Characterization of hyaluronan, hyaluronidase PH20, and HA synthase HAS2 in inflammation and cancer

Hyaluronan (HA, an extracellular matrix glycosaminoglycan) has been reported to have a variety of biological activities, including regulation of inflammation. While high molecular weight HA (HMW HA) is recognized to be anti-inflammatory, the activity of low molecular weight HA (LMW HA, the degradation catabolite of hyaluronidase) in inflammation is less clear. Because of reagent contamination of both hyaluronidase and HA used in many studies, many reported aspects of the associated biology need reinvestigation. There are many reports which have shown that LMW HA is pro-inflammatory; however, some recent publications raised serious doubts that LMW HA and hyaluronidase PH20 are not pro-inflammatory. Endotoxin and other contaminants in the reagents used in previous reports (i.e., bovine testicular hyaluronidase [BTH] Hyal type I-S and IV-S from Sigma) may be responsible for the observed inflammation. Further investigation has shown that the most purified BTH (type VI-S from Sigma) contains no endotoxin, but has substantial level of peptidoglycan, which is also pro-inflammatory. Caution should be taken when conducting studies of inflammation using HA and hyaluronidase that may contain endotoxin and peptidoglycan, as well as other pro-inflammatory contaminants. We have characterized HA affinity to its receptor CD44. While HMW HA binds to CD44 strongly, LMW HA degraded by PH20 hyaluronidase does not bind to CD44 on the cell surface. This may explain the observation that recombinant human hyaluronidase PH20 (rHuPH20) inhibits leukocyte migration upon inflammatory stimulation by interrupting CD44 interaction with HA, mediated by HMW HA. PEGPH20, a pegylated rHuPH20, has the same inhibitory activity on leukocyte transmigration as rHuPH20. Further investigation of leukocyte-inhibiting activity of rHuPH20 may reveal further clinical applications, such as wound healing and arthritis. Another report which is difficult to understand is that the ultra-high molecular weight HA produced by naked mole rat hyaluronan synthase-2 (nmrHAS2) contributes to cancer resistance, probably due to its specific anti-inflammatory activity. In our study, nmrHAS2 is cancer-promoting in human cancer cells, to a similar extent as human HAS2. The proposed cancer resistant activity of nmrHAS2 may apply selectively to its host animal species, the naked mole rat.