Development of three-dimensional culture system for bovine nucleus pulposus cells

Abstract

INTRODUCTION: Nucleus pulposus is a central component of the intervertebral disc and rich in extracellular matrix (ECM). In the pathology of degenerative disc disease (DDD), the changes in ECM may be related to changes in nucleus pulposus cells (NPCs). To study NPCs, it is important to culture them in vitro in an environment similar to the native tissue. Many factors such as the interactions between NPCs and ECM, the topological cues and the mechanical properties of the constructs should be considered. If a 3D NPCmatrix system mimicking the native NP is developed, it can be used in future studies in regenerative medicine and DDD as a model platform. OBJECTIVE: The objective is to maintain the survival and phenotype of NPCs in a 3D culture system, by resembling the native tissue microenvironment. Specifically, we compare two starting materials, namely collagen and collagen-GAG constructs. METHODOLOGY: bNPCs were either encapsulated in collagen microspheres or seeded on collagen-GAG constructs fabricated by coprecipitation. They were cultured up to two weeks before evaluation. Cell morphology, F-actin distribution and viability were revealed by staining. Phenotype and components in the ECM were analyzed by qRT-PCR, immunohistochemistry and histology. RESULTS: bNPCs can survive, proliferate and maintain some phenotypic markers, in our 3D systems. Collagen-GAG constructs may be better than collagen alone for long term culture of bNPCs. CONCLUSION: Collagen-GAG constructs may be a good starting material for culturing NPCs. Including other ECM components into the culture system may further help in supporting the cell phenotype.