微囊化大鼠胰岛异种移植的实验研究

目的　通过动物实验明确微囊化胰岛是否具有免疫隔离作用。方法　SD大鼠胰腺原位消化 ,Ficoll间断密度梯度离心法纯化、分离胰岛 ,气流吹喷制作海藻酸钠 /聚赖氨酸 /海藻酸钠(APA)微囊化大鼠胰岛 ,比较微囊化与未微囊化胰岛的胰岛素释放试验 ;将微囊化 (实验组 )与未微囊化 (对照组 )大鼠胰岛植入链脲佐菌素 (STZ)诱导的I型糖尿病小鼠中 ,作两组间血糖正常持续时间比较。结果　实验组与对照组的胰岛素释放试验差异无显著性 (P >0 .0 5 ) ;实验组血糖正常持续时间为 2 3～ 6 5d(平均 48d) ,对照组为 3～ 6d(平均 5d) ,两组差异有极显著性 (P <0 .0 1)。已排斥的实验组小鼠腹腔灌洗发现部分微囊化胰岛存活 ,部分已坏死 ,但微囊膜皆完整 ,囊壁无纤维化。结论　微囊具有良好的免疫隔离作用 ,可使胰岛移植物存活时间明显延长。同时推测微囊内移植物死亡与细胞因子、自由基作用或营养不足等有关。     

微囊化新生猪胰岛样细胞团异种移植

本实验研究新生猪与大鼠产的异种移植。结果显示，新生猪胰岛样细胞团可纠正糖尿病大鼠的高血糖状态；海灌酸钠－多聚赖氨酸微囊能有效保护移植物的存活，平均存活时间为４个月。而微囊作为一种新型防排斥手段，将应用于广泛的领域。     

微囊化胰岛细胞异种移植治疗小鼠糖尿病

我们进行空囊移植及胰岛、微囊化胰岛的异种移植 ,以期寻找影响胰岛存活的原因 ,现将结果报道如下。一、材料和方法1 .糖尿病小鼠的制备 :链脲菌素溶于枸橼酸钠溶液中 ( ｐＨ4.5)制成浓度1 0 ｇ/Ｌ溶液 ,2 2 0ｍｇ/ｋｇ体重小鼠腹腔注入。非空腹血糖连续 2次超过 3.5ｇ/Ｌ则定为已患糖尿病。剪尾以便携式血糖仪测定尾静脉血糖。2 .胰岛细胞的分离和纯化 :胰岛细胞的分离见文献 [1 ]。胰岛细胞纯化 :Ｆｉｃｏｌｌ离心。消化沉淀物和 2 5%Ｆｉｃｏｌｌ溶液混匀 ,其上依次加入 2 3.0 %、2 0 .5%、1 1 .0 %Ｆｉｃｏｌｌ各 2ｍｌ,80 0 ｇ离心 1 5…     

大鼠胰岛微囊化延长异种移植物在糖尿病小鼠体内生存期

本实验用胶原酶消化法分离新生大鼠胰岛，培养和Ｆｉｃｏｌｌ密度梯度离心纯化胰岛。用海藻酸－聚赖氨酸－海藻酸生物膜包裹胰岛并进行体外胰岛素分泌的能测定和体内异种移植实验。１０只链脲霉素诱导的糖尿病小鼠腹腔内移植微囊化胰岛３天后，小鼠血糖降至正常，体重逐渐增加，糖尿病得到纠正。维持最长者达１３０天，平均８５．２天。明显优于单纯胰岛植组，其功能维持不足９天。肾脏病理组织染色和透射电镜显示早期微囊化胰岛移植     

微囊化胰岛移植的研究进展

免疫隔离膜微囊化胰岛移植，通过一个人造屏障将胰岛与宿主的免疫系统隔离开业，为解决排斥反应开辟了新的思路和途径。在糖尿病动物模型上获得了令人鼓舞的效果，本文就海藻酸钠微囊的制备、免疫屏障作用、移植数量和移植后纤维化进行了探讨。     

改良的微囊化新生猪胰岛细胞移植治疗Ⅰ型糖尿病八例报告

在动物实验的基础上，对８例Ⅰ型糖尿病患者进行了改良的微囊化新生猪胰岛细胞移植治疗。８例患者中，３例移植于网膜囊，５例移植于三角肌，于移植前及移植后１、６、１２个月观察空腹血糖（ＦＢＧ）、糖基化血红蛋白Ａ１ｃ（ＨｂＡ１ｃ）、Ｃ－酞、ＣＤ４阳性细胞、ＣＤ８阳性细胞、可溶性白细胞介素２受体和胰岛素用量（ＩＮＳ）的变化。结果显示，移植前后Ｔ细胞亚群无明显变化，提示无排斥反应发生；结果还显示移植的猪胰岛数量决定了疗效：移植量少于１０个猪胰的胰岛（约３２６１８０±１２１３０个胰岛），移植后ＩＮＳ减量少于４０％，Ｃ－酞升高不明显，移植量达到１０个猪胰的胰岛，术后ＩＮＳ减量＞７５％，Ｃ－酞峰值是术前的２．５６倍。提示此种方法移植的必需量为１０个新生猪胰的胰岛。     

海藻酸-壳聚糖-聚乙烯乙二醇微囊化异种胰岛移植治疗小鼠糖尿病

目的比较海藻酸-聚赖氨酸-海藻酸微囊化(APA)大鼠胰岛和海藻酸-壳聚糖-聚乙烯乙二醇微囊化(ACP)大鼠胰岛移植对小鼠糖尿病的治疗作用.方法链脲霉素220 mg/kg体重腹腔内注射制作小鼠糖尿病动物模型.成模后分成APA微囊组、ACP微囊组和未微囊化胰岛组,每只小鼠腹腔内分别植入200个相应胰岛,检测监测移植前后小鼠的血糖.结果移植后第1天,3组小鼠血糖均较移植前明显降低.未微囊化胰岛移植组小鼠术后第2天血糖均恢复到术前水平;APA微囊组和ACP微囊组小鼠血糖分别维持正常(57.00±14.61)d和(41.67±16.73)d,此两组间差异无显著性(P＞0.05),但均较未微囊化胰岛移植组明显延长(P＜0.05).结论 ACP微囊与APA微囊一样具有免疫隔离作用;ACP微囊化异种胰岛移植可纠正小鼠糖尿病.     

微囊化大鼠胰岛异种移植治疗小鼠实验性糖尿病的研究

目的 研究海藻酸钠－聚赖氨酸－海藻酸钠包裹胰岛进行移植的效果。方法 将Ｗｉｓｔａｒ大鼠的胰腺先行胶原酶胰管内注射消化,然后分离,纯化,所得胰岛经培养后制成微囊包膜的胰岛,微囊直径为０．４￣０．５ｍｍ,每个微囊内包１个胰岛。     

微囊包膜猪胰岛异种移植的实验与临床研究

目的 探讨微囊化新生猪胰岛作为临床异种胰岛移植供体来源治疗Ⅰ型糖尿病的可能性。方法 利用微囊技术包膜新生猪胰岛细胞后体外培养，比较微囊化及未微囊化新生猪胰岛细胞的胰岛素分泌能力和胰岛素刺激释放试验。采用腹腔镜技术，对３例Ⅰ型糖尿病患者进行微囊新生猪胰岛异种移植治疗，观察移植前后空腹血糖、Ｃ肽和胰岛素用量的变化。结果 微囊与未微囊化新生猪胰岛在体外培养中各项检测指标差异无显著意义，两者均具有良好的生物活性。临床移植术后，２例受体的Ｃ肽峰值较移植前分别升高１１及２３倍，其中１例胰岛素用量减少６３％，１例完全停用胰岛素，成为胰岛素不依赖者。此２例的移植胰岛在受体内保持有功能存活已达８０ｄ。结论 海藻酸钠－聚赖氨酸－海藻酸钠微囊化新生猪胰岛生物活性良好，能在人体内功能存活并产生疗效，将可能是临床上异种胰岛移植的一个较好     

腹腔镜微囊化猪胰岛小网膜腔内异种移植的临床研究

目的 探讨腹腔镜技术应用于海藻酸钠－聚赖氨酸－海藻酸钠（APA）微囊包裹猪胰岛小网膜腔内异种移植术的可行性。方法 采用腹腔镜技术对5例6人次I型糖尿病病人进行了APA微囊新生猪胰岛小网膜腔内异种移植治疗，观察病人移植前后空腹血糖，C肽和胰岛素用量变化。结果 全部病例均未出现手术并发症。术后受体C肽较术前升高3－23倍，胰岛素用量明显减少，其中1例完全停用胰岛素，成为胰岛素不依赖者达31个月。结论 腹腔镜技术用于临床微囊化新生猪胰岛小网膜腔移植术治疗I型糖尿病安全可行，有较高的临床应用价值。     

经腹腔镜微囊化猪胰岛异种移植术

目的：探讨腹腔镜技术应用于海藻酸钠－聚赖氨酸－海藻酸钠（APA）微囊包裹猪胰岛小网膜腔内异种移植术的可行性。方法：采用腹腔镜技术对5例6人次I型糖尿病患者进行了APA微囊新生猪胰岛小网膜腔内异种移植治疗，观察患者移植前后空腹血糖、C肽和胰岛素用量变化。结果：全部病例均未出现手术并发症。术后受体C肽较术前升高3－23倍，胰岛素用量减少，基中1例完全停用胰岛素，成为胰岛素不依赖者达31个月。结论：腹腔镜技术用于临床微囊化新生猪胰岛小网膜腔移植术治疗I型糖尿病安全可行。     

Reluctance of French patients with type 1 diabetes to undergo pig pancreatic islet xenotransplantation

INTRODUCTION: Type 1 diabetes could possibly be treated by transplantation of pig pancreatic islets. In addition to medical difficulties and ethical problems, social hurdles may need to be overcome. We have evaluated the attitude of patients with type 1 diabetes to the xenotransplantation of pig pancreatic islets and to the potential risks associated with such treatment. METHODS: A survey of 214 patients with type 1 diabetes was carried out in France based on a multiple-choice questionnaire. RESULTS: At first, 52.0% of these patients indicated that they would agree to receive pig islet xenografts. The main sources of reluctance were the 'risk of disease transmission' (55.5%) and 'risks not yet identified' (48.7%). After they were told of the risk of cancer or infection associated with immunosuppression, 74.9% of the respondents chose to refuse the transplant, compared with 48.0% before they heard of such risks. A 68.1% would refuse the xenotransplant if it would not exempt them completely from being treated by insulin injections. Discontinuing insulin injections was the most important priority for diabetic patients (73.5%), rather than limitation of diabetes-related complications (52.5%) or increase in life expectancy (44.0%). After they were informed of all of the risks associated with the procedure, 70.5% of the respondents decided they would rather not take any risks, and said they would refuse pig islet transplantation. CONCLUSION: When diabetic patients learned about potential infectious risks and other risks associated with immunosuppression, reluctance to undergo xenotransplantation gained in significance or even led to refusal of the procedure.     

建立豚鼠至大鼠异种肝移植模型的实验研究

目的 建立豚鼠至大鼠异种肝移植动脉化动物模型。方法 利用血管套技术和显微外科技术进行了30例豚鼠至大鼠肝移植。观察受者存活时间和移植肝组织学及超微结构变化。结果移植大鼠存活时间平均为（98.00±26.38）min,肝细胞发生弥漫性水样变性,中央静脉和小叶间静脉淤血。肝细胞内糖原颗粒消失,细胞器水肿。结论 成功地建立了非协调性异种肝移植动脉化模型。能观察移植肝的超急性排斥反应变化。     

Parameters favouring successful adult pig islet isolations for xenotransplantation in pig-to-primate models

Dufrane D, D'hoore W, Goebbels R‐M, Saliez A, Guiot Y, Gianello P. Parameters favouring successful adult pig islet isolations for xenotransplantation in pig‐to‐primate models.
Xenotransplantation 2006; 13: 204–214. © Blackwell Munksgaard, 2006 Abstract: Background: In the near future, adult porcine islets of Langerhans appear as an unlimited source of insulin‐producing cells which could play a major role for treating diabetes mellitus. There is, however, an obvious lack of pre‐clinical results and data in the pig‐to‐primate model. One of the main hurdles of this model is certainly related to the difficulty of reproducing regularly successful porcine islet isolation. This experimental work was designed to provide guidelines applicable in pig pancreas procurement and islet isolation for successful islet xenotransplantation into primates. Methods: Pancreases were harvested from adult Belgium Landrace pigs (n = 79) in a single centre. The impact on islet yield of (1) pancreas procurement (blood exsanguination and warm ischaemia time (WIT)), (2) cold storage solutions (classic UW and modified UW (without hydroxyethyl starch and inverse K⁺/Na⁺ concentration)), (3) a dynamic or static method of pancreas digestion, and (4) the endotoxin content and enzymatic activity from five different batches of Liberase PI was studied. In addition, pancreatic biopsies (n = 18), performed before isolation, were retrospectively analyzed to study the impact of histomorphometry on porcine islet yield. Finally, two diabetic cynomolgus monkeys were transplanted without immunosuppression with 15 000 pig islet equivalents/kg body weight of recipient to assess in vivo the function of freshly isolated islets. Univariate and multivariate analyses were performed. Results: By multiple linear regression, the most significant variables that significantly improved islet yield were, firstly, the presence of <30 EU (endotoxin units) of endotoxin in Liberase batches, followed by a WIT under 10 min and the use of blood exsanguination before pancreas harvesting (P<0.005). In contrast, isolation method (dynamic vs. static) and the solution used for storage (short‐term) (UW vs. modified UW) did not significantly influence islet yield. The correlation of retrospective histomorphometry analysis of native pancreas and extemporaneous biopsy before isolation clearly determined a positive relationship between isolated islet number and the number of islets/cm² (r = 0.708, P<0.01) or with the percentage of large islets (r = 0.680, P<0.01) found in pancreas biopsies. Pig pancreases containing more than 82 islets/cm² and more than 42% of large islets (>100 μm) thus enabled more than 120 000 islet equivalents to be obtained in 90% of the cases, which is an ideal amount of islets to transplant into a primate of 4 to 5 kg. In vivo, a reduction of blood glucose (<200 mg/dl), associated with porcine C‐peptide production, was observed in two primates after transplantation with adult pig islets. At day 7 post‐transplantation, however, loss of islet function was associated with graft destruction and immune reaction. Conclusions: Morphological screening of the pig pancreas before isolation, optimal blood exsanguination, WIT <10 min, and an endotoxin content <30 EU/mg in Liberase PI batches determine successful pig islet isolation for xenotransplantation in primates.     

微囊化新生猪甲状旁腺细胞异种移植的实验研究

目的 探讨微囊化新生猪甲状旁腺细胞异种移植治疗大鼠甲状旁腺功能低下症的效果。方法 应用微囊化技术，制备微囊化（海藻酸钠－聚赖氨酸－海藻酸钠生物微胶囊）新生猪甲状旁腺细胞，32只去甲状旁腺的Wistar大鼠随机分成微囊组、非微囊组、空囊组和对照组，分别移植微囊化新生猪甲状旁腺细胞、甲状旁腺细胞、空微囊及生理盐水。移植后监测血钙及甲状旁腺素水平40周，40周后回收移植物，透射电镜检查。结果 移植后，微囊组大鼠血钙及甲状旁腺素水平恢复到正常范围内，直至观察结束时（40周），透射电镜检查显示移植物存活良好；非微囊组、空囊组和对照组大鼠的血钙及甲状旁腺素水平无改善。结论 微囊化新生猪甲状旁腺细胞异种移植在不用免疫抑制剂情况下，可以在大鼠体内存活，且有功能；海藻酸钠－聚赖氨酸－海藻酸钠生物微胶囊对免疫活性细胞及抗体具有屏蔽作用。