PH20 is not expressed in murine CNS and oligodendrocyte precursor cells

Abstract

Abstract Objective Expression of Spam1 / PH 20 and its modulation of high/low molecular weight hyaluronan substrate have been proposed to play an important role in murine oligodendrocyte precursor cell ( OPC ) maturation in vitro and in normal and demyelinated central nervous system ( CNS ). We reexamined this using highly purified PH 20. Methods Steady‐state expression of mRNA in OPC s was evaluated by quantitative polymerase chain reaction; the role of PH 20 in bovine testicular hyaluronidase ( BTH ) inhibition of OPC differentiation was explored by comparing BTH to a purified recombinant human PH 20 ( rHuPH 20). Contaminants in commercial BTH were identified and their impact on OPC differentiation characterized. Spam1 / PH 20 expression in normal and demyelinated mouse CNS tissue was investigated using deep RNA sequencing and immunohistological methods with two antibodies directed against recombinant murine PH20. Results BTH , but not rHuPH 20, inhibited OPC differentiation in vitro. Basic fibroblast growth factor ( bFGF ) was identified as a significant contaminant in BTH , and bFGF immunodepletion reversed the inhibitory effects of BTH on OPC differentiation. Spam1 mRNA was undetected in OPC s in vitro and in vivo; PH 20 immunolabeling was undetected in normal and demyelinated CNS. Interpretation We were unable to detect Spam1 / PH 20 expression in OPC s or in normal or demyelinated CNS using the most sensitive methods currently available. Further, “ BTH ” effects on OPC differentiation are not due to PH 20, but may be attributable to contaminating bFGF . Our data suggest that caution be exercised when using some commercially available hyaluronidases, and reports of Spam1 / PH 20 morphogenic activity in the CNS may be due to contaminants in reagents.