Can Treatment with Human Mesenchymal Stem Cells Rescue the Degenerative Disc Phenotype? An in Vitro Pilot Study of Induced Cytokine Expression.

Abstract

Background context: Given the relatively low cell density in degenerative discs, strategies intended to bolster disc cellularity through stem cell injections have come into clinical use. Stem cell therapy is meant to provide a source of viable disc cells that can promote a healthy disc phenotype. Nevertheless, there is a limited understanding of the mechanisms through which stem cell therapy impacts degeneration.

Purpose: The objectives of this pilot study were: 1) to evaluate gene expression changes associated with an in vitro induced degenerative phenotype in human nucleus pulposus (NP) cells, 2) to co-culture these degenerative NP cells with human mesenchymal stem cells (hMSCs) and investigate the impact this has on gene expression, 3) to investigate possible mechanisms by which hMSCs may impact the degenerative phenotype.

Study design: Laboratory study.

Methods: NP cells were isolated and cultured from patients undergoing anterior lumbar interbody fusion for degenerative disc disease. A degenerative phenotype was induced in cultured NP cells by treatment with an inflammatory protocol (10pg/ml IL-1β and 100pg/ml TNF-α) for 7 days. Gene expression of Treated NP cells was compared to Untreated NP cells via reverse transcriptase polymerase chain reaction. NP cells were then co-cultured with hMSCs in vitro and treated with the inflammatory protocol. Gene expression of Treated NP cells co-cultured with hMSCs was compared to Treated NP cells alone. Preliminary co-culture data demonstrated that IL-10 was uniquely and dramatically upregulated. Therefore, gene expression of Treated NP cells exposed to IL-10 for 24 hours was compared to Treated NP cells alone.

Results: Treated NP cells compared to Control NP cells showed upregulation of numerous pro-inflammatory cytokines, including CXCL5, IL-8, and IL-6 and downregulation of several anti-inflammatory cytokines, including IL-10. After co-culture of Treated NP cells with hMSCs, a significant increase in gene expression was identified in IL-10 (+15.34 fold), BMP-6 (+2.32 fold), and LIF (+2.14 fold). A significant decrease in gene expression (p < 0.05) was seen in CCL7 (-2.03) and CXCL12 (-1.67). Exposure of Treated NP cells to IL-10 resulted in upregulation of COL-2 (+1.55 fold, p=0.013) and downregulation of IL-8 (-1.4 fold), CXCL-5 (-1.58 fold,), and MMP-3 (-2.02 fold).

Conclusion: This in vitro pilot study shows that co-culture of degenerative phenotype NP cells with hMSCs produces multiple gene regulatory changes associated with an anti-inflammatory phenotype. Additionally, exposure of degenerative phenotype NP cells to IL-10 produces gene regulation associated with both anti-inflammatory and pro-extracellular matrix effects.

Clinical significance: These findings provide mechanistic support for the use of stem cell therapy as a strategy to decrease the pro-inflammatory molecular environment associated with disc degeneration. Additionally, given the challenges with the viability of hMSCs in the disc microenvironment, IL-10 may be another potential candidate for future targeted therapies for disc degeneration.