RNA in situ hybridization characterization of non-enzymatic derived bovine intervertebral disc cell lineages suggests progenitor cell potential

Abstract

Degeneration of the intervertebral disc (IVD) is a meritorious target for therapeutic cell based regenerative medicine approaches, however, controversy over what defines the precise identity of mature IVD cells and lack of single cell based quality control measures is of concern. Bos taurus and human IVDs are histologically more similar than is Mus musculus. The mature bovine IVD is well suited as model system for technology development to be translated into therapeutic cell based regenerative medicine applications. We present a reproducible non-enzymatic protocol to isolate cell progenitor populations of three distinct areas of the mature bovine IVD. Bovine specific RNA probes were validated in situ and employed to assess fate changes, heterogeneity, stem cell characteristics and differentiation potential of the cultures. Quality control measures with single cell resolution like RNA in situ hybridization to assess culture heterogeneity (PISH) followed by optimization of culture conditions could be translated to human IVD cell culture to increase the safety of cell based regenerative medicine.

Keywords: Annulus fibrosus; Nucleus pulposus; RNA in situ hybridization; Regenerative medicine; Single cell quality control.