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背景:能否获取到足够的具有生物活性的胰岛细胞是影响胰岛细胞移植成功与否的主要原因之一,细胞冻存是收集足够胰岛的可行手段,但冻存复苏过程可引起细胞的凋亡。目的:观察银杏叶提取物对大鼠胰岛获取及冻存后功能的影响。设计、时间及地点:随机对照动物实验,于2007-10/12在西安交通大学医学院环境与疾病相关基因教育部重点实验室完成。材料:选用成年SD大鼠40只用于分离提取胰岛细胞。方法:按随机数字表法将40只大鼠分为2组(n=20),银杏叶提取物组取胰腺前经尾静脉注射银杏叶提取物25mg/kg,获取胰岛后冻存,复苏后分别在含银杏叶提取物的培养基中培养3,7,14,21d(n=5)。对照组尾静脉注射等量盐水,冻存、培养时间同银杏叶提取物组。主要观察指标:电镜下观察胰岛细胞形态;应用原位末端标记法检测细胞凋亡情况;以免疫组化法检测Bcl-2表达。结果:①银杏叶提取物组胰岛细胞复苏损失率明显低于对照组(P<0.05)。②与对照组相比,银杏叶提取物组胰岛细胞复苏培养3d时内分泌颗粒及酶原颗粒增多明显。③银杏叶提取物组复苏培养21d后胰岛细胞仍可保持45%左右的存活率,对照组胰岛细胞存活率﹤35%,两组比较差异具有显著性意义(P<0.05)。④复苏培养7,14d,银杏叶提取物组胰岛细胞凋亡率低于对照组(P<0.05),Bcl-2表达高于对照组(P<0.05)。结论:银杏叶提取物能够减少胰岛冻存复苏的损失,阻止胰岛细胞的凋亡,从而保护了胰岛细胞的活性。
BACKGROUND: The availability of sufficient islet cells with biological activity is one of the main factors affecting the success of transplantation of islet cells. Cell cryopreservation is a feasible method for collecting sufficient islets, but cryopreservation and resuscitation can cause apoptosis. . OBJECTIVE: To observe the effect of Ginkgo biloba extract on the function of rat islets after harvest and cryopreservation. DESIGN, TIME AND SETTING: A randomized controlled animal experiment was performed at the Key Laboratory of Environmental and Disease Related Genes, Ministry of Education, Medical College of Xi’an Jiaotong University between October 2007 and December 12th. MATERIALS: Forty adult Sprague-Dawley rats were used to isolate and extract islet cells. Methods: Forty rats were divided into two groups according to the random number table (n=20). Ginkgo biloba extract group was injected with 25 mg/kg ginkgo biloba extract via the tail vein before the pancreas. They were cultured for 3, 7, 14 and 21 days (n=5) in a medium containing Ginkgo biloba extract. In the control group, the same amount of saline was injected into the tail vein, and the cryopreservation and culture time were the same as those of the Ginkgo biloba extract group. MAIN OUTCOME MEASURES: Islet cell morphology was observed under electron microscope; apoptosis was detected by in situ end labeling; Bcl-2 expression was detected by immunohistochemistry. RESULTS: 1The recovery rate of islet cells in Ginkgo biloba extract group was significantly lower than that in the control group (P<0.05). 2Compared with the control group, the secretion of endocrine granules and zymogen granules increased significantly in the Ginkgo biloba extract group when the islet cells were resuscitated for 3 days. 3 Ginkgo biloba extract group after 21 days of resuscitation culture islet cells can still maintain a survival rate of about 45%, the survival rate of islet cells in the control group <35%, the difference between the two groups was significant (P <0.05). 4 On 7 and 14 days after resuscitation, the apoptosis rate of islet cells in Ginkgo biloba extract group was lower than that in control group (P<0.05), and the expression of Bcl-2 was higher than that in control group (P<0.05). Conclusion: Ginkgo biloba extract can reduce the loss of islet resuscitation and prevent the apoptosis of islet cells, thus protecting the activity of islet cells.