Interaction of human mescenchymal stem cells with disc cells

Abstract Purpose of study: This study characterizes the potential role of human mesenchymal stem cells (MSCs) in regulating synthesis of extracellular matrix by disc cells derived from human degenerative discs. Methods used: Human nucleus pulposus cells (NPCs) and annulus fibrosis cells (AFCs) were isolated from degenerated disc tissues of patients and cultured independently in monolayer and pellet culture environment. Morphology, size and shape of pellets, proliferation rate and immunohistochemical staining characteristics of the cells were followed with time. After 2 weeks of culture in monolayer, freshly trypsinized disc cells and MSCs were cocast (cocultured) at a 50:50 ratio in pellet culture environment. Individual pellet cultures of NPCs, AFCs, chondrocytes and fibroblasts were used as controls. Proteoglycan and glycosaminoglycan contents were measured by spectrophotometer assay. After isolation of RNA from each sample, Type II collagen and proteoglycan were also confirmed by reverse transcriptase polymeric chain reaction. Coculture values were compared with the predicted values represented by the sum of individual control cultures. of findings: NPCs, AFCs, MSCs, chondrocytes and fibroblasts were identified histomorphologically by their size and shape. NPCs or AFCs could be readily distinguished from chondrocytes and fibroblasts in the pellet culture. The DNA content increased steadily with time in all populations. After 14 days of culture in monolayer, a yield of 5×106 cells per sample (NPCs, AFCs, NPCs+MSCs, AFCs+MSCs, chondrocytes, fibroblasts) were harvested for pellet culture. The rate of proteoglycan synthesis by NPCs remained fairly constant during the 14 days of pellet culture, whereas that by AFCs showed a marked increase with time. Cocultures of isolated cells from AFCs and MSCs in pellets resulted in a significant increase in proteoglycan and glycosaminoglycan synthesis compared with the predicted values, represented by the sum of the individual control cultured cells assuming a 50:50 ratio of the cells in coculture. In contrast, the mixed population of NPCs and MSCs cocultured in pellets did not result in higher synthesis of proteoglycan and glycosaminoglycan than the individual separate cultures. Purified NPCs synthesized less proteoglycan and glycosaminoglycan than either purified AFCs alone or mixed population of NPCs and MSCs cocultured in pellets. Relationship between findings and existing knowledge: There is very little existing literature currently available. Overall significance of findings: This study strongly suggests the possibility of cross-talk between disc cells and MSCs. Our initial observation that MSCs cocultured with AFCs derived from degenerative discs can upregulate the synthesis of proteoglycan by disc cells supports the hypothesis that disc cells from degenerative discs may be stimulated to provide the crucial functions of normal disc cells given the appropriate cellular environment. It also raises the intriguing possibility that in coculture disc cells may influence the differentiation of MSCs to express disc cell phenotypes. Disclosures: No disclosures. Conflict of interest: No conflicts.