胰高血糖素样肽-2联合凋亡抑制剂对大鼠急性放射性肠损伤的治疗作用
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摘要
目的探讨胰高血糖素样肽-2(GLP-2)联合凋亡抑制剂对于大鼠急性放射性肠损伤的治疗作用及机制。方法 Wistar大鼠75只,随机将其分为正常对照组、模型对照组、GLP-2组、GLP-2联合凋亡激动剂PAC-1组(GLP-2+PAC-1组)和GLP-2联合凋亡抑制剂Ac-DEVD-CHO组(GLP-2+Ac-DEVD-CHO组)共5组,采用60Coγ射线全腹部单次15Gy照射建立急性放射性肠损伤模型,照射后立即给药。于照射后1、3、5和14 d处死取材,分析小肠长度及质量,HE染色观察大鼠空肠组织学结构变化,并分析绒毛长度及隐窝深度,免疫印迹法检测空肠组织中活化型caspase-3(cleaved caspase-3)的表达情况。结果模型对照组大鼠在照射后第9天开始出现死亡;与模型对照组比,GLP-2组死亡率较高,出现死亡时间较早,GLP-2+PAC-1组死亡率在所有组中最高,且最早出现死亡,GLP-2+Ac-DEVD-CHO组无死亡。模型对照组大鼠照射后第5天小肠长度和小肠质量均显著降低(P<0.01),至第14天小肠质量显著增加(P<0.05);与模型对照组比,GLP-2组、GLP-2+PAC-1组和GLP-2+Ac-DEVD-CHO用药的3组大鼠小肠长度和质量均无显著差异。肠道病理学结果表明,照射后第1天模型对照组大鼠空肠组织隐窝部分出现较多凋亡及坏死细胞,至第3天绒毛出现严重脱落,绒毛长度及隐窝深度均显著降低(P<0.01);与模型对照组比,GLP-2组病变较轻,第3天绒毛长度及隐窝深度均显著升高(P<0.01),GLP-2+Ac-DEVD-CHO组空肠病变程度最轻,照射后第1、3、5、14天绒毛长度和隐窝深度均显著升高(P<0.01)。照射后第1天模型对照组空肠组织中cleaved caspase-3表达升高;与模型对照组比,照射后第3天用药的3组空肠组织中cleaved caspase-3表达均显著降低(P<0.01)。结论 GLP-2联合凋亡抑制剂Ac-DEVD-CHO对急性放射性肠损伤有保护作用,效果优于单用GLP-2。
Objective To investigate the therapeutic effect of glucagon-like peptide-2(GLP-2)combined with apoptosis inhibitors on acute radiation-induced intestinal injury in rats. Methods Seventy-five Wistar rats were randomly divided into five groups:normal control group,model group,GLP-2 group,GLP-2 combined apoptosis agonist PAC-1 group(group GLP-2+PAC-1)and GLP-2 combined apoptosis inhibitor Ac-DEVD-CHO group(group GLP-2+Ac-DEVD-CHO). The model of the acute radiation-induced intestinal injury was made by single-pass 15 Gy irradiation with60 Co γ-rays on abdomen. The general condition and weight of the rats were recorded daily. The rats were sacrificed at 1 d,3 d,5 d and 14 d after irradiation. The length and weight of the small intestine were analyzed. The histological changes of the jejunum were observed by HE staining. The length of the villi and the depth of the crypt were analyzed. The expression of cleaved caspase-3 in the jejunum was detected by Western blotting. Results After irradiation,death of rats in model group occurred on the 9 th day. Compared with the model group,the rats in the group GLP-2 had higher mortality and earlier time of death,while the mortality of the rats in the group GLP-2+PAC-1 was highest among all groups. In contrast,none was dead in the GLP-2+Ac-DEVD-CHO group. In the model group,the length and weight of small intestine were significantly decreased on the 5 th day after irradiation(P<0.01),while the weight of small intestine was significantly increased on the 14 th day(P<0.05). Compared with the model group,the length and quality of the small intestine of rats in GLP-2 group,GLP-2+PAC-1 group and the GLP-2+Ac-DEVD-CHO group had no significant difference. The histopathological results of intestine showed that there were many apoptic and necrotic cells in the crypt of rats in the model group on the 1 st day after irradiation. On the 3 rd day after irradiation,the villi sloughed off severely,and the length of the villi and the depth of the crypt were significantly decreased(P<0.01). The pathological change of jejunal structure of the GLP-2 group were lighter than that of the model group,and the length of the villi and the depth of the crypt were significantly increased on the 3 rd day(P<0.01). The structural damage of the GLP-2+Ac-DEVD-CHO group was the lightest and the recovery was best,while the length of the villus and the depth of the crypt were significantly increased on the 1 st,3 rd,5 th and 14 th day after irradiation(P<0.01). On 1 st day after irradiation,the expression of cleaved caspase-3 was increased in the jejunum of the model group. Compared with the model group,the expression of cleaved caspase-3 in the jejunum tissues of GLP-2 group,GLP-2+PAC-1 group and GLP-2+Ac-DEVD-CHO group was decreased on the 3 rd day after irradiation. Conclusion GLP-2 combined with apoptosis inhibitor has protective effect on the acute radiation-induced intestinal injury,and the efficiency is better than that of GLP-2.
Objective To investigate the therapeutic effect of glucagon-like peptide-2(GLP-2)combined with apoptosis inhibitors on acute radiation-induced intestinal injury in rats. Methods Seventy-five Wistar rats were randomly divided into five groups:normal control group,model group,GLP-2 group,GLP-2 combined apoptosis agonist PAC-1 group(group GLP-2+PAC-1)and GLP-2 combined apoptosis inhibitor Ac-DEVD-CHO group(group GLP-2+Ac-DEVD-CHO). The model of the acute radiation-induced intestinal injury was made by single-pass 15 Gy irradiation with60 Co γ-rays on abdomen. The general condition and weight of the rats were recorded daily. The rats were sacrificed at 1 d,3 d,5 d and 14 d after irradiation. The length and weight of the small intestine were analyzed. The histological changes of the jejunum were observed by HE staining. The length of the villi and the depth of the crypt were analyzed. The expression of cleaved caspase-3 in the jejunum was detected by Western blotting. Results After irradiation,death of rats in model group occurred on the 9 th day. Compared with the model group,the rats in the group GLP-2 had higher mortality and earlier time of death,while the mortality of the rats in the group GLP-2+PAC-1 was highest among all groups. In contrast,none was dead in the GLP-2+Ac-DEVD-CHO group. In the model group,the length and weight of small intestine were significantly decreased on the 5 th day after irradiation(P<0.01),while the weight of small intestine was significantly increased on the 14 th day(P<0.05). Compared with the model group,the length and quality of the small intestine of rats in GLP-2 group,GLP-2+PAC-1 group and the GLP-2+Ac-DEVD-CHO group had no significant difference. The histopathological results of intestine showed that there were many apoptic and necrotic cells in the crypt of rats in the model group on the 1 st day after irradiation. On the 3 rd day after irradiation,the villi sloughed off severely,and the length of the villi and the depth of the crypt were significantly decreased(P<0.01). The pathological change of jejunal structure of the GLP-2 group were lighter than that of the model group,and the length of the villi and the depth of the crypt were significantly increased on the 3 rd day(P<0.01). The structural damage of the GLP-2+Ac-DEVD-CHO group was the lightest and the recovery was best,while the length of the villus and the depth of the crypt were significantly increased on the 1 st,3 rd,5 th and 14 th day after irradiation(P<0.01). On 1 st day after irradiation,the expression of cleaved caspase-3 was increased in the jejunum of the model group. Compared with the model group,the expression of cleaved caspase-3 in the jejunum tissues of GLP-2 group,GLP-2+PAC-1 group and GLP-2+Ac-DEVD-CHO group was decreased on the 3 rd day after irradiation. Conclusion GLP-2 combined with apoptosis inhibitor has protective effect on the acute radiation-induced intestinal injury,and the efficiency is better than that of GLP-2.
引文
[1] Monti P,Wysocki J,Vander MA,et al. The contribution of radiation-induced injury to the gastrointestinal tract in the development of multi-organ dysfunction syndrome or failure[J]. BJR Suppl,2005,27(4):89-94.
[2]魏开建.葛根芩连汤治疗放射性肠炎的临床疗效观察[J].福建中医学院学报,2009,19(6):13-14.
[3] Gu J,Liu S,Mu N,et al. A DPP-Ⅳ-resistant glucagon-like peptide-2 dimer with enhanced activity against radiation-induced intestinal injury[J]. J Control Release,2017,260:32-45.
[4] Kalaiselvan R,Theis VS,Dibb M,et al. Radiation enteritis leading to intestinal failure:1994 patient-years of experience in a national referral center[J]. Eur J Clin Nutr,2014,68(2):166-170.
[5] Ester M,Wiesendanger-Wittmer,Nanna M,et al. Systematic review of the role of a belly board device in radiotherapy delivery in patients with pelvic malignancies[J]. Radiother Oncol,2012,102(3):325-334.
[6] Liu M,Liu B,Wang H,et al. Dosimetric comparative study of3 different postoperative radiotherapy techniques(3D-CRT,IMRT,and RapidArc)forⅡ-Ⅲstage rectal cancer[J]. Medicine(Baltimore),2015,94(1):e372.
[7] Liang W,BJ Leibowitz,Jian Y,et al. Inhibition of CDK4/6 protects against radiation induced intestinal injury in mice[J]. J Clin Invest,2016;126(11):4076-4087.
[8]王兴文,林晓燕,毕迎惠,等.小牛血去蛋白提取物对急性放射性肠炎大鼠小肠黏膜的修复作用及凋亡相关基因的影响[J].中华肿瘤杂志,2009,31(10):742-745.
[9]范崇桂,郭胜.银杏叶提取物对急性脑梗死大鼠凋亡基因的影响[J].中国医院药学杂志,2015,35(10):902-907.
[10]朱丽娜,武文辉,廖玮浩,等.放射性肠炎的发病机制及其治疗进展[J].临床医学研究与实践,2018,3(7):196-198.
[11]李春芳,孙力.高压氧联合药物灌肠治疗慢性放射性肠炎的疗效分析[J].西南国防医药,2018,28(6):572-573.
[12]常鹏宇.脂肪间充质干细胞修复放射性肠损伤的实验研究[D].长春:吉林大学,2014.
[13] Huang YH,Guo FL,Yao DH,et al. Surgery for chronic radiation enteritis:outcome and risk factors[J]. J Surg Res,2016,204(2):335-343.
[14]朱俊东,粟永萍,程天民.胰高血糖素样肽-2对放射损伤小鼠肠上皮恢复的影响[J]第三军医大学学报,2001,23(3):293-295.
[15] Sigalet DL,Wallace LE,Holst JJ,et al. Enteric neural pathways mediate the anti-inflammatory actions of glucagon-like peptide-2[J]. Am J Physiol Gastrointest Liver Physiol,2007,293(1):G211-G221.
[16]李思聪,李玉明.肠复康方对急性放射性肠炎大鼠小肠黏膜细胞凋亡基因的影响[J].安徽中医药大学学报,2016,35(4):67-73.
[17] Jiang H,Zhao PJ,Su D,et al. Paris saponinⅠinduces apoptosis via increasing the Bax/Bcl-2 ratio and caspase-3 expression in gefitinib-resistant non-small cell lung cancer in vitro and in vivo[J]. Mol Med Rep,2014,9(6):2265-2272.
[18]张丹,吕亮,申兴斌,等.大肠癌中Survivin、NF-κB、I-κB及Caspase-3的表达及意义[J].中国老年学杂志,2017,37(3):549-552.
[2]魏开建.葛根芩连汤治疗放射性肠炎的临床疗效观察[J].福建中医学院学报,2009,19(6):13-14.
[3] Gu J,Liu S,Mu N,et al. A DPP-Ⅳ-resistant glucagon-like peptide-2 dimer with enhanced activity against radiation-induced intestinal injury[J]. J Control Release,2017,260:32-45.
[4] Kalaiselvan R,Theis VS,Dibb M,et al. Radiation enteritis leading to intestinal failure:1994 patient-years of experience in a national referral center[J]. Eur J Clin Nutr,2014,68(2):166-170.
[5] Ester M,Wiesendanger-Wittmer,Nanna M,et al. Systematic review of the role of a belly board device in radiotherapy delivery in patients with pelvic malignancies[J]. Radiother Oncol,2012,102(3):325-334.
[6] Liu M,Liu B,Wang H,et al. Dosimetric comparative study of3 different postoperative radiotherapy techniques(3D-CRT,IMRT,and RapidArc)forⅡ-Ⅲstage rectal cancer[J]. Medicine(Baltimore),2015,94(1):e372.
[7] Liang W,BJ Leibowitz,Jian Y,et al. Inhibition of CDK4/6 protects against radiation induced intestinal injury in mice[J]. J Clin Invest,2016;126(11):4076-4087.
[8]王兴文,林晓燕,毕迎惠,等.小牛血去蛋白提取物对急性放射性肠炎大鼠小肠黏膜的修复作用及凋亡相关基因的影响[J].中华肿瘤杂志,2009,31(10):742-745.
[9]范崇桂,郭胜.银杏叶提取物对急性脑梗死大鼠凋亡基因的影响[J].中国医院药学杂志,2015,35(10):902-907.
[10]朱丽娜,武文辉,廖玮浩,等.放射性肠炎的发病机制及其治疗进展[J].临床医学研究与实践,2018,3(7):196-198.
[11]李春芳,孙力.高压氧联合药物灌肠治疗慢性放射性肠炎的疗效分析[J].西南国防医药,2018,28(6):572-573.
[12]常鹏宇.脂肪间充质干细胞修复放射性肠损伤的实验研究[D].长春:吉林大学,2014.
[13] Huang YH,Guo FL,Yao DH,et al. Surgery for chronic radiation enteritis:outcome and risk factors[J]. J Surg Res,2016,204(2):335-343.
[14]朱俊东,粟永萍,程天民.胰高血糖素样肽-2对放射损伤小鼠肠上皮恢复的影响[J]第三军医大学学报,2001,23(3):293-295.
[15] Sigalet DL,Wallace LE,Holst JJ,et al. Enteric neural pathways mediate the anti-inflammatory actions of glucagon-like peptide-2[J]. Am J Physiol Gastrointest Liver Physiol,2007,293(1):G211-G221.
[16]李思聪,李玉明.肠复康方对急性放射性肠炎大鼠小肠黏膜细胞凋亡基因的影响[J].安徽中医药大学学报,2016,35(4):67-73.
[17] Jiang H,Zhao PJ,Su D,et al. Paris saponinⅠinduces apoptosis via increasing the Bax/Bcl-2 ratio and caspase-3 expression in gefitinib-resistant non-small cell lung cancer in vitro and in vivo[J]. Mol Med Rep,2014,9(6):2265-2272.
[18]张丹,吕亮,申兴斌,等.大肠癌中Survivin、NF-κB、I-κB及Caspase-3的表达及意义[J].中国老年学杂志,2017,37(3):549-552.