Wide spectrum activity of a high molecular weight Hyaluronic Acid.

Background Hyaluronic acid (HA), a non-sulphated glycosaminoglycan, is present in synovial fluid, vitreous humour and many connective tissues. Pharmaceutical preparations of HA are used in clinical practice for wound healing, joint pain, kerato-conjunctivitis, asthma, mouth care, oesophageal-reflux, and gastritis. Moreover, it is used as a filler to counteract aging and facial lypoatrophy. Here we investigated the antiviral activity in vitro of a high molecular weight (1,8KD) HA used in aesthetic medicine.Methods The MTT test was used to investigate the cytotoxicity of HA on the different cell lines used for virus growth: VERO, MDCK, PK15, JJHAN, MARC145. With the aim of investigating whether HA may affect cell membrane stabilization, experiments were carried out in which VERO cells were pre-treated with HA and then exposed to two lysis solutions, HCl and Triton X-100. The antiviral activity of HA was assessed by viral yield assays against Coxsackievirus B5 (COXB5), Herpes simplex virus-1 (HSV-1), Mumps virus (MV), Adenovirus (ADV), Influenza Virus (WSN33-AH1N1), Human Herpesvirus-6 (HHV-6), Porcine Parvovirus (PPV), Porcine Respiratory Reproductive Syndrome Virus (PRRSV). The virucide activity was assessed by exposing the different viral inocula to HA for 30’ at room T° and then their residual infectivity was titrated on cells. Time course experiments were carried out with COXB5 and HSV-1 within a replication single cycle by adding. HA at different time points.Results The MTT test showed that HI displayed no significant toxicity up to 4mg/ml. In lysis experiments, HA was able to protect VERO cells from lysis by both TritonX-100 and HCl. HSV-1, COXB5, MV, WSN33, PPV were inhibited by HA, the most effective inhibition being observed against COXB5 (3,5 Log reduction) and MV (1.7Log reduction). No virucidal activity by HA was ever observed against any of the viruses tested. Kinetic experiments showed that both COXB5 and HSV-1 were consistently inhibited independently upon the time of HA addition during the virus replication cycle. Conclusions The wide spectrum of antiviral activity exhibited by HA against both RNA and DNA viruses, known to have different structures (with or without envelope) and replication strategies, suggests a non specific mechanism of action, likely involving cell membrane-virus interaction steps either in virus entry or exit. The results of the kinetic experiments and of the cell protection from lysis support this hypothesis.