Supplementary Material for: Activation of Peroxisome Proliferator-Activated Receptor δ Inhibits Angiotensin II-Induced Activation of Matrix Metalloproteinase-2 in Vascular Smooth Muscle Cells

We investigated the role of peroxisome proliferator-activated receptor (PPAR) δ on angiotensin (Ang) II-induced activation of matrix metalloproteinase (MMP)-2 in vascular smooth muscle cells (VSMCs). Activation of PPARδ by GW501516, a specific ligand for PPARδ, attenuated Ang II-induced activation of MMP-2 in a concentration-dependent manner. GW501516 also inhibited the generation of reactive oxygen species in VSMCs treated with Ang II. A marked increase in the mRNA levels of tissue inhibitor of metalloproteinase (TIMP)-2 and -3, endogenous antagonists of MMPs, was also observed in GW501516-treated VSMCs. These effects were markedly reduced in the presence of siRNAs against PPARδ, indicating that the effects of GW501516 are PPARδ dependent. Among the protein kinases inhibited by GW501516, suppression of phosphatidylinositol 3-kinase/Akt signaling was shown to have the greatest effect on activation of MMP-2 in VSMCs treated with Ang II. Concomitantly, GW501516-mediated inhibition of MMP-2 activation in VSMCs treated with Ang II was associated with the suppression of cell migration to levels approaching those in cells not exposed to Ang II. Thus, activation of PPARδ confers resistance to Ang II-induced degradation of the extracellular matrix by upregulating expression of its endogenous inhibitor TIMP and thereby modulating cellular responses to Ang II in vascular cells.

本研究探讨了过氧化物酶体增殖物激活受体δ（PPARδ）在血管紧张素Ⅱ（AngⅡ）诱导的血管平滑肌细胞（VSMCs）中基质金属蛋白酶-2（MMP-2）活化过程中的作用。采用PPARδ的特异性配体GW501516激活PPARδ，可呈浓度依赖性抑制血管紧张素Ⅱ诱导的基质金属蛋白酶-2活化。GW501516还可抑制血管紧张素Ⅱ处理的血管平滑肌细胞内活性氧（ROS）的生成。经GW501516处理的血管平滑肌细胞中，作为基质金属蛋白酶内源性拮抗剂的组织金属蛋白酶抑制剂（TIMP）-2和-3的mRNA水平显著升高。当使用靶向PPARδ的小干扰RNA（siRNA）干预时，上述效应显著减弱，表明GW501516的作用依赖于PPARδ。在GW501516抑制的多种蛋白激酶中，磷脂酰肌醇3-激酶（PI3K）/Akt信号通路的抑制，对血管紧张素Ⅱ处理的血管平滑肌细胞中基质金属蛋白酶-2的活化影响最为显著。与此同时，GW501516介导的血管紧张素Ⅱ处理的血管平滑肌细胞中基质金属蛋白酶-2活化的抑制作用，与细胞迁移能力的抑制相关，可使迁移水平接近未暴露于血管紧张素Ⅱ的细胞。综上，激活PPARδ可通过上调其内源抑制剂TIMP的表达，调节血管细胞对血管紧张素Ⅱ的细胞应答，进而抵抗血管紧张素Ⅱ诱导的细胞外基质降解。