老龄SD大鼠胰岛β细胞形态功能研究

Morphological and Functional Characteristics of Pancreatic Islet Beta Cells in Natural Aging SD Rats

目的观察3组不同月龄SD大鼠糖脂代谢指标变化，了解在基础及葡萄糖刺激下胰岛β细胞胰岛素分泌、结构变化与老龄糖耐量减退的关系，探讨老龄糖耐量受损的机制。方法采用免疫组化、细胞体视学、透视电镜等方法观察5～6、11～12、20～24月龄SD大鼠胰岛及胰岛β细胞形态及胰岛素分泌水平；将胰岛β细胞分离后与两种不同质量浓度的葡萄糖培养，观察葡萄糖刺激下老龄大鼠β细胞胰岛素分泌水平。结果20～24月龄大鼠体质量、空腹血糖及血脂水平有增高；11～12、20～24月龄大鼠血清中游离脂肪酸（FFA）水平显著增高；对葡萄糖刺激出现峰值降低、达峰时间延迟；胰岛β细胞体积增大和细胞内胰岛素分泌量减少。老龄胰岛β细胞内线粒体和内质网肿胀，空泡变性。从11～12月龄大鼠开始，胰腺中平均胰岛面积随着年龄的增加而减少；但每个胰岛中平均胰岛β细胞的面积比例却有增加。结论老龄鼠在空腹血糖正常时即已具备胰岛素抵抗的某些特征；老龄大鼠胰岛β细胞虽然有面积比例增大，但β细胞基础和葡萄糖刺激的胰岛素分泌功能减低。提示一旦糖代谢受损出现糖尿病，应尽早考虑胰岛素治疗同时改善胰岛素抵抗。

Objective To evaluate the effect of aging on metabolic profile and the changes in morphology and secreting function of β-cell. Methods SD rats with different ages （5-6, 11-12, 20-24 Mon. ） were used in this experiment to detect fasting blood glucose, lipid profile and free fatty acids （FFA） as metabolic parameters. Islet population and β-cell shape from different age groups were assessed by immunohistochemistry and electron microscope （EM）, respectively. Moreover, isolated rat islets from different ages were purified and cultured with two culture medium containing 5.6 mmol/L and 16. 7 mmol/L glucose over 24 hours, respectively. The secreted levels of insulin at 2, 4, 6, 24 hours were measured by radioimmunoassay （RIA）. Results （1） The significantly higher concentration of FFA was found in 11-12, 20-24 months groups（386.02±88.08;349. 43±93.47 vs. 263.93± 68.77 mmol/L, respectively, P〈0. 05）, with no difference in WT, FBG, lipid profile among three groups ; （2） Insulin secretion by glucose （5. 5 mmol/L） stimulating old rats showed lower peak value than younger counterparts did [6.40±2.7 vs. 13. 97±1, 23（6 Mon. ） ＆ 12.08±0. 76（12 Mon. ） mU/L · 30 islets] and so did another glucose （16.7 mmol/L） group [6.75±4. 69 vs. 13.41±0. 72（6 Mon. ） ＆. 13.56±0. 60（12 Mon. ）]. （3） The ratio of β-cell mass to islet was slightly larger in old rats than in younger rats [72.99±29. 65% vs. 67.76± 23. 48%（12 Mon.）＆ 61. 69±30. 12% （6 Mon. ）, P〉 0. 05]; The expression of insulin measured by immunohistochemistry was dramatically decreased in old rat islet compared with 5-6, 11-12 month rats [242.71± 95.37 （24 Mon. ）,309±83. 99（12 Mon. ） vs. 558±69.94（6 Mon. ）,P〈0. 05]. Conclusion Older rats showed higher FFA and age-related impairment of insulin secretion with or without the glucose stimulation. These findings suggest that insulin therapy should be taken into account for elder earlier once type 2 diabetes is diagnosed.