DIDS attenuates staurosporine-induced cardiomyocyte apoptosis by PI3K/Akt signaling pathway: activation of eNOS/NO and inhibition of Bax translocation.
OBJECTIVE
4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS), a non-selective chloride channel blocker, has been shown to prevent cell apoptosis, however, the underlying mechanisms remain undefined, thus the present study was to explore whether phosphatidylinositol 3'-kinase (PI3K)/Akt and its downstream molecules are involved in the cytoprotective effect of DIDS.


METHODS
Neonatal rat cardiomyocytes were exposed to staurosporine (STS) in the presence or absence of DIDS. Cell viability, apoptosis and expressions of Akt, phospho-Akt (p-Akt), eNOS, phospho-eNOS (p-eNOS), Bcl-2/Bax and nitric oxide (NO) production were determined, and Bax translocation was assessed by double immunofluorescence labeling and Western blotting.


RESULTS
DIDS markedly improved cell viability and exerted an anti-apoptotic effect on STS-exposed cardiomyocytes. DIDS resulted in a 2.1-fold increase of p-Akt over control levels, prevented the reduction in eNOS expression and phospho-eNOS levels induced by STS and significantly increased NO production (all P<0.01 vs. STS alone). Treatment with LY294002, a selective PI3K inhibitor, abolished DIDS-induced increases in p-Akt, eNOS, p-eNOS and NO production, and completely abrogated the DIDS-induced anti-apoptotic effect (P<0.01). Treatment with L-NAME, a non-selective NOS inhibitor similarly inhibited the increased NO but only partly abolished protective effects of DIDS (P<0.05). In addition, DIDS effectively inhibited STS-induced Bax translocation to mitochondria, which was also reversed by LY294002.


CONCLUSIONS
DIDS protects cardiomyocytes against STS-induced apoptosis via activating PI3K/Akt signaling pathway, including increasing eNOS phosphorylation and the subsequent NO production and inhibiting Bax translocation.
Liu
An-Heng
AH
Department of Geriatrics, Xijing Hospital, Fourth Military Medical University, Xi'an, China.
Cao
Ya-Nan
YN
Liu
Hong-Tao
HT
Zhang
Wei-Wei
WW
Liu
Yan
Y
Shi
Tang-Wang
TW
Jia
Guo-Liang
GL
Wang
Xiao-Ming
XM