxorubicin, ridaforolimus, and dalotuzumab was also evaluated. Results revealed that
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the decellularizated constructs simulated in vivo-like tumor architecture and allows the
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46 investigation of the roles of microenvironmental elements in ES tumor growth, cancer
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48 cell morphology, and induction of resistant cell phenotypes. The proposed 3D
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50 constructs are potentially useful to culture patient biopsies, test for anticancer
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53 compounds and to create a personalized cancer treatment.
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55 Among the spinning techniques, microfluidic spinning has emerged as one of the
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57 most promising approaches to produce in vitro models, since the mild process
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60 conditions afford a simple and safety encapsulation of cells onto fibers as well as
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1
2 provides an appropriate environment in terms of structure and composition for cell
3
4 seeding and culture31. Furthermore, by integrating microfluidic spinning with 3D
5
6
7 printing system or textile technologies such as weaving, braiding, and embroidering
8
9 methods, highly controlled hierarchical 3D microsized cellular constructs can be
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11 prepared143,159. Recently, 3D graphene oxide (GO) microfibrous scaffolds with an
12
13
14 adjustable fiber length, fiber diameter (100 – 200 µm), and scaffold structure by
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16 integrating the microfluidic spinning technology with a programmable 3D printing
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18 system was proposed, as illustrated in Fig. 15159. By using hydrothermal reduction,
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20
reduced graphene oxide (rGO) structures were also prepared. Human neuroblastoma
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23 cell line (SH-SY5Y), a commonly used cell line for in vitro neurological studies such as
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25 neuronal differentiation, metabolism, neurotoxicity, and neuroprotection, were seeded
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27 and cultured onto 3D scaffolds. It was demonstrated an oriented proliferation of SH-
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30 SY5Y cells onto the graphene microfibrous scaffolds, therefore indicating that these 3D
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32 cellular constructs are potential for electroactive tissue regeneration and drug-screening
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34 applications.
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Fig 15 - Schematic illustration of the microfluidic spinning integrated to a 3D printing
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58 system for fabricating graphene electroactive microfibrous scaffolds. rGO microfibrous
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60 scaffolds were obtained via hydrothermal reduction. 3D cellular constructs were
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1
2 prepared by seeding and culturing human neuroblastoma cell line (SH-SY5Y) on the
3
4 scaffolds. Reprinted with permission from ref (159). Copyright 2020 American Chemical
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7 Society.
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9
10
11 Despite numerous improvements that have been achieved by using 3D spun
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14 fibrous materials as a scaffold in disease modeling and pharmaceutical research, the
15
16 validation of these in vitro models to ensure that the biological functions reproduced are
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18 representative of the native tissue remains a challenge257,259. Furthermore, the
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fabrication of 3D cellular constructs model involves multiple assembly steps, which
21
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23 poses scale-up concerns. Overall, the opportunities for generating 3D cellular constructs
24
25 using spun micro/nanofibers scaffolds are immense, and therefore the simplified and
26
27 translational strategies should be a target in the future for researchers.
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30
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32 3.4 Sensing and biosensing
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34 Early disease diagnosis and monitoring play a key role in the treatment of
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37 patients, where the development of effective and sensitive sensors for disease markers
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39 detection is a critical component in this process5. Over the past years, nanofibers have
40
41 attracted great interest in the construction of sensing platforms because of their large
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surface area-to-volume ratio, high porosity as well as versatility in terms of structure
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46 and composition24,274,275. In order to improve the performance of nanofiber-based
47
48 sensors, functionalization strategies have been adopted including the incorporation of
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50 carbon nanostructures and metal nanoparticles within nanofiber structures and chemical
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53 or physical surface modification4. The modification of nanofibers greatly improve their
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55 sensitivity toward diverse analytes correlated to diseases such as cancers276,277,
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57 cardiovascular disease278, renal dysfunction279, diabetes280, among others. For example,
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60 Mercante et al prepared an electrochemical sensor relied on an electrospun polyamide
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2 6/poly(allylamine hydrochloride) (PA6/PAH) nanofibers functionalized with
3
4 multiwalled carbon nanotubes to detect the dopamine (DA), a neurotransmitter which
5
6
7 abnormal levels have been associated with various diseases such as Schizophrenia and
8
9 Parkinson’s disease274. The nanofiber platform displayed an excellent detection limit as
10
11 well as high sensitivity, stability and selectivity, even in the presence of interfering
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14 substances commonly present in biological fluids. In another study, the same group
15
16 fabricated an electrochemical immunosensor through covalent immobilization of anti-
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18 CA19-9 antibodies on electrospun polyamide 6/ poly(allylamine hydrochloride)
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nanofibers coated with gold nanoparticles (AuNPs) to detect the pancreatic cancer
21
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23 biomarker CA19-9281. The immunosensor showed high sensitivity toward CA19-9
24
25 biomarker and was able to selectively detect this compound in real patient samples, thus
26
27 demonstrating great potential to be applied on diagnosis of pancreatic cancer at early
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30 stages.
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32 Wearable nanofiber-based sensors have emerged as a promising approach to
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34 real-time monitoring of physiological signals such as pulse rate282, respiration283,
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37 muscle movements284, breath humidity285, and glucose level286. The renewed interest on
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39 these nanostructured materials is attributed to their facile and low-cost preparation,
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41 biocompatibility, biodegradability, high sensitivity, reliability, and stability besides
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43
appropriate mechanical properties and fast response time287. Jin et al.288 investigated
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46 electrospun polyvinylidene fluoride (PVDF) nanofibers incorporated with Poly(methyl
47
48 methacrylate) microbeads (Figure 16a) as a pressure sensor (Figure 16b). The authors
49
50 found that developed platforms displayed high flexibility, sensitivity, and stability as
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53 well as fast response time. In addition, the sensor demonstrated an excellent
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55 performance to detect heart rate and respiratory signals, as illustrated in Figures 16 c,d
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57 and 16 e,f, respectively.
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40 Fig. 16 - (a) SEM micrograph of the electrospun polyvinylidene fluoride (PVDF)
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42 nanofibers incorporated with poly(methyl methacrylate) microbeads; (b) Image of a
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45 nanofiber/microbead pressure sensor. (c) photograph of pressure sensor integrated in a
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47 face mask (d) real-time response of sensors in function of respiratory intensity during
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49 patient relaxation and exercise. (e) photograph of pressure sensor fixed to human neck
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skin by using a commercial adhesive tape. (f) Real-time response of sensor during pulse
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54 detection, where the inset corresponds to a typical curve of single pulse wave. Reprinted
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56 with permission from ref (288). Copyright 2020 American Chemical Society.
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As discussed, diverse studies have indeed demonstrated that nanofibers hold
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great potential for disease monitoring and diagnosis. However, the use of nanofibers for
57
1
2 such applications have experienced greater challenges in commercial adoption due to
3
4 the absence of strategies that converges manufacturing speed and precision to produce
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7 cost-effective, sensitive, and selective sensors. In addition, further studies are still
8
9 necessary to evaluate the long-term stability of such nanofiber systems.
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14 4. Summary and future perspectives
15
16 An overview of the spinning techniques to produce polymeric fibrous platforms
17
18 useful for biomedical purposes was reported in this review. We discussed the progress
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of innovative spinning strategies for the preparation of micro/nanofibrous structures
21
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23 from a wide range of natural and synthetic polymers and active materials to produce
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25 functional composite nanofibers with unique properties aiming at fulfilling the required
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27 features of a given biomedical application. Some of the approaches used to fabricate
28
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30 functional polymer nanofibers have experienced remarkable improvements in the last
31
32 years. For instance, solution blow spinning has demonstrated to display several
33
34 advantages over the conventional electrospinning technique, namely: no need to apply
35
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37 an electrical field, no dependence on dielectric constant of the solvent, ease of
38
39 fabrication and scaling, faster production rates, lower cost and applicability to any type
40
41 of surface, in particular to living systems.
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The future perspectives of spun fibrous materials in biomedical field are
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46 optimistic and promising, but there is room yet to improve the current fabrication
47
48 methods, as well as to develop new strategies to produce nanofibers at larger and
49
50 industrial scale, with greater control of the properties at the nanoscale and possibly up to
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53 the molecular level. Another important challenge for the use of micro/nanofibers for
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55 biomedical applications consists in the design of 3D-nanostructured materials exhibiting
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57 precise spatial and temporal control. The evolution of spinning technology to prepare
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60 3D fiber constructs with high-throughput, simplicity, and at a low cost will enable the
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2 faster translation of technologies into the clinic. To date, electrospinning and
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4 microfluidic spinning are the most promising techniques to produce 3D constructs.
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6
7 However, it is crucial that efforts should be devoted to the development of such
8
9 technologies in a way that ensures robust and cost-effective manufacturing scale-up.
10
11 Future directions for addressing this particular issue will be the convergence of
12
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14 conventional spinning techniques and 3D printing or textile technologies including
15
16 weaving, braiding, and embroidering methods. Overall, although some challenges for
17
18 the clinical application of spun fibrous platforms still remain, it is believed that these
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structures will become commercially available in the near future.
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22
23
24
25 5. Acknowledgments
26
27 The authors would like to thank the financial support from FAPESP (Fundação
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29
30 de Amparo à Pesquisa do Estado de São Paulo – grant numbers 2017/20973-4,
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32 2017/12174-4, 2018/22214-6), CNPQ (Conselho Nacional de Desenvolvimento
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34 Científico e Tecnológico), FAPEMIG (Fundação de Amparo a Pesquisa de Minas
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37 Gerais – grant numbers APQ-01505-15, APQ-00906-17) MCTI-SisNano
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39 (CNPq/402.287/2013-4), CAPES (Coordenação de Aperfeiçoamento de Pessoal de
40
41 Nível Superior) Financial Code 001 and Rede Agronano-Embrapa from Brazil. The
42
43
authors also acknowledge support and interaction with current and previous
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45
46 collaborators and students.
47
48
49
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