## 3 Chemical Modification of Hyaluronan
### Oxidation of Hyaluronan
The pro-inflammatory cytokines (TNF-\(\alpha\), IL-1\(\beta\), and IL-6) were down-regulated, while the expression of growth-promoting cytokines (TGF-\(\beta\)1, EGF, and VEGF) was up-regulated, which demonstrated angiogenesis in the diabetic mouse wound model [129]. [PERSON] et al. reported a Schiff base hydrogel combining chitosan grafted with chlorogenic acid (CS-CGA) and HA-di-CHO for encapsulating deferoxamine (DFO) for wound healing. The hydrogel showed tube formation, which confirmed angiogenesis and extended new vascularization in the tissue as confirmed by CD31 (platelet-endothelial cell adhesion molecule-1) immunostaining. The expressions of IL-1\(\beta\), IL-6, TNF-\(\alpha\), and C-reactive protein (CRP) in the hydrogel group were down-regulated by the hydrogel compared to a control group for both diabetic and bacterial-infected wounds. The expression of IL-10 was up-regulated demonstrating anti-inflammatory properties. The hydrogel promoted cell migration as confirmed by scratch assays in vitro on mouse fibroblasts (NIH-3T3) and HUVECs, showing antibacterial activity, and allowed a 96.5 \(\pm\) 1.5% wound closure after 14 days. The wound closure of bacterial-infected wounds treated was 68.5 \(\pm\) 2.4%, compared to 42.5 \(\pm\) 3.2% for the control group [130]. Unfortunately, the authors have not reported the Mw of oxidized HA. Figure 12: Hyaluronan oxidation effected at (**a**) hydroxyl of the position C6 in water or (**b**) DMSO [122]; (**c**) amidation and oxidation [127; 128]. (**d**) Oxidation at the skeletal mediated by NaIO\({}_{4}\) (several authors). [PERSON] et al. modified chitosan (1-3 \(\times\) 10\({}^{5}\) g/mol) with tann acid (TA) by ionic interactions and complexed it with HA-di-CHO for the preparation of injectable hydrogels. The oxidation of HA (4-7 \(\times\) 10\({}^{5}\) g/mol) was performed with sodium periodate. The two polymers crosslinked within 2 min of injection via a dynamically reversible Schiff base reaction under mild conditions. The hydrogel presented a compression strength (G') of 383 Pa, with swelling up to 1200%. The degradation of the hydrogel was faster at pH 7.4 than at pH 8.5. At pH 5.5 only 20% of the hydrogel remained after 24 h. The hemolysis rate of CS/OHA was determined to be 1.6% and increased to 3.3% in the hydrogels containing TA. The hydrogels stopped the bleeding as demonstrated in vivo in a rat liver injury model. A wound closure was observed after 13 days [71]. However, the Mw of oxidized HA was unreported to establish the biological effect of HA. [PERSON] et al. reported the oxidation of HA with NaIO\({}_{4}\) via ring opening of the polysaccharide at 50 \({}^{\circ}\)C for 24 h. Furthermore, the carboxyl moiety of HA was activated by EDC/NHS and reacted with dopamine for its polymerization. The derivative was crosslinked with carboxymethyl chitosan (CMCS, degree of carboxylation \(\geq\) 80%) and poly(vinyl alcohol). The reagents 1,1-diphenyl-2-picrylhydrazyl radical (DPPH), 2,2\({}^{\prime}\)-Azino-bis (3-ethylbenzothazoline-6-sulfonic acid) diammonium salt (ABTS), and 1,1-diphenyl- and 2-phenyl-4,4,5,5-tetramethylimidazololine-1-oxyl 3-oxide radical (PITO) were used to evaluate the antioxidant properties of the hydrogel. The adhesive properties of the material were demonstrated by the lap-shear strength test and showed a value of 105.45 \(\pm\) 1.47 kPa. The immunomodulatory capacity of the hydrogel was performed through flow cytometry on RAW 264.7 cells. The M1 polarization of macrophages was promoted to M2 after treatment with hydrogel. The release of NO from macrophages was reduced. The hydrogel showed antibacterial activity against _E. coli_ and _S. aureus_. An in vivo model of burn wound healing was tested, showing fast regeneration [72]. However, the Mw of native HA and modified HA were not reported in the work. [PERSON] et al. developed hydrogels composed of monoaldehyde HA and carbohydrate-modified gelatin (GL-CDH).