Nuclear factor kappaB in proliferation, activation, and apoptosis in rat hepatic stellate cells.
Nuclear factor kappaB in proliferation, activation, and apoptosis in rat hepatic stellate cells.
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发表时间：
2000
影响因子：
25.7
通讯作者：
A. Lang;R. Schoonhoven;S. Tuvia;D. Brenner;R. Rippe
中科院分区：
文献类型：
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作者：
A. Lang;R. Schoonhoven;S. Tuvia;D. Brenner;R. Rippe
在活化的肝星状细胞（hsc）中已证实转录因子NFkappaB的激活。我们研究了NFkappaB在造血干细胞增殖、激活和tnfalpha诱导的凋亡中的作用。方法使用表达IkappaB显性阴性蛋白（Ad5IkappaB）的腺病毒，在静止和活化的hsc中抑制NFkappaB的激活。静止hsc感染Ad5IkappaB或表达β -半乳糖苷酶（Ad5LacZ）的腺病毒。细胞培养7天。通过细胞形态、平滑肌α -肌动蛋白（α -sma）表达和α - 1(I)胶原蛋白稳态mRNA水平（Western blot和RNase保护实验）来确定hsc的活化。通过3h -胸腺嘧啶掺入和直接细胞计数测定培养激活的hsc的增殖情况。用Ad5IkappaB或Ad5LacZ感染静止或活化的hsc，然后用TNFalpha处理，分析凋亡情况。通过测定细胞数量、核形态、TUNEL检测和caspase 3活性等方法证实细胞凋亡。结果在培养7天后，Ad5IkappaB-和ad5lacz感染的细胞在形态学、α -sma表达和α 1(I)胶原mRNA水平上没有差异。Ad5IkappaB感染不改变活化造血干细胞的增殖。TNFalpha仅在ad5ikappab感染的活化hsc中诱导凋亡，而不是静止hsc。暴露于TNFalpha 12小时后，细胞开始凋亡。TNFalpha处理24 h后，60%的活化hsc发生凋亡。结论：造血干细胞的增殖或活化不需要NFkappaB活性；然而，NFkappaB保护活化的hsc免受tnfalpha诱导的凋亡。
BACKGROUND/AIMS Activation of the transcription factor NFkappaB has been demonstrated in activated hepatic stellate cells (HSCs). We investigated the role of NFkappaB in proliferation, in activation, and in TNFalpha-induced apoptosis of HSCs. METHODS NFkappaB activation was inhibited using an adenovirus expressing an IkappaB dominant negative protein (Ad5IkappaB) in both quiescent and activated HSCs. Quiescent HSCs were infected with Ad5IkappaB or an adenovirus expressing beta-galactosidase (Ad5LacZ). The cells were cultured for 7 days. HSCs activation was determined by cell morphology, smooth muscle alpha-actin (alpha-sma) expression, and steady-state mRNA levels of alpha1(I) collagen as assessed by Western blot and RNase protection assay, respectively. Proliferation was determined in culture-activated HSCs by 3H-thymidine incorporation and direct cell counting. Apoptosis was analyzed by infecting quiescent or activated HSCs with Ad5IkappaB or Ad5LacZ, and then treating with TNFalpha. Apoptosis was demonstrated by determining cell number, assessing nuclear morphology, TUNEL assay and caspase 3 activity. RESULTS After 7 days in culture no differences were noted between the Ad5IkappaB- and the Ad5LacZ-infected cells in the morphology, alpha-sma expression or in alpha1(I) collagen mRNA levels. Ad5IkappaB infection did not modify proliferation in activated HSCs. TNFalpha induced apoptosis only in Ad5IkappaB-infected activated, but not quiescent HSCs. Apoptosis was initially demonstrated 12 h after exposure to TNFalpha. Twenty-four h after the TNFalpha treatment, 60% of the activated HSCs were apoptotic. CONCLUSION NFkappaB activity is not required for proliferation or activation of HSCs; however, NFkappaB protects activated HSCs against TNFalpha-induced apoptosis.