Tissue engineering and biomaterials are seen as important ways to develop regenerative grafts to repair bone defects. Tissue engineering aims to combine biomaterials, cells and bioactive molecules to restore or improve the biological functions of damaged or diseased tissues. In order to improve cell viability, attachment, proliferation and homing, osteogenic differentiation, vascularisation, host integration and load bearing, tissue engineering has investigated a variety of scaffold materials and found that chondroitin sulphate (CS) has potential efficacy in bone regeneration and repair. CS is a type of sulphated anionic acidic mucopolysaccharide with numerous biological properties. This article aims to review the biological properties of CS, its role in osteogenic repair and its application in bone tissue engineering.
Figure 1. Types of nanomaterials as drug delivery systems in bone repair and remodelling. (Mani Divya, et al.; 2024)
Biological Properties of Chondroitin Sulphate
Biological properties of chondroitin sulphate CS is composed of D⁃glucuronic acid (GlcA) and N⁃acetyl⁃D⁃galactosamine (GalNAc) linked by repeated β-1,3 glycosidic linkages, generally containing 50 to 70 disaccharide units, and are widely distributed in the extracellular matrix and cell surfaces of tissues. In organisms they are mostly found in the form of proteoglycans and are an important component of connective tissues such as animal cartilage, tendons, trachea, laryngeal bones and skin. According to the different positions of the sulphate group in N-acetyl-D-galactosamine, it can be divided into chondroitin sulphates O, A, C, D, E, etc., namely CS⁃O, CS⁃A, CS⁃C, CS⁃Ｄ, CS⁃E, etc. In addition to the variability in the position of the sulphate group, different sulphate groups also have significantly different biological activities. For example, the sulphated fucose residues of fucosylchondroitin sulphate have anti-inflammatory, haematopoietic, etc. activities. CS has important biological properties. Studies have shown that CS has the effects of promoting cartilage regeneration, anti-inflammatory and antioxidant effects. CS works in two main ways: firstly, its own charged ionic groups interact with other substances; secondly, it regulates biomolecules in the signalling pathway and triggers reactions in the body.
Since CS is a type of sulfated anionic acidic mucopolysaccharide, it has the potential to attract ions with negative charge. Its sulfonic acid and carboxylate groups can react strongly with positively charged groups, such as calcium ions, through ionic bonds to form covalent bonds, thereby separating calcium ions in the mineralization center and controlling the crystal growth during bone tissue calcification. In addition, studies have shown that CS can also covalently bind to proteins through ionic interactions, such as binding to various growth factors and components of the extracellular matrix (ECM), some of which may have sensitization and/or intolerance, which can trigger the body's immune response and give CS antioxidant properties, reducing bone destruction, chondrocyte death and matrix component decomposition.
- Regulating Signal Molecules
Studies have found that CS can act as a co-receptor for soluble ligands, participate in the Wnt signaling pathway (Wnts), interact with fibroblast growth factors (FGFs), transforming growth factor β (TGF-β), bone morphogenetic proteins (BMPs) and other cytokines, regulate cell morphogenesis, and participate in osteogenesis. Researchers applied CS to osteoarthritis (OA) and found that CS inhibited the activation and nuclear translocation of nuclear factor-κB (NF-κB) subunit 1 in chondrocytes and synovial cells. NF-κB is a key regulatory factor that regulates the expression of many genes involved in the pathophysiology of tissue inflammation and cell recruitment. It is inferred that CS can regulate inflammatory pathways, reduce the level of inflammatory factors, reduce chondrocyte damage, and thus regulate the remodeling of subchondral bone.
The role of chondroitin sulfate in promoting osteoblastic repair Bone formation is a series of complex biochemical development processes, which can be divided into three periods: osteoblast proliferation, extracellular matrix aggregation, and mineralization of bone tissue. In this process, ECM molecules play a key role in tissue growth and cell differentiation, and CS is an important component of ECM molecules. Studies have found that CS can promote bone repair and new bone formation through multiple pathways including regulating osteoblast differentiation, immunomodulation, promoting cell proliferation and biomineralization.
- Regulating Osteogenic Differentiation
Cytokine expression plays a key role in osteogenic differentiation. Research has shown that CS and BMP-2 synergistically promote osteogenic differentiation of stem cells and upregulate the expression of alkaline phosphatase (ALP), osteocalcin (OCN) and collagen type I (COLI). ALP is a biomarker of osteoblast matrix maturation and OCN is a vitamin K-dependent calcium-binding protein. Both are important components involved in osteogenic differentiation and ECM mineralisation. In addition, CS can also signal through the Smad1-Smad5-Smad8 pathway to increase the expression of Runt-related transcription factor 2 (Runx2). Runx2 is a key transcription factor in the osteoblast differentiation process and is important at the onset of osteogenic differentiation of mesenchymal and other stem cells. It acts on downstream signals to promote the expression of key osteoblast proteins. In addition, based on the paracrine studies of osteoblasts on osteoclastogenesis, it has been found that CS can down-regulate the expression of osteoclast differentiation factor (RANKL), reduce the occurrence of osteoclasts and promote new bone formation. From the above, it can be concluded that CS regulates osteogenesis by promoting osteogenesis markers and/or reducing the expression of osteoclast factors.
Studies have shown that immune regulation plays an important role in the osteogenesis process, and CS has significant immunoregulatory functions. It can regulate bone regeneration by the osteogenic differentiation of stem cells by reshaping the local immune microenvironment, inhibiting fibroplasia, and upregulating osteogenic markers. Immune cells enter the site of bone destruction immediately after bone injury and promote the initial stages of healing by recruiting helper cells to the injury site. There is evidence that activated T lymphocytes can promote the maturation of osteoblasts through the production of soluble factors (such as RANKL), which is beneficial to bone repair. In fact, following bone injury, an inflammatory response occurs. Studies have found that the accumulation of inflammatory cytokines can mediate oxidative stress damage, promote osteoclast proliferation, and increase bone resorption, thus leading to osteoporosis. It can be seen that if the incidence and development of inflammation can be reduced, the destruction of bone tissue can be reduced, thereby promoting bone regeneration. Studies have shown that CS can regulate the phenotypic conversion of macrophages from M1 to M2, promote the expression of repair cytokines such as interleukin (IL)-4, IL-10 and TGF-β, change the local immune microenvironment from inflammatory to anti-inflammatory, promote stem cell recruitment, adhesion and proliferation, upregulate the expression of osteogenic markers and achieve bone repair. In addition, CS effectively reduces the increase in levels of inflammatory factors such as metalloproteinase-9 (MMP-9), IL-1β, IL-6 and tumour necrosis factor α (TNF-α), reduces the accumulation of inflammation and helps to balance bone homeostasis]. Obviously, the reduction in the expression and/or activity of these inflammatory molecules may explain the mechanism by which CS exerts its immunomodulatory effect. However, immune-regulated osteogenesis is a very complex process that requires a local immune response to activate the expression of cellular osteogenic markers and new bone formation under stimulation of signals. The specific role of CS in this process needs to be further investigated.
- Regulate biomineralization
During bone regeneration, biomineralization is an important process of sclerotic tissue. The most common biomineralizers are calcium and phosphate, which combine with organic polymers to form apatite crystals to provide structural support for bones. As one of the non-collagenous proteoglycans, CS plays a vital role in regulating the biomineralization of cartilage and bone tissue. CS has the potential to accumulate ions and can induce cell biomineralization to promote osteogenesis.
- Promote cell proliferation
Bone repair is inseparable from osteoblast proliferation. Some scholars have found that CS activates the TGF-β/Smads pathway, induces an increase in intracellular calcium ion levels, affects the cell cycle, and has a proliferative effect on chondrocytes; CS can affect the fixation of growth factors and other cytokines, and interact with bone cells (such as osteoblasts and osteoclasts) through integrins or other specific receptors, directly or indirectly affecting the adhesion, migration, growth, proliferation and differentiation of these cells.
In addition to the above effects, CS also has selective protein binding resistance, which can reduce platelet adhesion, promote the adhesion of endothelial cells and mesenchymal cells, and then promote cell differentiation, which is beneficial to bone remodeling.
Application of Chondroitin Sulfate in Bone Tissue Engineering
As a biomolecule with rich resources and excellent performance, CS is widely used in the field of bone tissue engineering medicine because of its significant bone repair effect, especially in craniofacial and oral medicine. Bone tissue engineering uses scaffold materials to induce bone formation in surrounding tissue, or uses scaffold materials as carriers or templates for implanting bone cells or other drugs. Research shows that CS is widely used in cell scaffolds, surface coatings, bioadhesives, and drug delivery systems in various forms. The researchers prepared a composite scaffold combining CS with chitosan and nano-bioglass, and found that CS promoted the expression and biomineralization of COLI and enhanced ALP activity, confirming the potential of the composite scaffold to promote tissue regeneration. In addition, someone has prepared CS glycosaminoglycan scaffold hydrogel and studied its suitability for the delivery of recombinant human bone morphogenetic protein, confirming that it can regulate bone TGF-β1 and BMPs signals and mediate critical size bone Defect regeneration and enhanced osteoblast mineralization. In addition, some scholars introduced CS into calcium phosphate cement (CPC) and found that CS-CPCs accelerated the preferential adsorption of fibronectin, up-regulated the expression of osteopontin (OPN) and ALP in BMSCs, and improved BMSCs's adhesion, proliferation, and osteogenic differentiation. Some scholars have used chondroitin sulfate in ECM bioinks for cartilage regeneration, which provides an ideal chemical and mechanical microenvironment for chondrogenic differentiation of BMSCs. More and more studies have shown that CS can be used with polymer compounds and bioactive molecular substances to prepare biomimetic material composites, confirming the role of CS in osteogenesis and repair.
References
- Mani Divya, et al.; A Short Review on Chondroitin Sulphate and Its Based Nanomaterials for Bone Repair and Bone Remodelling Applications. J. Compos. Sci. 2024, 8(1), 6.
- Mikami T, Kitagawa H. Biosynthesis and function of chondroitin sulfate. Biochim Biophys Acta. 2013, 1830(10):4719-33.
