本文選題：尿酸 + 人腎小球內(nèi)皮細(xì)胞�。� 參考：《天津醫(yī)科大學(xué)》2017年碩士論文

【摘要】：背景及目的:近年來隨著我國人民物質(zhì)生活條件的改善和生活方式的改變,高尿酸血癥的患病率呈現(xiàn)升高趨勢。高尿酸血癥(Hyperuricemia)是由嘌呤代謝障礙所致的一種代謝性疾病,高尿酸血癥不僅僅是痛風(fēng)的病因,而且與慢性腎臟病(chronic kidney disease,CKD)、心血管疾病、高血壓、代謝綜合征(Metobolic syndrom)的發(fā)生密切相關(guān)。二十世紀(jì)七十年代Ross等人提出了“內(nèi)皮細(xì)胞功能障礙”假說,他們提出血管內(nèi)皮細(xì)胞功能障礙是血管內(nèi)皮受損之后,內(nèi)皮細(xì)胞合成的各種細(xì)胞因子發(fā)生改變,局部炎癥反應(yīng)活躍,血管通透性增加,血管的收縮/舒張失衡。血管內(nèi)皮功能障礙在高血壓、心血管疾病和慢性腎臟病的發(fā)生中起到了重要的作用。但是高尿酸血癥對血管內(nèi)皮細(xì)胞的影響及其具體致病機(jī)制尚未明確,本研究擬探討不同尿酸濃度對腎小球內(nèi)皮細(xì)胞的影響及其可能的機(jī)制,為臨床上定期監(jiān)測,早期預(yù)防及治療高尿酸血癥提供理論基礎(chǔ)。方法:1、以人的腎小球內(nèi)皮細(xì)胞(HRGEC)為實(shí)驗(yàn)對象,經(jīng)CCK-8細(xì)胞毒性實(shí)驗(yàn)檢測,選擇尿酸的作用濃度。2、不同濃度(0、4、8、16 mg/dl)的尿酸作用于HRGEC 24小時(shí),Real-timePCR法、Western blotting法分別檢測eNOS、NF-κB p65、MCP-1、ICAM-1的表達(dá),細(xì)胞免疫熒光的方法檢測eNOS、NF-κB p65蛋白的表達(dá)。3、不同濃度(0、4、8、16 mg/dl)的尿酸作用于HRGEC 24小時(shí),利用熒光探針DCFH-DA,在37℃作用于HRGEC 30分鐘,倒置熒光顯微鏡下進(jìn)行細(xì)胞內(nèi)活性氧(Reactive oxygen species,ROS)檢測。結(jié)果:1、與對照組相比較,4 mg/dl尿酸組HRGEC的eNOS mRNA及蛋白的表達(dá)無顯著性差異(P0.05),8 mg/dl及16 mg/dl尿酸組HRGEC的eNOS mRNA及蛋白的表達(dá)顯著減少(P0.05);與4 mg/dl尿酸組相比較,8 mg/dl及16 mg/dl尿酸組HRGEC的eNOS mRNA及蛋白的表達(dá)顯著減少(P0.05)。2、與對照組相比較,4 mg/dl尿酸組HRGEC的ROS表達(dá)無顯著性差異(P0.05),8 mg/dl及16mg/dl尿酸組HRGEC的ROS表達(dá)顯著增多(P0.05);與4 mg/dl尿酸組相比較,8 mg/dl及16mg/dl尿酸組HRGEC的ROS表達(dá)顯著增多(P0.05)。3、與對照組相比較,4 mg/dl尿酸組HRGEC的NF-κB p65 mRNA及蛋白的表達(dá)無顯著性差異(P0.05),8 mg/dl及16 mg/dl尿酸組HRGEC的NF-κB p65mRNA及蛋白的表達(dá)顯著增多(P0.05);與4 mg/dl尿酸組相比較,8 mg/dl及16 mg/dl尿酸組HRGEC的NF-κB p65 mRNA及蛋白的表達(dá)顯著增多(P0.05)。4、與對照組相比較,4 mg/dl尿酸組HRGEC的MCP-1、ICAM-1 mRNA及蛋白的表達(dá)無顯著性差異(P0.05),8 mg/dl及16 mg/dl尿酸組HRGEC的MCP-1、ICAM-1 mRNA及蛋白的表達(dá)顯著增多(P0.05);與4 mg/dl尿酸組相比較,8 mg/dl及16 mg/dl尿酸組HRGEC的MCP-1、ICAM-1 mRNA及蛋白的表達(dá)顯著增多(P0.05)。結(jié)論:1、高濃度的尿酸可能通過氧化應(yīng)激途徑導(dǎo)致腎小球內(nèi)皮細(xì)胞損傷,進(jìn)而造成腎臟功能損傷。2、高濃度的尿酸可能通過激活NF-κB經(jīng)典炎癥信號通路導(dǎo)致腎小球內(nèi)皮細(xì)胞損傷。
[Abstract]:Background and objective: with the improvement of material living conditions and changes in lifestyle, the prevalence of hyperuricemia has increased in recent years. Hyperuricemia is a metabolic disease caused by purine metabolic disorder. Hyperuricemia is not only the cause of gout, but also is closely related to the occurrence of chronic kidney disease, cardiovascular disease, hypertension and metabolic syndrome. Ross et al. Put forward the hypothesis of "endothelial cell dysfunction" in the 1970s. They proposed that vascular endothelial cell dysfunction is caused by changes in various cytokines synthesized by endothelial cells after vascular endothelial damage. The local inflammatory reaction is active, the vascular permeability is increased, and the vasoconstriction / relaxation is out of balance. Vascular endothelial dysfunction plays an important role in the development of hypertension, cardiovascular disease and chronic kidney disease. However, the effect of hyperuricemia on vascular endothelial cells and its specific pathogenesis have not been clarified. This study aims to explore the effects of different concentrations of uric acid on glomerular endothelial cells and its possible mechanisms for regular clinical monitoring. Early prevention and treatment of hyperuricemia provide a theoretical basis. Methods the human glomerular endothelial cells (HRGECs) were used as experimental subjects. By CCK-8 cytotoxicity test, the concentration of uric acid was chosen as 0.2 and the concentration of uric acid at different concentrations was 816 mg / dl). The expression of eNOSNFB p65MCP-1 ICAM-1 was detected by real-time PCR and Western blotting. The expression of eNOS- 魏 B p65 protein was detected by cellular immunofluorescence. The uronic acid of different concentrations of eNOS- 魏 B p65 was applied to HRGEC for 24 hours. The fluorescence probe DCFH-DAwas applied to HRGECs for 30 minutes at 37 鈩，

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