Lysophosphatidic acid upregulates connective tissue growth factor expression in osteoblasts through the GPCR/PKC and PKA pathways
Lysophosphatidic acid upregulates connective tissue growth factor expression in osteoblasts through the GPCR/PKC and PKA pathways
复制标题
溶血磷脂酸通过 GPCR/PKC 和 PKA 途径上调成骨细胞中结缔组织生长因子的表达
DOI：
10.3892/ijmm.2016.2450
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发表时间：
2016-02-01
影响因子：
5.4
通讯作者：
Li, Zu-Bing
中科院分区：
文献类型：
--
作者：
Yu, Zi-Li;Li, Dian-Qi;Li, Zu-Bing
溶血磷脂酸（LPA）是一种高效的生物活性磷脂，参与各种生物过程。在这项研究中，LPA诱导结缔组织生长因子（CTGF/CCN 2）的表达和潜在的机制进行了研究，使用MC 3 T3-E1细胞系。用LPA受体的抑制剂、蛋白激酶C（PKC）和蛋白激酶A（PKA）的激活剂和抑制剂刺激MC 3 T3-E1细胞指定的时间段。使用RT-qPCR和蛋白质印迹分析来测量CCN 2的表达水平。免疫荧光染色观察PKC转位情况。LPA刺激后CCN 2 mRNA表达水平增加; LPA瞬时诱导CCN 2 mRNA表达; LPA刺激后2 h观察到最大表达水平。这种增加伴随着CCN 2蛋白的合成。LPA受体（1/3）被LPA特异性抑制剂Ki 16425抑制;结果，LPA诱导的CCN 2表达增加被消除。LPA还可诱导成骨细胞内PKC的膜转位，增强PKC的活性。用星形孢菌素预处理成骨细胞可阻止LPA诱导的CCN 2表达的增加，而佛波酯12-肉豆蔻酸酯13-乙酸酯（PMA）激活PKC可增强CCN 2的表达，表明PKC通路参与了LPA诱导的CCN 2表达的增加。LPA对PKA信号通路的干扰也可诱导CCN 2的表达。这些数据表明LPA通过激活PKC和PKA增加CCN 2表达。因此，PKC和PKA途径的调节功能与LPA诱导的CCN 2表达增加有关。
Lysophosphatidic acid (LPA) is an efficient, bioactive phospholipid involved in various biological processes. In this study, LPA-induced connective tissue growth factor (CTGF/CCN2) expression and the underlying mechanisms were investigated using the MC3T3-E1 cell line. The MC3T3-E1 cells were stimulated with an inhibitor of LPA receptors, an activator and inhibitor of protein kinase C (PKC) and protein kinase A (PKA) for indicated periods of time. RT-qPCR and western blot analyses were used to measure the expression levels of CCN2. Immunofluorescence staining was used to observe the translocation of PKC. The mRNA expression level of CCN2 was increased following stimulation of the cells with LPA; LPA transiently induced the mRNA expression of CCN2; maximum expression levels were observed 2 h following stimulation with LPA. This increase was accompanied by CCN2 protein synthesis. LPA receptor(1/3) was inhibited by Ki16425, a specific inhibitor of LPA(1/3); as a result, the LPA-induced increase in CCN2 expression was abrogated. LPA also induced the membrane translocation of PKC and enhanced PKC activity in the osteoblasts. Pre-treatment of the osteoblasts with staurosporine prevented the increase in CCN2 expression by induced by LPA, and the activation of PKC by phorbol 12-myristate 13-acetate (PMA) enhanced CCN2 expression, indicating that the PKC pathway is involved in the LPA-induced increase in CCN2 expression. The interference of PKA signaling also led to the induction of CCN2 expresion by LPA. These data indicate that LPA increases CCN2 expression through the activation of PKC and PKA. Thus, the regulatory functions of the PKC and PKA pathways are implicated in the LPA-induced increase in CCN2 expression.