Institute of Metabolism and Endocrinology, Second Xiang-Ya Hospital, Central South University, Changsha, Hunan, People's Republic of China.
PLoS One. 2011;6(12):e29037. doi: 10.1371/journal.pone.0029037. Epub 2011 Dec 15.
Several reports have shown that circulating insulin level is positively correlated with arterial calcification; however, the relationship between insulin and arterial calcification remains controversial and the mechanism involved is still unclear. We used calcifying vascular smooth muscle cells (CVSMCs), a specific subpopulation of vascular smooth muscle cells that could spontaneously express osteoblastic phenotype genes and form calcification nodules, to investigate the effect of insulin on osteoblastic differentiation of CVSMCs and the cell signals involved. Our experiments demonstrated that insulin could promote alkaline phosphatase (ALP) activity, osteocalcin expression and the formation of mineralized nodules in CVSMCs. Suppression of receptor activator of nuclear factor κB ligand (RANKL) with small interfering RNA (siRNA) abolished the insulin-induced ALP activity. Insulin induced the activation of extracellular signal-regulated kinase (ERK)1/2, mitogen-activated protein kinase (MAPK) and RAC-alpha serine/threonine-protein kinase (Akt). Furthermore, pretreatment of human osteoblasts with the ERK1/2 inhibitor PD98059, but not the phosphoinositide 3-kinase (PI3K) inhibitor, LY294002, or the Akt inhibitor, 1L-6-hydroxymethyl-chiro-inositol 2-(R)-2-O-methyl-3-O-octadecylcarbonate (HIMO), abolished the insulin-induced RANKL secretion and blocked the promoting effect of insulin on ALP activities of CVSMCs. Recombinant RANKL protein recovered the ALP activities decreased by RANKL siRNA in insulin-stimulated CVSMCs. These data demonstrated that insulin could promote osteoblastic differentiation of CVSMCs by increased RANKL expression through ERK1/2 activation, but not PI3K/Akt activation.
已有多项报告表明，循环胰岛素水平与动脉钙化呈正相关；然而，胰岛素与动脉钙化之间的关系仍存在争议，其涉及的机制尚不清楚。我们使用钙化血管平滑肌细胞（CVSMCs）作为研究对象，CVSMCs 是血管平滑肌细胞的一个特定亚群，能够自发表达成骨细胞表型基因并形成钙化结节，以研究胰岛素对 CVSMCs 成骨分化的影响及其涉及的细胞信号通路。我们的实验表明，胰岛素可以促进碱性磷酸酶（ALP）活性、骨钙素表达和 CVSMCs 矿化结节的形成。用小干扰 RNA（siRNA）抑制核因子 κB 配体受体激活剂（RANKL）可消除胰岛素诱导的 ALP 活性。胰岛素诱导细胞外信号调节激酶（ERK1/2）、丝裂原活化蛋白激酶（MAPK）和 Rac-α 丝氨酸/苏氨酸蛋白激酶（Akt）的激活。此外，ERK1/2 抑制剂 PD98059 预处理人成骨细胞，但不使用磷酸肌醇 3-激酶（PI3K）抑制剂 LY294002 或 Akt 抑制剂 1L-6-羟甲基-手性肌醇 2-（R）-2-O-甲基-3-O-十八烷基碳酸（HIMO），可消除胰岛素诱导的 RANKL 分泌，并阻断胰岛素对 CVSMCs 的 ALP 活性的促进作用。重组 RANKL 蛋白可恢复由 RANKL siRNA 降低的胰岛素刺激的 CVSMCs 中的 ALP 活性。这些数据表明，胰岛素可通过激活 ERK1/2 增加 RANKL 表达来促进 CVSMCs 的成骨分化，而不是通过 PI3K/Akt 激活。
