Amiloride slows down calpain-mediated ABCA1 degradation through in-hibition of hypoxia-induced NHE1 expression
10.3969/j.issn.1000-4718.2015.11.011
- VernacularTitle:Amiloride 通过抑制缺氧诱导的 NHE1表达而延缓 calpain 介导的 ABCA1降解
- Author:
Xiangang MO
;
Li ZHANG
;
Luochao ZHANG
;
Long WANG
;
Ning XIANG
;
Juan YANG
;
Xiang SONG
- Publication Type:Journal Article
- Keywords:
Adenosine triphosphate-binding cassette transporter A1;
Amiloride;
Sodium-hydrogen exchanger 1;
Calpain
- From:
Chinese Journal of Pathophysiology
2015;(11):1992-1997
- CountryChina
- Language:Chinese
-
Abstract:
AIM:To examine the effects of hypoxia on sodium-hydrogen exchange 1 (NHE1) expression, in-tracellular Ca2+concentration ( [ Ca2+] i ) and calpain activity, and to explore the effect of amiloride on adenosine triphos-phate-binding cassette transporter A1 (ABCA1) degradation and its calpain-related mechanism.METHODS:RAW264.7 cells were exposed to hypoxia for 0 h, 12 h, 24 h and 48 h.The cell viability was measured by MTT assay and the expres-sion of NHE1 at mRNA and protein levels was detected by real-time PCR and Western blot.[ Ca2+] i was analyzed by flow cytometry.Calpain activity was assessed by the method of Suc-LLVY-aminoluciferin.Furthermore, the protein levels of ABCA1 in the RAW264.7 cells exposed to hypoxia for 24 h were determined after 6 h or 12 h treatment with NHE1 inhibi-tor amiloride in the presence of cycloheximide.ABCA1 protein levels and calpain activity were detected after 12 h incuba-tion with calpain inhibitor ALLN or intracellular calcium-chelating agent BAPTA.RESULTS: Hypoxia inhibited the cell viability in a time-dependent manner.Hypoxia up-regulated the mRNA and protein expression of NHE1, and increased [ Ca2+] i and calpain activity.Hypoxia increased the degradation of ABCA1 and amiloride slowed down the ABCA1 degra-dation.ALLN or BAPTA increased ABCA1 protein level and decreased calpain activity.CONCLUSION:NHE1 inhibitor amiloride attenuates the calpain-mediated degradation of ABCA1, indicating that hypoxia-induced NHE1 might, at least in part, participate in the ABCA1 degradation.
