本文選題：氯胺酮 + 藥物暴露　； 參考：《重慶醫(yī)科大學(xué)》2016年碩士論文

【摘要】：目的:探討胚胎早期氯胺酮暴露對(duì)爪蛙心臟發(fā)育的影響。方法:將爪蛙胚胎從第8期暴露氯胺酮至第21期。(1)利用活體胚胎、心臟組織成像,免疫熒光(immunofluorescence)以及整體胚胎原位雜交(whole-mount RNA in situ hybridization,WMISH)觀察胚胎及其心臟形態(tài)的變化,利用透射式電子顯微鏡(transmission electron microscope,TEM)觀察胚胎心臟超微結(jié)構(gòu)的改變。(2)計(jì)數(shù)胚胎心率及計(jì)算心室縮短分?jǐn)?shù)(ventricular shortening fraction,VSF),以評(píng)估胚胎的心臟功能。(3)實(shí)時(shí)熒光定量PCR(Real-time Quantitative PCR,RT-qPCR)檢測(cè)心臟發(fā)育相關(guān)基因mRNA的表達(dá)。(4)蛋白質(zhì)印跡技術(shù)(western blot,WB)檢測(cè)XMLC2(Xenopus Myosin Light Chain 2)及pH3(phospho-histone H3)蛋白的表達(dá)。結(jié)果:隨氯胺酮暴露濃度升高,爪蛙胚胎的死亡率逐漸增加,體長(zhǎng)逐漸縮短(P0.05)。當(dāng)暴露濃度為0.5mg/ml時(shí),氯胺酮可導(dǎo)致以心臟增大為主的胚胎畸形,其他畸形還包括:腸道缺陷、體軸彎曲及體長(zhǎng)縮短等;還導(dǎo)致胚胎的心肌超微結(jié)構(gòu)中Z線的連續(xù)性中斷甚至部分缺失;pH3蛋白在胚胎心臟中的相對(duì)表達(dá)量增加(P0.05);胚胎的心率減慢(P0.05)及心室縮短分?jǐn)?shù)下降(P0.05);XMLC2的mRNA及蛋白的相對(duì)表達(dá)量在所檢測(cè)的四個(gè)時(shí)期均明顯下調(diào)(P0.05)。結(jié)論:早期氯胺酮暴露對(duì)爪蛙胚胎有呈濃度依賴性的致死性。當(dāng)暴露濃度為0.5mg/ml時(shí),氯胺酮可導(dǎo)致以心臟增大為主的胚胎畸形,并引起心肌超微結(jié)構(gòu)異常、心臟細(xì)胞增殖增加及心臟功能受損。氯胺酮可能通過下調(diào)XMLC2在mRNA及蛋白水平的表達(dá),導(dǎo)致心臟增大、心肌超微結(jié)構(gòu)異常及心臟收縮力的改變。這些結(jié)果為孕早期氯胺酮暴露可能引起的胎兒缺陷提供了一個(gè)新的理論依據(jù)。
[Abstract]:Objective: to investigate the effects of early embryonic ketamine exposure on cardiac development of Rana claw. Methods: Frog embryos were exposed to ketamine from stage 8 to phase 21. The morphologic changes of the embryos and their hearts were observed using in vivo embryos, cardiac tissue imaging, immunofluorescence fluorescence) and in situ hybridization of whole embryos, whole-mount RNA in situ hybridization (WMISH). Using transmission electron microscope (TEM) to observe the ultrastructural changes of the embryonic heart, count the heart rate of the embryo and calculate the ventricular shortening fraction of ventricular shortening fractionation, to evaluate the cardiac function of the embryo. 3) real-time fluorescence quantitative PCR(Real-time Quantitative PCR RT qPCR) was used to detect cardiac hair. The expression of mRNA and pH3(phospho-histone H3) were detected by Western blotblotWB). Results: with the increase of ketamine concentration, the embryo mortality and body length of Rana claw increased. When the exposure concentration is 0.5mg/ml, ketamine can lead to fetal malformation with heart enlargement, other malformations include intestinal defects, body axis bending and shortening of body length. It also caused the continuity of Z line in the myocardial ultrastructure of the embryo and even the partial deletion of the pH 3 protein in the embryonic heart. The relative expression of P0.05 protein in the embryonic heart increased; the embryo heart rate slowed down P0.05) and the ventricular shortening fraction decreased P0.05 / xmlC2 mRNA and protein phase. The expression of P0.05 was significantly down-regulated in the four periods. Conclusion: early ketamine exposure is a concentration-dependent lethal effect on the embryos of Rana claw. When the exposure concentration was 0.5mg/ml, ketamine could lead to fetal malformation, cardiac ultrastructure abnormality, cardiac cell proliferation and cardiac function damage. Ketamine may induce cardiac enlargement, myocardial ultrastructure abnormality and cardiac contractility by down-regulating the expression of XMLC2 in mRNA and protein. These results provide a new theoretical basis for possible fetal defects caused by ketamine exposure in early pregnancy.
【學(xué)位授予單位】：重慶醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2016
【分類號(hào)】：R614

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