Interaction between STIM1 and Orai1 in calcium-sensing receptor-media-ted calcium influx and nitric oxide generation
10.3969/j.issn.1000-4718.2017.10.007
- VernacularTitle:钙离子感受蛋白STIM1与Orai1在钙敏感受体介导钙内流及NO生成中的相互作用
- Author:
mei La WANG
1
;
Hua ZHONG
;
Na TANG
;
juan Li PANG
;
jun Chun ZHANG
;
Fang HE
Author Information
1. 石河子大学医学院新疆地方病与民族高发病教育部重点实验室
- Keywords:
Stromal interaction molecule 1;
Calcium release-activated calcium channel protein 1;
Ca2+-sen-sing receptor;
Nitric oxide
- From:
Chinese Journal of Pathophysiology
2017;33(10):1773-1780
- CountryChina
- Language:Chinese
-
Abstract:
AIM:To investigate the interaction of Ca 2+-sensing proteins , stromal interaction molecule 1 (STIM1) and calcium release-activated calcium channel protein 1 (Orai1), in Ca2+-sensing receptor (CaSR)-mediated extracellular Ca2+ influx and production of nitric oxide ( NO).METHODS: Human umbilical vein endothlial cells (HUVECs) were incubated with CaSR agonist spermine [activating store-operated calcium channels (SOC) and receptor-operated calcium channels ( ROC) ] alone or combined with CaSR negative allosteric modulator Calhex 231+ROC analogue TPA (activating ROC, blocking SOC), protein kinase C (PKC) inhibitor Ro 31-8220, or PKCα/β1 selective inhibitor Go 6976 (activate SOC, blocking ROC).The protein expression of STIM1 and Orai1 was determined by the method of immu-nofluorescence .The interaction between STIM 1 and Orai1 was examined by co-immunoprecipitation .The second to third passages of HUVECs were divided into STIM 1 and Orai1 short hairpin RNA group ( shSTIM1+shOrai1 group ) , vehicle-STIM1+vehicle-Orai1 group and control group , and then incubated with the 4 different treatments above .The intracellular Ca2+concentration ( [ Ca2+] I ) was detected using the fluorescent Ca 2+indicator Fura-2/AM.The production of NO was also determined by DAF-FM DA fluorescent probe .RESULTS:The protein expression of STIM 1 and Orai1 was located in the cytoplasm.Compared with control group , the localization of STIM1 and Orai1 in the cytoplasm was reduced after the HUVECs were incubated with Calhex 231+TPA, Ro 31-8220 or Go 6976, and the interaction of STIM1 and Orai1 was de-creased significantly .The [ Ca2+] I and the net NO fluorescence intensity in shSTIM 1+shOrai1 group were significantly re-duced after the 4 different treatments (P<0.05).CONCLUSION:STIM1 and Orai1 are components of SOC and ROC in store-and receptor-operated Ca 2+entry and NO generation .
