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Desai, N.; Rana, D.; Salave, S.; Gupta, R.; Patel, P.; Karunakaran, B.; Sharma, A.; Giri, J.; Benival, D.; Kommineni, N. Biomedical Applications of Chitosan. Encyclopedia. Available online: (accessed on 19 June 2024).
Desai N, Rana D, Salave S, Gupta R, Patel P, Karunakaran B, et al. Biomedical Applications of Chitosan. Encyclopedia. Available at: Accessed June 19, 2024.
Desai, Nimeet, Dhwani Rana, Sagar Salave, Raghav Gupta, Pranav Patel, Bharathi Karunakaran, Amit Sharma, Jyotsnendu Giri, Derajram Benival, Nagavendra Kommineni. "Biomedical Applications of Chitosan" Encyclopedia, (accessed June 19, 2024).
Desai, N., Rana, D., Salave, S., Gupta, R., Patel, P., Karunakaran, B., Sharma, A., Giri, J., Benival, D., & Kommineni, N. (2023, July 04). Biomedical Applications of Chitosan. In Encyclopedia.
Desai, Nimeet, et al. "Biomedical Applications of Chitosan." Encyclopedia. Web. 04 July, 2023.
Biomedical Applications of Chitosan

Chitosan, a biocompatible and biodegradable polysaccharide derived from chitin, has surfaced as a material of promise for drug delivery and biomedical applications. Different chitin and chitosan extraction techniques can produce materials with unique properties, which can be further modified to enhance their bioactivities. Chitosan-based drug delivery systems have been developed for various routes of administration, including oral, ophthalmic, transdermal, nasal, and vaginal, allowing for targeted and sustained release of drugs. Additionally, chitosan has been used in numerous biomedical applications, such as bone regeneration, cartilage tissue regeneration, cardiac tissue regeneration, corneal regeneration, periodontal tissue regeneration, and wound healing. Moreover, chitosan has also been utilized in gene delivery, bioimaging, vaccination, and cosmeceutical applications. Modified chitosan derivatives have been developed to improve their biocompatibility and enhance their properties, resulting in innovative materials with promising potentials in various biomedical applications.

chitosan drug delivery tissue regeneration wound healing

1. Drug Delivery Applications

Drug delivery is a crucial aspect of modern medicine as it determines the efficacy and safety of a therapeutic substance. Targeted drug delivery within the body is a complex process that requires overcoming several barriers, such as the removal of drugs by the liver/kidneys, rapid degradation in the bloodstream, and limited permeation across biological membranes. A suitable carrier or vehicle is necessary to protect the drug from degradation, increase its circulation time, and enhance its localization at the target site. The ideal drug delivery vehicle should possess several key attributes, such as biocompatibility, biodegradability, and controlled release characteristics [1]. Biocompatibility ensures that the delivery vehicle does not produce adverse effects on the body, while biodegradability ensures that the delivery vehicle can be safely metabolized and eliminated from the body after its function is complete. Controlled release properties ensure that the drug is released in a controlled manner over a defined period of time, allowing for sustained therapeutic levels of the drug and reducing the frequency of dosing. Chitosan has shown great promise for drug delivery applications due to its unique properties. On account of its outstanding biocompatible and biodegradability, this polymer has been classified by the FDA as “Generally Recognized as Safe” (GRAS) and is approved for use in tissue engineering and drug delivery applications [2]. The LD50 of chitosan in oral administration to mice is more than 16 g/kg, and results from multiple acute toxicity studies have established chitosan to be safe for systemic use [3]. The repeating glycosidic residue in chitosan has a primary amino group that acts as a site for chemical bonding and gives the molecule a positive charge. This positive charge gives chitosan pH sensitivity, allowing it to form ionic complexes with drugs and control drug release. One of the key benefits of chitosan is its versatility, as it can be modified into various dosage forms, each with its own unique properties and applications.

1.1. Oral Delivery

Oral drug delivery involves the administration of medication through the mouth, usually in the form of tablets, capsules, or liquids, to be absorbed by the digestive system and eventually into the bloodstream. It is the most frequent and convenient routes of drug administration, as it is non-invasive and easily accessible. It is often the most practical and preferred route for patients, as it can be self-administered and does not require healthcare professionals. Additionally, oral delivery permits for the sustained release of medication over prolonged time, which can increase patient compliance and improve therapeutic outcomes. The extensive mucosal surface area of the gastrointestinal tract facilitates drug absorption, making it an exceedingly favored route for administering drugs. Many drugs cannot be administered orally owing to their susceptibility to degradation by gastrointestinal secretions, including enzymes, and the variable pH conditions. Physiological factors like transit time, blood perfusion, the presence of food, and limited absorption windows for several drugs can further hinder their absorption when administered orally [4]. Chitosan possesses favorable properties for use in oral drug delivery systems. Its cationic properties allow it to interact easily with negatively charged cell membranes, enhancing cellular uptake. Unlike many water-insoluble polymers, chitosan can partially dissolve in water, forming a hydrogel in aqueous environments. This unique quality enables it to adhere to mucosal surfaces, making it an excellent choice for oral drug delivery. Additionally, some chitosan derivatives, such as glycol chitosan solution or glycol chitosan-based nanoparticles, can inhibit the P-glycoprotein efflux pump (increasing the likelihood of absorption into the bloodstream) [5]. Some interesting applications of chitosan-based systems for oral delivery are discussed below.
N-cinnamyl substituted O-amine functionalized chitosan (cinnamyl-chitosan) was examined as a novel excipient by Ren et al. [6]. Directly compressed tablets were prepared using microcrystalline cellulose (as binder/filler), magnesium stearate (as a lubricant), acetaminophen (as drug substance), and either chitosan or the modified cinnamyl-chitosan. As the contents of other ingredients were kept the same for all formations, any difference in terms of flowability, compression behavior, and disintegration characteristics of the tableting blends was due to the chitosan type. While the plastic behavior of the 20% cinnamyl-chitosan blend was similar to the control group, their larger particle size provided them with higher elasticity, as evident by the result of the force-displacement profile analysis. This resulted in easy deformation of cinnamyl-chitosan, facilitating high compaction of the blend at relatively low levels of compressive pressures. With bigger particle sizes, cinnamyl-chitosan mixtures display good flowability and compaction characteristics, devoid of tableting defects. In comparison to MCC, they also exhibit good acetaminophen release properties as excipients. In addition to superior mechanical strength and drug delivery performance, the chemically modified cinnamyl-chitosan had enhanced antibacterial and antioxidative effects, presenting it as a possible alternative excipient for direct compression. In a similar study, Yasufuku et al. utilized modified low molecular weight chitosan’s antioxidant properties to create chitosan tablets for oral consumption with extended-release capabilities. The chitosan exhibited scavenging activity in response to hydroxyl and superoxide radicals’ oxidative stress, which allowed it to endure breakdown and gradually release theophylline (a model drug) over a duration exceeding 12 h in a controlled manner [7].
Electrospinning is a useful technique for creating quick-dissolving oral films, but producing nanofibers from pure chitosan can be difficult owing to its polycationic properties in solution, rigid structure, and intra-molecular interactions. Qin et al. inspected the possibility of improving the electro spinnability of chitosan by blending it with pullulan, an extracellular polysaccharide derived from Aureobasidium pullulans, which can increase solution viscosity, reduce conductivity, and lower surface tension. To prepare the chitosan/pullulan blend for electrospinning, separate solutions of pullulan and chitosan were made by dissolving their powders in 50% (v/v) aqueous acetic acid at various concentrations. The blend was then created by mixing these solutions in varying ratios while keeping the total polymer content fixed at 10% w/v. After stirring for 12 h at room temperature and degassing for an additional hour, the solutions were electrospun. The ratio of chitosan to pullulan had an impact on both the properties of the solution and the morphology of the resulting nanofibers. Specifically, as the amount of chitosan increased, the viscosity and conductivity of the solutions also increased. Scanning electron microscopy (SEM) revealed that the diameter of the nanofibers reduced initially followed by an increase. Hydrogen bond interactions between chitosan and pullulan molecules were indicated by Fourier transform infrared spectra, whereas X-ray diffraction analysis confirmed that the electrospinning process reduced the overall crystallinity of the materials. Finally, wetting studies demonstrated that the material completely dissolved within 1 min, and aspirin encapsulation tests confirmed the potential use of the material in the oral mucosal release [8].

1.2. Ophthalmic Delivery

Ocular drug delivery presents a significant challenge in pharmaceutical research, mainly due to the unique pharmacokinetic and pharmacodynamic properties of the eye. The eye’s distinctive structure restricts the entry of drug molecules, making it hard to maintain an effective drug concentration at the required site of action. To achieve therapeutic concentrations of active agents in oral treatment for ocular disorders, substantial doses of active agents are required which may cause severe side. In addition, systemic access is impeded by the blood–retinal, blood–aqueous, and blood–vitreous barriers [9]. Therefore, topical administration of ophthalmic dosage forms remains the only practical option. Formulations like drops, gels, and creams are available to treat surface disorders (such as conjunctivitis, blepharitis, and keratitis sicca) as well as to deliver intraocular therapy through the cornea (for diseases such as glaucoma or uveitis) [10]. However, poor pre-corneal retention, poor penetration across the cornea, and efficient drug elimination mechanisms, such as tear drainage, protein binding, and induced tear production, limit the ocular bioavailability of topical preparations to less than 5% [11]. Attempts have been made to address the issue of poor bioavailability and the consequential repeated drug administration by focusing on enhancing corneal residence time by employing viscosity enhancers, mucoadhesive polymers, and in situ gel-forming systems. Chitosan inherently possesses all these attributes. Its biocompatibility, anti-bacterial nature, pH neutrality, and ability to undergo degradation in response to lysozyme (also known as muramidase which is an enzyme found in tears that is part of the innate immune system) make it a potential candidate to develop controlled release ocular platforms [12].
Nanoparticles are the most extensively studied among chitosan-based delivery systems for the eyes. They are typically produced through ionic gelation, in which tripolyphosphate (TPP) is employed as a cross-linking agent. During this process, the positively charged amino groups are utilized as they react with the negatively charged TPP groups, resulting in a decrease in mucoadhesive capability [13]. Dubashynskaya et al. reported mucoadhesive cholesterol–chitosan self-assembled particles for topical ocular delivery of dexamethasone (Dex). To synthesize these particles, the researchers utilized a carbodiimide-mediated coupling reaction with a succinyl linker to create cholesterol–chitosan conjugates. They then optimized the synthesis by adjusting the molar ratios of reactants to produce cholesterol–chitosan conjugates with varying degrees of substitution (ranging from 1.2 to 5.8%). The resulting submicron particles had hydrodynamic diameters of 700–900 nm and a ζ-potential greater than 30 mV, which provided enough repulsive force to ensure optimal physical colloidal stability and superior mucoadhesive properties. The Dex-loaded particles were determined to be non-toxic, had membrane-stabilizing properties, and demonstrated sustained anti-inflammatory activity [14].
Rather than preparing entire particles out of chitosan, its solution can be added to other nanoparticles that have actually been made or are in the process of being developed to export some of its advantages [15]. Eid et al. developed chitosan-coated niosomes for enhancing the precorneal residence period, eye permeation, and bioavailability of azithromycin (for the treatment of bacterial conjunctivitis). Drug-loaded niosomes were prepared through modified thin-film hydration strategy, followed by coating with chitosan solution. The optimized particles had a mean diameter of 376 nm, entrapment of 74.2%, the surface charge of 32.1 mV, and a muco-adhesion force of 3114 dyne/cm2. Compared to commercial drops, the platform displayed a three-fold increase in azithromycin concentration in the rabbit eyes, attributed to the enhanced permeability coefficient of chitosan-coated niosomes and sustained release of entrapped drug [16].
Superior therapeutic outcomes are observed with in situ hydrogels and film/membrane-based ocular delivery systems. When instilled into the eye, a typical in situ gelling system is initially in a liquid state. However, in response to various stimuli, it rapidly transforms into a viscoelastic gel within the conjunctiva sac of the eye. This results in a significantly prolonged residence time of the gel, which gradually releases the encapsulated drugs over an extended period of time. As a result, this sustained release provides improved bioavailability of the drug and reduces the frequency of administration. Various types of in situ gelling systems have been proposed as novel ophthalmic formulations, including those that are temperature-, pH-, and ionic-activated [17].

1.3. Transdermal Delivery

The skin, being the body’s largest and most expansive organ, is a highly advantageous avenue for administering numerous medications. Compared to intravenous and oral delivery methods, transdermal administration offers a variety of benefits, including avoidance of first-pass metabolism, consistent blood levels, and both localized and systemic delivery. It is also suitable for treating a wide range of conditions, such as chronic pain, hormonal imbalances, and cardiovascular diseases [18]. However, the effectiveness of transdermal delivery is often impeded by the stratum corneum. It is composed of keratin filaments enclosed in a cornified envelope and covered by a multilamellar lipid bilayer, which acts as a physical barrier to drug molecules. Despite the commercial availability of several transdermal treatments (such as patches, ointments, spreads, and sprays), drawbacks like poor bioavailability, low drug penetration, undesirable side effects (such as skin irritation), and poor patient compliance remain a concern [19].
With the aim to promote effective transdermal delivery, Shekh et al. developed a novel scaffold carrier by functionalizing polyacrylonitrile nanofibers with oxidized chitosan followed by grafting of antiviral drug acyclovir onto the nanofibers to achieve a sustained and controllable release profile. The modification of oxidized chitosan was verified by SEM, where the average diameter of nanofibers increased from 218 to 354 nm, with increased surface roughness. In vitro drug release study showed that the acyclovir nanofibers modified by oxidized chitosan exhibited a more sustained release curve (following the Korsmeyer-Peppas model with a Fickian diffusion mechanism) than acyclovir nanofibers without modification. This behavior may be explained by the formation of Schiff bases between the drug molecules on the nanofibers and the oxidized chitosan. 
Chitosan can be fabricated into nanoparticles having highly tunable size and surface charge, which enables them to have extraordinary specific surface area and great permeability in the skin. Several nanoparticle-based systems have been looked into for the treatment of skin-related ailments. Riezk et al. conducted an investigation into the potential application of chitosan nanoparticles as a drug delivery system for the management of cutaneous leishmaniasis. The researchers prepared two different types of chitosan nanoparticles: one that was positively charged using tripolyphosphate sodium and another that was negatively charged with dextran sulphate. When Amphotericin B (AmB) was loaded onto both types of nanoparticles, they demonstrated in vitro activity against Leishmania major intracellular amastigotes that was comparable to that of unencapsulated AmB. 

1.4. Nasal Delivery

Nasal delivery of drugs refers to the administration of medication through the nasal cavity. This method takes advantage of the rich blood supply present in the mucous membranes of the nasal cavity to facilitate the rapid absorption and distribution of drugs throughout the body. Historically, the nasal route has been utilized to administer topical formulations for the management of local diseases in the upper respiratory tract, like nasal allergies, infections, and congestion. More recently, the nasal route has been increasingly used for the systemic delivery of small molecular weight drugs to treat chronic diseases such as diabetes and obesity. In particular, researchers are exploring the potential of the nasal route for the management of chronic neurodegenerative conditions of the central nervous system (CNS), including Parkinson’s disease and Alzheimer’s disease, as well as for faster treatment of disorders such as migraine and convulsions [20]. The nasal route offers advantages such as avoidance of first-pass metabolism, elimination of needle phobia associated with the parenteral route, and possible self-administration, leading to improved patient convenience. However, the nasal route is not without its challenges. The high nasal secretion rate can limit the residence time of drugs in the nasal cavity, and the volume of an administered formulation is often restricted.

1.5. Vaginal Delivery

Apart from its role as a genital organ involved in reproduction, the vagina has also been identified as a potential route for drug administration. The vaginal lumen is susceptible to various pathological conditions such as bacterial, fungal, protozoal, and viral infections, which have traditionally been treated with localized administration of antimicrobial/antiviral drugs within the cervicovaginal region [21]. This route of administration has several advantages over oral administration, including avoiding first-pass metabolism, reducing gastrointestinal side effects, and ease of application. The development of any vaginal drug delivery system must take into account the unique microenvironment of the vagina, including its pH range (typically between 4.0 and 5.0), microbiota, and cyclical changes [22]. While vaginal administration of therapeutic agents for local infections has demonstrated comparable efficacy to oral administration, commercial products for topical vulvovaginal treatment have low patient acceptability as they are associated with discomfort and short drug retention time [23]. Therefore, the development of mucoadhesive systems is necessary to optimize the acceptability and therapeutic efficacy of such therapies [24]. Chitosan is a promising candidate for the development of vaginal drug-delivery systems due to its antimicrobial properties and muco-adhesion/mucopenetration feature. Its use in such systems may improve the retention time of drugs and increase patient compliance [25].
Mucoadhesive matrix tablets, which create an in situ gelling effect due to their interaction with the vaginal fluid, are among the most preferred vaginal formulations due to their ease of preparation, low cost, and high stability. Fitaihi et al. conducted a study to investigate the use of chitosan, in combination with other bioadhesive polymers such as hydroxypropyl methylcellulose, guar gum, sodium carboxymethyl cellulose, and polyvinylpyrrolidone, to develop vaginal tablets with sustained-release properties of fluconazole. The tablets were designed to release the drug in acidic vaginal fluid (pH 4.8) for prolonged periods. The team evaluated 17 different formulations with varying ratios of chitosan and the other polymers and selected two formulations for further study. The first formulation contained chitosan, guar gum, and polyvinylpyrrolidone in a ratio of 1:2:1, while the second was formulated with chitosan and sodium carboxymethyl cellulose in a ratio of 1:2. Both formulations demonstrated acceptable powder flowability and tablet characteristics, including desirable hardness and friability. The drug release profiles at pH 4.8 showed sustained-release characteristics for both formulations, and the tablets exhibited good bioadhesion to the mouse peritoneum membrane [26].

2. Biomedical Applications

2.1. Bone Regeneration

Bone regenerative therapy offers the potential to treat various skeletal disorders/injuries, such as fractures, osteoporosis, and congenital defects, which can cause chronic pain, disability, and reduced quality of life. Conventional treatments for these conditions, such as surgery and drugs, have limitations and often result in incomplete recovery. In this context, chitosan-based platforms can be employed to promote bone regeneration by direct delivery of growth factors/drugs to the site of injury or by providing a biomimetic template for the growth of new bone tissue. Chitosan promotes the production of extracellular matrix (ECM) components, such as collagen, which are essential for bone growth and repair. Additionally, its antimicrobial properties reduce the risk of infection, and its biodegradable nature makes it an attractive option for use in long-term applications [27]. While chitosan is a promising material for various medical applications, it is important to acknowledge its limitations when it comes to bone tissue engineering.
On account of their osteoconductivity, resorbability, and similarity to natural bone minerals, calcium phosphate (and its derivatives) is widely used in combination with other biomaterials for bone regeneration. Chen et al. synthesized three different types of calcium phosphates, namely calcium-deficient hydroxyapatite (CDHA), beta-tricalcium phosphate (TCP), and biphasic calcium phosphate (BCP) using a reverse emulsion method followed by calcination. They then compared the efficacy of these materials in promoting bone regeneration by blending them with chitosan to produce porous hybrid membranes. All membranes possessed interconnected pores with porosity and pore size ranging from 91 to 95% and from 102 to 147 µm, respectively. 
ECM-mimicking materials are useful in bone regeneration because they mimic the native ECM of bones, which provides a template for cells to organize and form new tissue. These materials are designed to have similar chemical composition and microarchitecture as the natural ECM. By doing so, they can create a supportive environment that promotes cell adhesion, proliferation, and differentiation, leading to enhanced bone regeneration [28]. Additionally, they also are designed to have specific mechanical properties and degradation rates that match the desired time course of bone healing. This can help to optimize the delivery of growth factors/drugs and reduce the risk of implant failure.

2.2. Cartilage Tissue Regeneration

Cartilage regeneration is a challenging area due to the cumulative existence of several factors. Firstly, cartilage is a complex structure that consists of chondrocytes, ECM components, and signaling molecules. This complexity makes it difficult to recreate the natural environment of cartilage in vitro, which is necessary for successful regeneration. Secondly, it has a limited blood supply, making it difficult for cells to access the nutrients and oxygen needed to survive and function. Avascularity also hinders the effective delivery of therapeutic agents to the site of tissue damage. Lastly, cartilage is subjected to high loads and repetitive motions, so it is tricky to find materials that have the appropriate mechanical properties while also being biologically compatible [29]. The therapeutic success of chondrocyte transplantation and/or the quality of neocartilage production is highly dependent on the cell-carrier material used. By tailoring the properties of chitosan and its derivatives, it is possible to address some of the challenges associated with cartilage regeneration. Several platforms have been reported in recent years which focus on addressing the inherently low regenerative capacity of cartilages by utilizing the ECM-mimicking approach or by functionalization with signaling molecules that enhance the chondrocyte’s proliferation and differentiation [30]. Building upon the success of traditional chitosan-based approaches, new innovations are emerging that offer even greater potential for cartilage regeneration.
Osteochondral tissue (commonly found in weight-bearing joints, such as the knee and ankle) combines articular cartilage and subchondral bone within a single structure. In case of damage, its repair is extremely complex owing to a biphasic structure. The use of multizonal scaffolds that mimic the gradient transitions from the cartilage surface to the subchondral bone can open new avenues for osteochondral regeneration. To address this, Pitrolino et al. combined freeze-drying and porogen-leach-out methods to produce a bioresorbable multi-layered chitosan scaffold with controlled porosity in distinct but integrated layers. The mean pore size for the cartilage phase, interphase, and subchondral phase were 160 ± 12.3 μm, 248 ± 33.2 μm, and 275 ± 31.7 μm, respectively. The authors reported the incorporation of 70% (by weight) nano-hydroxyapatite to provide additional strength to the bone-like layer. Under compressive loading, the scaffold demonstrated a prompt mechanical recovery and remained intact under tensile loading. Human mesenchymal stem cells showed successful attachment and growth on the scaffold, with cells adopting a typical adherent morphology on the bone layer and a rounded shape on the chondrogenic layer. In vitro studies revealed that mesenchymal stem cells differentiate into both osteogenic and chondrogenic cells in specific layers of the scaffold, influenced by the unique pore gradient and composition of the material [31].
Articular cartilage is a type of soft tissue that covers the ends of bones in synovial joints and provides a smooth, low-friction surface for movement. This type of cartilage plays a crucial role in absorbing shock and reducing friction between the bones in the joint. Approaches to regenerate it have been widely explored as the articular cartilage is prone to get damaged due to osteoarthritis or trauma (direct injuries to the joint, such as a fall or a sports-related injury). 

2.3. Cardiac Tissue Regeneration

The global burden of cardiovascular disease is a major health challenge, with heart disease being the leading cause of death worldwide. As the demand for heart transplants far outweighs the supply of suitable donor organs, there is a growing need for alternative approaches to treat or replace damaged heart tissue. Cardiac tissue engineering offers a promising solution by combining the principles of biology, materials science, and engineering to develop functional cardiac tissue. There are several strategies for cardiac tissue engineering, including cell injection, implantation of three-dimensional tissue constructs or patches, acellular material injection, and valve replacement [32]. To achieve physiological function, the materials used in these approaches must possess mechanical and electrical properties similar to those of native cardiac tissue, which is composed of cardiomyocytes, smooth muscle cells, and endothelial cells. Additionally, the materials must be designed in a way that promotes tissue integration and functional connectivity with the surrounding cells and blood vessels.

2.4. Corneal Regeneration

The cornea, a critical component of the eye, relies on corneal epithelial cells to maintain its transparency. Unfortunately, these cells are unable to regenerate in humans, which can result in vision loss due to factors such as aging, trauma, or disease. In extreme cases, a significant reduction of corneal epithelial cells can lead to blindness. According to the World Health Organization, the corneal disease is a leading cause of blindness globally, second only to cataracts [33]. Given the high demand for corneal tissue and the limited availability of donors, the field of corneal tissue engineering has become increasingly important for the treatment of ocular cells [34]. The cornea plays a crucial role in ensuring the clarity of vision but is also susceptible to injury. Currently, treatment for corneal injuries often involves a transplant of an amniotic membrane, but this approach carries the risk of infection and rejection. To address this issue, the use of biomaterial-based scaffolds or membranes has been widely explored [35]. Chitosan is a popular choice due to its biocompatibility and anti-inflammatory properties, but it has limitations, such as poor mechanical strength. To address this, chitosan can be blended with other polymers to enhance its scaffold characteristics. An ideal cornea tissue scaffold must have similar mechanical and optical properties to the natural cornea and must be able to support cells and maintain high adhesion. In addition, optical transparency is a critical requirement for corneal implants, so extra care must be taken to ensure that the materials and processing methods used to preserve this important characteristic [36]. The scaffold or membrane should also be flexible and resilient enough to withstand surgical manipulation. The subsequent part highlights some of the exceptional research work centered on chitosan and corneal regeneration.
The current treatment methods for corneal stromal defects often result in inadequate clinical outcomes due to the irregular shapes of the defects. To address this problem, Feng et al. created a new type of biomimetic scaffold within situ-forming properties. This was achieved through dendronizing chitosan with oligoethylene glycol-based dendrons, resulting in a novel type of dendronized chitosan (DC) with thermo-gelling properties. The DCs have specific radial amphiphilicity, which allows them to self-assemble into long fibers and form highly transparent hydrogels through thermal aggregation and gelation. The authors found that they could easily modulate the gelling points of the hydrogel by varying the grafting ratios of dendrons. When tested in physiological conditions (PBS at 37 °C), the DC hydrogel displayed enhanced mechanical strengths. The biocompatibility of the platform was demonstrated using rabbit corneal stromal cells, which showed that it could promote keratinocyte proliferation and migration. The DC hydrogel’s potential for corneal tissue engineering was further suggested by the migration of cells into the microporous material. In a corneal stromal defect model in rabbits, the in vivo performance of the platform was evaluated. Two weeks after implantation, the results of H&E showed that there was hardly any observed inflammatory response in the stromal layer of the cornea in the DC hydrogel group. This indicates that the hydrogel can fill the matrix while also reducing the inflammatory response. Four weeks after implantation, migration of resident keratocytes to the gel matrix was observed, suggesting that the platform is suitable for the growth of corneal stromal cells [37].

2.5. Periodontal Tissue Regeneration

The tissues that support teeth, known as the periodontium, are made up of the gingiva, cementum, periodontal ligament, and alveolar bone. An inflammatory disease called periodontitis leads to the deterioration of these tissues and eventually results in tooth lost. Currently, treatments for periodontitis center around removing plaque and controlling local inflammation through scaling and root planning and other surgical procedures. Although these treatments alleviate symptoms and slow the progression of the disease, they are unable to restore the connection between the periodontal tissues and teeth to their original state [38]. This means that even after treatment, the functionality of teeth and dentition remains impacted. Efforts have been made to regrow these tissues through methods such as bone grafts. However, the results of these approaches are inconsistent. Therefore, it is crucial to find alternative strategies for restoring the structure and function of periodontal tissues in periodontitis patients [39].
The process of tooth development involves niche-residing dental follicle cells transforming into cementoblasts, fibroblasts, and osteoblasts, which create cementum, periodontal ligament, and alveolar bone, respectively. However, once tooth development is complete, the niche that plays a crucial role in forming the supporting tissues is no longer present, making it challenging to repair damaged or lost periodontium. This is where biomaterial-assisted periodontal tissue engineering comes in, offering the possibility of recreating the microenvironment and regenerating functional tissues [40]. Chitosan is a favorable material for dental applications due to its bioactivity, natural antimicrobial properties, and biocompatibility. While it works well on its own, combining chitosan with compatible organic or inorganic materials can lead to improved results in periodontal regeneration [41]. This is because the periodontal microenvironment is complex and requires a multi-component platform to meet the varied compositional and mechanical needs for successful regeneration. The platform must also be able to regulate the formation of periodontal tissues with the proper structural features [42].

2.6. Wound Healing

Wounds are disruptions to the integrity of the skin resulted due to external physical/thermal injury or internal pathological conditions. They can be categorized into two types: acute and chronic. Acute wounds heal entirely in 8–12 weeks with little scarring; however, chronic wounds may reoccur and have a recovery duration that exceeds 12 weeks [43]. The healing process can be hindered by underlying physiological conditions, making it necessary to properly manage these wounds. Examples of chronic wounds comprise of venous leg ulcers, pressure ulcers (bed sores), diabetic foot ulcers, and non-healing surgical wounds. Skin layers and affected areas serve as the basis for the gradation of wounds. The epidermal skin surface is only involved in superficial wounds. Partial thickness wounds are injuries involving the epidermis, deeper dermal layers, blood vessels, hair follicles, and sweat glands. When the subcutaneous fat or deeper tissue, as well as the epidermal and dermal layers, are wounded, full-thickness wounds result [44].
Wound healing is a complex and multifaceted process that requires a conducive environment to support its acceleration. The body’s normal reaction to damage consists of a sequence of interconnected stages such as hemostasis, inflammation, proliferation, and remodeling that aim to restore the skin’s integrity. This process involves numerous cell types, enzymes, cytokines, proteins, and hormones working in concert. Following injury, hemostasis is triggered to form blood clots and constrict blood vessels, followed by the secretion of proinflammatory cytokines and growth factors that promote inflammation. Macrophages, neutrophils, and lymphocytes are then recruited to facilitate reepithelization and angiogenesis, leading to the proliferation of fibroblasts and keratinocytes and eventual differentiation of fibroblasts into myofibroblasts, which deposit extracellular matrix [45]. To aid the healing process, a proper wound dressing must be applied to protect the wound site from external mechanical and microbial stress. Traditional dressings like cotton, bandages, and gauze often fail to provide a moist and supportive environment and can cause pain upon removal due to wound drainage [46].
An ideal wound dressing must have several key properties to facilitate the healing process. Firstly, it should maintain a moist environment to allow for proper gas exchange and prevent dryness of the wound. Secondly, it should be able to absorb any exudates produced by the wound to maintain a clean and hygienic environment. Thirdly, it should support cell proliferation and inhibit bacterial growth by preventing infections and promoting healing [47]. Studies have shown that chitosan is a promising option for use in wound care. Its unique set of biological features, such as antibacterial, hemostatic, and mucoadhesive characteristics, make it an attractive option for wound dressings. 

3. Miscellaneous Applications

3.1. Gene Delivery

Gene delivery is the process of introducing genetic material, such as DNA or RNA, into cells to modify their function or behavior. Gene delivery has become a crucial tool in modern biotechnology and medicine as it allows for the targeted modification of cells and tissues, opening up new possibilities for disease treatment, gene therapy, and genetic engineering. It involves the use of gene delivery techniques to treat diseases caused by genetic mutations [48]. These mutations can be corrected by the introduction of normal genes into the patient’s cells, either to replace a defective gene or to supplement its function. Gene delivery therapy has shown great promise in the treatment of a wide range of genetic disorders, including cystic fibrosis, sickle cell anemia, muscular dystrophy, and various cancers. Challenges like susceptibility of gene degradation by nucleases in the bloodstream, lack of specificity towards targeted cells, and the inability of negatively charged genes to enter negatively charged cellular membranes make it impractical to directly deliver therapeutic genes (neither systemically nor locally) in the absence of a delivery vector [49].
Gene delivery vectors can be either viral or non-viral, each with its own advantages and limitations. Viral gene delivery vectors use modified viruses that have evolved to efficiently infect cells to deliver therapeutic genetic material. The have high transfection efficiency and provide stable gene expression, but their utilization is not preferred due to their significant safety concerns (unwanted inflammatory and immune responses, toxicity, immunogenicity, and substantial cost) [50]
The unique physiochemical and biological properties of modified chitosan derivatives have made them a popular option as non-viral vectors for gene delivery. Chitosan’s polycationic nature enables the formation of complexes with negatively charged nucleic acids via electrostatic interactions, providing protection from nucleases [51]. As cellular and nuclear membranes are also negatively charged, they interact with positively charged chitosan, allowing the complex to be taken up and relocated to the nucleus. Additionally, chitosan’s amino groups contribute to endosome escape through the “proton sponge effect”, which facilitates the intracellular release of the nucleic acid complex [52]. To enable efficient and effective gene delivery, critical chitosan characteristics must be tuned. First, for appropriate stabilization/protection and intracellular gene release after uptake, the optimal molecular range of chitosan should be between 65 and 170 kDa [53]. Secondly, the degree of deacetylation should be high as it translates to more primary amines, thereby increasing positive charge and eventually promoting transfection [54]. Lastly, the chitosan-based gene delivery system should have a high N/P ratio (fraction of chitosan nitrogen per gene phosphate). A higher N/P ratio improves the stability of the chitosan-gene complex and promotes better interaction with cells resulting in better transfection efficiency [55]. Keeping these considerations in mind, the subsequent section discussed some cutting-edge gene delivery platforms based on chitosan.

3.2. Bioimaging

Molecular imaging provides the capacity to assess biological and metabolic processes in live organisms without intrusive methods. Many non-invasive procedures have been developed for uses ranging from clinical diagnostics to cellular biology research and drug development since the advent of X-ray technology. Throughout clinical and preclinical drug research, these cutting-edge technologies have the potential to enhance our understanding of illnesses and therapeutic action. In contrast to traditional readouts such as immunohistochemistry, molecular imaging techniques may be done in the intact organism with an excellent spatial and temporal resolution, making them ideal for in vivo studies of biological processes [56].
Aggregation-induced emission (AIE) is a photophysical effect in which non-luminescent molecules in solution can become brightly luminescent upon aggregation owing to the restricted intramolecular motions. This means that when the molecules come together, the nonradiative decay pathways that usually cause low fluorescence quantum yields are suppressed, leading to an increase in radiative decay and a higher quantum yield of fluorescence [57]. Several chitosan-based systems working on the AIE effect have been explored for biological imaging. Shi et al. developed a redox-responsive polymeric nanocarrier with AIE-mediated bioimaging ability. The system was fabricated from a biotinylated chitosan-modified amphiphilic polymer, wherein the hydrophilic chitosan was associated via a hydrophobic tetraphenylethylene (responsible for AIR) unit through a disulfide bond. The polymer self-assembles into a nanosphere (TPE-bi(SS-CS-Bio)). The presence of biotin enhanced the nanocarrier’s cellular uptake, and subsequent exposure to a high level of glutathione caused rapid disassembly and release of constituents. AIE feature allowed tracking upon distribution into cells with a time-dependent increase in observed fluorescence [58].

4. Other Applications

4.1. Vaccination

Chitosan derivatives, specifically trimethyl chitosan, are widely hypothesized to function as vaccine adjuvants. It can activate the innate immune system through the stimulation of Toll-like receptors and other pattern-recognition receptors. This results in the production of cytokines and chemokines that promote the recruitment of immune cells to the site of antigen exposure. Additionally, it can modulate the immune system by promoting a Th1 response and inhibiting a Th2 response. This leads to a stronger cellular immune response and a reduced risk of allergic reactions [59].
In one study, Wang et al. explored the use of chitosan hydrochloride salt stabilized emulsion as a vaccine adjuvant. Since conventional emulsion adjuvants are stabilized by non-ionic surfactants, their electroneutrality limits the loading efficiency of negatively charged antigens. The chitosan salt facilitated the loading of negatively charged Ovalbumin (adsorption rate of up to 97.99 ± 0.33%) and significantly enhanced humoral immunity by boosting recognition/uptake by APCs (via charged interaction) [60]. Chitosan-based systems have also been widely investigated as vectors for intranasal vaccine delivery. Mosafer et al. developed alginate-coated chitosan or trimethyl chitosan nanoparticles loaded with inactivated PR8 influenza virus (via direct coating) for nasal immunization. When tested in BALB/c mice, PR8-trimethylchitosan-alginate formulation elicited a higher ratio of IgG2a/IgG1 antibody titer (which promotes Th1 immune response) for immunization against the influenza virus [61].

4.2. Cosmeceuticals

Cosmetics refer to a range of products and substances that are applied to any part of the body for the purpose of enhancing, maintaining, or altering one’s appearance. Although synthetic compounds are often used as an active ingredient in cosmetic products, their long-term use can lead to skin irritation, itching, phototoxicity, and photo allergy. Due to current international regulations and increased demand for eco-friendly products, the cosmetic industry has shifted towards “green cosmetics” and has been conducting extensive research to find natural ingredients to replace traditional petrochemical-derived ones [62]. Chitosan, owing to its natural source and biocompatibility, has attracted significant attention in the cosmetic industry due to its unique qualities such as a natural humectant and moisturizer, a rheology modifier, and a formulation stability enhancer. Additionally, chitosan reduces the requirement of preservatives. Of late, some interesting cosmetic applications of the chitosan-based system have been reported [63].


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