Early assessment of apoptosis in isolated islets of langerhans
Early assessment of apoptosis in isolated islets of langerhans
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DOI：
10.1097/00007890-200104150-00006
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发表时间：
2001-04-15
期刊：
影响因子：
6.2
通讯作者：
Inverardi, L
中科院分区：
文献类型：
--
作者：
Cattan, P;Berney, T;Inverardi, L
背景。有大量证据表明胰岛移植后早期移植物丢失与分离诱导的胰岛细胞凋亡有关，通过膜联蛋白V结合检测caspase 3活性和检测丢失的细胞膜不对称性，是分析早期凋亡事件的新方法。在这项研究中，我们将这些测试与末端脱氧核苷酸转移酶（TdT）介导的dUTP缺口末端标记（TUNEL）测试和凝胶后DNA片段分析进行了比较。用白细胞介素-1 β、干扰素γ和肿瘤坏死因子α（已知的诱导胰岛细胞损伤的细胞因子）治疗前后，新鲜分离的大鼠胰岛的电泳结果。用caspase 3活性和膜联蛋白V结合作为检测，分离后第1天观察到可测量的凋亡水平，尽管TUNEL试验和DNA凝胶电泳未检测到明显的DNA片段。基线caspase 3活性为0.8 +/- 0.3 U/100胰岛当量，细胞因子刺激后3小时增加到1.4+/-0.45 U/100胰岛当量(P
Background. There is substantial evidence to link early graft loss after islet transplantation to isolation-induced islet cell apoptosis, Measurement of caspase 3 activity and detection of the lost cell membrane asymmetry, revealed by annexin V binding, are newly available assays that allow the analysis of early events of apoptosis,Methods. In this study, we compared these tests with the terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end labeling (TUNEL) assay and analysis of DNA fragmentation after gel. electrophoresis in freshly isolated islets obtained from rats, before and after treatment with interleukin-1 beta, interferon gamma, and tumor necrosis factor alpha, cytokines known to induce islet cell damage.Results. A measurable level of apoptosis was observed the day after isolation when caspase 3 activity and annexin V binding were used as assays, although no substantial DNA fragmentation was detected with TUNEL assay and DNA gel electrophoresis. Baseline caspase 3 activity was 0.8 +/- 0.3 U/100 islet equivalents and it increased to 1.4+/-0.45 U/100 islet equivalents 3 hr after cytokine stimulation (P