CD58对妊娠早期山羊子宫局部细胞因子的调节及其与EPF关系的研究

【作者】 马勇江；

【导师】 曹斌云； 靳亚平；

【作者基本信息】 西北农林科技大学 ， 动物遗传育种与繁殖， 2001， 硕士

【摘要】 试验研究分为二个部分，第一部分主要从三个方面进行研究：（1）采用IL-2的生物学测定方法和应用IL-4、TGF-β1、TGF-β2试剂盒标准测定方法，对正常妊娠组的早期妊娠山羊（第15～19d）子宫局部的细胞因子IL-2、IL-4、TGF-β1、TGF-β2水平（活性）变化规律进行了研究。结果表明：妊娠早期山羊子宫两侧局部细胞因子水平（活性）无差异；妊娠早期山羊子宫局部IL-2活性先呈上升变化，后出现下降趋势；IL-4水平一直呈上升变化；TGF-β1水平较高，但呈不规则曲线变化；TGF-β2先呈上升变化，后趋于平稳，并保持在较高水平。据此认为：妊娠早期山羊子宫局部细胞因子分泌及活性有一动态变化规律。（2）将CD58处理后山羊妊娠第16～18d子宫局部上述细胞因子水平（活性）与同期正常妊娠的山羊进行了比较分析，结果表明：CD58可极显著地抑制IL-2的产生（P＜0.01），而明显地促进IL-4的产生（P＜0.05），并对TGF-β2产生具有一定促进作用（P＞0.05），而对TGF-β1无影响（P＞0.05）。据此认为：CD58对妊娠早期山羊子宫局部细胞因子的产生具有调节作用。（3）对正常妊娠组与CD58处理组山羊妊娠第15～19d子宫液进行SDS-PAGE电泳后发现：正常妊娠组与CD58处理组山羊妊娠第16～18d子宫液中出现一分子量约为17KD的胚源性特异蛋白，第15、19d不出现。该特异蛋白的出现与CD58处理与否无关，而CD58对其产生量的影响难以作出判断。试验研究的第二部分是研究CD58与山羊早孕因子（EPF）的关系，主要应用猪淋巴细胞Ea花环抑制试验，检测了山羊妊娠早期血清中EPF与人工制备CD58的活性，并对EPF与CD58的关系进行了探讨。试验结果证明：（1）猪淋巴细胞Ea花环抑制试验可应用于山羊EPF活性的测定；（2）妊娠早期（1~21d）山羊血清中存在EPF活性，妊娠第1，8d出现活性峰值（RIT值分别为18.8±1.1，21.2±1.8），其余时间EPF活性相对稳定，并呈现一规律性变化；（3）CD58（1：100）孵育淋巴细胞后，RIT值显著高于HBSS液对照组(P<0.05)，CD58（1：1，1：10）孵育淋巴细胞后，RIT值极显著高于HBSS液对照组（P<0.01）。据此认为CD58活性可以用Ea花环抑制试验进行测定并具有与EPF协同ALS使RIT值升高作用相似的功能；（4）妊娠山<WP=4>羊血清经CD58，CD2，HBSS液等处理后孵育淋巴细胞，经CD58处理组的RIT值极显著高于HBSS液处理组（P＜0.01），经CD2处理组RIT值极显著低于HBSS液处理组（P＜0.01）而与HBSS液对照组差异不显著（P＞0.05）。试验表明CD58与EPF可共同协同ALS抑制Ea花环形成，CD2对EPF有一定程度的封闭作用。据此认为CD58与EPF具有一定程度的抗原交叉性。更多还原

【Abstract】 This dissertation is divided into two parts. The first part is: (1) a study on the changes of the level or activity of the local-uterine cytokines such as IL-2,IL-4,TGF-β1 and TGF-β2 Using a bioassay for IL-2,IL-4 kit ,TEG- β1kit and TGF-β2 kit, respectively, in the 15~19th d of the pregnant goat(control group). The main results are: There is no different in the levels of the above cytokines between right and left uterinal horn during early pregnancy; The activity of IL-2 increases firstly, and then decreases; TGF-β1 has a high level irregularly during early pregnancy; TGF-β2 increases firstly and then remains in high level. These results show that there is a rule of dynamic changes in the secretion and activity of the cytokines during early pregnancy in goat.(2) a study on the changes of the above cytokines has been researched after the pregnant goats were treated by CD58(treatment group). The results are: CD58 can significantly inhibit the production of IL-2(P<0.01), however, accelerate the production of IL-4 significantly (p<0.05); CD58 can accelerate the production of TGF-β2 to a certain degree, but has no influence on TGF-β1. These results indicate that CD58 has a regulatory function in secretion of the local-uterine cytokines <WP=5>during early pregnancy.(3) after the uterine flushings in the 15~19th d of early pregnancy in goats of control group and treatment group was electrophorsed by SDS-PAGE,a special protein (Molecular mass is approximate 17KD) derived from embryo is present in the 16~18th d of the pregnant goats of control and treatment group, but not in the 15,19th d. This result suggests that the presence of this special protein has nothing to CD58, and it is not clear for CD58 to influence the production of the protein. The second part is studies on the relationship between CD58 and EPF in goat. The early active rosette inhibition test of pig lymphocyte be used to detect the activities of early pregnancy factor(EPF) in goat serum during early pregnancy and CD58 refined before the test, and study the relationship between EPF and CD58 .Available evidence shows that: (1) The activity of EPF in goats can be assayed by the early active rosette inhibition test of pig lymphocytes; (2) EPF s present in 1~21th d of the early pregnancy in goat, and there are two peaks of activity in the 1st and 8th d of pregnancy ( the value of the resotte inhibition titers (RITs) is 18.1±1.1 and 21.2±1.8, respectively), but the activity remains steadily during other days. (3) According to what the value of RITs is Significantly higher in pig lymphocytes incubated by CD58 than that of in pig Lymphocytes incubated by HBSS, CD58 (1∶100) (p<0.05) and CD58 (1∶1,1∶10)(p<0.01),two conclusions are drawed: the early active rosette inhibition test can be used to detect the activity of CD58, and CD58 has a similar function to EPF which has a coordinate effect on raising the value of RITs with antilymphocyte serum (ALS). (4) The value of RITs is much greater in pooled sera from early pregnant goat using CD58 than that of in those using HBSS (p<0.01), and significantly lower in pooled sera using CD2 than in those using HBSS(p<0.01), but has no difference between treatment group by CD2 and HBSS control group .These results show that CD58 and EPF had a coordinate effect on reducing the formment of the early active resottes induced by ALS together, and CD2 can inhibit the activity of EPF to a certain degree. In a word, there is a character of antigen cross between CD58 and EPF.更多还原