Hu Rui, He Mu-Lan, Hu Hao, Yuan Bing-Xiang, Zang Wei-Jin, Lau Chu-Pak, Tse Hung-Fat, Li Gui-Rong
Department of Medicine and Research Centre of Heart, Brain, Hormone and Healthy Aging, The University of Hong Kong, Pokfulam, Hong Kong SAR, China.
J Cell Physiol. 2009 Sep;220(3):765-70. doi: 10.1002/jcp.21823.
Intracellular free Ca2+ (Ca(i)2+) is an important regulator of many cellular activities; however, Ca2+ signaling is not well studied in human preadipocytes. The purpose of the present study was to characterize Ca2+ signal pathways using a confocal scanning technique and RT-PCR. It was found that spontaneous Ca(i)2+ oscillations were observed in 12.1% preadipocytes, and number of cells with Ca2+ oscillations was increased to 47.9% by 1% fetal bovine serum. Ca(i)2+ oscillations were dependent on Ca2+ entry mainly via stored-operated Ca2+ (SOC) entry. They were suppressed by the SOC entry channel blocker La3+, the phospholipase C (PLC) inhibitor U73122, the inositol trisphosphate receptor (IP3R) blocker 2-amino-ethoxydiphenyl borate, or the sarcoplasmic/endoplasmic reticulum Ca2+ pump (SERCA) inhibitors thapsigargin and cyclopiazonic acid, but not by ryanodine. The IP3R activator thimerosal increased Ca(i)2+ oscillations. In addition, the plasma membrane Ca2+ pump (PMCA) inhibitor carboxyeosin and Na+--Ca2+ exchanger (NCX) inhibitor Ni2+ both suppressed Ca2+ oscillations. RT-PCR revealed that the mRNAs for IP3R1-3, SERCA1,2, NCX3 and PMCA1,3,4, Ca(V)1.2, and TRPC1,4,6, STIM1 and Orai1 (for SOC entry channels) were significant in human preadipocytes. The present study demonstrates that multiple Ca2+ signal pathways are present in human preadipocytes, and provides a basis for investigating how Ca2+ signals regulate biological and physiological activities of human preadipocytes.
细胞内游离钙离子（Ca(i)2+）是多种细胞活动的重要调节因子；然而，钙离子信号在人前脂肪细胞中的研究尚不充分。本研究的目的是利用共聚焦扫描技术和逆转录-聚合酶链反应（RT-PCR）对钙离子信号通路进行表征。研究发现，12.1%的前脂肪细胞中观察到自发的Ca(i)2+振荡，1%的胎牛血清可使出现钙离子振荡的细胞数量增加至47.9%。Ca(i)2+振荡主要依赖于通过储存-操作性钙离子（SOC）内流的钙离子进入。它们受到SOC内流通道阻滞剂La3+、磷脂酶C（PLC）抑制剂U73122、肌醇三磷酸受体（IP3R）阻滞剂2-氨基乙氧基二苯硼酸盐或肌浆网/内质网钙离子泵（SERCA）抑制剂毒胡萝卜素和环匹阿尼酸的抑制，但不受ryanodine的抑制。IP3R激活剂硫柳汞增加了Ca(i)2+振荡。此外，质膜钙离子泵（PMCA）抑制剂羧基曙红和钠-钙交换体（NCX）抑制剂Ni2+均抑制了钙离子振荡。RT-PCR显示，IP3R1-3、SERCA1、2、NCX3和PMCA1、3、4、Ca(V)1.2以及TRPC1、4、6、STIM1和Orai1（用于SOC内流通道）的信使核糖核酸（mRNA）在人前脂肪细胞中显著表达。本研究表明人前脂肪细胞中存在多种钙离子信号通路，为研究钙离子信号如何调节人前脂肪细胞的生物学和生理学活动提供了依据。
