Lower back pain (LBP) prevalence is widespread due to an aging population and the limited efficacy of current treatments. As LBP is strongly associated with intervertebral disc (IVD) degeneration, it is thought that cell-based therapies could alleviate LBP by repairing IVD tissue. Various natural and synthetic biomaterials have been investigated as potential IVD tissue engineering scaffolds. Self-assembling peptide hydrogels (SAPHs) combine advantages of both natural and synthetic biomaterials; for example they are biocompatible and have easily modifiable properties. The present study demonstrated that a de novo SAPH had comparable strength to the native tissue, was injectable, restored the IVD cell phenotype and stimulated deposition of appropriate matrix components. Results illustrated the promise of SAPHs as scaffolds for IVD tissue engineering.
AimsVascular calcification is a major cause of morbidity and mortality. Fibroblast growth factor-2 (FGF-2) plays an instructive role in osteogenesis and bone development, but its role in vascular calcification was unknown. Therefore, we investigated the involvement of FGF-2 in vascular calcification and determined the mechanism by which it regulates this process.Methods and resultsWe demonstrate that FGF-2 expression is increased in vascular smooth muscle cells (VSMCs) induced to deposit a mineralized matrix by incubation with β-glycerophosphate. FGF-2 is also localized to sites of calcification within human atherosclerotic plaques. The expression of syndecan-4, a heparan sulfate proteoglycan which regulates FGF-2 signalling, is also increased in mineralizing VSMCs and co-localizes with FGF-2 in human calcified atherosclerotic plaques. Exogenous FGF-2 inhibits VSMC mineralization, and this inhibition is reduced when syndecan-4 expression is knocked-down using siRNA. Biochemical inhibition of FGFR signalling using a pan FGFR inhibitor (BGJ398) or knocking-down syndecan-4 expression in VSMCs using siRNA increases VSMC mineralization. These increases are prevented by inhibiting transforming growth factor-β (TGFβ) signalling with SB431542, suggesting cross-talk between FGF-2 and TGFβ signalling is crucial for the regulation of VSMC mineralization. Syndecan-4 can also regulate FGF-2 signalling directly via protein kinase Cα (PKCα) activation. Biochemical inhibition of PKCα activity using Gö6976, or siRNA-mediated suppression of PKCα expression increases VSMC mineralization; this increase is also prevented with SB431542. Finally, the ability of FGF-2 to inhibit VSMC mineralization is reduced when PKCα expression is knocked-down.ConclusionThis is the first demonstration that syndecan-4 promotes FGF-2 signalling, and in turn, suppresses VSMC mineralization by down-regulating TGFβ signalling. Our discoveries that FGF-2 and syndecan-4 expression is increased in mineralizing VSMCs and that PKCα regulates FGF-2 and TGFβ signalling in VSMCs suggests that the syndecan-4/FGF-2/TGFβ signalling axis could represent a new therapeutic target for vascular calcification.
BackgroundVascular calcification is associated with increased cardiovascular morbidity and mortality in patients with atherosclerosis, diabetes and chronic kidney disease. However, no viable treatments for this condition have been identified. This study aimed to determine whether farnesyl transferase inhibitors (FTIs) can reduce vascular calcification and the mechanism by which this reduction occurs.ResultsWe demonstrate that FTI-277 significantly inhibits phosphate-induced mineral deposition by vascular smooth muscle cells (VSMC) in vitro, prevents VSMC osteogenic differentiation, and increases mRNA expression of matrix Gla protein (MGP), an inhibitor of mineralization. FTI-277 increases Akt signaling in VSMC in short-term serum-stimulation assays and in long-term mineralization assays. In contrast, manumycin A has no effect on Akt signaling or mineralization. Co-incubation of VSMC with FTI-277 and SH6 (an Akt inhibitor) significantly reduces the inhibitory effect of FTI-277 on mineralization, demonstrating that FTI-277 inhibits calcification by activating Akt signaling. Over-expression of the constitutively active p110 sub-unit of PI3K in VSMC using adenovirus activates Akt, inhibits mineralization, suppresses VSMC differentiation and significantly enhances MGP mRNA expression. FTI-277 also inhibits phosphate-induced activation of caspase 3 and apoptosis of VSMC, and these effects are negated by co-incubation with SH6. Finally, using an ex vivo model of vascular calcification, we demonstrate that FTI-277 inhibits high phosphate-induced mineralization in aortic rings derived from rats with end-stage renal failure.ConclusionsTogether, these results demonstrate that FTI-277 inhibits VSMC mineral deposition by up-regulating PI3K/Akt signaling and preventing apoptosis, suggesting that targeting farnesylation, or Akt specifically, may have therapeutic potential for the prevention of vascular calcification.
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