Pro-inflammatory activity of contaminating DNA in hyaluronic acid preparations

Abstract
Hyaluronic acid (HA), an abundant non‐sulfated glycosaminoglycan component of the extracellular matrix, has applications in drug delivery, tissue engineering and as an ingredient in cosmetics. HA preparations containing high‐molecular‐weight polymers are also used in the treatment of inflammatory disorders such as arthritis and interstitial cystitis. Low‐molecular‐weight fragments derived from HA have been reported to induce pro‐inflammatory cytokines such as IL‐12 and TNF‐α, and could therefore potentially exacerbate existing inflammation. We therefore examined the pro‐inflammatory activity of HA preparations, since inflammatory reactions are known to occur following administration of HA. We tested low‐molecular‐weight fragments obtained from seven different HA preparations, either by sonication (≅ 3 times 105 Da) or by hyaluronidase digestion (≅ 1 times 104 Da), for the ability to induce the synthesis of IL‐12 and TNF‐α by human monocytic cells. We found that two of the seven HA preparations tested stimulated the synthesis of IL‐12 and TNF‐α by human monocytic cells. We unexpectedly found that the induction of IL‐12 and TNF‐α by these HA preparations was not due to their degradation to low‐molecular‐weight fragments, since their native high‐molecular‐weight forms possessed the same ability to stimulate IL‐12 and TNF‐α synthesis, but was due to the presence of contaminating DNA. Treatment of these two HA preparations with deoxyribonuclease I abrogated or reduced the induction of IL‐12 and TNF‐α. It is clear from this study that HA preparations can induce the synthesis of pro‐inflammatory cytokines by monocytes. The ability of HA to act as a pro‐inflammatory mediator may not, however, be related to the presence of low‐molecular‐weight HA fragments, but to the presence of DNA. The presence of pro‐inflammatory DNA in HA preparations should be evaluated before its use, not only for the treatment of patients with inflammatory disorders, but also before many other applications.