人胚胎干细胞体外定向诱导分化胰腺前体细胞及胰岛细胞移植治疗NOD/SCID糖尿病小鼠

目的 探讨人胚胎干细胞（human embryonic stem cells,hESCs）体外定向诱导分化胰腺前体细胞及胰岛细胞移植治疗非肥胖糖尿病/严重联合免疫缺陷（non-obese diabetic/severe combined immunodeficient,NOD/SCID）小鼠的可行性.方法 体外分4阶段诱导hESCs定向分化为胰岛细胞：①诱导分化形成定型内胚层;②诱导胰腺细胞定向分化;③扩增胰腺前体细胞;④促进胰岛细胞成熟.观察诱导各阶段细胞形态变化、免疫荧光鉴定胰十二指肠同源异型盒基因（PDX-1）、胰高糖素（glucagon）、胰岛素（insulin）、C肽（C-peptide）、葡萄糖转运子2（Glut-2）的表达.3阶段及4阶段分化形成的细胞分别植入链脲菌素（streptozotocin,STZ）诱导形成的NOD/SCID糖尿病小鼠一侧附睾脂肪垫内,观察血糖变化.结果 hESCs诱导4阶段细胞表达胰高血糖素、胰岛素、Glut-2;共表达PDX-1和C肽;流式鉴定诱导4阶段胰岛素阳性细胞为17.1％;体外检测有葡萄糖刺激的胰岛素释放反应;3阶段及4阶段分化形成的细胞分别植入NOD/SCID 糖尿病小鼠体内可逆转其高血糖至少12周.结论 体外定向诱导hESCs分化形成的胰腺前体细胞及胰岛细胞分别植入NOD/SCID小鼠可逆转其高血糖.     

小鼠胰岛α细胞的缺失对移植胰岛功能的影响

目的　探讨胰岛α细胞的丢失对胰岛素分泌功能的影响。方法　在人胰岛分离过程中 ,经胶原酶的过度消化造成胰岛周边α细胞缺失 ,用免疫组化及胰岛素 /胰高糖素含量分析予以证实。在体外 ,检测葡萄糖刺激引起的实验性糖尿病小鼠胰岛素的分泌 ;在体内 ,评价胰岛移植后对糖尿病小鼠血糖的调节 ,并研究胰高糖素对α细胞缺失胰岛功能的影响。结果　胶原酶的过度消化引起胰岛周边的α细胞丢失 ,使分离胰岛内胰岛素 /胰高糖素比值明显升高。与正常胰岛相比 ,α细胞缺失胰岛对葡萄糖刺激产生的胰岛素明显降低 ,胰岛素释放在正常胰岛和α细胞缺失胰岛分别为2 2 2 7uU/ml± 32 1uU/ml和 12 4 6uU/ml± 12 6uU/ml(P <0 0 1)。在体内 ,移植α细胞缺失的胰岛到糖尿病的C5 7/BL小鼠后不能有效地纠正动物的高血糖 ,胰岛移植后的平均血糖浓度在正常胰岛组和α细胞缺失胰岛组分别为 :8 9mmol/L± 1 98mmol/L和 2 1 3mmol/L± 2 2mmol/L(P <0 0 1) ,而给予外源性的胰高糖素可以在体外及体内明显改善α细胞缺失胰岛的胰岛素分泌功能。结论　胰岛α细胞的缺失明显降低胰岛的胰岛素分泌功能 ,用外源性的胰高糖素可以改善α细胞缺失胰岛的胰岛素分泌功能。     

Somatostatin receptor expression and biological functions in endocrine pancreatic cells: review based on a doctoral thesis

Type 1 diabetes is resulting from the selective destruction of insulin-producing betacells within the pancreatic islets. Somatostatin acts as an inhibitor of hormone secretion through specific receptors (sst1-5). All ssts were expressed in normal rat and mouse pancreatic islets, although the expression intensity and the co-expression pattern varied between ssts as well as between species. This may reflect a difference in response to somatostatin in islet cells of the two species. The Non-Obese Diabetic (NOD) mouse model is an experimental model of type 1 diabetes, with insulitis accompanied by spontaneous hyperglycaemia. Pancreatic specimens from NOD mice at different age and stage of disease were stained for ssts. The islet cells of diabetic NOD mice showed increased islet expression of sst2-5 compared to normoglycemic NOD mice. The increase in sst2-5 expression in the islets cells may suggest either a contributing factor in the process leading to diabetes, or a defense response against ongoing beta-cell destruction. Somatostatin analogues were tested on a human endocrine pancreatic tumour cell line and cultured pancreatic islets. Somatostatin analogues had an effect on cAMP accumulation, chromogranin A secretion and MAP kinase activity in the cell line. Treatment of rat pancreatic islets with somatostatin analogues with selective receptor affinity was not sufficient to induce an inhibition of insulin and glucagon secretion. However, a combination of selective analogues or non-selective analogues via costimulation of receptors can cause inhibition of hormone production. For insulin and glucagon, combinations of sst2 + sst5 and sst1 + sst2, respectively, showed a biological effect. In summary, knowledge of islet cell ssts expression and the effect of somatostatin analogues with high affinity to ssts may be valuable in the future attempts to influence beta-cell function in type 1 diabetes mellitus, since down-regulation of beta-cell function may promote survival of these cells during the autoimmune attack.     

肾上腺髓质素对自发性高血压大鼠胰岛分泌功能影响的体外实验

目的　探讨肾上腺髓质素 (AM)对自发性高血压大鼠 (SHR)和 Wistar- Kyoto(WKY)鼠胰岛分泌功能的影响。方法　分别在含 2 .8mm ol/ L 及 11.1mm ol/ L 葡萄糖的 RPMI16 4 0培养基中加不同浓度 AM(0、10 - 8、10 - 7和 10 - 6 m ol/ L )培养 SHR和 WKY大鼠胰岛 ,用放射免疫分析 (RIA )法测定两次培养液的胰岛素及胰升糖素含量。结果　加入不同浓度 AM培养 WKY鼠胰岛 2 4 h后 ,培养液中胰岛素含量呈剂量依赖性降低 ;高浓度 AM(10 - 6 mol/ L )使 SHR和胰岛培养液中胰岛素含量明显降低 (P<0 .0 5 )。SHR、WKY鼠的各组培养液中胰升糖素含量无明显差异 ,SHR与 WKY鼠相比亦无差异。结论　 AM对 WKY鼠的胰岛β细胞的分泌功能具有剂量依赖性抑制作用 ,高浓度 AM对 SHR胰岛β细胞的分泌功能具有抑制作用。 AM对胰岛α细胞分泌功能无影响     

Fas配体表达对同种胰岛移植的影响

目的探究睾丸细胞FasL表达能否对共移植的胰岛移植物提供免疫豁免作用以及胰岛细胞FasL基因转染对同种胰岛移植的影响。方法　将同种大鼠胰岛及睾丸细胞同时移植于糖尿病受体，重组腺病毒AdV-FasL感染胰岛细胞后移植，观察移植物存活情况、胰岛功能，并检测移植物内浸润淋巴细胞以及基因转染胰岛细胞凋亡情况。结果　单纯移植胰岛组平均存活期为(6.3±0.6)d。与胰岛细胞同时移植的睾丸细胞数增加至1×107时，存活期大于50d（P<0.05）。表达FasL的睾丸细胞在移植物内诱导浸润淋巴细胞凋亡。FasL基因转染组出现排斥加速，存活期缩短至(3.4±0.2)d。FasL转染的胰岛细胞在移植后24h见FasL表达，48h表达增强，移植后FasL转染胰岛细胞凋亡。结论　表达FasL的睾丸细胞与胰岛同时移植可诱导活化的淋巴细胞凋亡，使胰岛移植物获得免疫豁免、存活期延长，但通过FasL基因转染使胰岛细胞直接表达FasL引起胰岛细胞凋亡和粒细胞浸润，导致排斥加速。     

The effect of FasL expression on pancreatic islet allografts

目的　探究睾丸细胞FasL表达能否对共移植的胰岛移植物提供免疫豁免作用以及胰岛细胞FasL基因转染对同种胰岛移植的影响。方法　将同种大鼠胰岛及睾丸细胞同时移植于糖尿病受体 ,重组腺病毒AdV FasL感染胰岛细胞后移植 ,观察移植物存活情况、胰岛功能 ,并检测移植物内浸润淋巴细胞以及基因转染胰岛细胞凋亡情况。结果　单纯移植胰岛组平均存活期为 (6 3± 0 6 )d。与胰岛细胞同时移植的睾丸细胞数增加至 1× 10 7时 ,存活期大于 6 0d(P <0 0 5 )。表达FasL的睾丸细胞在移植物内诱导浸润淋巴细胞凋亡。FasL基因转染组出现排斥加速 ,存活期缩短至 (3 4± 0 2 )d。FasL转染的胰岛细胞在移植后 2 4h见FasL表达 ,4 8h表达增强 ,移植后FasL转染胰岛细胞凋亡。结论　表达FasL的睾丸细胞与胰岛同时移植可诱导活化的淋巴细胞凋亡 ,使胰岛移植物获得免疫豁免、存活期延长 ,但通过FasL基因转染使胰岛细胞直接表达FasL引起胰岛细胞凋亡和粒细胞浸润 ,导致排斥加速。     

Role and mechanism of rosiglitazone on the impairment of insulin secretion induced by free fatty acids on isolated rat islets

Background Prolonged exposure of pancreatic β-cells to fatty acids increases basal insulin secretion but inhibits glucose-stimulated insulin secretion. Rosiglitazone is a new antidiabetic agent of the thiazolidinediones. However, the relationship between thiazolidinediones and insulin secretion is highly controversial. The aim of this study is to explore the effect and mechanism of rosiglitazone on insulin secretion of islets under chronic exposure to free fatty acids （FFA）. Methods Pancreatic islets were isolated from the pancreata of male Sprague-Dawley rats by the collagenase digestion and by the dextran gradient centrifugation method. The purified islets were cultured in the presence or absence of rosiglitazone and palmitate for 48 hours. The insulin secretion was measured by radioimmunoassay. The mRNA level of peroxisome proliferator-activated receptor y, uncoupling protein 2 0dCP-2） and insulin were determined by real-time polymerase chain reaction （PCR）. The cell cytotoxicity assay was measured by cell counting kit-8. Results Islets exposed to elevated palmitate for 48 hours showed an increased basal and a decreased glucose-stimulated insulin secretion （P〈0.01）. The mRNA level of UCP-2 was increased by 3.7 fold in the 0.5 mmol/L concentration of palmitate. When islets were cultured with palmitate （0.5 mmol/L） in the presence of rosiglitazone （1.0 pmol/L）, both basal and glucose-stimulated insulin secretion reversed to a pattern of control islets （P〈0.05, P〈0.0 1）. The addition of rosiglitazone in the culture medium decreased the mRNA level of UCP-2 by 2.2 fold, having a statistically significant difference （P〈0.05） as compared with islets cultured with palmitate alone. The cell viability was not affected. Conclusion The protective effects of rosiglitazone on insulin secretion of isolated pancreatic islets under chronic exposure to palmitate might be mediated through the downregulation of UCP-2 expression.     

Pulsatility of insulin release – a clinically important phenomenon

Abstract

The mechanisms and clinical importance of pulsatile insulin release are presented against the background of more than half a century of companionship with the islets of Langerhans. The insulin-secreting β-cells are oscillators with intrinsic variations of cytoplasmic ATP and Ca²⁺. Within the islets the β-cells are mutually entrained into a common rhythm by gap junctions and diffusible factors (ATP). Synchronization of the different islets in the pancreas is supposed to be due to adjustment of the oscillations to the same phase by neural output of acetylcholine and ATP. Studies of hormone secretion from the perfused pancreas of rats and mice revealed that glucose induces pulses of glucagon anti-synchronous with pulses of insulin and somatostatin. The anti-synchrony may result from a paracrine action of somatostatin on the glucagon-producing α-cells. Purinoceptors have a key function for pulsatile release of islet hormones. It was possible to remove the glucagon and somatostatin pulses with maintenance of those of insulin with an inhibitor of the P2Y₁ receptors. Knock-out of the adenosine A₁ receptor prolonged the pulses of glucagon and somatostatin without affecting the duration of the insulin pulses. Studies of isolated human islets indicate similar relations between pulses of insulin, glucagon, and somatostatin as found during perfusion of the rodent pancreas. The observation of reversed cycles of insulin and glucagon adds to the understanding how the islets regulate hepatic glucose production. Current protocols for pulsatile intravenous infusion therapy (PIVIT) should be modified to mimic the anti-synchrony between insulin and glucagon normally seen in the portal blood.     

Influence of microenvironment on engraftment of transplanted β-cells

Pancreatic islet transplantation into the liver provides a possibility to treat selected patients with brittle type 1 diabetes mellitus. However, massive early β-cell death increases the number of islets needed to restore glucose homeostasis. Moreover, late dysfunction and death contribute to the poor long-term results of islet transplantation on insulin independence. Studies in recent years have identified early and late challenges for transplanted pancreatic islets, including an instant blood-mediated inflammatory reaction when exposing human islets to the blood microenvironment in the portal vein and the low oxygenated milieu of islets transplanted into the liver. Poor revascularization of remaining intact islets combined with severe changes in the gene expression of islets transplanted into the liver contributes to late dysfunction. Strategies to overcome these hurdles have been developed, and some of these interventions are now even tested in clinical trials providing a hope to improve results in clinical islet transplantation. In parallel, experimental and clinical studies have, based on the identified problems with the liver site, evaluated the possibility of change of implantation organ in order to improve the results. Site-specific differences clearly exist in the engraftment of transplanted islets, and a more thorough characterization of alternative locations is needed. New strategies with modifications of islet microenvironment with cells and growth factors adhered to the islet surface or in a surrounding matrix could be designed to intervene with site-specific hurdles and provide possibilities to improve future results of islet transplantation.     

移植部位对移植胰岛长期存活的影响

目的 观察移植部位对糖尿病小鼠移植胰岛长期存活的影响.方法 采用胶原酶P溶液胰管灌注和Ficoll-400不连续密度梯度纯化法获取供体小鼠胰岛.受体糖尿病小鼠分别接受同基因和异基因肾被膜下、小网膜囊和腋窝胰岛移植,监测受体移植后的血糖变化;根据血糖情况取移植胰岛排斥标本,观察不同部位移植胰岛的组织学变化.结果 成功分离、纯化供体小鼠胰岛,纯度为( 94±5)％或(90±5)％,存活率为(92±3)％.胰岛移植后均能使受体小鼠高血糖逆转至正常.同基因或异基因胰岛移植短期存活状态:肾被膜组与小网膜囊组的血糖降低值比较差异无统计学意义(P＞0.05),移植后肾被膜组血糖值显著低于小网膜囊组(P＜0.05);腋窝组血糖降低值和血糖值与肾被膜组和小网膜囊组比较,差异均有统计学意义(P＜0.05).同基因或异基因胰岛移植长期存活状态:肾被膜组与小网膜囊组血糖降低值比较差异有统计学意义(P＜0.05),且肾被膜组血糖值较低,移植胰岛存活率较高;腋窝组移植胰岛不能达到长期存活;在同一部位,同基因与异基因胰岛移植受体的血糖降低值比较,差异有统计学意义(P＜0.05).组织学检查发现:肾被膜组和小网膜组胰岛完整,有少量炎症细胞浸润;腋窝组胰岛细胞被破坏,大量炎症细胞浸润.结论 肾被膜下胰岛移植降血糖效果迅速、稳定,能达到长期存活的目的,可视为胰岛移植的理想部位.     

宫内生长迟缓大鼠胰腺ghrelin和胰岛素表达变化

目的：探讨宫内生长迟缓大鼠胰岛ghrelin表达规律，阐明ghrelin是否参与宫内生长迟缓所致的胰岛素抵抗的进程。方法：建立宫内生长迟缓（Ⅰ组）和正常出生体重（N组）大鼠模型；测定血糖、血ghrelin、血胰岛素水平、胰腺ghrelin含量和高糖刺激后胰腺胰岛素分泌量；RTPCR检测胰腺ghrelin和胰岛素mRNA水平；免疫荧光和共聚焦技术检测胰岛ghrelin和胰岛素的阳性细胞数。结果：Ⅰ组1d、3d和7d的血浆胰岛素水平均低于N组（P〈0．05），胰岛素抵抗指数（HOMA—IR）也低于N组相应时间点（P〈0．05）。两组血糖、血浆胰岛素、HOMA—IR和血浆ghrelin水平有随时间变化的趋势（一’：4．12～125．97，P均〈0．001）。I组血浆ghrelin水平与HOMA—IR呈正相关（r=0．553，P=0．000）。两组胰腺ghrelin含量、ghrelinmRNA、insulin’细胞和ghrelin’细胞比例均随日龄增长呈下降趋势（F=-49．29～427．28，P均〈0．001），两组间ghrelin含量和mRNA变化差异显著（F=-69．98～169．22，P均〈0．05）；Ⅰ组和N组胰岛素分泌水平分别与其胰腺ghrelin含量呈负相关（r=-0．895，P=0．000；r=-0．458，P=0．002），胰岛ghrelin’细胞比例分别与其胰腺insulin’细胞比例呈负相关（r=-0．810，P=0．000；r=-0．714，P=0．000）。两组大鼠胰岛ghrelin’细胞分布不同。结论：宫内生长迟缓对血浆ghrelin水平、胰岛素分泌的影响在生后持续存在，ghrelin参与了宫内生长迟缓者发生胰岛素抵抗的进程。     

骨髓干细胞在大鼠移植肝中的诱导分化

目的 研究骨髓干细胞在大鼠移植肝中的诱导分化情况及对大鼠长期存活的影响.方法 雌性受体大鼠随机分3组:空白对照组(A组)、D-hanks液组(B组)、骨髓干细胞组(C组).观察大鼠的中位生存时间、肝组织病理变化,基因Sry原位杂交和甲胎蛋白免疫组化双检测观察骨髓干细胞的分化情况.结果 C组中位生存时间大于180 d(P＜0.05);C组无明显的急性排斥反应.基因Sry原位杂交阳性,并且表达甲胎蛋白.结论 骨髓干细胞门静脉输注能减轻急性排斥反应,诱导大鼠肝移植术后长期存活;并能在移植肝的环境中诱导分化为肝细胞,表达甲胎蛋白,并逐渐替代移植肝本身的细胞.     

成年大鼠胰腺导管干细胞的体外分离?培养及鉴定

目的:建立成年大鼠胰腺导管干细胞的体外分离、培养及鉴定的方法.方法:V型胶原酶溶液灌注消化成年大鼠胰腺组织,并经过Ficoll密度梯度离心去除胰岛组织,培养于含10%胎牛血清的RPMI1640培养液,而后加入表皮生长因子(EGF)和碱性成纤维细胞生长因子(bFGF)继续培养,7天可形成单层细胞,用0.25%胰酶-EDTA消化并传代培养.取第2代细胞利用免疫荧光染色和RT-PCR方法检测CK19、Pdx-1、Nestin、insulin及glucagon的表达.结果:经胶原酶灌注消化胰腺导管上皮,再经过Ficoll密度梯度离心去除胰岛组织,可使胰腺导管细胞得到较好的纯化.免疫荧光结果示:胰腺导管细胞CK19、Pdx-1和Nestin染色呈阳性,阳性细胞率分别为(88.6±6.2)%、(84.6±8.6)%和(79.3±10.5)%,而insulin及glucagon染色为阴性.RT-PCR结果显示该细胞表达CK19、Pdx-1和Nestin基因.结论:该方法可较好的分离出胰腺导管细胞.经鉴定该法培养所获细胞具有胰腺干细胞的特性.     

白细胞介素-1对大鼠离体胰岛B细胞分泌功能的影响

目的 :观察白细胞介素 1(IL 1)对大鼠胰岛B细胞分泌功能的影响并探讨其机制。方法 :采用胰岛细胞培养的方法 ,观察IL 1对胰岛素分泌和MTT还原的影响 ;通过分子杂交检测c fosmRNA并分析蛋白质合成的变化。结果 :IL 1作用后胰岛素分泌和MTT还原量明显减少 ,且有时间依赖性 ;IL 1诱导c fosmRNA ,从而使细胞蛋白质表达发生变化。结论 :IL 1可能通过改变胰岛细胞内基因表达 ,抑制细胞代谢 ,从而使胰岛素分泌减少     

高脂饮食肥胖大鼠胰岛细胞胰岛素抵抗机理的探讨

目的探讨高脂饮食诱导的肥胖大鼠在具有外周胰岛素抵抗(IR)的情况下,胰岛胰岛素、胰高血糖素的分泌和合成功能,高糖刺激下胰高血糖素和胰岛素的分泌以及胰岛内胰岛素信号转导分子的改变。方法30只雄性Wistar大鼠随机分为高脂饲料喂养的肥胖组和普通饲料喂养的对照组,每组各15只,共喂养20周。采用胰腺组织匀浆,检测胰岛素和胰高血糖素的含量;胰岛细胞表面灌注检测高糖状态胰岛素和胰高血糖素的动态分泌变化;免疫组织化学染色及图像分析检测胰岛素受体(IRc)及胰岛素受体底物1、2(IRS1、IRS2)在两组大鼠胰岛的表达。结果(1)肥胖组胰岛素敏感指数(ISI)明显低于对照组,肥胖组血胰高血糖素水平和胰岛内的胰高血糖素水平均显著高于对照组(362pg/ml±58pg/mlvs291pg/ml±35pg/ml,P<0.05;442pg/ml±56pg/mlvs287pg/ml±48pg/ml,均P<0.05)。(2)肥胖组葡萄糖刺激的胰岛素分泌(GSIS)受损,16.7mmol/L葡萄糖可显著抑制对照组胰岛α细胞胰高血糖素的分泌,而在肥胖组这种抑制作用消失。(3)胰岛存在IRc、IRS1和IRS2的表达。肥胖组胰岛IRc、IRS2的表达较对照组胰岛分别低28%和22%(均P<0.01)。结论高脂饮食诱导的肥胖大鼠胰岛细胞的胰岛素信号转导通路受损,即在有外周IR的同时也具有胰岛内的IR,这可能是肥胖状态下胰岛细胞功能障碍的内在机制之一。     

重症急性胰腺炎胰腺内分泌功能的实验研究

目的:观察重症急性胰腺炎(SAP)大鼠胰腺内分泌功能的变化。方法:雄性Wistar大鼠48只,随机分为假手术组(SO组)、重症急性胰腺炎组(SAP组),每组24只。胆胰管逆行注射5%牛磺胆酸钠制备SAP模型。术后(3、6、12)h分批剖杀大鼠,每个时间点8只。肉眼观察胰腺形态学变化,取胰头部胰腺组织行病理学检查并评分。生化法检测血清淀粉酶(AMY),放射免疫分析法测定血清胰岛素(INS)、C-肽(CP)和胰高血糖素(GG)的变化。结果:SAP大鼠造模成功后,随时间延长胰腺外分泌部损伤逐渐加重,并在12h达高峰,但内分泌部胰岛仅在12h出现形态学变化;SAP组3,6和12h组大鼠血清INS和CP水平与SO组相比升高,差异有统计学意义(P<0.05)。SAP组6和12h大鼠血清胰高血糖素与SO组相比明显升高,差异有统计学意义(P<0.05)。结论:5%牛磺胆酸钠制备的SAP模型中,出现了胰腺内分泌部的损伤,表现为其分泌激素水平的紊乱。     

Glucose control of glucagon secretion—‘There’s a brand-new gimmick every year’

Glucagon from the pancreatic α-cells is a major blood glucose-regulating hormone whose most important role is to prevent hypoglycaemia that can be life-threatening due to the brain’s strong dependence on glucose as energy source. Lack of blood glucose-lowering insulin after malfunction or autoimmune destruction of the pancreatic β-cells is the recognized cause of diabetes, but recent evidence indicates that diabetic hyperglycaemia would not develop unless lack of insulin was accompanied by hypersecretion of glucagon. Glucagon release has therefore become an increasingly important target in diabetes management. Despite decades of research, an understanding of how glucagon secretion is regulated remains elusive, and fundamentally different mechanisms continue to be proposed. The autonomous nervous system is an important determinant of glucagon release, but it is clear that secretion is also directly regulated within the pancreatic islets. The present review focuses on pancreatic islet mechanisms involved in glucose regulation of glucagon release. It will be argued that α-cell-intrinsic processes are most important for regulation of glucagon release during recovery from hypoglycaemia and that paracrine inhibition by somatostatin from the δ-cells shapes pulsatile glucagon release in hyperglycaemia. The electrically coupled β-cells ultimately determine islet hormone pulsatility by releasing synchronizing factors that affect the α- and δ-cells.     

氧化应激与β细胞脂性凋亡关系及机制研究

目的:观察长期高脂饮食对大鼠胰岛β细胞凋亡影响,探讨氧化应激相关因子及基因表达与β细胞脂性凋亡关系及可能机制.方法:41只SD雄性大鼠随机分为高脂饲料喂养的肥胖组(n=21)和普通饲料喂养的正常对照组(n=20),喂养28周后,检测血浆及胰腺组织中的丙二醛(MDA)、还原型谷胱甘肽(GSH)浓度;静脉葡萄糖耐量试验评价早相胰岛素分泌反应(AIR);TUNEL法观察胰岛β细胞凋亡情况并计数每个胰岛中β细胞凋亡率;采用荧光定量PCR(Real-time PCR)比较两组大鼠胰岛细胞解偶联蛋白-2(UCP-2)mRNA表达的情况.结果:28周时高脂饲养组血浆及胰腺组织中MDA高于正常饲养组(P<0 05),而高脂饲养组血浆及胰腺组织中GSH低于正常饲养组(P<0 01).与基础值比较,高脂饲养组糖负荷后胰岛素增高的幅度低于正常饲养组(3 0倍vs 5 7倍,P<0 05).高脂饲养组1 min后血糖浓度下降缓慢,3、5、10 min血糖水平均高于正常饲养组(P<0 05).高脂饲养组胰岛β细胞凋亡率比正常饲养组增加40 0%(P<0 01).高脂饲养组胰岛细胞UCP-2基因mRNA表达与正常饲养组相比升高22 4%(P<0 01).结论:高脂喂养28周后SD大鼠胰岛β细胞凋亡率明显增加,血浆及胰岛细胞中MDA升高、GSH降低, 而且胰岛细胞UCP-2基因mRNA表达升高,提示氧化应激可能参与了高脂引起的β细胞脂性凋亡.     

L—盐酸赖氨酸对脑损伤大鼠胰高血糖素和血糖含量的影响

目的　研究 Ｌ盐酸赖氨酸对脑损伤后大鼠的胰高血糖素、胰岛素及血糖含量的影响。方法　采用落体法致大鼠脑损伤;伤后立刻一次腹腔注射 Ｌ盐酸赖氨酸,２４ｈ 后断头处死,用免疫组织化学 Ａ Ｂ Ｃ 法结合图像分析技术,观察 Ｌ盐酸赖氨酸（６２１ ．５ｍｇ·ｋｇ － １ 及３１０ ．８ｍｇ·ｋｇ － １） 对脑损伤后大鼠胰岛 Ａ 细胞内胰高血糖素（ Ｇｌｕ） 及 Ｂ 细胞内胰岛素（ Ｉｎｓ） 免疫反应强度和光密度值的影响;用生化分析仪测定血糖含量。结果　 Ｌ盐酸赖氨酸能显著增强 Ａ 细胞内 Ｇｌｕ 免疫反应强度,提高平均光密度值,增加 Ｇｌｕ 的含量,并降低血糖水平,但对 Ｂ 细胞内 Ｉｎｓ 的免疫反应强度及光密度值无明显影响。结论　 Ｌ盐酸赖氨酸可抑制 Ｇｌｕ 的释放并产生显著的降血糖作用。     

T细胞分化群40补体免疫球蛋白基因修饰骨髓间充质干细胞在肝移植排斥反应中的作用

目的探讨T细胞分化群40补体免疫球蛋白(CD40LIg)基因修饰骨髓间充质干细胞(MSCs)肝移植排斥反应中的作用。方法采用双袖套法建立DA-Lewis大鼠原位肝移植模型,经门静脉输注CD40LIg基因转染的MSCs,观察大鼠肝移植术后肝功能的变化、生存情况和移植肝病理形态学的改变,检测肝移植大鼠干扰素(INF-γ)和白细胞介素4(IL-2)水平的变化。结果(1)基因转染MSCs组生存时间显著长于对照组、单纯MSCs组和基因转染组,而单纯MSCs组和基因转染组又较对照组显著延长(P<0.05)。(2)对照组谷丙转氨酶(ALT)、总胆红素(TBil)、IL-2和INF-γ水平在肝移植术后急剧升高,各时段均显著高于3个治疗组(P<0.05);(3)肝脏组织病理检查,对照组呈重度排斥反应,单纯MSCs组和基因转染组为轻度排斥反应,而基因转染MSCs组无明显排斥反应。结论 CD40LIg基因修饰MSCs可以延长肝移植大鼠的存活时间,抑制大鼠肝移植的排斥反应。