title: Overexpression of TRPC3 increases apoptosis but not necrosis in response to ischemia-reperfusion in adult mouse cardiomyocytes.
abstract: An increase in cytosolic Ca2+ via a capacitative calcium entry (CCE)-mediated pathway, attributed to members of the transient receptor potential (TRP) superfamily, TRPC1 and TRPC3, has been reported to play an important role in regulating cardiomyocyte hypertrophy. Increased cytosolic Ca2+ also plays a critical role in mediating cell death in response to ischemia-reperfusion (I/R). Therefore, we tested the hypothesis that overexpression of TRPC3 in cardiomyocytes will increase sensitivity to I/R injury. Adult cardiomyocytes isolated from wild-type (WT) mice and from mice overexpressing TRPC3 in the heart were subjected to 90 min of ischemia and 3 h of reperfusion. After I/R, viability was 51 +/- 1% in WT mice and 42 +/- 5% in transgenic mice (P < 0.05). Apoptosis assessed by annexin V was significantly increased in the TRPC3 group compared with WT (32 +/- 1% vs. 21 +/- 3%; P < 0.05); however, there was no significant difference in necrosis between groups. Treatment of TRPC3 cells with the CCE inhibitor SKF-96365 (0.5 microM) significantly improved cellular viability (54 +/- 4%) and decreased apoptosis (15 +/- 4%); in contrast, the L-type Ca2+ channel inhibitor verapamil (10 microM) had no effect. Calpain-mediated cleavage of alpha-fodrin was increased approximately threefold in the transgenic group following I/R compared with WT (P < 0.05); this was significantly attenuated by SKF-96365. The calpain inhibitor PD-150606 (25 microM) attenuated the increase in both alpha-fodrin cleavage and apoptosis in the TPRC3 group. Increased TRPC3 expression also increased sensitivity to Ca2+ overload stress, but it did not affect the response to TNF-alpha-induced apoptosis. These results suggest that CCE mediated via TRPC may play a role in cardiomyocyte apoptosis following I/R due, at least in part, to increased calpain activation.
authors.affiliation: Division of Cardiovascular Disease, Dept. of Medicine, Univ. of Alabama at Birmingham, Birmingham, AL 35294-0005, USA.
authors.forename: Dan
authors.initials: D
authors.lastname: Shan
authors.forename: Richard B
authors.initials: RB
authors.lastname: Marchase
authors.forename: John C
authors.initials: JC
authors.lastname: Chatham
mesh_terms: D000179:Acrylates / Q000494:pharmacology; D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D002118:Calcium / Q000378:metabolism; D002121:Calcium Channel Blockers / Q000494:pharmacology; D020746:Calcium Channels, L-Type / Q000378:metabolism; D020013:Calcium Signaling / Q000187:drug effects; D002154:Calpain / Q000037:antagonists & inhibitors / Q000378:metabolism; D002352:Carrier Proteins / Q000378:metabolism; D002470:Cell Survival; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D007093:Imidazoles / Q000494:pharmacology; D008297:Male; D051379:Mice; D008822:Mice, Transgenic; D008840:Microfilament Proteins / Q000378:metabolism; D015428:Myocardial Reperfusion Injury / Q000378:metabolism* / Q000473:pathology; D032383:Myocytes, Cardiac / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism* / Q000473:pathology; D009336:Necrosis; D050052:TRPC Cation Channels / Q000235:genetics / Q000378:metabolism*; D013997:Time Factors; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism; D015854:Up-Regulation; D014700:Verapamil / Q000494:pharmacology; D013049:Spectrin
chemical_list: D000179:Acrylates; D002118:Calcium; D002121:Calcium Channel Blockers; D020746:Calcium Channels, L-Type; D002154:Calpain; D002352:Carrier Proteins; D015853:Cysteine Proteinase Inhibitors; D007093:Imidazoles; D008840:Microfilament Proteins; D050052:TRPC Cation Channels; D014409:Tumor Necrosis Factor-alpha; D014700:Verapamil; D013049:Spectrin; C100442:PD 150606; C104603:TRPC3 cation channel; C096351:transient receptor potential cation channel, subfamily C, member 1; C063159:1-(2-(3-(4-methoxyphenyl)propoxy)-4-methoxyphenylethyl)-1H-imidazole