Dual effect of thapsigargin on cell death in porcine aortic smooth muscle cells.
A sustained increase in the cytosolic Ca(2+) concentration ([Ca(2+)](i)) can cause cell death. In this study, we found that, in cultured porcine aortic smooth muscle cells, endoplasmic reticulum (ER) stress, triggered by depletion of Ca(2+) stores by thapsigargin (TG), induced an increase in the [Ca(2+)](i) and cell death. However, the TG-induced death was not related to the [Ca(2+)](i) increase but was mediated by targeting of activated Bax to mitochondria and the opening of mitochondrial permeability transition pores (PTPs). Once the mitochondrial PTPs had opened, several events, including collapse of the mitochondrial membrane potential, cytochrome c release, and caspase-3 activation, occurred and the cells died. TG-induced cell death was completely inhibited by the pan-caspase inhibitor Z-VAD-fmk and was enhanced by the Ca(2+) chelator 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA), suggesting the existence of a Ca(2+)-dependent anti-apoptotic mechanism. After TG treatment, Ca(2+)-sensitive mitogen-activated protein kinase (MAPK) activation was induced and acted as a downstream effector of phosphatidylinositol 3-kinase (PI 3-kinase). The protective effect of Z-VAD-fmk on TG-induced cell death was reversed by BAPTA, PD-098059 (an MAPK kinase inhibitor), or LY-294002 (a PI 3-kinase inhibitor). Taken together, our data indicate that ER stress simultaneously activate two pathways, the mitochondrial caspase-dependent death cascade and the Ca(2+)-dependent PI 3-kinase/MAPK anti-apoptotic machinery. The Bax activation and translocation, but not the [Ca(2+)](i) increase, may activate mitochondrial PTPs, which, in turn, causes activation of caspases and cell death, whereas Ca(2+)-dependent MAPK activation counteracts death signaling; removal of Ca(2+) activated a second caspase-independent death pathway.
Chin
Ting-Yu
TY
Department of Biochemistry, National Defense Medical Center, 161 Min-Chuan East Road Section 6, Taipei, Taiwan, Republic of China.
Lin
Hsiu-Chen
HC
Kuo
Ju-Ping
JP
Chueh
Sheau-Huei
SH
D000818:Animals; D001011:Aorta; D017209:Apoptosis / Q000187:drug effects* / Q000502:physiology; D001692:Biological Transport; D002118:Calcium / Q000172:deficiency / Q000378:metabolism; D020169:Caspases / Q000378:metabolism; D002478:Cells, Cultured; D004721:Endoplasmic Reticulum / Q000187:drug effects / Q000378:metabolism; D004789:Enzyme Activation / Q000502:physiology; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D007425:Intracellular Membranes / Q000378:metabolism; D008928:Mitochondria / Q000201:enzymology; D033681:Mitochondrial Membrane Transport Proteins / Q000378:metabolism; D000083162:Mitochondrial Permeability Transition Pore; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D009131:Muscle, Smooth, Vascular / Q000187:drug effects / Q000502:physiology*; D032389:Myocytes, Smooth Muscle / Q000187:drug effects / Q000502:physiology*; D009994:Osmolar Concentration; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases / Q000037:antagonists & inhibitors; D015398:Signal Transduction / Q000502:physiology; D013552:Swine; D019284:Thapsigargin / Q000494:pharmacology*; D051028:bcl-2-Associated X Protein / Q000378:metabolism
D002118:Calcium; D020169:Caspases; D004791:Enzyme Inhibitors; D033681:Mitochondrial Membrane Transport Proteins; D000083162:Mitochondrial Permeability Transition Pore; D020928:Mitogen-Activated Protein Kinases; D019869:Phosphatidylinositol 3-Kinases; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases; D019284:Thapsigargin; D051028:bcl-2-Associated X Protein