Regeneration of islet β-cells in tree shrews and rats

Backgroud: Current understanding of injury and regeneration of islet β-cells in diabetes is mainly based on rodent studies. The tree shrew is now generally accepted as being among the closest living relatives of primates, and has been widely used in animal experimentation. However, there are few reports on islet cell composition and regeneration of β-cells in tree shrews.Methods: In this study, we examined the changes in islet cell composition and regeneration of β-cells after streptozotocin(STZ) treatment in tree shrews compared with Sprague-Dawley rats. Injury and regeneration of islet β-cells were observed using hematoxylin and eosin(HE) staining and immunohistochemical staining for insulin, glucagon, somatostatin and PDX-1.Results: Our data showed that in rats islet injury was most obvious on day 3 after injection, and islet morphologies were significantly restored by day 21. Regeneration of islet β-cells was very pronounced in rats, and mainly involved regeneration of centro-acinar cells and transformation of extra-islet ductal cells. In tree shrews, the regeneration of islet β-cells was not as significant. On days 3 and 7, only scattered regenerated cells were observed in the remaining islets. Further, no regeneration of centro-acinar cells was observed.Conclusion: The results suggest that the repair mechanism of islet β-cells in tree shrews is similar to that of humans.

Effects of mycophenolate mofetil vs cyclosporine administration on graft survival and function after islet allotransplantation in diabetic rats

AIM: To develop an experimental model of islet allotransplantation in diabetic rats and to determine the positive or adverse effects of MMF as a single agent. METHODS: Thirty-six male Wistar rats and 18 male Lewis rats were used as recipients and donors respectively. Diabetes was induced by the use of streptozotocin (60 mg/kg) intraperitoneally. Unpurified islets were isolated using the collagenase digestion technique and transplanted into the splenic parenchyma. The recipients were randomly assigned to one of the following three groups: group A (control group) had no immunosuppression; group B received cyclosporine (CsA) (5 mg/kg); group C receivedmycophenolate mofetil (MMF) (20 mg/kg). The animalswere killed on the 12th d. Blood and grafted tissues were obtained for laboratory and histological assessment. RESULTS: Median allograft survival was significantly higher in the two therapy groups than that in the controls (10 and 12 d for CsA and MMF respectively vs 0 d for the control group, P＜0.01). No difference in allograft survival between the CsA and MMF groups was found. However,MMF had less renal and hepatic toxicity and allowed weight gain.CONCLUSION: Monotherapy with MMF for immunosu ppression was safe in an experimental model of islet allotransplantation and was equally effective with cyclosporine, with less toxicity.

ISLET FORMATION AND REGENERATION

Objective To explore the mechanisms of differentiation and development of pancreatic endocrine cells as well as pancreatic regeneration.Methods Human embryonic pancreatic tissue at 7-14 weeks of gestation was collected.Diabetes mellitus rat model was induced with 65 mg/kg of streptozotocin.Insulin, glucagon, somatostatin, nestin, and cytokeratin 19 (CK19) of pancreatic tissues were observed by immunohistochemistry.Results At 9 weeks of gestation, pancreatic epithelial cells began to co-express insulin, glucagon, somatostatin, and CK19 before migration.Islet cells gradually congregated along with the increase of aging, and at 14 weeks of gestation histological examination showed islet formation.At 12 weeks of gestation, nestin-positive cells could be seen in the pancreatic mesenchyme.During early embryogenesis, islet cells of pancreatic ducts co-expressed insulin, glucagon, and somatostatin.During pancreatic regeneration after damage, nestin expression of islet cells increased.Conclusion In the early stage of embryogenesis, islet cells of primary pancreatic ducts can be differentiated to multipotential endocrine cells before migration.During tissue regeneration, pancreatic stem cells may differentiate and proliferate to form pancreatic islet.

Regenerative medicine of pancreatic islets

The pancreas became one of the first objects of regenerative medicine,since other possibilities of dealing with the pancreatic endocrine insufficiency were clearly exhausted.The number of people living with diabetes mellitus is currently approaching half a billion,hence the crucial relevance of new methods to stimulate regeneration of the insulin-secretingβ-cells of the islets of Langerhans.Natural restrictions on the islet regeneration are very tight;nevertheless,the islets are capable of physiological regeneration viaβ-cell self-replication,direct differentiation of multipotent progenitor cells and spontaneousα-toβ-orδ-toβ-cell conversion(trans-differentiation).The existing preclinical models ofβ-cell dysfunction or ablation(induced surgically,chemically or genetically)have significantly expanded our understanding of reparative regeneration of the islets and possible ways of its stimulation.The ultimate goal,sufficient level of functional activity ofβ-cells or their substitutes can be achieved by two prospective broad strategies:β-cell replacement andβ-cell regeneration.The“regeneration”strategy aims to maintain a preserved population ofβ-cells through in situ exposure to biologically active substances that improveβ-cell survival,replication and insulin secretion,or to evoke the intrinsic adaptive mechanisms triggering the spontaneous non-β-toβ-cell conversion.The“replacement”strategy implies transplantation ofβ-cells(as non-disintegrated pancreatic material or isolated donor islets)orβ-like cells obtained ex vivo from progenitors or mature somatic cells(for example,hepatocytes orα-cells)under the action of small-molecule inducers or by genetic modification.We believe that the huge volume of experimental and clinical studies will finally allow a safe and effective solution to a seemingly simple goal-restoration of the functionally activeβ-cells,the innermost hope of millions of people globally.

Study of the immunoisolating effects of barium-alginate microencapsu lation on rat islets allograft survival

Objective: To evaluate the immunoisola ti ng effects of barium-alginate microencapsulation on islets allograft survival. Methods: The nonmicroencapsulated and microencapsulated islets w ere transplanted under the kidney capsule or intraperitoneally into Wistar rat w ith STZ-induced diabetes. The blood glucose and insulin secretion of grafts wer e observed. Graft function was tested by oral glucose tolerance test (OGTT). Results: ①Five diabetic rats became normoglycemic for 48 to 72 h after microencapsulated islets transplantation. The survival of transplanted i slets was on an average of 6 W. ②The normalization of the glycemia and insulin in the transplanted rats was associated with normal glucose and insulin profiles in response to OGTT. Conclusion: Microencapsulation with barium -alginate membrane can prolong islet survival and protect islets against allore jection.

Pancreatic islet transplantation

Type 1 diabetes mellitus is an autoimmune disease,which results in the permanent destruction of β-cells of the pancreatic islets of Langerhans.While exogenous insulin therapy has dramatically improved the quality of life,chronic diabetic complications develop in a substantial proportion of subjects and these complications generally progress and worsen over time.Although intensive insulin therapy has proven effective to delay and sometimes prevent the progression of complications such as nephropathy,neuropathy or retinopathy,it is difficult to achieve and maintain long term in most subjects.Reasons for this diff iculty include compliance issues and the increased risk of severe hypoglycemic episodes,which are generally associated with intensification of exogenous insulin therapy.Clinical studies have shown that transplantation of pancreas or purified pancreatic islets can support glucose homeostasis in type 1 diabetic patients.Islet transplantation carries the special advantages of being less invasive and resulting in fewer complications compared with the traditional pancreas or pancreas-kidney transplantation.However,islet transplantation efforts have limitations including the short supply of donor pancreata,the paucity of experienced islet isolation teams,side effects of immunosuppressants and poor long-term results.The purpose of this article is to review recent progress in clinical islet transplantation for the treatment of diabetes.

Protective effect of glucocorticoid-free immunosuppressive regimen in allogenic islet transplantation

BACKGROUND: The most common complication after allogenic islet transplantation is rejection. This study was to evaluate the effect of anti-rejection of glucocorticoid-free immunosuppressive regimen on allogenic islet transplantation. METHODS: Tacrolimus(FK506)+mycophenolate mofetil (MMF) and FK506+MMF+prednisone (Pred) were administered respectively for 2 weeks to inhibit rejection after allogenic islet transplantation in rats, which were compared with the control group. The concentrations of blood glucose, insulin and C-peptide were determined dynamically in recipients and the sites of transplantation were observed morphologically. RESULTS: As compared with the control group without immunosuppressive agents, FK506+MMF and FK506+MMF+Pred could prolong the survival time of grafts significantly. There were many morphologically intact islets in the liver of recipients 2 months after transplantation. Group FK506+MMF kept normal levels of blood glucose, insulin and C-peptide beyond 60 days after transplantation. In contrast, group FK506+MMF+Pred secreted less C-peptide(P<0.05) and maintained a higher level of blood glucose concentration (P<0.01) after the operation. There was no significant difference in insulin concentrations between the two groups. The level of blood glucose beyond the first 2 weeks after drug withdrawal in group FK506+MMF+Pred decreased obviously (P<0.05), and the secretion of insulin and C-peptide increased. These results were compared with those the first 2 weeks after transplantation and the first 2 weeks after drug withdrawal. CONCLUSIONS: Both regimens of FK506+MMF and FK506+MMF+Pred could provide effective immunosup-pression. Moreover the combined glucocorticoid-free immunosuppressive strategy of low-dose FK506 and MMF could protect islet grafts in islet transplantation without diabetogenic side-effects.

Nutritional programming of pancreatic β-cell plasticity

Nutritional insufficiency during pregnancy has been shown to alter the metabolism of the offspring and can increase the risk of type 2 diabetes. The phenotype in the offspring involves changes to the morphology and functional capacity of the endocrine pancreas, and in the supporting islet microvasculature. Pancreatic β-cells possess a plastic potential and can partially recover from catastrophic loss. This is partly due to the existence of progenitors within the islets and the ability to generate new islets by neogenesis from the pancreatic ducts. This regenerative capacity is induced by bone marrow-derived stem cells, including endothelial cell progenitors and is associated with increased angiogenesis within the islets. Nutritional insults in early life, such as feeding a low protein diet to the mother, impair the regenerative capacity of the β-cells. The mechanisms underlying this include a reduced ability of β-cells to differentiate from the progenitor population, changes in the inductive signals from the microvasculature and an altered presence of endothelial progenitors. Statin treatment within animal models was associated with angiogenesis in the islet microvasculature, improved vascular function and an increase in β-cell mass. This demonstrates that reversal of the impaired β-cell phenotype observed following nutritional insult in early life is potentially possible.

Recent progress in pancreatic islet transplantation

Diabetes mellitus remains a major burden.More than 200 million people are affected worldwide,which represents 6%of the world’s population.Type 1 diabetes mellitus is an autoimmune disease,which induces the permanent destruction of theβ-cells of the pancreatic islets of Langerhans.Although intensive insulin therapy has proven effective to delay and sometimes prevent the progression of complications such as nephropathy,neuropathy or retinopathy,it is difficult to achieve and maintain long term in most subjects.The successes achieved over the last few decades by the transplantation of whole pancreas and isolated islets suggest that diabetes can be cured by the replenishment of deficientβcells.However,islet transplantation efforts have various limitations,including the limited supply of donor pancreata,the paucity of experienced islet isolation teams,side effects of immunosuppressants and poor long term results.The purpose of this article is to review the recent progress in clinical islet transplantation for the treatment of diabetes and to describe the recent progress on pancreatic stem/progenitor cell research,which has opened up several possibilities for the development of new treatments for diabetes.

Annona muricata fruit extract protects against diethylnitrosamine-induced hepatocellular cancer in rats

Objective: To evaluate the anticancer potentials of Annona muricata fruit by in vitro and in vivo methods. Methods: The ethanolic extract of Annona muricata fruit was prepared by Soxhlet extraction method and further fractionated with petroleum ether, ethyl acetate and chloroform. The fractions were tested for cytotoxicity, apoptosis, scratch wound assay, and cell cycle analysis. IC50, apoptotic index and percentage cell migration were determined using HepG2 cells. For the in vivo studies, hepatocellular carcinoma was induced by administering 0.01% diethylnitrosamine(DEN) in drinking water in Wistar rats. In pre-treatment, rats were co-administered 200 mg/kg of fruit extract with DEN for 14 weeks. In post-treatment, the extract was co-administered after 8-weeks of DEN-induction for 14 weeks. Liver function test, haematological test, oxidative stress markers, relative liver weight, number of cancer nodules and histopathological parameters were determined.Results: Annona muricata fruit extract =significantly lowered cell proliferation counts. The chloroform-fraction possessed higher activity (IC50=(53.7±4.3) μg/mL)The chloroform fraction inhibited cell migration, which was significant compared to curcumin. Further investigations regarding the mode of anticancer activity revealed that the chloroform fraction induced apoptosis. The cell cycle analysis indicated that cells were being arrested at G0/G1. In the in vivo studies, the DEN-control group showed a significant decrease in body weights with increased mortality rate, hepatic nodules, and impairment of liver function compared to normal rats. The rats pre-treated and post-treated with the extract showed positive results with significant improvement in the parameters that were adversely affected by DEN. In addition, other adverse effects of DEN, such as blood dyscrasias and hepatic endogenous antioxidant, were significantly attenuated by Annona muricata fruit extract.Conclusions: The Annona muricata fruit extract has anticancer activity when tested by in vitro and in vivo hepatocellular cancer models.

Cyclic nucleotide phosphodiesterase 3B is connected to osteopontin and protein kinase CK2 in pancreatic <i>β</i>-cells

Islets from RIP-PDE3B mice, exhibiting β-cell specific overexpression of the cAMP/cGMP-degrading enzyme phosphodiesterase 3B (PDE3B) and dysregulated insulin secretion, were subjected to microarray analysis. We show that osteopontin (OPN) mRNA is increased in a dose-dependent manner in islets from RIP-PDE3B mice, as compared to wild-type islets. In addition, in silico analysis shows that PDE3B and OPN are interacting. Furthermore, OPN interacts with protein kinase CK2 ina distinct submodule of the protein-protein interaction network. We studied PDE3B and OPN proteins and, in some cases, also PDE1B and PDE4C, under conditions of relevance for insulin secretion. In the presence of forskolin, PDE inhibitors, insulin, or a protein kinase CK2 inhibitor, similar alterations in protein levels of PDE3B and OPN are shown. In summary, results from using a number of strategies demonstrate a connection between PDE3B and OPNas well as a role for protein kinase CK2 inpancreatic β-cells.

Forkhead box O1 / pancreatic and duodenal homeobox 1 intracellular translocation is regulated by c-Jun N-terminal kinase and involved in prostaglandin E-2-induced pancreatic beta-cell dysfunction

Prostaglandin E-2(PGE(2)) is a well-known mediator of beta-cell dysfunction in both type 1 and type 2 diabetes.We recently reported that down-regulation of the Akt pathway activity is implicated in PGE(2)-induced pancreatic beta-cell dysfunction.The aim of this study was to further dissect the signaling pathway of this process in pancreatic beta-cell line HIT-T15 cells and primary mouse islets.We found that PGE(2) time-dependently increased the c-Jun N-terminal kinase(JNK) pathway activity.JNK inhibition by the JNK-specific inhibitor SP600125 reversed PGE(2)-inhibited glucose-stimulated insulin secretion(GSIS).PGE(2) induced dephosphorylation of Akt and FOXO1, leading to nuclear localization and transactivation of FOXO1.Activation of FOXO1 induced nuclear exclusion but had no obvious effect on the whole-cell protein level of pancreatic and duodenal homeobox 1(PDX1).However, these effects were all attenuated by JNK inhibition.Furthermore, adenovirus-mediated overexpression of dominant-negative(DN)FOXO1 abolished whereas constitutively active(CA)-FOXO1 mimicked the effects of PGE(2) on GSIS in isolated mouse islets.In addition, we demonstrated that DN-JNK1 but not DN-JNK2 or CA-Akt abolished the PGE(2)-induced AP-1 luciferase reporter activity, whereas DN-JNK1 and CA-Akt but not DN-JNK2 reversed the effect of PGE(2) on FOXO1 transcriptional activity, and overexpression of DN-JNK1 rescued PGE(2)-impaired GSIS in mouse islets.Our results revealed that activation of the JNK is involved in PGE(2)induced beta-cell dysfunction.PGE(2)-mediated JNK1 activation, through dephosphorylation of Akt and FOXO1, leads to nuclear accumulation of FOXO1 and nucleocytoplasmic shuttling of PDX1, finally resulting in defective GSIS in pancreatic beta-cells.

丹蛭降糖胶囊对糖尿病性骨质疏松大鼠的治疗作用及机制

【目的】探讨丹蛭降糖胶囊对糖尿病性骨质疏松(DOP)大鼠的治疗作用及机制。【方法】将54只大鼠随机分为假手术组,模型组,中药低、中、高剂量组及吡格列酮组,每组9只。除假手术组外,其余组别大鼠均建立DOP模型,建模过程中感染死亡3只。成功建模1 d后,中药低、中、高剂量组大鼠分别灌胃丹蛭降糖胶囊0.54、1.08、2.16 g·kg^(-1)·d^(-1),吡格列酮组灌胃盐酸吡格列酮10 mg·kg^(-1)·d^(-1),模型组和正常组大鼠灌胃等体积生理盐水,连续8周。给药结束后,检测性激素代谢水平,包括雌二醇(E2)、促卵泡生成素(FSH)、促黄体生成素(LH);检测胰岛功能指标,包括空腹血糖(FPG)、空腹胰岛素(FINS)、稳态模式评估法测定的胰岛素抵抗指数(HOMA-IR)及胰岛素敏感指数(ISI);检测骨代谢指标,包括骨钙素(OC)、血钙(Ca)、血磷(P)和碱性磷酸酶(ALP);苏木素-伊红(HE)染色法观察股骨组织病理形态;末端脱氧核苷酸转移酶介导的dUTP缺口末端标记(TUNEL)法检测胰腺组织胰岛β细胞凋亡;Western Blot法检测股骨组织胰岛素样生长因子1受体(IGF-1R)、Wnt1、β-catenin蛋白表达。【结果】与假手术组比较,模型组E2水平降低,FSH、LH水平升高,胰岛功能指标FPG、FINS及HOMA-IR值升高,ISI值降低,骨代谢指标OC及ALP含量升高,胰岛细胞凋亡率升高,股骨组织IGF-1R蛋白表达水平升高,Wnt1、β-catenin蛋白表达水平降低(均P<0.05),HE染色结果显示骨质疏松特征病理表现;与模型组比较,中药低、中、高剂量组及吡格列酮组E2水平升高,FSH、LH水平降低,胰岛功能指标FINS、FPG及HOMA-IR值降低,ISI值升高,骨代谢指标OC及ALP含量降低,胰岛β细胞凋亡率降低,IGF-1R蛋白表达水平降低,Wnt1、β-catenin蛋白表达水平升高(均P<0.05),HE染色结果显示骨组织病理形态明显改善。中药高剂量组上述各指标与吡格列酮组比较,差异均无统计学意义(P>0.05)。【结论】丹蛭降糖胶囊具有抗糖尿病性骨质疏松作用,其机制可能与改善胰岛功能,调节性激素紊乱,抑制股骨组织IGF-1R表达、激活Wnt1和β-catenin表达,改善骨代谢有关。

邓老御膅膏对2型糖尿病大鼠的影响

目的观察邓老御膅膏(玉米须、黄精、茯苓、山药等)对2型糖尿病大鼠的影响。方法将SD大鼠随机分为正常组、模型组、消渴丸组(阳性对照药,0.78 g·kg^(-1))及邓老御膅膏低、中、高剂量组(5.63、11.25、22.50 g·kg^(-1)),每组12只。采用高脂饮食联合一次性腹腔注射链脲佐菌素(STZ,30 mg·kg^(-1))制备2型糖尿病大鼠模型。造模成功后灌胃给药,每天1次,连续8周。每周记录各组大鼠24 h平均饮水量;给药第2、4、6、8周检测大鼠空腹血糖(FBG);进行糖耐量(OGTT)试验,测定灌服葡萄糖后0、0.5、1、2 h的血糖值,计算时间-血糖曲线下面积(AUC);采用ELISA法测定血清胰岛素(FINS),计算胰岛素抵抗指数(HOMA-IR)与胰岛素敏感性指数(IAI);采用ELISA法测定全血糖化血红蛋白(HbA1c)及血清胰岛素样生长因子1(IGF-1)、C肽(C-P)水平;采用全自动生化分析仪测定血清总胆固醇(TCHO)、甘油三酯(TG)水平;苏木精-伊红(HE)染色观察大鼠胰腺组织病理变化,进行胰腺组织病理学半定量评分。结果与正常组比较,模型组大鼠出现多饮、多食、多尿现象,反应迟钝,第1~8周的饮水量均显著升高(P<0.01);给药第2~8周期间的FBG水平显著升高(P<0.01);口服葡萄糖后不同时间点的血糖水平及AUC值显著升高(P<0.01);血液中的HbA1c含量显著升高(P<0.01);各组大鼠的血清FINS含量无明显差异(P>0.05),模型组大鼠的HOMA-IR显著升高(P<0.01),IAI显著降低(P<0.01);血清IGF-1水平明显降低(P<0.05),C-P、TCHO、TG水平显著升高(P<0.05,P<0.01);胰岛腺泡形态不规则,体积缩小,腺泡细胞核增多,胰岛数量减少,部分胰岛见少量空泡变和胰腺淋巴细胞局灶性浸润,胰腺组织病理评分显著升高(P<0.01)。与模型组大鼠比较,邓老御膅膏给药组大鼠“三多一少”症状及精神状态得到一定改善,第1~7的周饮水量有不同程度降低(P<0.05,P<0.01);给药第2~8周期间的FBG水平有不同程度降低(P<0.05,P<0.01);口服葡萄糖后不同时间点的血糖水平及AUC值有一定程度降低,但差异无统计学意义(P>0.05);血液中的HbA1c含量显著降低(P<0.01);HOMA-IR有一定程度降低,但差异无统计学意义(P>0.05);血清C-P、TG水平明显降低(P<0.05,P<0.01),IGF-1水平有升高趋势,但差异无统计学意义(P>0.05);TCHO水平有下降趋势,但差异无统计学意义(P>0.05);胰岛腺泡形态不规则情况有所改善,胰岛数量有所增加,胰岛空泡变和淋巴细胞浸润减少,胰腺组织病理评分显著降低(P<0.01)。结论邓老御膅膏对2型糖尿病大鼠具有一定降糖作用,可能与其调节脂质代谢、修复胰岛损伤有关。

大鼠胰岛的分离纯化方法改进与功能鉴定

目的通过改进胰腺消化和分离的技术条件,提高成年大鼠胰岛分离纯化产率和质量。方法用胶原酶Ⅺ液灌注消化成年SD大鼠胰腺,对胰岛分离纯化方法加以改进:以4种比重的Euro-Ficoll(F1:D=1.132,F2:D=1.108,F4:D=1.069)Hank’s(F5:D=1.023)不连续密度梯度离心,以离心半径15cm,2000r/min于4℃缓慢升降离心20min,收集位于F1和F2界面的胰岛。双硫腙特异染色法鉴定胰岛纯度;二醋酸酯荧光素/碘化丙啶染色法计算胰岛成活率;放射免疫分析法检测葡萄糖刺激的胰岛素分泌量,计算刺激指数。将胰岛当量(islets equivalent quantity,IEQ)为1000的胰岛移植于同品系糖尿病大鼠肾包膜下,9d内隔日观察动物血糖的变化,评价胰岛功能。比较分离条件优化前后收获胰岛的产率和质量。结果改进纯化方法后每只大鼠胰岛收获量为(920±122)IEQ,胰岛纯度>90%,胰岛细胞成活率为91%±2%。胰岛细胞功能良好,在低糖和高糖刺激后培养液中胰岛素浓度分别为(18.25±0.32)mU/L和(36.70±3.57)mU/L,刺激指数为2.01±0.15。1000 IEQ胰岛移植于糖尿病大鼠肾包膜下,观察期内可维持动物血糖水平正常。结论改进后的胶原酶灌注消化和不连续梯度离心方法提高了胰岛的产率,保证了胰岛的高纯度及高成活率。

山药多糖对体外培养大鼠胰岛细胞活性及胰岛素分泌的影响

为评价山药多糖对胰岛细胞功能的影响,以不同质量浓度葡萄糖刺激胰岛素释放,通过四甲基偶氮唑盐(MTT)法检测胰岛β细胞活性,同时以反转录-聚合酶链式反应(RT-PCR)检测胰岛细胞的抗凋亡基因bcl-2的表达,研究山药多糖与大鼠胰岛细胞共培养对胰岛细胞活性与胰岛素分泌水平的影响,并对其影响机制进行探讨.结果发现,1600 mg/L组胰岛细胞活性稍有下降,但是与对照组没有显著差异(p>0.05);其余各组随培养液中山药多糖质量浓度的增加,胰岛细胞活性增加.低糖或高糖质量浓度环境中实验组与对照组的胰岛素分泌没有显著差异(p>0.05),RT-PCR发现实验组的bcl-2表达显著高于对照组(p<0.05).800 mg/L山药多糖明显提高胰岛细胞的存活率,改善胰岛功能.

多种分离纯化大鼠胰岛细胞的实验方法比较

【目的】采用不同的分离或消化方法 ,探讨如何提高大鼠胰岛细胞分离纯化后的收获量和质量。【方法】将 2 5只供体SD雄性大鼠依胰腺分离或消化的方法不同随机地分为 5组 :①A组 :胆总管灌注胶原酶P ;②B组 :胆总管灌注Hanks液 ;③C组 :胆总管不灌注液体 ;A、B、C 3组胰腺直接分次消化 ;④D组 :胆总管不灌注 ,胰腺剪碎后分次消化 ;⑤对照组 :胆总管灌注胶原酶P ,胰腺直接单次消化。将分离的胰岛沉淀物用Ficoll 4 0 0纯化并培养 ,再把胰岛细胞移植于糖尿病裸鼠受体腹腔内。【结果】纯化后的胰岛细胞在收获量上A和B组明显多于对照组或C组 ,而D组最低 ;A、B和D 3组的纯度均高于对照组或C组 ;在胰岛成活率方面 ,A、B、C 3组均高于对照组或D组。移植于糖尿病裸鼠受体内的大鼠胰岛细胞均可逆转高血糖状态。【结论】经胆总管灌注胶原酶后的大鼠胰腺用分次消化的方法可提高胰岛细胞的收获量、纯度和活性。

桑叶多糖MLPⅡ的基本结构及对糖尿病模型大鼠的降血糖作用

从桑叶中分离纯化获得一种具有降血糖活性的均一多糖组分MLPⅡ。高效液相色谱分析桑叶多糖MLPⅡ主要由甘露糖(man)、鼠李糖(rha)、葡萄糖(glc)、木糖(xyl)和阿拉伯糖(ara)等5种单糖组成,其摩尔比为n(man)∶n(rha)∶n(glc)∶n(xyl)∶n(ara)=8.73∶1.04∶6.53∶2.13∶1.00;红外光谱显示桑叶多糖MLPⅡ的主要连接键为β-糖苷键。桑叶多糖MLPⅡ对糖尿病模型大鼠的降血糖试验结果表明,150 mg/kg MLPⅡ治疗组糖尿病模型大鼠的体质量与对照组糖尿病模型大鼠相比提高了26.22%,而空腹血糖(FBG)浓度降低了52.89%,达到极显著水平,并且治疗组糖尿病模型大鼠的糖化血红蛋白(HbA1c)、空腹胰岛素(FINS)和C-肽(C-P)水平得到改善,损伤胰岛细胞得到修复,胰岛β细胞合成和胰岛素分泌的能力增强。研究结果提示,桑叶多糖MLPⅡ可有效控制糖尿病模型大鼠的体质量和空腹血糖水平,并改善糖耐量,而促进糖尿病模型大鼠胰岛β细胞合成及胰岛素分泌可能是桑叶多糖MLPⅡ降血糖作用机制之一。

大鼠实验性胃溃疡自愈期间胰岛胃泌素和生长抑素细胞的免疫组织化学研究

目的 探讨大鼠实验性胃溃疡自愈期间 ,胰岛胃泌素免疫反应细胞 (gastrin ,G)和生长抑素细胞(somatostatin ,SSorD)的变化。 方法 免疫组织化学ABC技术。 结果 大部分G细胞免疫反应深浅不一 ;溃疡术后 4、10d ,胰岛G细胞面数密度增高 ,与正常或盐水组相比P <0 0 5。D细胞面数密度于 4d增加 ,P <0 0 5。 结论 成年大鼠胰岛细胞呈胃泌素免疫反应阳性 ;胰岛G细胞和D细胞可能以内分泌或旁分泌调节的途径间接或直接参与大鼠实验性胃溃疡修复的过程。

大鼠胰岛内γ-干扰素样免疫反应阳性物质的表达

In order to study the correlation between the neuroendocrine and immune systems, we observed the distribution and expression of interferon γ(IFN-γ) in cells of SD rat pancreas islets using the streptavidin-perosidase immunohistochemical method. A number of IFN-γ-like positive substances were found in the in the cytoplasm of pancreas islet A cells, but the nucleus could not be stained. Most of the IFN-γ-like positive cells were distributed around the periphery of the pancreas islet, the cells were round or oval in shape and the cell body was relatively large. Results suggest IFN-γ may play a role in the regulation of the function of pancreas islet cells and act as a neuro-immune medium .