Vasu Srividya, McClenaghan Neville H, Flatt Peter R
From the SAAD Centre for Pharmacy and Diabetes, University of Ulster, Coleraine, Northern Ireland.
Pancreas. 2016 Oct;45(9):1320-9. doi: 10.1097/MPA.0000000000000645.
Mechanisms of toxicity and cell damage were investigated in novel clonal human pancreatic beta cell line, 1.1B4, after exposure to streptozotocin, alloxan, ninhydrin, and hydrogen peroxide.
Viability, DNA damage, insulin secretion/content, [Ca]i, and glucokinase/hexokinase, mRNA expression were measured by MTT assay, comet assay, radioimmunoassay, fluorometric imaging plate reader, enzyme-coupled photometry, and real-time polymerase chain reaction, respectively.
Chemicals significantly reduced 1.1B4 cell viability in a time/concentration-dependent manner. Chronic 18-hour exposure decreased cellular insulin, glucokinase, and hexokinase activities. Chemicals decreased transcription of INS, GCK, PCSK1, PCSK2, and GJA1 (involved in secretory function). Insulin release and [Ca]i responses to nutrients and membrane-depolarizing agents were impaired. Streptozotocin and alloxan up-regulated transcription of genes, SOD1 and SOD2 (antioxidant enzymes). Ninhydrin and hydrogen peroxide up-regulated SOD2 transcription, whereas alloxan and hydrogen peroxide increased CAT transcription. Chemicals induced DNA damage, apoptosis, and increased caspase 3/7 activity. Streptozotocin and alloxan decreased transcription of BCL2 while increasing transcription of BAX. Chemicals did not affect transcription of HSPA4 and HSPA5 and nitrite production.
1.1B4 cells represent a useful model of human beta cells. Chemicals impaired 1.1B4 cell secretory function and activated antioxidant defense and apoptotic pathways without activating endoplasmic reticulum stress response/nitrosative stress.
研究新型克隆人胰腺β细胞系1.1B4在暴露于链脲佐菌素、四氧嘧啶、茚三酮和过氧化氢后毒性和细胞损伤的机制。
分别通过MTT法、彗星试验、放射免疫分析法、荧光成像酶标仪、酶联光度法和实时聚合酶链反应测量细胞活力、DNA损伤、胰岛素分泌/含量、细胞内钙浓度（[Ca]i）以及葡萄糖激酶/己糖激酶的mRNA表达。
化学物质以时间/浓度依赖的方式显著降低了1.1B4细胞活力。18小时慢性暴露降低了细胞胰岛素、葡萄糖激酶和己糖激酶活性。化学物质降低了胰岛素（INS）、葡萄糖激酶（GCK）、前蛋白转化酶枯草溶菌素1（PCSK1）、前蛋白转化酶枯草溶菌素2（PCSK2）和连接蛋白43（GJA1）（参与分泌功能）的转录水平。胰岛素释放以及对营养物质和膜去极化剂的[Ca]i反应受损。链脲佐菌素和四氧嘧啶上调了超氧化物歧化酶1（SOD1）和超氧化物歧化酶2（SOD2）（抗氧化酶）基因的转录。茚三酮和过氧化氢上调了SOD2转录，而四氧嘧啶和过氧化氢增加了过氧化氢酶（CAT）转录。化学物质诱导了DNA损伤、细胞凋亡并增加了半胱天冬酶3/7活性。链脲佐菌素和四氧嘧啶降低了B细胞淋巴瘤-2（BCL2）转录水平，同时增加了Bax蛋白（BAX）转录。化学物质不影响热休克蛋白A4（HSPA4）和热休克蛋白A5（HSPA5）转录以及亚硝酸盐生成。
1.1B4细胞代表了人β细胞的有用模型。化学物质损害了1.1B4细胞的分泌功能，激活了抗氧化防御和凋亡途径，而未激活内质网应激反应/亚硝化应激反应。
