發(fā)布時(shí)間：2018-05-17 03:25

  本文選題：高密度脂蛋白 + 氧化型低密度脂蛋白��； 參考：《山西醫(yī)科大學(xué)》2017年碩士論文

【摘要】：目的:通過細(xì)胞實(shí)驗(yàn)探究早發(fā)冠心病(PCAD)患者高密度脂蛋白(HDLPCAD)與健康人群HDL(HDLhealth)抗人臍靜脈內(nèi)皮細(xì)胞(HUVECs)凋亡作用是否有區(qū)別及其可能機(jī)制。方法:1.PCAD患者和與之相匹配的健康人血樣的采集;2.使用Lipoprint脂蛋白分析儀分析HDLhealth和HDLPCAD亞組分(HDL1-HDL10)分布情況;3.PCAD患者和與之相匹配的健康人血樣HDL的分離、提取、鑒定;4.分別用0μg/ml、50μg/ml、100μg/ml、150μg/ml、200μg/ml的氧化低密度脂蛋白(ox-LDL)處理HUVECs 24小時(shí),MTT檢測細(xì)胞存活度,明確ox-LDL引起HUVECs凋亡的合適濃度;5.首先分別用0μg/ml、50μg/ml、100μg/ml、150μg/ml、200μg/ml的HDLhealth預(yù)處理HUVECs 18小時(shí),接著再用最適濃度的ox-LDL處理HUVECs 24小時(shí),MTT檢測細(xì)胞存活度,明確HDLhealth預(yù)處理HUVECs的最適濃度;6.首先用最適濃度HDLhealth分別預(yù)處理HUVECs 0小時(shí)、6小時(shí)、12小時(shí)、18小時(shí)、24小時(shí),接著再用最適濃度的ox-LDL處理HUVECs24小時(shí),MTT檢測細(xì)胞存活度,明確HDLhealth預(yù)處理HUVECs的最適時(shí)間;7.用最適濃度HDLhealth、HDLPCAD對HUVECs進(jìn)行最適作用時(shí)間的預(yù)處理,再用合適濃度的ox-LDL處理HUVECs 24小時(shí),MTT檢測細(xì)胞存活度;流式細(xì)胞儀檢測細(xì)胞凋亡;Western blot檢測caspase-3、caspase-9蛋白的表達(dá);用試劑盒測定活性氧(ROS)活性。結(jié)果:1.HDLPCAD亞組分大顆粒含量(HDL1-HDL3)為(28.5±5.74),較HDLhealth(46.8±15.2)低,而小顆粒(HDL8-HDL10)含量為(21.4±7.75),較HDLhealth(10.9±5.38)高;2.100μg/ml ox-LDL處理HUVECs 24小時(shí)后細(xì)胞存活率為60.34%,存活率較空白處理組出現(xiàn)顯著降低(P0.05);3.200μg/ml HDLhealth預(yù)處理HUVECs18h后,細(xì)胞存活率為82.01%,可以明顯減弱ox-LDL對細(xì)胞的損傷;4.200μg/ml HDLhealth顯著抑制100μg/ml ox-LDL誘導(dǎo)的HUVECs凋亡及caspase-3、caspase-9蛋白表達(dá)和ROS的產(chǎn)生;5.200μg/ml HDLPCAD預(yù)處理HUVECs 18小時(shí)后,細(xì)胞存活率為65.5%,其作用較HDLhealth(80.28%)減弱;6.200μg/ml HDLPCAD可抑制100μM ox-LDL誘導(dǎo)HUVECs凋亡、caspase-3、caspase-9蛋白表達(dá)及ROS產(chǎn)生,但作用較HDLhealth減弱。結(jié)論:HDLPCAD與HDLhealth比較,可能由于其亞組分大顆粒含量(HDL1-HDL3)較HDLhealth低,小顆粒較HDLhealth(HDL8-HDL10)高,導(dǎo)致其抗氧化功能減弱,抑制ox-LDL誘導(dǎo)內(nèi)皮細(xì)胞凋亡的功能亦減弱,從而減弱或喪失抗As的作用。
[Abstract]:Aim: to investigate the difference between HDLPCAD (high density lipoprotein) (HDLPCAD) and HDLhealth (healthy) in patients with premature coronary heart disease (CHD) and healthy subjects (HDL) on human umbilical vein endothelial cells (HUVECs) apoptosis and its possible mechanism. Methods: 1. Blood samples from PCAD patients and matched healthy people were collected. The distribution of HDLhealth and HDLPCAD subfractions HDL1-HDL10 was analyzed by Lipoprint lipoprotein analyzer. 3. Isolation, extraction and identification of HDL from blood samples of PCAD patients and matched healthy people. HUVECs was treated with 0 渭 g / ml 50 渭 g / ml ~ (100 渭 g / ml) of oxidized low density lipoprotein 200 渭 g / ml for 24 hours, respectively. The cell viability was determined by MTT assay, and the appropriate concentration of ox-LDL induced HUVECs apoptosis was determined. HUVECs was pretreated with 0 渭 g / ml ~ 50 渭 g / ml ~ (50 渭 g / ml) ~ 100 渭 g / ml ~ (-1) HDLhealth for 渭 g / ml ~ (-1) g/ml for 18 hours, then HUVECs was treated with ox-LDL at the optimal concentration for 24 hours to determine the cell viability, and the optimal concentration of HUVECs pretreated by HDLhealth was 6 ~ (6). The optimal concentration of HDLhealth was used to pretreat HUVECs for 0 hours, 6 hours, 12 hours, 18 hours and 24 hours respectively. Then, the optimal concentration of ox-LDL was used to detect the viability of HUVECs24 cells, and the optimal time for HDLhealth pretreatment of HUVECs was determined. HUVECs was pretreated with the optimal concentration of HDLhealth HUVECs for 24 hours, then HUVECs was treated with appropriate concentration of ox-LDL for 24 hours to detect cell viability, flow cytometry was used to detect apoptosis and Western blot was used to detect the expression of caspase-3 and caspase-9 protein. The reactive oxygen species (Ros) activity was determined with the kit. Results 1. The content of HDL1-HDL3 in HDL1-HDL3 was lower than that of HDLhealth(46.8 鹵15.2in HDL1-HDL3, while the content of HDL8-HDL10 in HDL8-HDL10 was 21.4 鹵7.75, which was higher than that in HDLhealth(10.9 鹵5.38. The survival rate of HUVECs treated with 2.100 渭 g/ml ox-LDL for 24 hours was 60.344.The survival rate of HDL1-HDL3 was significantly lower than that of control group, and the survival rate of HDL8-HDL10 was significantly lower than that of control group after pretreatment with HUVECs18h of 3.200 渭 g/ml HDLhealth. The cell survival rate was 82.01, which could significantly attenuate the damage induced by ox-LDL (4.200 渭 g/ml HDLhealth), inhibit the HUVECs apoptosis induced by 100 渭 g/ml ox-LDL, the expression of caspase-3 and caspase-9 protein, and the production of ROS (5.200 渭 g/ml HDLPCAD) for 18 hours after pretreatment with HUVECs. The cell survival rate was 65.5, the effect of which was lower than that of HDLhealth80.28) 6.200 渭 g/ml HDLPCAD could inhibit the expression of caspase-3 caspase-9 protein and the production of ROS in HUVECs induced by 100 渭 M ox-LDL, but the effect was weaker than that of HDLhealth. Conclusion compared with HDLhealth, it is possible that the content of large particles of HDLhealth is lower than that of HDLhealth, and the small particles are higher than HDL8-HDL10, which results in the decrease of antioxidant function and the inhibition of apoptosis induced by ox-LDL in endothelial cells, thus weakening or losing the anti-arsenic effect.
【學(xué)位授予單位】：山西醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號】：R541.4

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