A systematic re-evaluation of enzymatic isolation methodology for nucleus pulposus cells

Abstract

INTRODUCTION: The nucleus pulposus (NP) is the central part of IVD and understanding its function is essential for studying IVD degeneration. Enzymatic digestion by collagenase is used to release cells from NP tissues for cell phenotype characterization or further in vitro cell culture studies. For cell isolation from NP of different species, various protocols exist and vary greatly in terms of enzyme concentration, digestion duration, and type of precollagenase treatments such as digestion by pronase, which is a mixture of different proteases. In contrast to NP from other species, the cells in bovine or human mature NP have fewer cell-cell contacts and are surrounded by less gel-like matrix rich in proteoglycan and collagen. Thus we hypothesized that the pronase dig...