| Lower back pain affects 70 million people in the United States. Annually, nearly 300,000 individuals seek surgical treatment and receive lumbar spinal fusion of an intervertebral disc (IVD). Current techniques are invasive, painful, and may require revision. Understanding the processes controlling mineralization in this region could lead to improved treatment and more effective surgical and non-surgical fusion.;Mineralization is a complex process with regulation at both the cellular and chemical level. A class of proteins called bone morphogenetic proteins (BMPs) coordinate the cell-based process; BMP-2 and -7 act in bone to guide the architecture and formation of mineralized matrix. The Nucleus Pulposus (NP) of the IVD is typically uncalcified and has low expression of BMPs and other mineral biomarkers. However, mineral deposition has been reported during degenerative disc disease (DDD). Typically during DDD there is an increase in inflammatory cytokines such as Tumor Necrosis Factor (TNF) and Interleukin-1 (IL-1)?. In various cell types, both cytokines have been shown to regulate the expression of the progressive ankylosis gene product (ANKH). This mechanism of regulation is not fully understood.;ANKH is a channel protein involved in the regulation of extracellular inorganic pyrophosphate (PPi), a potent chemical inhibitor of mineralization. Along with ANKH, Ectonucleotide pyrophosphatase/phosphodiesterase (ENPP)1 increases the extracellular PPi concentration by hydrolyzing more complex polyphosphates, thus maintaining a soft tissue phenotype. Conversely, Tissue Nonspecific Alkalkine Phosphatase (TNAP), hydrolyzes PPi and creates a milieu favorable for mineral deposition. TNAP can also hydrolyze complex organic phosphates to generate free inorganic phosphate (Pi) to be incorporated into new mineral. TNAP requires a phosphate source to initiate mineralization. Mineral is typically deposited physiologically in gaps in collagen I substrate, though NP tissue is largely collagen II.;The interplay of cytokines, PPi-generating proteins like ANKH and ENPP1, and the mineral-inducing protein TNAP has not been studied in NP cells or tissue. The mechanisms through which inflammatory cytokines affect gene expression has not yet been described. We show that both TNF and IL-1? can negatively regulate ANKH and ENPP1 in nucleus pulposus cells grown in vitro. Furthermore, we propose ascorbic acid/Vitamin C alters the temporal nature of these gene expression responses to cytokine stimulation.;Experimenting with the introduction of BMP2, BMP7, and TNAP and the suppression of ANKH and ENPP1 with cytokines could lead to a more rapid mineralization of the NP tissue space and augment or even replace current surgical techniques. This work suggests the possibility of over-riding the soft tissue phenotype of the NP of the IVD, driving it towards a mineralized phenotype. |
