大鼠胰岛素瘤实验模型的建立及其应用

目的 通过移植大鼠胰岛素瘤INS-1细胞至糖尿病小鼠肾包膜下使之形成胰岛素瘤,并诱导其发生凋亡,建立可用于研究胰岛β细胞凋亡机制的动物模型.方法 将5×106 INS-1细胞接种于链脲霉素(STZ)造模的糖尿病小鼠左肾包膜下,监测动物空腹血糖和血清胰岛素水平的变化,当血糖趋近正常后摘取动物左侧肾脏并检测其血糖和血清胰岛素水平的变化；固定包埋摘取的肾脏,进行HE染色以及胰岛素的免疫组化染色,确定胰岛素瘤模型的建立.在胰岛素瘤动物模型中,腹腔给予毒胡萝卜素(TG)或软脂酸钠(PA),监测给药后动物空腹血糖的变化,当血糖浓度出现逆转时,摘取动物左侧肾脏,通过TUNEL原位染色法检测移植瘤细胞的凋亡.结果 将INS-1细胞移植到糖尿病小鼠肾包膜下后,从第9天开始,动物空腹血糖进行性降低,血清胰岛素水平逐渐升高,当动物血糖接近至正常时,摘取动物左肾导致动物血糖显著升高,在摘取的左肾可见明显的移植瘤,免疫组织化学染色显示移植瘤细胞为胰岛素阳性.在胰岛素瘤动物模型给予TG或PA刺激后,动物空腹血糖出现逆转,显著升高,血清中胰岛素含量明显降低,摘取动物左侧肾脏后,TUNEL原位染色发现移植瘤内有明显的细胞凋亡.结论 大鼠胰岛素瘤INS-1细胞肾包膜下移植可以建立胰岛素瘤动物模型,应用此动物模型可以在体内研究胰岛β细胞凋亡的机制.

Establishment and application of rat insulinoma model in nude mice

Objective To establish an animal model to investigate pancreatic 13 cell apoptosis through transplanting insulinoma INS-1 cells into the renal capsules of streptozotocin （STZ） -treated diabetic mice. Methods We transplanted 5×10^6INS-1 cells into the left renal capsules of STZ-treated diabetic mice. After transplantation, the changes of fasted glycemia and fed insulin levels were monitored. When the glycemia reached to normal level, the left kidneys of the mice were removed. After that, the fasted glycemia and fed insulin levels were detected and the left kidneys were immediately fixed and subsequently examined by means of routine HE staining and insulin immuohistochemical staining to confirm establishment of insulinoma model. The insulinoma mice were i.p. injected once daily with palmitate （PA, 100 mg/kg per day） or thapsigargin （TG,5 mg/kg per day） for 9 days, and the changes of fasted glyeemia were monitored. When the glycemia was reversed, the left kidneys of the mice were removed, and TUNEL staining was employed to investigate 13 cell apoptosis in the xenografts. Results 9 days after transplantation of INS- 1 cells into the left renal capsules of STZ-treated diabetic mice, the fasting glycemia of the mice began to decline, and the fed insulin levels began to increase gradually. When the glycemia reached to normal level, removal of the left kidneys led to a significant increase in fasting glycemia. HE staining showed INS-1 cells can form tumors after subrenal capsules transplantation. Immunohistochemical staining showed insulin expression in the INS-1 cell xenografts. In insulinoma model, treatment of insulinoma mice with TG or PA led to increased fasting glycemia and decreased fed insulin levels. TUNEL staining showed that those treatments can cause significant cell apoptosis in xenografts. Conclusion Insulinoma animal model can be established by transplanting of rat insulinoma INS-1 cells into the renal capsules, and this model can be used to investigate the mechanism of the pancreatic 13 cell apoptosis in vivo.