Synthesis Of Novel Hyaluronic Acid Derivatives And The Evaluation Of Anti-inflammatory,Anti-cancer And Repairpromoting Activities

Inflammatory reaction is the stress defense response to pathogen infection and tissue damage,which plays an important role in maintaining the body's health.However,the inflammatory response also provides the Double-Edged Sword effects on the body,as a long-term chronic inflammation can lead to other diseases.Studies have shown that long-term chronic inflammation is an important cause of tumor development.Although many achievements have been made in the development of anti-inflammatory compounds and their biological formulations,due to the complexity of the mechanism of chronic inflammatory diseases,the response of drugs in the inflammatory site is still unsatisfactory and cannot reverse the progress of the disease.Current anti-inflammatory drugs in clinics showed diffiierence in the levels of side effects,which has become a problem for the prescription to chronic inflammation treatment.Hyaluronic acid(HA)is a macromolecular carbohydrate polymer synthesized endogenously.It is an important component of the extracellular matrix and is widely presented in the human eye,skin,joint cavity and synovial fluid.In recent years,because of its non-immunogenicity,non-toxicity,and its special physicochemical properties and physiological functions,HA has been widely used in the fields of arthritis treatment,ophthalmic surgery and regenerative medicine.It acts as the agents for joint lubrication,pain relief,molecular barrier,epidermal and intraluminal filling,and also showed biological effects to promote the repair of endogenous tissue damage.Therefore,the development of new HA derivatives with anti-inflammatory activity has become a new direction in the research of discovering highly effective and low toxicity antiinflammatory drugs.In this study,a series of HA derivatives were synthesized through chemical reaction modification using natural HA as raw materials.The synthesized HA derivatives were characterized via mass spectrometry(ESI-TOF-MS and MALDI-TOF-MS)and nuclear magnetic resonance(NMR)analysis techniques.In the meantime,HA derivatives(TFHA and HA-NP)obtained from two different synthetic methods were selected,respectively and their biological activities were explored through in vitro cell experiments and the rat arthritis model.The MALDI-TOF-MS analysis showed that the molecular weight distribution of HA derivatives obtained by optimized synthesis methods was relatively uniform,30-60 kDa,and mainly distributed in the range of 39-42 kDa.ESI-TOF-MS / NMR analysis showed that the percentage modifications of HA derivatives DHA,BHA,TFHA,HANP and HA-MTX were 35.1%,34.5%,29.0%,66.0% and17.0%,respectively.The biochemical experiments have indicated that TFHA promoted the proliferation,migration,and tube formation in human umbilical vein vascular endothelial cells(HUVEC).In addition,TFHA inhibited the expression of inflammatory cytokines IL-1?,IL-6 and TNF-? in mouse macrophages(RAW264.7)model induced by lipopolysaccharide(LPS)to various degrees.Furthermore,the experiments with arthritic rat model demonstrated that TFHA effectively reduced the inflammatory cells infiltration in tissue and inhibited the expression of IL-6 inflammatory cytokine in tissues.In the meantime,biochemical studies of HA-NP showed that it has significant cytotoxicity to cancer cell lines,such as the non-small cell lung cancer cells(A549),human breast cancer cells(MCF-7),and human liver cancer cells(SMMC-7221 and HepG2).HA-NP inhibited the proliferation of cancer cells,while,it was less toxic to the highly invasive human breast cancer cells(MDA-MB-231)and highly proliferative human cervical cancer cells(Hela).In contrast,HA-NP has no obvious cytotoxicity to a normal human cell,HUVEC..Moreover,HA-NP inhibited the expression of inflammatory factors IL-1?,IL-6 and TNF-? in LPS-induced RAW264.7 mouse macrophage model.In summary,the synthetic HA derivative TFHA showed both biological effects to promote tissue repair and anti-inflammatory.HA-NP showed anti-inflammatory and anti-cancer activities.This study shed light on the research of the development of novel HA derivatives with therapeutic potential.