Gouni-Berthold I, Berthold H K, Weber A A, Ko Y, Seul C, Vetter H, Sachinidis A
Medizinische Universitäts-Poliklinik, Bonn, Germany.
Naunyn Schmiedebergs Arch Pharmacol. 2001 Feb;363(2):215-21. doi: 10.1007/s002100000352.
Recent evidence suggests that apoptosis may be involved in the control of vascular smooth muscle cell (VSMC) number in atherosclerotic lesions. The peroxisome proliferator-activated receptor gamma (PPARgamma) ligands thiazolidinediones have been reported to induce apoptosis in macrophages and in a variety of tumor cell lines. To evaluate whether these agents also induce apoptosis in VSMC, cultured rat VSMC were treated with increasing doses of the thiazolidinedione analogues troglitazone (TRO) and rosiglitazone (RSG). Both ligands induced cell death in a concentration-dependent manner (EC50 12.1+/-3.3 microM and 1.43+/-0.39 microM, respectively), causing almost complete cell death at the highest concentrations (100 microM and 10 microM for TRO and RSG, respectively), along with an expected parallel decrease in [3H]thymidine uptake into cell DNA (EC50 6.7+/-2.4 microM and 0.75+/-0.19 microM, respectively). The cell count was determined by the coulter counter principle. Furthermore two apoptotic markers were measured, the caspase 3 activity and the cytoplasmic histone-associated DNA fragments, both of which were significantly increased when the aforementioned high concentrations were used. This indicates that apoptosis is involved in the TRO- and RSG-induced VSMC growth suppression. The same concentrations of TRO and RSG caused an unexpected stimulation of the extracellular signal-regulated response kinases 1 and 2 (ERK1/2) and stimulated the p38 mitogenic-activated protein (MAP) kinase as determined by Western blotting. In order to establish whether the proapoptotic effects of TRO and RSG are mediated through ERK1/2 activation, we used the selective MAP kinase kinase (MEK) inhibitor PD98059 (20 microM), which suppressed the TRO- and RSG-induced ERK1/2 activation but did not abolish their proapoptotic effects. We conclude that the thiazolidinedione analogues TRO and RSG induce cell death due to apoptosis in VSMC through an ERK1/2-independent pathway.
近期证据表明，细胞凋亡可能参与动脉粥样硬化病变中血管平滑肌细胞（VSMC）数量的调控。据报道，过氧化物酶体增殖物激活受体γ（PPARγ）配体噻唑烷二酮可诱导巨噬细胞及多种肿瘤细胞系发生凋亡。为评估这些药物是否也能诱导VSMC凋亡，用递增剂量的噻唑烷二酮类似物曲格列酮（TRO）和罗格列酮（RSG）处理培养的大鼠VSMC。两种配体均以浓度依赖方式诱导细胞死亡（EC50分别为12.1±3.3微摩尔和1.43±0.39微摩尔），在最高浓度时（TRO和RSG分别为100微摩尔和10微摩尔）几乎导致细胞完全死亡，同时细胞DNA中[3H]胸腺嘧啶核苷摄取量也呈预期的平行下降（EC50分别为6.7±2.4微摩尔和0.75±0.19微摩尔）。细胞计数采用库尔特计数器原理。此外，还检测了两种凋亡标志物，即半胱天冬酶3活性和细胞质组蛋白相关DNA片段，当使用上述高浓度药物时，二者均显著增加。这表明细胞凋亡参与了TRO和RSG诱导的VSMC生长抑制。相同浓度的TRO和RSG意外地刺激了细胞外信号调节激酶1和2（ERK1/2），并通过蛋白质免疫印迹法检测到其刺激了p38丝裂原活化蛋白（MAP）激酶。为确定TRO和RSG的促凋亡作用是否通过ERK1/2激活介导，我们使用了选择性MAP激酶激酶（MEK）抑制剂PD98059（20微摩尔），其抑制了TRO和RSG诱导的ERK1/2激活，但并未消除它们的促凋亡作用。我们得出结论，噻唑烷二酮类似物TRO和RSG通过ERK1/2非依赖途径诱导VSMC因凋亡而发生细胞死亡。
