International Journal of Biological Macromolecules 219 (2022) 545–557

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International Journal of Biological Macromolecules

journal homepage: www.elsevier.com/locate/ijbiomac

Review

Chitosan-based high-strength supramolecular hydrogels for 3D bioprinting

Jiaqi Xu, Manyue Zhang, Wenzhen Du, Jiuhong Zhao, Guixia Ling *, Peng Zhang *
Shenyang Pharmaceutical University, No. 103, Wenhua Road, Shenyang 110016, China

A R T I C L E I N F O A B S T R A C T

Keywords: The loss of tissues and organs is a major challenge for biomedicine, and the emerging 3D bioprinting technology
Chitosan has brought the dawn for the development of tissue engineering and regenerative medicine. Chitosan-based
Supramolecular hydrogels supramolecular hydrogels, as novel biomaterials, are considered as ideal materials for 3D bioprinting due to
3D bioprinting
their unique dynamic reversibility and fantastic biological properties. Although chitosan-based supramolecular
hydrogels have wonderful biological properties, the mechanical properties are still under early exploration. This
paper aims to provide some inspirations for researchers to further explore. In this review, common 3D bio­
printing techniques and the properties required for bioink for 3D bioprinting are firstly described. Then, several
strategies to enhance the mechanical properties of chitosan hydrogels are introduced from the perspectives of
both materials and supramolecular binding motifs. Finally, current challenges and future opportunities in this
field are discussed. The combination of chitosan-based supramolecular hydrogels and 3D bioprinting will hold
promise for developing novel biomedical implants.

1. Introduction proliferation and affects the adhesion of hydrogels to surrounding bio­

The severe shortage of tissues and organs remains a major global
challenge [1]. In recent years, the gradual rise of 3D bioprinting has
brought light to the development of tissue engineering and regenerative
medicine. 3D bioprinting is the process of converting 3D digital models
into tissues or organs printed with cytocompatible bioink [2], which has
the advantages including precise control of cell distribution, scalability,
high resolution, and low cost [3,4]. Therefore, 3D bioprinting technol­
ogy has received extensive attention over the years. But the develop­
ment of bioink suitable for printing remains a major problem to be
solved for 3D bioprinting.
Hydrogels are a kind of 3D network structure gel formed by physical
or chemical crosslinking. The high water content, porous structure,
excellent biocompatibility and biodegradability of hydrogels are similar
to natural extracellular matrix (ECM), providing a favorable environ­
ment for the survival of various cells [5]. Accordingly, hydrogels are
promising biomaterials, which have been widely used in drug delivery,
wound healing, 3D bioprinting and tumor therapy [6–11].
Generally, the conventional hydrogels rely on chemical crosslinking
between polymer chains, such as hydrogels based on chemical initiators,
crosslinking agents, and metal catalysts [12]. However, the covalently
crosslinked hydrogels exhibit brittleness, irreversibility and lack of self-
healing [13], which hinder cell differentiation, migration and
logical tissues [14,15]. Moreover, the hydrogels crosslinked by covalent
interactions are usually permanent and irreversible as well as poor
biocompatibility and biodegradability, which makes the network
structure unable to be rebuilt after being destroyed [12] and becomes an
obstacle to the diffusion of nutrients [16]. As a result, traditional
hydrogels still have certain limitations in 3D bioprinting, hindering the
widespread applications of 3D bioprinting in biomedicine.
Chitosan is a natural linear polysaccharide and the second most
abundant natural polymer after cellulose. Chitosan hydrogels have been
widely used in the field of biomedicine. For example, Deng et al. [17]
developed an adenine-modified chitosan hydrogel for promoting wound
healing, Wang et al. [18] designed a chitosan-based conductive hydrogel
as a flexible sensor, and Dziadek et al. [19] synthesized a chitosan-based
hydrogel with high antioxidant and anticancer activity. Compared with
traditional hydrogels, Chitosan supramolecular hydrogels based on non-
covalent interactions have dynamic reversibility as well as excellent
biocompatibility and biodegradability, which facilitates the recon­
struction of network structures. Additionally, similar to the natural
ECM, this interaction can provide a suitable living environment for cells
and promote the integration of surrounding tissues. Furthermore, su­
pramolecular hydrogels also have the characteristics of shear-thinning,
self-healing [16], and stimuli responsiveness, which meet the neces­
sary conditions for cells to survive in the human ECM. But the unique

* Corresponding authors.
E-mail addresses: pharlab@163.com (G. Ling), zhangpengspu@163.com (P. Zhang).

https://doi.org/10.1016/j.ijbiomac.2022.07.206
Received 2 June 2022; Received in revised form 21 July 2022; Accepted 25 July 2022
Available online 28 July 2022
0141-8130/© 2022 Published by Elsevier B.V.
J. Xu et al.
Fig. 1. Schematic diagram of four printing technologies. (a) Inkjet-based 3D bioprinting. (b) Extrusion-based 3D bioprinting. (c) Laser-based 3D bioprinting. (d)
Stereolithography 3D bioprinting.
dynamic reversibility also affects the mechanical properties of chitosan-
based supramolecular hydrogels, resulting in limited applications of
chitosan-based supramolecular hydrogels. To expand the applications of
chitosan supramolecular hydrogel-based 3D bioprinting in bioengi­
neering. Firstly, this review briefly introduces the commonly used 3D
bioprinting techniques and the conditions that 3D printed bioink need to
meet, so that readers can understand the background knowledge better.
Then, chitosan-based supramolecular hydrogels applied to 3D printing
are discussed from the perspective of improving mechanical properties,
which is helpful for researchers to select appropriate mechanisms and
materials to prepare biomaterials with different strengths. This review is
based on other authoritative reviews to which interested readers are
referred for a more comprehensive exploration [20,21].
2. 3D bioprinting technologies
2.1. Inkjet-based 3D bioprinting
Inkjet-based 3D bioprinting mainly uses heat or piezoelectric force to
eject the bioink from the nozzles [22]. As shown in Fig. 1a, thermal
inkjet bioprinting has the characteristics of fast printing speed and low
cost. However, thermal inkjet bioprinting produces droplets with a small
diameter range (30–80 μm), which can easily lead to nozzle clogging
[23]. In addition, it is difficult to print uniform and stable droplets due to
thermal and shear stress. Compared with thermal inkjet bioprinting,
piezoelectric inkjet bioprinting produces droplets in the range of
50–100 μm in diameter [24], which can more easily control droplet
formation and reduce nozzle clogging. However, it has been previously
demonstrated that piezoelectric forces can disrupt the integrity of cell
wall and cause cell lysis by sonication at frequencies of 15–25 kHz [25].
Therefore, thermal inkjet bioprinting can maintain higher cell viability
(90 %) compared to piezoelectric inkjet bioprinting. But inkjet-based 3D
bioprinting usually chooses printing materials with low viscosity (<10
mPa/s) and low cell density (<106 cells/s) due to the tendency of cell
deposition which causes nozzle clogging [26]. Additionally, printing
low viscosity bioinks can cause ink droplet spreading, spattering, and
deformation, affecting the resolution and the integrity of printed
structures [27]. These shortcomings limit the widespread application of
this technology.
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International Journal of Biological Macromolecules 219 (2022) 545–557
2.2. Extrusion-based 3D bioprinting
Compared to the ink droplets of inkjet-based 3D bioprinting,
extrusion-based 3D bioprinting extrudes cylindrical lines, as shown in
Fig. 1b, which can print high viscosity (30 mPa/s-above 6 × 107 mPa/s)
and high density (>106 cells/s) materials [28]. However, due to the high
shear stress requirements of bioinks with high viscosity, the cell viability
of extrusion-based 3D bioprinting is usually maintained at 40–80 %
[27], with low resolution (200–1000 μm) [2] and prone to nozzle
clogging during printing. In addition, excessive crosslinking can affect
cell proliferation, migration, and spreading [29]. Typically, extrusion-
based 3D printing can be divided into two categories: pneumatic and
mechanical (piston or rotating screw). 3D bioprinting under the action
of gas pressure is difficult to accurately control the deposited mass due to
the delay of compressed gas volume [30]. Higher shear stress is gener­
ated by mechanical action during bioprinting (rotating screw). In gen­
eral, extrusion-based printing techniques require the selection of high-
viscosity bioink to strengthen structures to avoid damage to cells
caused by shear stress [31]. However, high-viscosity materials can cause
nozzle clogging and affect the uniform distribution of cells, which can be
solved by using bioink with shear-thinning properties or secondary
crosslinking [32]. Although extrusion-based bioprinting has certain
limitations, this technique can print anatomically porous constructs,
which expands its biomedical applications [33].
2.3. Laser-based 3D bioprinting
Unlike inkjet-based and extrusion-based 3D bioprinting, as shown in
Fig. 1c, since there is no printing nozzle, laser-based 3D bioprinting can
avoid direct contact with cells and will not be affected by shear stress
[34]. Therefore, this printing technique can print a series of different
viscosity materials and maintain high cell viability (>90 %) [3]. In
addition, laser-based 3D bioprinting has a high resolution (20–50 μm),
which can mimic the natural tissue structure to print an accurate and
complex pattern [2]. This feature is related to the viscosity of the bioink
and various characteristics of the pulsed laser source [35]. However,
laser-based 3D bioprinting still faces challenges of high cost and
complexity. Some undeveloped parameters affect the size and shape of
the ink droplets [36]. Although the absence of nozzles breaks the
J. Xu et al. International Journal of Biological Macromolecules 219 (2022) 545–557

Table 1
Comparison of four 3D bioprinting technologies.
Parameters Inkjet-based 3D bioprinting Extrusion-based 3D bioprinting Laser-based 3D bioprinting Stereolithography 3D Ref.
bioprinting

Bioprinting Medium (mm/s) Slow (10–50 μm/s) Medium (mm/s) Fast (mm3/s) [45]
speed
7
Bioink 3.5–12 mPa/s 30 mPa/s-above 6 × 10 mPa/s 1–300 mPa/s No limitation [25,46,47]
viscosity
Cell viability >85 % 40–80 % >90 % >85 % [48,49]
Cell density Low (<106 cells/mL) High Medium (<108 cells/mL) Medium (<108 cells/mL) [25]
Resolution 20–100 μm 200–1000 μm 20–50 μm 20–50 μm [27]
Cost Low Medium High High [50]
Biomaterial Thermo/pH/photosensitive Thermo/photosensitive Photosensitive Photosensitive [51,52]
choice
Advantages Compatibility with low viscosity Multiple biomaterials and cell Wide range of printable Several photopolymerizable [22,26,53,54]
biomaterials (3.5–12 mPa/s); High types can be incorporated. viscosity; No nozzle; High materials can be used; Surface
throughput; Easy implementation. Homogeneous and heterogeneous resolution (20–50 μm) and smooth; Complex structure can
structures can be created; Good cell vitality (>90 %). be obtained.
cell sedimentation effect.
Disadvantages Nozzle clogging; Thermal stress and Nozzle clogging; Effect of shear Difficult to incorporate Poor cell sedimentation effects;
mechanical stress; Limited material; stress and viscosity on cells; multiple bioactive agents; Cytotoxicity of photosensitive
Poor cell sedimentation effect; Limited material. Poor cell sedimentation materials.
Complex 3D structures are difficult effects and cell
to implement. homogeneity.

technical limitations of using bioink with different viscosities, the
inability to print ink droplets to the exact spot will cause a certain
amount of waste. Furthermore, the materials applicable to laser-based
bioprinting are limited to photosensitive materials, and the use of UV
light and photoinitiators during photocuring can lead to cell damage and
cytotoxicity [37–39]. In order to achieve high shape fidelity, the overall
flow rate of this technology is relatively low, resulting in a decrease of
printing efficiency [40].
2.4. Stereolithography 3D bioprinting
Stereolithographic 3D bioprinting is another printing technique
without printing nozzles, which has a high resolution (<100 μm) and
high cell viability (>85 %) [41]. As shown in Fig. 1d, it is a photosen­
sitive bioink that selectively cures light in layer-by-layer printing.
Therefore, it is necessary to control the wavelength and intensity of light
and the concentration of the photoinitiator so as not to affect cell
viability [31]. Stereolithographic 3D bioprinting, which allows users to
control the resolution, porosity, and geometry of the print, has been
widely used in tissue engineering, such as bone tissue [42], neural tissue
[43], and vascular tissue [41]. Compared with laser-based 3D bio­
printing, it is worth noting that this is a 3D bioprinting technology with
the help of light projection, and its printing time depends on the
thickness of the material to be printed, rather than the size and
complexity of each layer [2]. This greatly reduces the complexity of
printing operations. But the technology should choose a low-viscosity
bioink, because during the photocuring process, the viscous bioink
will stay inside the structure, which is not conducive to the formation of
3D structures [31]. Although no nozzles are required like laser-based
printing, this technology is costly and the biodegradability and cyto­
toxicity of photosensitive materials cannot be ignored [44]. Table 1
summarizes the different printing techniques discussed above.
3. Bioink for 3D bioprinting
In short, 3D bioprinting is the transformation of printed structures
into tissues that can survive in the body. In order to achieve this trans­
formation, an ideal bioink should have three main functions. Firstly, the
bioink should have certain mechanical properties, which can protect
cells and maintain cell viability. Secondly, the bioink must be mechan­
ically compatible with the printing technology, which is mainly re­
flected in the printing technology based on inkjet and extrusion. Finally,
the bioink should have a stable structure after printing and still have
biological activity and specific biological functions in vivo [31]. As a
result, bioink should not only meet the requirements of biological ma­
terials but also meet the needs of biology [55]. The requirements of
biomaterials mainly include characteristics such as printability, easy
functional modification, and biodegradability [3]. Due to the limitations
of extrusion-based and inkjet-based 3D bioprinting, bioink for these two
printing techniques should consider both shear-thinning properties and
viscosity [34]. Biological properties mainly include biocompatibility,
cytocompatibility, and the cell activity after printing. Biocompatibility
refers to cells having good cell viability when cultured with bio­
materials. At the same time, it has cell behaviors such as proliferation,
adhesion, and spreading. To obtain an ideal bioink, the biological
functions and physicochemical properties of bioink can be designed, for
instance, introducing groups with certain characteristics, changing the
composition ratio of the original groups, or grafting biological small
molecules with specific functions. 2-ureido-4[1H]-pyrimidinone (UPy)
is a widely studied hydrogen bonding unit [56]. The introduction of Upy
in supramolecular hydrogels can induce the supramolecular properties
of biomaterials and improve the mechanical properties of the bio­
materials by changing hydrophobic/hydrophilic ratio, thereby obtain­
ing ideal bioinks [16]. Furthermore, the introduction of nitrobenzyl and
azobenzene can endow supramolecular hydrogels with photo-
responsiveness [57], while the introduction of sodium chloride and
vancomycin can drive the self-assembly of supramolecular hydrogels
[58,59]. Therefore, to satisfy these properties, bioink need to possess
reasonable mechanical properties, and researchers have conducted
extensive researches in this field in recent years [60,61]. At present, how
to balance these conditions to obtain an ideal bioink is the main chal­
lenge for 3D bioprinting.
4. Chitosan-based supramolecular hydrogels
Chitosan is a cationic polymer produced by partial deacetylation of
chitin, containing N-acetyl-D-glucosamine and deacetylated D-glucos­
amine units [62]. Chitosan is the most abundant natural polymer after
cellulose [30]. In addition, chitosan has excellent biological properties,
such as biodegradability, biocompatibility and non-toxicity [63]. Chi­
tosan is the only positively charged natural polysaccharide, which can
interact with negatively charged molecules such as polysaccharides,
proteins, lipids, etc. [30]. Due to the presence of cations, it can generate
ionic interactions with negatively charged cells to destroy bacterial cell
membranes, exhibiting excellent antibacterial properties [64]. Addi­
tionally, it can also interact with negatively charged blood cells to form

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Table 2
Comparison of four supramolecular interactions.
Interactions Strength Features
Host-guest Wide range of One or more hydrogen bonding, hydrophobic
interactions strengths interactions, electrostatic interactions, π-π stacking, and
van der Waals interactions
Hydrogen Weak (mostly Interaction between hydrogen atoms and electronegative
bonding 20 kJ/mol) atoms
Ionic interactions Relatively Electrostatic interactions between oppositely charged
strong ions or groups
Hydrophobic Medium Interactions between water-soluble polymers with water-
interactions strength insoluble end groups, side chains or monomers
thrombus and achieve hemostasis [65]. Based on the above character­
istics, Chitosan has been widely used in the fields of food, cosmetics and
biomedicine [66].
The concept of supramolecular chemistry was first proposed by
Nobel laureate Jean-Marie Lehn [67]. Supramolecular chemistry is a
specific but non-permanent interaction, i.e., non-covalent interactions.
So far, a variety of interactions have been developed to prepare supra­
molecular hydrogels, such as host-guest interactions, hydrogen bonding,
ionic interactions, and hydrophobic interactions [56,68]. Due to the
dynamic nature of non-covalent interactions, supramolecular hydrogels
exhibit responsiveness to various stimuli, such as temperature [69],
redox reactions [70], illumination [71], etc., which trigger molecular
self-assembly. This feature provides a view for studying sensors based on
external stimuli. Supramolecular hydrogels are comparable to or better
than traditional hydrogels in terms of their transient dynamic proper­
ties, stimuli responsiveness and the ability to combine multiple biolog­
ical functions [12]. In addition, supramolecular hydrogels have more
excellent tunability, biocompatibility and biodegradability [72]. More­
over, supramolecular hydrogels can spontaneously be degraded or
metabolized in vivo when subjected to external stimuli due to their
unique stimuli-responsiveness, thereby avoiding the harm to human
health caused by accumulation in the body [59]. Additionally, supra­
molecular hydrogels have better reversibility, which can still return to
its original state after external damage. The binding between supra­
molecular hydrogels relies on non-covalent interactions between mole­
cules, which are similar to the interactions between proteins and
carbohydrates in the ECM. In addition, it can also imitate ECM in me­
chanical properties, and can be used for 3D cell culture [73]. Under this
biological inspiration, supramolecular materials can be used to mimic
this dynamic interaction to form novel biomaterials. Consequently, the
biodegradability and biocompatibility of such hydrogels are generally
better compared to conventional hydrogels. Supramolecular hydrogels
can also be easily modified with various functional components to
achieve expected or even unexpected effects. These properties make
supramolecular hydrogels excellent bioink for 3D bioprinting. Although
supramolecular hydrogels are significantly superior to traditional
hydrogels in terms of reversibility, biocompatibility, biodegradability,
and ECM-mimicking properties, their mechanical properties are rela­
tively unsatisfactory [12], which limits their applications in tissue en­
gineering. Table 2 summarizes the different interactions discussed
above.
In general, chitosan-based hydrogels can be classified into covalently
crosslinked and non-covalently crosslinked. However, most of the
crosslinking agents used in covalent-based chitosan hydrogels are
cytotoxic or have unknown effects on the body [74]. Irreversible cova­
lency cannot reconstruct the damaged hydrogel structure. Compared
with covalent hydrogels, chitosan supramolecular hydrogels crosslinked
by non-covalent interactions are dynamically reversible. And the sol-gel
transition can be achieved by physical stimulation (e.g., pH, tempera­
ture, magnetic field) [75], so that the cytotoxicity of the chitosan -based
supramolecular hydrogels was negligible due to the absence of cross­
linking agents. Therefore, non-covalent based chitosan hydrogels with
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International Journal of Biological Macromolecules 219 (2022) 545–557
Examples Ref.
Cyclodextrins (CD) and adamantane; CD and alginate; [79,80]
cucurbiturils (CB) and hyaluronic acid; CB and carboxymethyl
cellulose
HA, chitosan, starch, carboxymethylcellulose, and hydroxypropyl [81,82]
chitosan
Alginate and Ca2+; alginate and chitosan; HA and chitosan [72]
chitosan and β-glycerophosphate; methylcellulose and N- [83–85]
isopropylacrylamide; carboxymethylcellulose and poly(N-
isopropylacrylamide)
higher safety and lower production cost have received extensive atten­
tion, such as drug delivery [76], wound dressings [77], and bioscaffolds
[78]. However, compared with strong covalent interactions, the me­
chanical properties of chitosan supramolecular hydrogels based on non-
covalent interactions are relatively unsatisfactory, thus limiting their
application in biomedicine.
5. Strategies to enhance the strength of chitosan hydrogels
The field of mechanics has demonstrated the importance of the
mechanical properties of ECM for cell growth and stability [86–89]. This
is also one of the main limiting factors for bioink suitable for 3D bio­
printing. However, the tunable mechanical properties of chitosan-based
supramolecular hydrogels are still in the early stage of exploration.
Consequently, it is necessary to understand how to improve the me­
chanical properties, which will facilitate the rational design of the su­
pramolecular structures of hydrogels for applications in 3D bioprinting.
In this part, we will discuss how to form hydrogels with certain me­
chanical properties from the aspect of binding motifs and materials of
chitosan supramolecular hydrogels (Table 2), of course, shear-thinning
and self-healing properties will also be mentioned.
5.1. Chitosan-based nanomaterials
Graphene, due to unique two-dimensional structure, has become an
ideal nanoscale material for the fabrication of supramolecular hydro­
gels. However, the strong π-π interaction between graphenes easily leads
to the agglomeration of sheet-like graphene in the hydrogel matrix,
therefore, a great deal of researches is mainly focused on graphene de­
rivatives. For example, reduced graphene oxide (rGO) with excellent
water dispersibility can be utilized as a crosslinking agent and rein­
forcing filler [90]. Furthermore, oxidation of graphene with a strong
oxidant can improve the solubility of graphene under physiological
conditions, resulting in the formation of graphene oxide (GO), which can
form a crosslinked network with hydrophilic polymers through physical
interactions [91]. The resulting material has excellent properties and
unique functionality. GO has high mechanical strength, hydrophilicity,
biocompatibility and biostability, which can significantly enhance the
mechanical strength of chitosan hydrogels [92]. Chitosan-GO nano­
composite hydrogels have been extensively explored as scaffolds for
tissue engineering. Marapureddy et al. [93] investigated a simple
chitosan-GO hydrogel without polymeric support and achieved tunable
mechanical strength of chitosan hydrogels only by changing the con­
centration of GO. Chitosan-GO scaffolds could be obtained by direct
write printing with a nozzle diameter <400 μm. The fine structure of the
scaffold helped to print complex structures and improve self-assembly
and communication between cells. The results showed that the addi­
tion of 0.5 wt% GO could significantly increase the storage modulus,
viscosity and yield stress of chitosan hydrogels.
Nanocellulose, a natural polysaccharide with excellent physico­
chemical properties, has been widely used in biomedical field as a
reinforcing agent. According to the structure, nanocellulose can be
J. Xu et al.
Fig. 2. (a) Storage modulus (solid lines) and loss modulus (without lines) of CS/SF/CNPs hydrogels. (b) Evaluation of the recovery strength of CS/SF/CNPs
hydrogels. (c–d) Degradation rates of CS/SF/CNPs scaffolds in PBS and protease media. (e) The obtained CT images of rats under different conditions [96].
further divided into cellulose nanofibrils (CNFs), cellulose nanocrystals
(CNCs) and cellulose nanoparticles (CNPs) [94]. Nanocellulose have
become ideal materials for improving the mechanical properties of
polymers due to their excellent mechanical strength and biocompati­
bility. Tamo et al. [95] developed a chitosan-CNF hydrogel in which the
concentrations of CNF and chitosan were 2.0 %–3.0 % (w/v) and 0.2 %–
0.4 % (w/v), respectively. In addition, the hydrogel did not require any
chemical modification or any chemical crosslinking agents during the
preparation process. The obtained chitosan-CNF constructs had excel­
lent mechanical properties (Young's modulus 3.0 MPa, stress at break
1.5 MPa, strain at break 75 %). This high-strength, environmentally
friendly biomaterial based on natural polymers had great potential for
repairing meniscus, cartilage, and intervertebral disc. Li et al. [96]
investigated a chitosan/silk fibroin/cellulose nanoparticle (CS/SF/
CNPs) scaffold. Compared with pure polymer scaffolds, CS/SF/CNPs
scaffolds exerted excellent rheological property, swelling potential (840
→ 3670 %) and recovery strength (Fig. 2a–b). In addition, CS/SF/CNPs
scaffolds also exhibited higher degradation rates than pure polymer
scaffolds, indicating that SF and CNPs could promote the degradation of
chitosan (Fig. 2c–d). The results of the study showed that CS/SF/CNPs
scaffolds could promote macrophages polarization (M1 → M2) and
osteo-immunomodulatory. Compared with the control group, the rats in
the CS/SF/CNPs group exhibited bone regeneration (Fig. 2e), which
may be related to the promotion of osteogenic-associated genes and
growth factors by the MAPK phosphorylation pathway. Ajdary et al.
[97] investigated methods to realize chitosan-CNF mesh implants based
on direct ink writing. It was found that chitosan-sorbed post-treatment
on 3D-printed CNF networks could obtain hydrogels with excellent
mechanical properties (683 ± 63 MPa modulus and 2.5 ± 0.4 MPa
tensile strength), which was related to the electrostatic interaction be­
tween chitosan and CNF.
Currently, more and more researchers increase the mechanical
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International Journal of Biological Macromolecules 219 (2022) 545–557
properties of chitosan supramolecular hydrogels by nanomaterials such
as graphene [93] and nanocellulose [98]. Although the graphene itself is
prone to agglomeration, rGO and GO have excellent water dispersibility
and biocompatibility, which can be used for the preparation of chitosan
supramolecular hydrogels. Nanocellulose with excellent mechanical
strength and biocompatibility have also become ideal materials for
improving the mechanical properties of chitosan hydrogels. Further­
more, different nanomaterials can also be combined to form high
strength chitosan supramolecular hydrogels with different functions
[95,96].
5.2. Chitosan-based titanium alloy materials
Titanium alloys are widely used implant materials with high me­
chanical strength and corrosion resistance. On the contrary, due to the
high mechanical properties, it is prone to stress shielding, which leads to
osteolysis [99]. 3D printed porous titanium alloy (pTi) scaffolds has a
porous structure that can reduce stiffness while possessing desired me­
chanical properties, and can tailor optimal implants according to human
physiological anatomy and clinical practical applications. Its unique
microporous structure is more compatible with human tissue and can
promote tissue reconstruction and functional recovery [100]. Never­
theless, such pTi implants also have certain limitations. Not only do they
not solve the infection problems, but the porous structure also provides a
suitable breeding ground for bacteria, which is more likely to cause
tissue infection. What's more, inorganic 3D printed implants are not
biologically active. Therefore, it is necessary to design reasonable bio­
coating materials to solve these problems.
In recent years, with the continuous exploration of biological mate­
rials, it was found that adding antibacterial agents such as cellulose,
chitosan, and alginate in inorganic or organic bioink could effectively
solve the problem of tissue infection in biomedicine [101]. Wu et al.
J. Xu et al. International Journal of Biological Macromolecules 219 (2022) 545–557

Fig. 3. (a) Preparation of supramolecular hydrogels. (b) pTi composite scaffolds filled with supramolecular hydrogels. (c) Osseointegration process. (d) Appearance
of the femurs. (e) Radiograph of the femurs. (f–g) Van-Gieson staining map of the bone defect area (black area represented scaffold, red area represented bone).
Abbreviations: S: pTi scaffold; SG: pTi scaffold filled with supramolecular hydrogel; SGC: pTi scaffold filled with BMSCs and supramolecular hydrogel; SGB: pTi
scaffold filled with BMP-2 and supramolecular hydrogel; SGCB: pTi scaffold filled with supramolecular hydrogel pTi scaffolds for BMSCs, BMP-2 and supramolecular
hydrogels [105].

[102] designed an antibacterial hydrogel based on chitosan and gelatin
(GT) as a coating material for improving the fixation of the artificial
joint prosthesis-bone interface and preventing infection. With the in­
crease of chitosan content, the mechanical properties of hydrogel
gradually increased. The stress of the antibacterial hydrogels containing
3 %, 5 % and 7 % chitosan was 0.030 MPa, 0.037 MPa and 0.057 MPa,
respectively, which was attributed to the change of crosslinking struc­
ture from linear to reticular structure caused by the increase of chitosan
content. The increase in the degree of crosslinking was mainly related to
the hydrogen bonding between chitosan and GT molecules and the
electrostatic interaction between chitosan and sodium citrate. The
elastic modulus and tensile strength of 7chitosan -10GT were 0.920 MPa
and 0.268 MPa, respectively. In addition, with the increase of chitosan
content, the bonding strength between chitosan-GT hydrogel and pTi
implants (3.36 MPa) also gradually increased. The porous structure of
chitosan-GT hydrogel allowed the loading of silver nanoparticles and
exhibited excellent antibacterial properties. The chitosan-GT antibac­
terial hydrogel could not only promote the fixation of the prosthesis-
bone interface, but also had excellent antibacterial properties and
biocompatibility, which provided the possibility for the replacement of
artificial joints.
Additionally, pTi implants, because of their smooth surface, are not
prone to adhesion of cells used to promote osteogenesis, especially in the
diseases of osteoporosis and rheumatoid arthritis [103,104]. However,
the introduction of chitosan supramolecular hydrogels with remarkable
self-healing ability and injectability into implants can effectively solve

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Fig. 4. (a) Crosslinking time of hydrogels with 2 % polysaccharide concentration. (b) Effect of composition at 24 h of gelation time. (c) Degradation rates of
hydrogels under different pH conditions after crosslinking for 24 h. (d) Self-healing properties of chitosan-HA hydrogels [115].
this trouble, and is beneficial to maintain the stability of the network
structure and the mechanical properties of hydrogels. Bai et al. [105]
prepared supramolecular polysaccharide hydrogels using N-carbox­
yethyl chitosan, adipic acid dihydrazide (ADH) and hyaluronic acid-
aldehyde (HA-ALD) as raw materials (Fig. 3a). The supramolecular
polysaccharide hydrogels had excellent self-healing, injectability, and
biodegradability, which could be completely degraded in vivo within 4
weeks without causing inflammation. Bone marrow mesenchymal stem
cells (BMSCs) and bone morphogenetic protein-2 (BMP-2) with the
function of promoting bone repair and regeneration [106] were subse­
quently encapsulated in supramolecular hydrogels to maintain their
biological activity (Fig. 3b). Finally, the prepared supramolecular
hydrogels were combined with pTi scaffolds to develop a 3D bioactive
surface with compressive strength of 48.0 ± 2.1 MPa and elastic
modulus of 1.63 ± 0.2 GPa for improving the microenvironment of bone
defects (Fig. 3c). Supramolecular polysaccharide hydrogels containing
BMSCs and BMP-2 filled in the pTi scaffold facilitated the integration of
pTi implants with surrounding bone tissue and bone ingrowth. This is
because the presence of BMSCs can improve bone microstructure
[107,108] and BMP-2 has a significant osteogenic effect [106]. On the
other hand, the pTi scaffold loaded with supramolecular polysaccharide
hydrogel provided certain mechanical support for bone regeneration
due to the high strength of the pTi scaffold itself. The results showed that
the composite scaffold with BMSCs and BMP-2 could effectively induce
bone growth (Fig. 3d–e), promote osseointegration with the surrounding
bone (Fig. 3f–g) and regulate the osteoporotic microenvironment.
Although 3D-printed pTi scaffolds can effectively promote bone
regeneration, their porous structures and smooth surfaces limit their
widespread use in biomedicine. Fortunately, combining with chitosan
supramolecular hydrogels with different functions can break this limi­
tation. As mentioned above, antimicrobial-based supramolecular
hydrogels can make up for the defects of bacterial breeding in the porous
structure. The self-healing properties of supramolecular hydrogels can
551
International Journal of Biological Macromolecules 219 (2022) 545–557
solve the problem of poor integration with surrounding tissues. But it is
worth noting that the osseointegration ability of scaffolds is also closely
related to the porosity, pore size, and distribution of scaffolds. In gen­
eral, the porosity of bone tissue scaffolds should be 60 %–70 % and the
pore size should be 50-500 μm, which is conducive to the proliferation
and adhesion of osteoblasts [109]. On the other hand, most of the cur­
rent antibacterial agents such as AgNPs are cytotoxic [110]. Environ­
mentally friendly antimicrobials based on natural polymers, such as
chitosan and cellulose, can be ideal alternatives [101]. It has been
proved that Si4+can induce the secretion of type I bone collagen, regu­
late the interaction between cells and stimulate the differentiation of
stem cells, thus promoting bone regeneration and bone integration
[111]. Therefore, it is necessary to add Si4+ into bone tissue implants.
5.3. Chitosan-based natural biomaterials
Natural biomaterials are widely used in the bioink formulations due
to their inherent biocompatibility, biodegradability and tunable prop­
erties [112]. Proteins, polysaccharides and peptides have become
excellent scaffold materials for tissue engineering due to their similarity
to ECM. Chitosan is the most common biomaterial used in the prepa­
ration of bioink. The mechanical strength and gelation kinetics of chi­
tosan hydrogels can be tuned by combining with different natural
biomaterials [113–115].
Pectin (PEC) is an anionic heteropolysaccharide with gelling prop­
erties, high tunability, good biodegradability and non-toxicity. Mil­
chailidou et al. [116] used PEC to stabilize the structure of chitosan
hydrogels because the amino groups of chitosan could form strong
electrostatic interactions with the carboxyl groups of PEC. This study
investigated the effect of different concentrations of PEC on the rheo­
logical properties of bioink through rheological analysis, and finally
determined that chitosan -PEC 5–10 % was the optimal printing condi­
tion. Furthermore, alginate is a polyanion with excellent
J. Xu et al.
biocompatibility and shear thinning properties that is commonly used in
3D bioprinting. However, the viscosity of alginate solutions is poor,
which makes it difficult to guarantee the structure and shape fidelity of
alginate-based 3D printed constructs. In order to solve this problem, Liu
et al. [114] prepared chitosan-alginate hydrogels by changing the pH
value. Chitosan was swelling but insoluble in the alginate solution,
which effectively increased the viscosity of the alginate solution (1.5–4
times). After treatment with hydrochloric acid solution, the change of
pH value triggered the electrostatic interaction between chitosan and
alginate, which promoted the gelation process. Compared with the
alginate hydrogel crosslinked with Ca2+, the chitosan -alginate hydrogel
exhibited better mechanical strength, with a compressive modulus of
1.4 MPa under 90 % strain.
In addition to the natural materials mentioned above, hyaluronic
acid (HA) has high hydrophilicity and viscoelasticity, which can pro­
mote the proliferation and adhesion of cells [117]. Based on this, Fer­
nandez et al. [115] proposed a chitosan-HA composite hydrogel with
potential mucoadhesive. This study mainly evaluated the relationship
between the synthesis conditions of chitosan-HA hydrogels and me­
chanical properties, rheological properties, mucoadhesive, and swelling.
The rheological measurement results showed that the storage modulus
was higher than the loss modulus when the crosslinking time was 2 h,
indicating the electrostatic interaction between chitosan and HA. When
the crosslinking time was 24 h and 48 h, the modulus difference between
hydrogels was not obvious, indicating that 24 h was a limiting time node
(Fig. 4a). Furthermore, the increase in modulus of the hydrogel with a
polysaccharide concentration of 2 % was not significant compared to the
other samples, indicating that the concentration of 2 % was a limiting
concentration (Fig. 4b). The results of adhesion test were consistent with
the rheological results, and the mucoadhesive properties were closely
related to the crosslinking time and but not to the polysaccharide con­
centration. To determine the stability and applicability of chitosan-HA
Fig. 5. (a) Tensile properties and self-healing properties of hydrogels with different SBMA contents, AAc contents and soaking time. (b) Preparation of DN hydrogels.
(c) Compression properties of original and self-healing hydrogels. (d) Schematic diagram of self-healing properties [129].
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International Journal of Biological Macromolecules 219 (2022) 545–557
hydrogels, the degradability of the hydrogels was evaluated in this
study (Fig. 4c). The results showed that the degradation rate at acidic pH
was significantly higher than that at neutral pH, which was attributed to
the hydrogen bonding among chitosan molecules and the electrostatic
interaction between chitosan and HA. In addition, the chitosan-HA
hydrogels also exhibited excellent self-healing ability (Fig. 4d).
As mentioned above, most of the binding of chitosan to natural
biomaterials requires electrostatic interactions. Such hydrogels formed
by electrostatic interactions among polyelectrolytes are called poly­
electrolyte complex (PEC) hydrogels [115]. PEC hydrogels based on
non-covalent interactions have self-healing properties and simple
preparation processes, which have attracted extensive attention of re­
searchers. However, the mechanical properties of PEC hydrogels are
unsatisfactory. The improvement of mechanical properties of chitosan-
based PEC hydrogels can be achieved by changing the synthesis condi­
tions, such as crosslinking time and polysaccharide concentration.
Additionally, different pH conditions also have certain effects on the
strength of PEC supramolecular hydrogels due to the hydrogen bonding
among chitosan molecules and the electrostatic interactions between
chitosan and other polyelectrolytes. A study showed that the degrada­
tion rate of PEC hydrogel based on chitosan-HA at pH 5 (80 %) was
significantly higher than that at pH 7 (<60 %), which was caused by
different electrostatic interactions between chitosan and HA at different
pH conditions [115]. Other natural biomaterials, such as proteins and
peptides, also need further development to form high-strength supra­
molecular hydrogels with chitosan. Moreover, the dual crosslinking
strategy is also an effective method to enhance the mechanical proper­
ties of PEC supramolecular hydrogels [118]. On the one hand, complex
supramolecular network structures can be formed among polymers with
potential dual crosslinking ability. On the other hand, the mechanical
properties of chitosan supramolecular hydrogels can also be enhanced
by secondary crosslinking. Common dual crosslinkers include Ca2+,
J. Xu et al.
β-glycerophosphate solution, methacrylates, and photocrosslinking
agents [119,120].
5.4. Chitosan-based synthetic biomaterials
5.4.1. Modification of chitosan
Compared with natural biomaterials, synthetic biomaterials have
relatively poor biocompatibility and biodegradability, but they possess
excellent mechanical properties and can be modified or functionalized
to obtain specific physicochemical characteristics [121]. Liu et al. [122]
performed phenol-functionalization of chitosan, and the phenol-
functionalized chitosan hydrogels exhibited fast gelling rate, excellent
self-healing properties and superior light-crosslinking ability, thus
further enhancing the mechanical properties of chitosan hydrogels. Shen
et al. [123] synthesized the photocurable chitosan by N-acylation of the
amino group of chitosan with methacrylic anhydride (MA), which could
be printed by digital light processing. It is worth noting that the cross­
linking between chitosan and MA did not use any chemical cross-linking
agents and chitosan obtained good water-solubility and UV-crosslinking
properties due to the introduction of MA. To ensure that the chitosan-
MA hydrogels had sufficient mechanical strength to stabilize its own
structure, this study explored the effect of the degree of substitution (DS)
of MA and the concentration of chitosan-MA hydrogels on the me­
chanical properties. The results showed that with the increase of DS, the
compressive modulus and stress at rupture of chitosan-MA hydrogels
increased significantly from 110 kPa to 315 kPa and from 18 kPa to 82
kPa, respectively. With the increase of chitosan-MA concentration, the
compressive modulus also increased significantly (from 315 kPa to 910
kPa). Zhang et al. [124] prepared a maleic chitosan (MCS) which was
highly substituted by acrylate groups, and then introduced thiol-
terminated poly (ethylene glycol) (TPEG) into the system to form a
novel chitosan hydrogel. TPEG, with a highly reactive thiol group, has
been extensively studied due to its biocompatibility and applicability
[125]. The results showed that the step-chain photopolymerization of
MCS and TPEG could effectively enhance the mechanical strength. With
the increase of thiol/acrylate molar ratio, the fracture length and elastic
modulus of chitosan hydrogels increased to 0.60 ± 0.04 MPa and 10.1
kPa, respectively, which was related to the complex supramolecular
interaction between MCS and TPEG molecules. In vitro degradation
experiments showed that the degradation rate of chitosan hydrogels was
closely related to the content of thiols, showing tunable biodegrad­
ability. With increasing thiol content, the degradation rates of MT1
(thiol: acrylate = 0.012:1), MT2 (thiol: acrylate = 0.024:1) and MT3
(thiol: acrylate = 0.048:1) were 83 %, 79 % and 60 %, respectively, after
11 days.
In general, modification of chitosan can produce biomaterials with
specific physicochemical properties. Modified chitosan can be combined
with specific materials by non-covalent interaction to obtain supramo­
lecular hydrogels with tunable mechanical properties. The tunability of
the mechanical properties of biomaterials is usually related to the den­
sity of crosslinks and the concentration of polymers [126]. The photo­
curing strategy can promote the rapid gelation of hydrogels and form
supramolecular hydrogels with high strength. However, the effects of
photocrosslinkers and UV light on cells need to be evaluated.
5.4.2. Double-network chitosan hydrogels
Besides modifying or functionalizing chitosan, combining chitosan
with some synthetic materials to form double-network (DN) hydrogels is
also an effective strategy to enhance mechanical properties. DN hydro­
gel is a tough network formed by the interaction between rigid network
and flexible network [127]. Under the action of stress, the rigid network
will produce certain fracture to dissipate energy, while the flexible
network maintains elasticity to keep the stability of the structure [128].
Zhang et al. [129] successfully prepared a dual physically crosslinked
DN hydrogel. The DN hydrogel was formed by the electrostatic inter­
action in the chitosan-citrate (CS-Cit) network and the ionic interaction
553
International Journal of Biological Macromolecules 219 (2022) 545–557
and hydrogen bonding in the poly (sulfobetaine-co-acrylic acid) (P
(SBMA-co-AAc) network (Fig. 5b). This study explored the effect of the
hydrogel composition on the mechanical properties and self-healing
properties. The results showed that supramolecular hydrogels with the
best mechanical strength and self-healing properties were obtained with
SBMA content of 3 M, AAc content of 5 mol%, and soaking time of 8 h
(Fig. 5a). At 85 % strain, the compressive stresses of the original
hydrogel and the self-healing hydrogel were 0.32 MPa and 0.28 MPa,
respectively, indicating that the hydrogel had excellent self-healing
properties (Fig. 5c–d). The hydrogel also exhibited high stretchability
(800 %) due to dynamic electrostatic interactions and hydrogen
bonding. Additionally, the hydrogel had good electrical conductivity
and reliable relative resistance change, which could be used to prepare
hydrogel sensors for detecting human motion.
It has been found that some synthetic polymers such as polyvinyl
alcohol (PVA) [130], polyethylene oxide [131], and poly­
vinylpyrrolidone [132] can significantly enhance the mechanical prop­
erties of natural polymer-based hydrogels. PVA-based hydrogels can be
prepared by physical methods without using chemical crosslinkers.
Moreover, PVA has great potential in biomedical applications due to its
ease of production, excellent biodegradability, biocompatibility and
mechanical properties. Liu et al. [133] designed a DN hydrogel scaffold
based on chitosan and PVA. DN hydrogels were considered to be one of
the most promising tough hydrogels. The mixing of chitosan and PVA
could form the first hydrogel network. Then genipin was added to the
chitosan-HA solution to further increase the viscosity of the solution.
Finally, the freeze-thaw cycles of chitosan-PVA hydrogels formed the
second hydrogel network of PVA crosslinking. With the change of the
mass ratio of chitosan and PVA, the rupture tensile stress of chitosan
hydrogels increased to 115 kPa. The freeze-thaw cycles of PVA formed a
dense crosslinked network, which reduced the swelling ratio of the
hydrogel and ensured the structural integrity and stability.
Different from the above preparation process, Pan et al. [134]
designed an ultrahigh strength chitosan-PVA hydrogels. The method
first blended chitosan and PVA solutions and printed the prescribed
structures by direct ink-writing technology. Then the cyclic free­
zing–thawing was performed. Finally, the structure was soaked in so­
dium citrate solution. The prepared chitosan-PVA hydrogels formed a
DN structure composed of crosslinking of PVA and chitosan ionic in­
teractions between amino and carboxyl groups. This study examined the
effects of freeze-thaw cycles and the concentration of Na3Cit aqueous
solution on the mechanical properties of hydrogels. The results showed
that the crosslinking density of the hydrogel increased with the increase
of the Na3Cit aqueous solution concentration under one freeze-thaw
cycle. At a strain of 302.27 ± 15.70 %, the hydrogel exhibited a ten­
sile strength of 12.71 ± 1.32 MPa and a Young's modulus of 14.01 ±
1.35 MPa. Furthermore, the hydrogel exhibited wonderful fatigue
resistance and self-healing due to dynamically reversible ionic in­
teractions and hydrogen bonding.
5.4.3. Chitosan hydrogels based on poly(ε-caprolactone) scaffolds
Poly (ε-caprolactone) (PCL) has been widely used in the preparation
of bone scaffolders due to its high mechanical strength, high thermal
stability and low melting temperature [135]. However, the biomedical
applications of synthetic materials are limited due to their lack of
cellular recognition sites and hydrophobicity [136]. Natural bio­
macromolecules (e.g., chitosan, cellulose) have excellent biocompati­
bility, which can interact with cell surface receptors and perform
specific functions. Based on this, Liang et al. [137] combined porous PCL
scaffolds with chitosan hydrogels to form composite scaffolds that could
be used for bone tissue engineering. The results showed that with the
gradual degradation of chitosan hydrogels, the bioactive molecules
encapsulated by chitosan hydrogels were gradually released, which
effectively promoted the regeneration of bone tissue. In addition, the
compressive strength of the composite scaffold was about 6.7 MPa
(similar to the mechanical strength of bone tissue), indicating that the
J. Xu et al. International Journal of Biological Macromolecules 219 (2022) 545–557

Table 3 nanomaterials to achieve cartilage regeneration. TFNA could effectively

High-strength chitosan hydrogels. absorb specific bioactive molecules and promote their proliferation and
Strategies Hydrogel Types of 3D Applications Ref. differentiation, thus realizing the repair and regeneration of cartilage
compositions printing defects.
Nanomaterials Chitosan and GO Extrusion- Tissue [93] The combination of chitosan hydrogels with PCL scaffolds is also an
based engineering effective approach to enhance mechanical properties. On the one hand,
Chitosan and CNF Extrusion- Repair of [95] natural biomacromolecules endow the composite scaffold with excellent
based intervertebral biocompatibility and promote cell proliferation and differentiation. On
discs, cartilage
and meniscus
the other hand, PCL endowe the composite scaffold with wonderful
Chitosan, SF, and Extrusion- Bone [96] mechanical properties. Besides PCL, hardystonite (HT) is also a potential
CNPs based regeneration biomaterial, and the combination of chitosan and HT will also be an
Chitosan and Extrusion- Biomedical [97] ideal bone replacement material [139]. Table 3 summarizes the current
oxidized CNF based implants
chitosan-based high-strength supramolecular hydrogels.
Chitosan and CNCs Extrusion- Bone tissue [98]
based engineering
scaffolds 6. Conclusion and prospects
Titanium alloy Chitosan and GT Extrusion- Artificial joint [102]
materials based replacement Chitosan-based supramolecular biomaterials have been used as ideal
N-CS, ADH, and Electron Bioactive [105]
HA-ALD beam interface
bioink for structural printing due to their high similarity to natural ECM
melting in various properties. However, unsatisfactory mechanical properties
Natural PEG, chitosan, Extrusion- Re-establishing [119] are the main challenge for chitosan-based supramolecular hydrogels as
biomaterial α-CD, and GT based complicated bioink. This review focuses on describing strategies to enhance the
tissues and
mechanical properties of chitosan-based supramolecular hydrogels from
organs
Chitosan and PEC Extrusion- Bioscaffolds [116] the perspective of materials and supramolecular binding motifs. The
based combination of chitosan-based supramolecular hydrogels with nano­
(pneumatic) materials with excellent mechanical properties and titanium alloy ma­
Chitosan and Extrusion- Tissue [114] terials can not only effectively enhance the strength, but also endow the
alginate based engineering
hydrogel with nano-characteristics or compensate for the defects of ti­
scaffolds
Chitosan and HA Extrusion- Drug release [115] tanium alloy materials. In addition, the combination of chitosan with
based different natural and synthetic materials can also significantly improve
Chitosan and poly Extrusion- 3D bioprinting [113] the mechanical properties, such as the formation of PEC hydrogels, the
(gamma-glutamic based
modification of chitosan, and the formation of DN hydrogels.
acid)
Chitosan and PEC Extrusion- Bioscaffolds [140] However, the field still faces some challenges. As mentioned above,
based and wound supramolecular hydrogels have fascinating dynamic properties, but few
healing non-covalently bound polymers have been explored, so further explo­
Synthetic Phenol- Extrusion- 3D bioprinting [122] ration of biomaterials with strong interactions is still needed. Moreover,
biomaterial functionalized based
how to balance the dynamic properties and mechanical properties of
chitosan and
dibenzaldehyde- chitosan-based supramolecular hydrogels is also a key point. If the
terminated hydrogel is too dynamic, it will degrade too quickly in vivo or in vitro,
telechelic poly but high viscosity hydrogels will affect 3D bioprinting. Currently, most
(ethylene glycol).
chitosan-based supramolecular hydrogels are applied in extrusion-based
Chitosan and MA Laser-based 3D bioprinting [123]
(DLP)
3D bioprinting. Considering that light-based 3D bioprinting can print
MCS and TPEG Extrusion- Tissue [124] high viscosity bioink, one promising field is the development of cell-
based engineering friendly photocrosslinkers, thereby avoiding the harm of photo­
and drug crosslinkers to the human body. Of course, these biocompatible photo­
delivery
crosslinkers can also be used for secondary crosslinking. Although some
Chitosan-Cit and P Extrusion- Strain sensors [129]
(SBMA-co-AAc) based progress has been made in chitosan-based supramolecular hydrogels for
Chitosan and PVA Extrusion- Drug release [133] 3D bioprinting, little is known about their effects on cell signal trans­
based and tissue ductions and the human immune system. Furthermore, materials for 3D
regeneration
bioprinting cannot change over time or as the environment changes.
Chitosan and PVA Extrusion- Engineering, [134]
based intelligent
Therefore, the further development of chitosan-based supramolecular
machine, and biomaterials combined with 4D printing is a promising direction, which
soft robotics will bring unprecedented surprises to the preparation of biomaterials
Chitosan and PCL Extrusion- Bone tissue [137] that respond to time or environmental changes.
scaffolds based engineering
Chitosan, PCL Extrusion- Repair and [138]
scaffolds, and based regeneration of Declaration of competing interest
TFNA cartilage
defects The authors declare that they have no known competing financial
Chitosan and HT Extrusion- Bone [139] interests or personal relationships that could have appeared to influence
based replacement
material
the work reported in this paper.

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