【作者】 吕剑光；

【导师】 霍贵成；

【作者基本信息】 东北农业大学 ， 食品科学， 2011， 硕士

【摘要】 胃肠黏膜及上皮损伤在临床上常见,严重的黏膜损伤会引发全身性的病理、生理、及内脏功能紊乱。初乳中的胰岛素生长因子-Ⅰ(IGF-Ⅰ)是一种在分子结构上与胰岛素类似的活性蛋白多肽物质,在牛初乳中大量存在,是牛初乳中最重要的促生长因子。本实验主要通过初乳IGF-Ⅰ提取物对体外细胞以及损伤大鼠肠黏膜的作用来了解其对胃肠黏膜损伤的修复作用。本研究主要以牛初乳为原料,利用超滤、离子交换层析技术提取IGF-Ⅰ。通过体外、体内试验研究探讨初乳中IGF-Ⅰ提取物对肠黏膜修复。体外试验主要通过人体肠道细胞系CaCO-2的增殖实验来研究初乳中IGF-Ⅰ提取物;体内试验采用腹腔内注射MTX的方法制备SD大鼠小肠炎模型。实验设正常对照组,MTX模型对照组(20μg/kg),牛初乳IGF-Ⅰ提取物修复组(75μg/kg),IGF-Ⅰ修复组(75μg/kg)。制模当天开始,每日按剂量灌胃一次,正常对照组和模型组灌正理盐水。分别在第3d和第6d处死大鼠,采用分光光度法检测大鼠血浆二氨氧化酶(DAO)活性和血浆D-乳酸水平,光镜下观察小肠组织病理形态学改变。体外试验结果显示随着IGF-Ⅰ提取物浓度的增加,对细胞增殖的促进作用逐渐增强,达到并超过一定浓度(100ng/mL)时,增殖作用达到饱和;用100ng/mLIGF-Ⅰ提取物处理MTX模型组细胞,发现IGF-Ⅰ提取物能够显著刺激受损的CaCO-2细胞增殖(P<0.01)。体内试验结果显示,与对照组相比,在第3d时模型组大鼠血浆DAO活性和血浆D-乳酸水平明显升高(P<0.01),小肠损伤程度明显高于对照组(P<0.01),牛初乳中IGF-Ⅰ提取物组和IGF-Ⅰ组可降低血浆DAO活性及D-乳酸水平(P<0.01),减轻小肠损伤程度(P<0.01);在第6d时,无论与第6d的MTX模型组还是与第3d的牛初乳IGF-Ⅰ提取物修复组及IGF-Ⅰ修复组DAO活性及D-乳酸水平都有明显的降低(P<0.01),小肠损伤程度也明显减轻(P<0.01)。本试验结果说明初乳中IGF-Ⅰ提取物对人体肠道细胞系CaCO-2有显著的促增殖作用,在超过100ng/mL时,作用趋向饱和,不抑制细胞增殖;初乳IGF-Ⅰ提取物对肠黏膜的通透性具有一定保护作用,可减轻炎细胞的浸润。通过该课题对IGF-Ⅰ与肠黏膜损伤修复及伤口愈合关系的研究,最终成果是开发能快速治疗肠黏膜损伤或伤口愈合的药物。更多还原

【Abstract】 The injury of gastrointestinal mucosa and epithelial is common in clinical, serious mucosal can cause systemic dysfunction on pathology, physiology and visceral. colostrum insulin-like growth factor-I (IGF-I) as a kind of active peptide substances is similar with insulin on molecular structure, which is abundance in bovine colostrum, and is the most important growth factors. In this study, to understand the role of IGF-I on the repair of gastrointestinal injury by vitro and vivo experiments.Take bovine colostrums as raw material, methods IGF-Ⅰwas purfied by ultrafiltration, ion exchange chromatography. This study explored the repair of colostrum IGF-Ⅰextraction on intestinal mucosal by vitro and vivo experments. In vitro, this article researched effect of colostum IGF-Ⅰextraction by measuring the impacts on CaCO-2 cell proliferation; in vivo, enteritis was induced in rats through peritoneal injection of methotrexate (20μg/kg). The experimental animals were randomly divided into four groups: normal group, MTX model group (20μg/kg), bovine colostrum IGF-I extraction recovery group (75μg/kg) and IGF-I recovery group (75μg/kg). The last two groups were administered in corresponding groups by gastric irrigation daily for 5 days, and the administered time had been from 2nd day to 5th day, the normal group and MTX model group were administered with Physiological Saline. The rats were died on 3nd and 6nd, Plasma diamine oxidase (DAO) and D-lactate level were measured, and histological changes in intestinal tissue were observed under the light microscope.In vitro, the rusults showed that CaCO-2 cell proliferation is gradually increased by the increased concentration of IGF-Ⅰextraction , when the concentration of IGF-Ⅰextraction exceed a certain concentration(100ng/mL), the proliferation is saturated. The cell model was treated by IGF-Ⅰextraction(100ng/mL), which had a significantly proliferation on the damaged CaCO-2 cell(P<0.01). In vivo, The results showed that the activity of DAO and D-lactate level of MTX model group were significantly increased compared with normal group on 3nd (P<0.01), and intestine injury was deeper than the normal group (P<0.01). The activity of DAO and D-lactate level of bovine colostrum IGF-I extraction recovery group were significantly decreased on 6 nd compared with model group on 6 nd and recovery group (P<0.01), and intestine injury was lighter than the normal group (P<0.01).The results of this study showed that colostrum IGF-I extract had significant proliferation promoting effect on human intestinal cells CaCO-2. When its concentration exceeded 100ng/mL, cell proliferation effect became saturated and did not indicate inhibitation effect. The colostrum IGF-Ⅰextract showed certain protective effect on the permeability of bowel mucosa and could reduce the inflammatory cells infiltration. The relationship between IGF-Ⅰand intestinal mucosa damage repair and wound healing was investigated in this research in order to develop new medicines which could deal with intestinal mucosa damage or wound healing quickly.更多还原