Nanofiber Technology

• Techniques of nanofibers for tissue

engineering
• Characteristics of tissue engineering
nanofibrous structures
• Electrospinning of synthetic and natural
polymers
3 techniques to achieve nanofibers for TE

Phase separation

Electrospinning
 Phase separation
• This process involves dissolving /
pelarutan of a polymer in a solvent at
a high temperature followed by a
liquid–liquid or solid–liquid phase
separation induced by lowering the
solution temperature
• Capable of wide range of geometry
and dimensions include pits, islands,
fibers, and irregular pore structures
• Simpler than self-assembly a) powder, b) scaffolds with continuous network, c) foam with closed
pores.4

SEM of nanofibrous scaffold with interconnected spherical macropores1

 Electrospinning
• This process involves the
ejection/pelepasan of a
charged polymer fluid onto an
oppositely charged surface.
• multiple polymers can be
combined
• control over fiber diameter and
scaffold architecture

A schematic of the electrospinning process to illustrate the basic

phenomena and process components1
 Electrospinning
Proses elektrospining dibangun
dengan menggunakan arus listrik
tegangan tinggi dan kemudian
larutan di charging dengan
tegangan tinggi tersebut.
Kemudian apabila daya dorong
mekanik dan listrik mampu
mengalahkan gaya tegangan
permukaan maka terbentuk
polimer jet.
Polimer jet ini bergerak kearah kolektor. Dalam
perjalan menuju kolektor terjadi pengurangan
diameter jet dan pada saat sampai dikolektor
polimer sudah hampir kering dan diameter serat
sudah dalam ukuran nano.
Scaffolds under Electrospinning
• Goal of scaffolds
• Properties
• Materials
– Synthetic
– Natural
• Limitations
 Goals/Properties
Goal of TE is to combine cell, scaffold
(artificial ECM) and bioreactor to design and
fabricate tissues and organs.

Design scaffold with maximum control over:

biocompatibility (chemical)
biodegradability (mechanical)
Scaffolds Properties

• Biocompatible • Promote growth • Maintain 3-D structure • Non-immunogenic

• Support tissue and cell forces

 Materials
• Synthetic • Natural
– Mimic mechanical – Cell recognition
properties (strength, – Biodegradable
elongation and knot – Difficult degradation
retention) control between host
– Mass production
– Degradation between
hosts is minimal
 Synthetic
• Polyglycolic acid (PGA)
– Highly crystalline, hydrophilic, byproduct is glycolic acid
• Polylactic acid (PLA)
– Hydrophobic, lower melting temperature, byproduct is lactic acid
• Polydioxanone (PDO)
– Highly crystalline
• Polycaprolactone (PCL)
– Semi-crystalline properties, easily co-polymerized, byproduct caproic
acid
• Blends
– PGA-PLA
– PGA-PCL
– PLA-PCL
– PDO-PCL
 Poly(glycolic acid) (PGA)
Advantages: Disadvantages:
• Biocompatible & biodegradable • fast degradation causes pH change
• bioabsorption (2-4 wks) • Tissue may require buffering
• electrospinning yields diameters ~ capacity
200 nm
• Good choice for high strength and
elasticity and fast degrading material

SEM showing the random fiber arrangement (left) and the

aligned fiber orientation (right) (1600× magnification).
 PGA
• Spinning orientation
affects scaffold elastic
modulus

• The overall results

exhibit a correlation
between the fiber
diameter and
orientation and the
elastic modulus and
strain to failure

Fig. summarizes mechanical properties in terms of the elastic

modulus and strain at failure of PGA in a uniaxial model.
 Poly(lactic acid) (PLA)
Advantages: Disadvantages:
• Biocompatible & biodegradable • Larger diameter fibers ~ microscale
• bioabsorption (30 wks)
• Good choice for drug delivery do to
predictable degradation

SEM showing the random oriented PLA from cholorform (left)

and the randomly oriented PLA from HFP (right)
 PGA + PLA blends (PLGA)
Group Tested copolymers of following Group found:
ratios:
• 75%PLA–25%PGA • Hydrophilicity proportional to
• 50%PLA–50%PGA, composition of copolymer
• blended PLA and PGA together in • Degradation rate proportional to
HFP at ratios of 100:0, 75:25, 50:50, composition of copolymer
25:75
 PLGA
Applications of PLGA:
• cardiac tissue in mice for
tissue regeneration
• individual
cardiomyocytes
attachment at seeding
• scaffold loaded with
antibiotics for wound
healing

mechanical properties, such as tangential modulus, peak stress, and strain to

failure, of these copolymers and blends appear to be controlled by the
fiber/polymer composition
 Polydioxanone (PDO)
Advantages: Disadvantages:
• Biocompatible & biodegradable • Lack on knot retention
• degradation rate between PGA/PLA • Lack of adaptability to developing tissue
• Shape memory
• Excellent flexibility
• Modulus comparable collagen and elastin
• Good source for future vascular grafts

SEM of 180 nm diameter randomly

Results of the fiber diameter analysis versus PDO concentration
oriented fibrous structures
illustrating the linear relationship between electrospinning
solution concentration and fiber
diameter.
 Polycaprolactone (PCL)
Advantages: Disadvantages:
• Biocompatible & biodegradable • No shape retention/penahanan
• Inexpensive (highly elastic)
• Highly elastic
• Slow degradation (1 – 2 yrs)
• Good choice for Human
mesenchymal stem cells (hMSCs)
seeding/penyemaian to induce
differentiation

Applications:
• Bone tissue strengthening
• Cardiac grafts
• Collagen and cellular interaction
• Differentiation with MSC cells
 PGA + PCL blend
Advantages:
• PGA – high stress tolerance
• PCL – highly elastic
• Optimum combination PCL/PGA ~ 1/3
• Longer degradation time ~ 3 months
(PCL-2 yrs, PGA 2-4 wks)
 PLA + PCL blends
Advantages: Disadvantages:
• Greater elasticity than PGA+PCL • Decreasing PLA+PCL ratios
• Similar tensile strength to PLA decreases strain capacity, optimized
• ~5% addition of PCL increased at 95:5
strain by 8 fold
• Overall best synthetic ECM for
cardiac applications

Strain to failure of a tissue matrix a function of both composition (varying blend ratios of PLA and PCL) and fiber
alignment
 PDO + PCL blends
Advantages:
• PCL high elasticity
• PDO shape memory

Peak stress values for various PCL concentrations versus PDO:PCL ratio Disadvantages:
and orientation of the testing specimen
• lower tensile capacity than
PDO
• lower elasticity than PDO
• larger fiber diameter

* Further investigation needed

Strain at break values for various PCL concentrations versus PDO:PCL ratio
and orientation of the testing specimen
 Natural
• Elastin
• Gelatin collagen
• Fibrillar collagen
• Collagen blends
• Fibrinogen
 Elastin
Advantages:
• Linearly elastic biosolid
• Insoluble and
hydrophobic
• Critical role in shape and
energy recovery for
organs

Disadvantages:
• Less elastic than native
elastin
SEM of electrospun elastin scaffold at 250 mg/ml.
• Needs to be combined
with PDO to increase
tensile strength
• Fiber ~300 nm (not as
small as PDO ~ 180 nm)
• Varying diameter
 Collagen
Gelatin (denatured collagen)
Advantages:
• Biocamptibale and
biodegradable
• Inexpensive
Disadvantages:
• Quick to dissolve

Fibril-forming (Types I, II, III)

• Most abundant natural polymers in body
• Important role in ECM
• Type I: principal structure in ECM
• Type II: pore size and fiber diameter easily
controlled
• Type III: still under investigation
 Collagen blends (1st attempts)
• Artery
– Intima: innermost layer, composed of single layer of endothelial
cells on basement membrane of elastin and Type IV collagen
– Media: thickest layer, several layers consisting of collagen type I
& III, elastin and proteoglycans
– Adventitia: made of fibroblast and collagen type I

(Left) Photograph of 2 and 4 mm ID electrospun scaffolds. (Middle) SEM of tubular electrospun composite. (Right) SEM
of electrospun 40:40:20 blend of collagen type I, collagen type III, and elastin with random orientation.
 Collagen blends (1st attempts)
• Collagen Type I & III + PDO
– indication that blends of PDO and collagen may
match mechanical and morphological requirements of
a blood vessel's microenvironment (similar to PDO
section).

Tangential modulus presented as a function of the ratio of PDO to collagen type I & III. collagen I
highest tensile capacity, optimal ratio for all collagens was 70:30 collagen-PDO.
 Globular proteins
• Fibrinogen (protein in blood plasma – wound healing)
– Low concentration produced fibers within range of fibrinogen
fibers in plasma clots (80, 310, 700 nm)
– High surface area to volume ratio: increases area available for
clot formation
– Stress capacity comparable to collagen (80-100 MPa)

the linear relationship between concentration and fiber diameter composing the structures produced.
 Globular proteins
• Hemoglobin & myoglobin
– Fiber sizes 2-3 µm & 490 – 990 nm
– Spun with fibrinogen for clotting and healing improvements
– High porosity means higher oxygenation
– Clinical applications:
• Drug delivery
• Hemostatic bandages
• Blood substitutes

SEM of electrospun hemoglobin in 2,2,2-Trifluoroethanol at 150 mg/ml (A), 175 mg/ml (B), and 200
mg/ml
 Summary
Scaffolds:
• Electrospinning viable for both synthetic and biological
scaffolds/mats

• synthetic polymers
– PGA, PLA and PLGA most commonly used
– PDO most similar to Elastin collagen blend (limited by shape memory)
– PCL most elastic and mixed frequenlty with other material
– Provide nanoscale physical features
• Natural polymers
– Collagen Type I & III + PDO: best possible match for blood vessels

Limitations on Scaffolds
• Mechanical material failure
• Immunogenic reaction to material