【作者】 李文；

【导师】 王庆宝； 王艺；

【作者基本信息】 泰山医学院 ， 外科学， 2016， 硕士

【摘要】 背景随着经济的发展及人们生活方式的改变,糖尿病病人数量呈逐渐增多趋势,成为威胁人们健康的慢性非传染性疾病。糖尿病患者因胰岛功能的损伤使其血糖水平持续处于高位,并由此引发一系列并发症。有研究表明^[1,2],糖尿病患者罹患恶性肿瘤的风险与正常人群相比明显增高,根据基础实验及流行病学研究,糖尿病所导致的代谢减慢及炎症结果被认为在癌症的起始阶段及早期进展中其重要促进作用。胰腺癌作为一种恶性程度很高的肿瘤之一,其发生发展及预后与糖尿病之间的关系受到广泛关注。大量的科学研究已经证实^[3],长期的糖尿病病史会加大患有胰腺恶性肿瘤的风险。许多研究同时也表明,大约20%-30%胰腺癌患者在癌症确诊前的2-4年内出现糖尿病的症状。在此情况下,虽尚未在临床上确诊胰腺恶性肿瘤,但可认为高血糖是其导致产生^[2,4]。在这些患者中,胰腺恶性肿瘤被认为是导致副肿瘤性高血糖的隐秘因素。同时在一些胰腺癌患者行胰腺部分切除术后新发糖尿病会消退^[5,6]。因此糖尿病与胰腺癌之间的关系错综复杂,值得进一步研究。文献报道的对糖尿病与胰腺癌关系的研究,大多数是从临床角度,通过病例分析及数据调查进行的,尚无从蛋白表达等角度探究糖尿病病情进展及其与胰腺癌发生之间关系的报道。本实验通过分子水平的研究,可以为胰腺癌的早期诊断、治疗和预后的判断提供新的思路。目的根据糖尿病大鼠基础代谢指标、血生化指标和组织学相关染色,判断糖尿病病情严重程度,进一步观察胰腺组织中apoptin蛋白在糖尿病不同病变程度中表达的亚细胞部位和含量变化差异;以期为胰腺癌的早期诊断提供一种新的检测方法。方法采用禁食后一次性腹腔内大剂量注射链脲佐菌素并配合高脂高糖饮食构建糖尿病大鼠模型,记录其进食量、饮水量、尿量及体重等基础代谢指标并搜集其血液、胰腺、肝脏及肾脏组织标本,采用HE染色、tunel染色及免疫组化等方法进行研究,并与同时期正常对照组大鼠进行比较。结果DM组与BC组相比:进食量、饮水量、尿量、空腹血糖（FBG）、MDA、TC、TG、LDL-C、血NGAL、肝脏NO、NOS水平明显增高（P<0.05）;基础体重、血清胰岛素（INS）、C肽含量、血清SOD、HDL-C、血NO明显降低（P<0.05）。胰腺HE染色及免疫组化结果显示:DM组与BC组相比,胰岛数量减少,体积减小,形态欠规则,胰岛数及胰岛内内分泌细胞数量减少,细胞排列紊乱,大部分细胞核染色较浅,少部分细胞呈现空泡状。免疫组化可见apoptin蛋白在DM组大鼠胰腺细胞胞质及胰腺外分泌腺细胞中表达,并随时间的变化表达量逐渐增加,而同期BC组大鼠胰腺中未发现明显的apoptin蛋白表达。TUNEL染色可见DM组大鼠胰腺在各检测时间点的细胞凋亡率与同时期的正常对照组相比明显增高。肾脏组织免疫组化可见NGAL表达的主要部位位于近端肾小管上皮细胞胞浆中,而在肾小球、肾间质及远端肾小管胞浆中未见明显阳性染色,随着时间的增长,NGAL在近端小管的表达量逐渐增多,染色逐渐加深,而同时期正常对照组大鼠的肾脏组织中未见明显染色。结论1、糖尿病大鼠胰腺组织中apoptin蛋白的表达量明显增多,且与病情严重程度密切相关,病情越重,含量越高。2、糖尿病大鼠胰腺组织中apoptin蛋白的表达亚细胞部位与病情严重程度无关联。3、apoptin蛋白未来可能成为胰腺癌早期诊断的新的指标。更多还原

【Abstract】 BackgroundWith the development of economy and the change of people’s life style,the number of diabetic patients is gradually increasing,which has become a chronic non communicable disease that threatens people’s health.Diabetic patients with islet function damage to their blood sugar levels remain at high levels,and thus lead to a series of complications.Studies have shown that,diabetic patients suffering from the risk of malignant tumor and normal population were significantly higher than that,on the basis of experimental and epidemiological studies,diabetes caused by a slowing of the metabolism and inflammation results were found to in the initial stage of cancer and early development of its important role in promoting.Pancreatic cancer is one of the highly malignant tumors.The relationship between the development and prognosis of pancreatic carcinoma and the relationship between the prognosis and the diabetes mellitus has received extensive attention.A large number of scientific studies have confirmed that,a long history of diabetes,will increase the risk of pancreatic cancer.Many studies also show that about 20%-30% of pancreatic cancer patients have symptoms of diabetes in the 2-4 years before the diagnosis of cancer.In this case,although it has not yet been clinically diagnosed with pancreatic cancer,but can be considered to be the cause of high blood sugar to produce.In these patients,malignant tumor of the pancreas is thought to lead to hidden factors paraneoplastic hyperglycemia.At the same time in some patients with pancreatic cancer after partial resection of pancreatic cancer will be the new onset of diabetes,Therefore,the relationship between diabetes and pancreatic cancer is complex,and it is worth further study.On the relationship between diabetes and pancreatic cancer research reported in the literature,most of them are from a clinical point of view,by means of case analysis and survey data,there is no from the protein expression of angle to explore the progression of diabetes and its report and the relationship between the occurrence of pancreatic cancer.In this study,we can provide a new idea for early diagnosis,treatment and prognosis of pancreatic cancer by studying the molecular level.Objective:According to the histological staining diabetes rat basal metabolic indexes,blood biochemical indexes and organization,judge the severity of diabetes,further observe the expression of apoptin protein in pancreatic tissue in diabetic patients with different severity of subcellular location and content changes of difference;to diabetes assessment provides a new detection method.MethodsBy fasting after intraperitoneal dose injection of streptozotocin(STZ)and with high fat and high sugar diet,construct the model of diabetic rats recorded feeding,water intake,urine volume and weight based metabolic indices and to collect blood,pancreas,liver and kidney tissue specimens,by HE staining,TUNEL staining and immunohisto chemical and method of study,and with the same period of normal control group rats were compared.Result:The DM group and the BC group compared with that of the food intake,water intake,urine volume,fasting blood glucose(FBG),MDA,TC,TG,LDL-C,serum NGAL,liver no,NOS levels increased significantly(P < 0.05);basis weight,serum insulin(INS),C peptide,serum superoxide dismutase(SOD),HDL-C,blood no decreased significantly(P < 0.05).Pancreatic HE staining and immunohistochemical results showed that the DM group and the BC group compared to reduce the number of islets,reduced size and irregular shape,reduce the number of the number of pancreatic islets and endocrine cells,the cells arranged in disorder,most nuclear staining shallow,less part of the cells presented vacuole.Immune group of visible apoptin protein expressed in DM group rats pancreatic cell matter and pancreatic acinar cells,and with the time changes of expression gradually increased,and the same period BC rats’ s pancreatic not found obvious apoptin protein expression.TUNEL staining showed that the apoptosis rate of DM group was significantly higher than that of the normal control group at different time points.Kidney immune group of visible expression of NGAL in the main part in proximal renal tubular epithelial cell cytoplasm,and in the glomerular,renal between matter and distal renal tubular cytoplasm had no obvious positive staining,with the growth time of NGAL in the proximal tubular expression gradually increased,the dye color gradually deepened,and the same period in normal control group rat kidney tissues showed no staining.Conclusion :1.the expression of apoptin protein in pancreatic tissue of diabetic rats increased significantly,and was closely related to the severity of the disease,the disease is more serious,the higher the content.2.The subcellular location of apoptin protein in pancreatic tissue of diabetic rats was not correlated with the severity of the disease.3.apoptin protein in the future may become a new indicator of early diagnosis of pancreatic cancer.更多还原