Bone marrow stem cells in response to intervertebral disc-like matrix acidity and oxygen concentration implications for cell-based regenerative therapy

Study Design. In vitro culture of porcine bone marrow stem cells (BMSCs) in varying pH microenvironments in a threedimensional hydrogel system. Objective. To characterize the response of BMSCs to varying pH environments (blood [pH 7.4], healthy intervertebral disc (IVD) (pH 7.1), mildly degenerated IVD (pH 6.8), and severely degenerated IVD (pH 6.5) in three-dimensional culture under normoxic (20%) and hypoxic (5%) conditions. Summary of Background Data. The IVD is an avascular organ relying on diffusion of essential nutrients through the cartilaginous endplates (CEPs) thereby creating a challenging microenvironment. Within a degenerated IVD, oxygen and glucose concentrations decrease further (<5% oxygen, <5mmol/L glucose) and matrix acidity (<pH 6.8) increases resulting in especially adverse conditions. This has major implications for injectable cell-based strategies as these adverse microenvironmental conditions might severely affect the survival and regenerative potential of transplanted cells. Methods. BMSCs were encapsulated in 1.5% alginate and ionically cross-linked in 102 mmol/L CaCl₂ solution to form beads (diameter=5 mm), which were cultured in different microenvironmental conditions (pH 6.5, 6.8, 7.1, and 7.4; oxygen: 5% and 20%). Results. This study demonstrated decreased DNA content, increased cell death and minimal sulphated-glycosaminoglycans (sGAG) and collagen accumulation at pH 6.5 with increased proliferation, sustained cell viability and increased sGAG and collagen accumulation in pH 6.8 or higher. These findings suggest that there is a threshold at pH 6.8, below which cells cannot survive and accumulate nucleus pulposus-like matrix components (sGAG and collagen). Conclusion. Translation into a multimodal protocol requires the survival of stem cells and their ability to function normally amidst the harsh microenvironment. This study demonstrates the critical implication of degeneration stage and suggests stratified targeting to identify suitable candidates through measurement of the local pH thereby maximizing the efficacy for IVD cellular regenerative interventions.