生长抑素减轻小鼠胰岛移植过程中胰腺外分泌细胞对移植胰岛的损伤
생장억소감경소서이도이식과정중이선외분비세포대이식이도적손상
Protective effects of somatostatin on mice islets injury after transplantation by pancreas exocrine cells
  • 万方数据
    中华器官移植杂志 중화기관이식잡지
    Chinese Journal of Organ Transplantation
    2015年 7期 417-423 ,共7页

    韩小乐%糜亮亮%周乐亮%张鹏%田磊

    한소악%미량량%주악량%장붕%전뢰

    胰岛移植%生长抑素%胰腺外分泌细胞%凋亡 胰島移植%生長抑素%胰腺外分泌細胞%凋亡
    이도이식%생장억소%이선외분비세포%조망
    Islet transplantation%Somatostatin%Pancreas exocrine cell%Apoptosis
    目的 观察与探讨生长抑素减轻小鼠胰岛移植过程中胰腺外分泌细胞对移植胰岛的损伤及其机制.方法 (1)体外实验:选取20只雄性BALB/C小鼠,采用随机数字表法分为生长抑素组和生理盐水组,每组10只.在获取胰岛手术麻醉前30 min,生长抑素组小鼠腹腔注射生长抑素10μg/g,生理盐水组注射等体积的生理盐水,然后分别收集两组小鼠的胰腺外分泌细胞及胰岛细胞,流式细胞仪检测两种细胞凋亡情况.(2)体内实验:获取小鼠胰岛及胰腺外分泌细胞,并对胰岛进行纯度及活性检测.应用链脲佐菌素建立小鼠糖尿病模型,然后进行胰岛移植,分别于小鼠左肾被膜上、下级同时移植胰岛250个及等体积的胰腺外分泌细胞.采用随机数字表法将成功建立胰岛移植模型的40只小鼠平均分为2组,实验组术后腹腔注射生长抑素,连续28 d,对照组术后注射等体积生理盐水,连续28 d.同时两组小鼠分别腹腔注射5乙炔基-2'-脱氧尿苷(EDU)5 μg/g,连续28 d.术后连续监测小鼠血糖;术后第8天,两组小鼠均进行葡萄糖耐量实验(IPGTT);术后10和28 d两组各随机选取10只小鼠分别切除左肾,采用免疫组织化学法检测左肾包膜下抗淀粉酶抗体的表达,采用免疫荧光染色法检测移植胰岛β细胞增殖情况.结果 (1)流式细胞检测结果显示,生长抑素组小鼠胰腺外分泌细胞凋亡率明显高于对照组(P＜0.05),两组间胰岛细胞凋亡率的差异无统计学意义(P＞0.05);(2)获取的小鼠胰岛纯度及活性均高于95％.胰岛移植后,两组小鼠血糖均恢复正常,对照组较实验组血糖恢复正常时间延迟(P＜0.05),对照组的血糖水平明显高于实验组;IPGTT实验结果显示移植物血糖调节功能正常;术后10d实验组和对照组移植物周围均可见抗淀粉酶抗体阳性颗粒,但实验组阳性颗粒数较对照组减少(P＜0.05);免疫荧光检测显示,实验组移植胰岛Insulin+ EDU+β细胞数较对照组明显增多(P＜0.05).术后28 d,实验组抗淀粉酶抗体阳性颗粒数较对照组明显减少(P＜0.05),但与术后10d相比两组均无明显变化(P＞0.05),实验组增值的β细胞数仍明显高于对照组(P＜0.05),但与术后10天相比两组增殖率均显著降低(P＜0.05).结论 生长抑素可以减轻小鼠胰岛移植过程中胰腺外分泌细胞对移植胰岛的损伤,这可能与生长抑素诱导小鼠胰腺外分泌细胞的凋亡,抑制胰腺外分泌细胞胰淀粉酶的分泌,以及促进胰岛β细胞增殖等因素有关.
    목적 관찰여탐토생장억소감경소서이도이식과정중이선외분비세포대이식이도적손상급기궤제.방법 (1)체외실험:선취20지웅성BALB/C소서,채용수궤수자표법분위생장억소조화생리염수조,매조10지.재획취이도수술마취전30 min,생장억소조소서복강주사생장억소10μg/g,생리염수조주사등체적적생리염수,연후분별수집량조소서적이선외분비세포급이도세포,류식세포의검측량충세포조망정황.(2)체내실험:획취소서이도급이선외분비세포,병대이도진행순도급활성검측.응용련뇨좌균소건립소서당뇨병모형,연후진행이도이식,분별우소서좌신피막상、하급동시이식이도250개급등체적적이선외분비세포.채용수궤수자표법장성공건립이도이식모형적40지소서평균분위2조,실험조술후복강주사생장억소,련속28 d,대조조술후주사등체적생리염수,련속28 d.동시량조소서분별복강주사5을결기-2'-탈양뇨감(EDU)5 μg/g,련속28 d.술후련속감측소서혈당;술후제8천,량조소서균진행포도당내량실험(IPGTT);술후10화28 d량조각수궤선취10지소서분별절제좌신,채용면역조직화학법검측좌신포막하항정분매항체적표체,채용면역형광염색법검측이식이도β세포증식정황.결과 (1)류식세포검측결과현시,생장억소조소서이선외분비세포조망솔명현고우대조조(P＜0.05),량조간이도세포조망솔적차이무통계학의의(P＞0.05);(2)획취적소서이도순도급활성균고우95％.이도이식후,량조소서혈당균회복정상,대조조교실험조혈당회복정상시간연지(P＜0.05),대조조적혈당수평명현고우실험조;IPGTT실험결과현시이식물혈당조절공능정상;술후10d실험조화대조조이식물주위균가견항정분매항체양성과립,단실험조양성과립수교대조조감소(P＜0.05);면역형광검측현시,실험조이식이도Insulin+ EDU+β세포수교대조조명현증다(P＜0.05).술후28 d,실험조항정분매항체양성과립수교대조조명현감소(P＜0.05),단여술후10d상비량조균무명현변화(P＞0.05),실험조증치적β세포수잉명현고우대조조(P＜0.05),단여술후10천상비량조증식솔균현저강저(P＜0.05).결론 생장억소가이감경소서이도이식과정중이선외분비세포대이식이도적손상,저가능여생장억소유도소서이선외분비세포적조망,억제이선외분비세포이정분매적분비,이급촉진이도β세포증식등인소유관.
    Objective To investigate the protective effects of somatostatin (SS) on mice islets injury after transplantation by pancreas exocrine cells and its mechanism.Method (1) In vitro, 20 male BALB/C mice were randomly divided into the SS group (n =10) and the control group (n = 10).The animals in SS group were injected with SS (10 g/g) by intraperitoneal injection (i.p) before 30 min, and those in the control group were given the same amount of normal saline (i.p).The pancreas exocrine cells and islet cells in two groups were extracted respectively, and the apoptosis was detected by flow cytometric.(2) The pancreases of mice were digested with collagenase, islets and pancreatic exocrine cells were collected, and the purity and activity of islet was detected.In vivo, 8-9-week old male BALB/C mice were induced into diabetic mice with Streptozocin (STZ) (190 mg/kg body weight, i.p).250 islets and the equal volume of pancreatic exocrine cells were transplanted into different regions of left kidney subcapsule.Forty mice were divided into two groups randomly.The experimental group was injected with SS (10 g/g, 3 times every day, i.p) for 28 days after operation, and the control group was injected with the same amount of normal saline (3 times every day, i.p) for 28 days.Then mice in two groups were injected with 5-Ethynyl-2'-deoxyuridine (EDU) (5 g/g, once every day, i.p) for 28 days.Blood glucose 1evel was monitored continually.Glucose tolerance test was performed after 8 days, and the left kidney was removed respectively after 10 days and 28 days.The expression of anti-amylase antibodies in subcapsule was detected by irnmunohistochemieal staining.The proliferation of islet beta cells was measured by immunofluorescence staining.Result (1) The apoptosis rate of pancreas exocrine cells in the experimental group was significantly higher than in the control group (P＜0.05).There was no significant difference in the apoptosis rate of islet cells between the experimental group and the control group (P＞0.05).The purity and activity of islet were above 95％.After islets transplantation, the blood glucose levels in control and experimental groups were normal, but experimental group had the advanced islet function in reversing diabetes.The average blood glucose level in control group was significantly higher than in experimental group, and the blood glucose regulating function of islet was normal.A large number of anti-amylase antibody-positive cells were found in renal subcapsule in the control group while little seen in the experimental group after 10 days.Immunofluorescence showed that the Insulin + EDU+ β cells of islet in the experimental group were more than those in the control group.The number of anti—amylase antibody-positive cells in the experimental group was significantly less than in the control group after 28 days, but showed no obvious difference from that at 10th day.The number of increased beta cells in the experimental group was still significantly greater than in the control group after 28 day, but the proliferation rate was reduced as compared with that at 10th day.Conclusion SS can reduce pancreas exocrine cells damage in the process of mice islets transplantation.SS can induce the apoptosis of damaged pancreas exocrine cells, inhibit pancreatic acinar cells from secreting pancreatic amylase and promote proliferation of islet beta cells.
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