Department of Cardiology, Affiliated Hospital of Jiangsu University, Zhenjiang 212001, China.
Department of Pathology, Affiliated Hospital of Jiangsu University, Zhenjiang 212001, China.
Biomed Pharmacother. 2017 Jul;91:1053-1064. doi: 10.1016/j.biopha.2017.05.031. Epub 2017 May 15.
Vascular calcification is established to be a critical factor in diabetes mellitus, which causes cardiovascular and amputation complication of diabetic patients. OPG/RANKL/RANK axis serves as a regulatory role in vascular calcification. Ghrelin, an endogenous ligand of growth hormone secretagogue receptor (GHSR), has been reported to exhibit potent cardiovascular protective effects. However, the role of ghrelin in the regulation of diabetic vascular calcification is still elusive. Here, we reported the role of ghrelin and its relationship with OPG/RANKL/RANK system in patients with diabetic foot amputation. In vivo and in vitro investigations were performed. Sixty type 2 diabetic patients with foot amputation were enrolled in vivo investigation, and they were divided into three groups through Doppler ultrasound: mild stenosis group (n=20), moderate stenosis group (n=20), and severe stenosis/occlusion group (n=20). Morphological analysis results showed diffused calcium depositions in the anterior tibial artery of diabetic amputees. Compared with the mild and moderate stenosis group, the severe stenosis/occlusion group had more spotty calcium depositions in atherosclerotic plaques. Western blot analysis indicated the expressions of osteoprotegerin (OPG) and ghrelin were downregulated, while the expression of receptor activator of nuclear factor kappa B ligand (RANKL) was upregulated with the vascular stenosis aggravation. Pearson correlation analysis revealed a negative correlation between calcium content and ghrelin levels (r=-0.58, P<0.001), as well as the ghrelin levels and sRANKL levels (r=-0.57, P<0.001). Meanwhile, OPG levels were positively correlated with ghrelin levels (r=0.63, P<0.001). From in vitro investigation, we found that the high-glucose (HG), high-lipid (HL), and β-glycerophosphate (β-GP) considerably increased the total calcium content, ALP activity, and expression of osteogenic markers in vascular smooth muscle cells (VSMCs). Ghrelin blunted calcification in a dose-dependent manner. In addition, ghrelin upregulated OPG expression and downregulated RANKL expression in VSMC calcification when anti-OPG antibody and RANKL were performed. Collectively, we therefore conclude serum ghrelin level may be a predictor of diabetic vascular calcification. The possible mechanism may be related with OPG/RANKL signal.
血管钙化被确定为糖尿病的一个关键因素，它会导致糖尿病患者心血管和截肢并发症。OPG/RANKL/RANK 轴在血管钙化中起调节作用。生长激素释放肽受体 (GHSR) 的内源性配体 ghrelin 已被报道具有强大的心血管保护作用。然而，ghrelin 在调节糖尿病血管钙化中的作用仍不清楚。在这里，我们报告了 ghrelin 的作用及其与糖尿病足截肢患者 OPG/RANKL/RANK 系统的关系。进行了体内和体外研究。60 例 2 型糖尿病足截肢患者纳入体内研究，根据多普勒超声将其分为三组：轻度狭窄组（n=20）、中度狭窄组（n=20）和重度狭窄/闭塞组（n=20）。形态分析结果显示，糖尿病截肢患者的胫骨前动脉有弥漫性钙沉积。与轻度和中度狭窄组相比，重度狭窄/闭塞组的粥样硬化斑块中存在更多点状钙沉积。Western blot 分析表明，随着血管狭窄加重，骨保护素 (OPG) 和 ghrelin 的表达下调，核因子 kappa B 配体受体激活剂 (RANKL) 的表达上调。Pearson 相关分析显示，钙含量与 ghrelin 水平呈负相关（r=-0.58，P<0.001），ghrelin 水平与可溶性 RANKL 水平呈负相关（r=-0.57，P<0.001）。同时，OPG 水平与 ghrelin 水平呈正相关（r=0.63，P<0.001）。从体外研究中，我们发现高葡萄糖（HG）、高脂质（HL）和β-甘油磷酸（β-GP）可显著增加血管平滑肌细胞（VSMCs）的总钙含量、碱性磷酸酶活性和成骨标志物的表达。ghrelin 呈剂量依赖性抑制钙化。此外，当使用抗 OPG 抗体和 RANKL 时，ghrelin 在 VSMC 钙化中上调 OPG 表达并下调 RANKL 表达。综上所述，我们得出结论，血清 ghrelin 水平可能是糖尿病血管钙化的预测因子。可能的机制与 OPG/RANKL 信号有关。
