## 3 Hyaluronic Acid in Treating the IVD: Structure, Synthesis and Turnover
#### The Key Function of Hyaluronic Acid in Repairing the Intervertebral Disc  
The biochemical properties of the HA molecule are entirely dependent on its size [69; 71; 72]. Within tissues, the HA molecule interacts with various proteins comprising of hyaladherins such as CD44, RHAMM (CD168) (Figure 3). In contrast, inside the ECM, it interacts with N-terminals of proteoglycans such as aggrecan (ACAN), versican, brevican, neurocan and TNF\(\alpha\)-stimulated gene-6  
Figure 3: The anti-inflammatory role of hyaluronic acid (HA). HA molecule (on the top) is represented according to its international symbols by repeated units of glucuronic acid (GlcA-\(\beta\)1-3) and N-acetylglucosamine (GlcNAc-\(\beta\)1-4) disaccharides. The anti-inflammatory role of HA is mediated through reacting with its several receptors such as CD44 and receptor for HA-mediated motility (RHAMM) preventing the cascades signalled by inflammatory cytokines such as IFN\(\alpha\), TNF\(\alpha\) and IL1\(\beta\) through deterring the contact with their receptors.  
(TSG-6) protein [69; 73]. These fundamental interactions are essential for HA function in terms of cell-to-cell communication and signalling [65; 74]. The mechanical properties of HA can be attributed to its high molecular weight structure. HA is capable of absorbing water approximately 10-10\({}^{4}\) times its mass as it is negatively charged and is considered an osmotically active molecule [63; 64; 75; 76]. Due to its large volume while hydrated, it can fill large spaces, therefore absorbing shocks and acting like a lubricant [75; 77].  
HA-CD44 interactions mediate endocytic activity and activate Rho and Rac1 GTPases to regulate cytoskeletal organization [78]. CD44 activation also activates src-related tyrosine kinases to induce cell proliferation through NF-kB signalling [79]. RHAMM activates pp60c-src tyrosine kinase to modulate focal adhesions for RHAMM-mediated cell mobility [80]. Its ability to form large extracellular networks through cell-matrix binding inhibits cytokine binding to prevent downstream signalling (Figure 3) [75; 81]. Therefore, HMW HA is capable of inhibiting the signalling of inflammatory cytokines and matrix-degrading enzymes (TNF-\(\alpha\), IL-8, iNOS, agrecanase 2, matrix metalloproteinases (MMP)s) that were evident upon treating chondrocytes with HMW HA, which also resulted in inhibition of phagocytosis and macrophage activation [82; 83]. In tissue injury and remodelling, enzymes break down HMW HA resulting in fragments of LMW HA which can induce phosphorylation of p38 or p42 or p44 mitogen-activated protein kinase (MAPK) and NF-kB which are associated with toll-like receptor (TLR)4 signalling [63; 84]. Depending on the tissue type and the size of LMW HA fragments (f-HA)s, pro-inflammatory cytokines can be induced [63; 81]; therefore, angiogenesis and tissue remodelling is stimulated [69; 71]. It was discovered that human disc cells treated with 6-12 disaccharide fragments of HA can dramatically up-regulate catabolic enzymes such as IL-1\(\beta\), IL-8, IL-6, MMP1, MMP13, and cyclooxygenase (COX2). Furthermore, it was identified that TLR-2 mediated the role of f-HAs in stimulating the protein expression of IL-6, and both IL-6 and MMP-3 were supported by the MAPK signalling pathway [85].