子宫内膜MMP-2和TIMP-2表达的变化对奶牛慢性子宫内膜炎的影响

发布时间：2018-04-05 22:31

本文选题：基质金属蛋白酶　切入点：组织抑制剂　出处：《华中农业大学》2015年硕士论文

【摘要】：基质金属蛋白酶2(MMP-2)是一种蛋白水解酶;它能够水解细胞外基质中的多种成分,其主要功能是组织和重塑ECM。MMP-2的自然组织抑制剂为TIMP-2,它可以结合不管是有活性的还是无活性的MMP-2,从而使MMP-2的活动中止,继而有效的使MMP-2的胶原和明胶水解的活性降低了,最终抑制了癌细胞或子宫内膜细胞的转移和侵袭。试验调查显示,在正常情况下,在MMP-2和TIMP-2的相互作用下,它们是处于平衡协调的关系。白细胞介素-10是经过Th2分泌的大分子,能够降低Th1的功能,具有很强的抗炎作用,为抗炎因子。前列腺素E2(PGE2)是一种促炎细胞因子,它不仅能促进Th1细胞和Th1型T细胞的分化增加,进而促进炎症反应;而且可以抑制免疫反应和细胞吞噬、中性粒细胞聚集与迁移。研究显示,IL-10和PGE2与哺乳动物体内MMP-2、TIMP-2存在密切的联系,奶牛子宫内膜ECM的降解和重组受到MMP-2与TIMP-2的密切调控。因此,研究MMP-2、TIMP-2对奶牛子宫内膜炎的影响,可以调控奶牛子宫内膜炎的发生发展及其疾病的防治提供新的理论基础和实验依据。本实验选用健康荷斯坦奶牛的对照组和患有子宫内膜炎荷斯坦奶牛的试验组各10头,采取血液以及子宫内膜组织。用ELISA方法检测血清白细胞介素-10与前列腺素E2。所取的子宫内膜组织一部分用于提取总RNA,并在反转录之后采用荧光定量PCR法检测基质金属蛋白酶-2与基质金属蛋白酶组织抑制剂-2的m RNA表达水平;另一部分是用于免疫组化法测定基质金属蛋白酶-2与基质金属蛋白酶组织抑制剂-2在蛋白水平上的分泌。运用SPSS Statistics 17.0分析实验结果。实验结果显示:1.健康荷斯坦奶牛组中PGE2浓度为1.473103±80.29ng/L,IL-10含量为48.41±5.45 ng/L;而患有子宫内膜炎的荷斯坦奶牛组中PGE2的浓度为0.933103±21.99ng/L,IL-10含量为78.45±2.55 ng/L。实验组中PGE2浓度极显著低于对照组(P0.01);而IL-10含量显著高于对照组(P0.01)。2.试验组子宫内膜组织中基质金属蛋白酶-2与基质金属蛋白酶组织抑制剂-2m RNA水平均极显著低于对照组(P0.01)。3.健康组子宫内膜组织中基质金属蛋白酶-2与基质金属蛋白酶组织抑制剂-2的蛋白水平用IOD值表达,分别为(10.452±1.231)3103、(8.367±1.324)3103;而患有子宫内膜炎组分别为(5.432±0.587)3103、(4.287±0.536)3103。因此,健康组中MMP-2与TIMP-2在蛋白水平表达都高于对照组,差异极显著(P0.01)。实验结果表明,子宫内膜炎病牛中IL-10的含量升高、PGE2的含量降低,子宫内膜中MMP-2、TIMP-2低表达,从而引起ECM供给奶牛子宫内膜细胞的平衡稳态得到了破坏,促进了对子宫内膜炎的产生和发展。说明它们之间存在着密切的联系,这还需要进一步的实验予以证明。
[Abstract]:Matrix metalloproteinase 2 (MMP-2) is a proteolytic enzyme that hydrolyzes many components of extracellular matrix.Its main function is that the natural tissue inhibitor of tissue remodeling of ECM.MMP-2 is TIMP-2, which can combine both active and inactive MMP-2, thus suspending the activity of MMP-2 and effectively reducing the hydrolysis activity of collagen and gelatin in MMP-2.It eventually inhibits the metastasis and invasion of cancer cells or endometrial cells.The experimental results show that MMP-2 and TIMP-2 are in a balanced and coordinated relationship under normal conditions.Interleukin-10 (IL-10) is a macromolecule secreted by Th2, which can reduce the function of Th1. It has a strong anti-inflammatory effect and is an anti-inflammatory factor.Prostaglandin E _ 2 (PGE _ 2) is a pro-inflammatory cytokine, which can not only increase the differentiation of Th1 cells and Th1 T cells, but also inhibit the immune response, phagocytosis, neutrophil aggregation and migration.The results showed that IL-10 and PGE2 were closely related to MMP-2 and TIMP-2 in mammalian tissues. The degradation and recombination of ECM in the endometrium of dairy cows were closely regulated by MMP-2 and TIMP-2.Therefore, to study the effect of MMP-2 and TIMP-2 on endometritis in dairy cows can provide a new theoretical basis and experimental basis for regulating the occurrence and development of endometritis and the prevention and treatment of the disease in dairy cows.Blood and endometrial tissue were collected from 10 healthy Holstein cows and 10 endometritis Holstein cows respectively.Serum interleukin-10 and prostaglandin E _ 2 were detected by ELISA.Part of the endometrial tissue was used to extract total RNAs, and the m RNA expression levels of matrix metalloproteinase-2 and tissue inhibitor of matrix metalloproteinase-2 were detected by fluorescence quantitative PCR after reverse transcription.The other part is used to detect the secretion of matrix metalloproteinase-2 and tissue inhibitor of matrix metalloproteinase-2 at the protein level by immunohistochemical method.The experimental results were analyzed by SPSS Statistics 17.0.The result of the experiment shows that 1: 1.The concentration of PGE2 in the experimental group was significantly lower than that in the control group, while the content of IL-10 was significantly higher than that in the control group.The levels of matrix metalloproteinase-2 (MMP-2) and tissue inhibitor of matrix metalloproteinase (MMPs) -2m RNA in endometrium of the experimental group were significantly lower than those in the control group (P 0.01 路3).Therefore, the expression of MMP-2 and TIMP-2 in the healthy group was significantly higher than that in the control group (P 0.01).The results showed that the content of IL-10 in endometritis cattle increased and the content of PGE2 decreased, and the expression of MMP-2 and TIMP-2 in endometrium decreased, which resulted in the disruption of the homeostasis of ECM supply to the endometrial cells of dairy cows.Promote the production and development of endometritis.This indicates that there is a close relationship between them, which needs to be proved by further experiments.
【学位授予单位】：华中农业大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：S858.23

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