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（peroxisome proliferator-activated receptor, PPARδ）在血管紧张素Ⅱ（angiotensin Ⅱ, Ang Ⅱ）诱导的血管平滑肌细胞（vascular smooth muscle cells, VSMCs）中基质金属蛋白酶（matrix metalloproteinase, MMP）-2激活过程中的作用。采用PPARδ特异性配体GW501516激活PPARδ后，可呈浓度依赖性减弱Ang Ⅱ诱导的VSMCs中MMP-2的激活。此外，GW501516还可抑制Ang Ⅱ处理的VSMCs中活性氧的生成。经GW501516处理的VSMCs中，基质金属蛋白酶内源性拮抗剂组织金属蛋白酶抑制剂（tissue inhibitor of metalloproteinase, TIMP）-2与-3的mRNA水平显著升高。上述效应在靶向PPARδ的小干扰RNA（siRNA）存在时显著减弱，表明GW501516的作用依赖于PPARδ。在GW501516抑制的蛋白激酶中，抑制磷脂酰肌醇3-激酶/Akt信号通路对Ang Ⅱ处理的VSMCs中MMP-2的激活影响最为显著。同时，GW501516介导的Ang Ⅱ处理VSMCs中MMP-2激活的抑制作用，可使细胞迁移能力降至接近未受Ang Ⅱ处理的细胞水平。综上，激活PPARδ可通过上调其内源抑制剂TIMP的表达，从而调控血管细胞对Ang Ⅱ的应答，进而抵抗Ang Ⅱ诱导的细胞外基质降解。