Division of Cardiovascular Sciences, School of Medical Sciences, Faculty of Biology, Medicine and Health, University of Manchester, Manchester Academic Health Science Centre, Manchester, UK.
Stem Cell Glycobiology Group, School of Materials, University of Manchester, Manchester, UK.
Cardiovasc Res. 2017 Nov 1;113(13):1639-1652. doi: 10.1093/cvr/cvx178.
Vascular 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.
We 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.
This 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.
血管钙化是发病率和死亡率的主要原因。成纤维细胞生长因子-2（FGF-2）在成骨和骨发育中发挥指导作用，但它在血管钙化中的作用尚不清楚。因此，我们研究了 FGF-2 参与血管钙化的情况，并确定了它调节该过程的机制。
我们证明，在β-甘油磷酸孵育诱导血管平滑肌细胞（VSMCs）沉积矿化基质时，FGF-2 的表达增加。FGF-2 也定位于人动脉粥样硬化斑块内的钙化部位。硫酸乙酰肝素蛋白聚糖（syndecan-4）的表达也在矿化的 VSMCs 中增加，syndecan-4 是一种调节 FGF-2 信号的肝素硫酸蛋白聚糖，它与人钙化的动脉粥样硬化斑块中的 FGF-2 共定位。外源性 FGF-2 抑制 VSMC 矿化，当使用 siRNA 敲低 syndecan-4 表达时，这种抑制作用降低。使用泛 FGFR 抑制剂（BGJ398）或使用 siRNA 在 VSMCs 中敲低 syndecan-4 表达来抑制 FGFR 信号转导，会增加 VSMC 矿化。用 SB431542 抑制转化生长因子-β（TGFβ）信号转导可预防这些增加，这表明 FGF-2 和 TGFβ 信号转导之间的交叉对话对于调节 VSMC 矿化至关重要。syndecan-4 还可以通过蛋白激酶 Cα（PKCα）激活直接调节 FGF-2 信号转导。使用 Gö6976 抑制 PKCα 活性的生化作用，或使用 siRNA 抑制 PKCα 表达会增加 VSMC 矿化；用 SB431542 也可预防这种增加。最后，当敲低 PKCα 表达时，FGF-2 抑制 VSMC 矿化的能力降低。
这是第一个证明 syndecan-4 通过下调 TGFβ 信号转导来促进 FGF-2 信号转导，进而抑制 VSMC 矿化的研究。我们发现，在矿化的 VSMCs 中 FGF-2 和 syndecan-4 的表达增加，PKCα 调节 VSMCs 中的 FGF-2 和 TGFβ 信号转导，这表明 syndecan-4/FGF-2/TGFβ 信号轴可能成为血管钙化的新治疗靶点。
