早發(fā)冠心病患者高密度脂蛋白抗炎作用的機制研究
發(fā)布時間：2018-10-25 10:59
【摘要】：目的:探討早發(fā)冠心病患者高密度脂蛋白(High Density Lipoprotein,HDL)與健康受試者HDL相比對氧化型低密度脂蛋白(Oxidized Low-Density Lipoprotein,ox-LDL)誘導的人臍靜脈內(nèi)皮細胞(Human Umbilical Vein Endothelial Cells,HUVECs)炎癥反應影響的區(qū)別及可能的機制,進一步明確早發(fā)冠心病的發(fā)病機制,為早發(fā)冠心病的臨床防治提供新思路。方法:1.選取符合實驗條件的健康受試者(男性55歲,女性65歲)及相應年齡的早發(fā)冠心病患者(男性55歲,女性65歲),采集血樣,密度梯度離心法分離HDL;2.ox-LDL處理HUVECs不同時間,通過Elisa法檢測細胞上清液中的炎癥因子如高遷移率族蛋白B1(High Mobility Group Protein Box 1,HMGB-1)、腫瘤壞死因子-α(Tumor Necrosis Factor-α,TNF-α)、細胞間黏附分子-1(Intercellular Cell Adhesion Molecule-1,ICAM-1)的表達情況,通過Western Blot檢測炎癥通路分子HMGB-1及晚期糖基化終末產(chǎn)物受體(Receptor for Advanced Glycation End Product,RAGE)的表達情況,明確炎癥反應最顯著的時間點;3.不同濃度的HDL孵育HUVECs相應時間后,檢測上述炎癥分子及炎癥通路分子的表達情況,明確HDL抗炎作用最顯著的濃度;4.比較早發(fā)冠心病患者HDL和健康受試者HDL抗炎作用是否具有差異。結(jié)果:1.50mg/L ox-LDL可以誘導HUVECs產(chǎn)生炎癥反應,隨著時間的延長細胞上清液中HMGB-1、TNF-α及ICAM-1的水平升高(P0.05),在48小時達到高峰;細胞HMGB-1、RAGE蛋白表達量升高,在24小時達到高峰,48小時表達量降低,但仍較正常細胞表達量高,RAGE表達量一直處于升高狀態(tài),在48小時達到高峰(P0.05)。2.HDL對ox-LDL誘導的HUVECs的炎癥反應具有抑制作用,細胞上清液中HMGB-1、TNF-α、ICAM-1的水平及細胞HMGB-1、RAGE蛋白表達量隨著HDL濃度的增加而降低(P0.05),當HDL濃度達到200mg/L時上述炎癥分子的表達量最低,說明此時HDL對炎癥反應的抑制作用最顯著(P0.05)。3.早發(fā)冠心病患者HDL組與健康受試者HDL組相比,細胞上清液中HMGB-1、TNF-α、ICAM-1的水平及細胞HMGB-1、RAGE蛋白表達量較高,但與模擬組相比上述分子水平較低,與空白組相比上述分子水平較高(P0.05)。結(jié)論:1.HDL可以通過抑制人臍靜脈內(nèi)皮細胞HMGB-1/RAGE炎癥通路發(fā)揮抗炎作用。2.早發(fā)冠心病患者HDL與健康受試者HDL相比,其抗炎作用減弱,可能與其對HMGB-1/RAGE炎癥通路的抑制作用減弱有關(guān)。提示HMGB-1/RAGE炎癥通路可能作為早發(fā)冠心病防治的靶點。
[Abstract]:Objective: to investigate the effect of high density lipoprotein (High Density Lipoprotein,HDL) on oxidative low density lipoprotein (Oxidized Low-Density Lipoprotein,ox-LDL) -induced inflammation of human umbilical vein endothelial cells (Human Umbilical Vein Endothelial Cells,HUVECs) and its possible mechanism in patients with premature coronary heart disease (CHD) compared with healthy subjects (HDL). To further clarify the pathogenesis of early coronary heart disease and provide a new idea for clinical prevention and treatment of early onset coronary heart disease. Methods: 1. Blood samples were collected from healthy subjects (male 55 years old, female 65 years old) and premature coronary heart disease patients (55 years old for males and 65 years old for women). HDL;2.ox-LDL was separated by density gradient centrifugation to treat HUVECs for different time. The expression of inflammatory factors, such as high mobility group protein B1 (High Mobility Group Protein Box 1 (HMGB-1), tumor necrosis factor- 偽 (Tumor Necrosis Factor- 偽 (TNF- 偽) and intercellular adhesion molecule-1 (Intercellular Cell Adhesion Molecule-1,ICAM-1), was detected by Elisa assay. The expression of HMGB-1 and advanced glycosylation end product receptor (Receptor for Advanced Glycation End Product,RAGE) was detected by Western Blot to determine the most significant time point of inflammatory reaction. 3. After HUVECs was incubated with different concentrations of HDL, the expression of the above inflammatory molecules and inflammatory pathway molecules was detected, and the most significant anti-inflammatory effect of HDL was determined. 4. To compare the anti-inflammatory effects of HDL and HDL in patients with early coronary heart disease (CHD). Results: 1.50mg/L ox-LDL could induce the inflammatory reaction of HUVECs, and the levels of HMGB-1,TNF- 偽 and ICAM-1 in the supernatant of cells increased with time (P0.05) and reached the peak at 48 hours, the expression of HMGB-1,RAGE protein increased, reached the peak at 24 hours, and decreased at 48 hours. However, the expression of RAGE was still higher than that of normal cells, and the expression of RAGE reached its peak at 48 hours (P0.05). 2.HDL could inhibit the inflammatory response of HUVECs induced by ox-LDL. The levels of HMGB-1,TNF- 偽, ICAM-1 and the expression of HMGB-1,RAGE protein in the supernatant decreased with the increase of HDL concentration (P0.05). The lowest expression of the above inflammatory molecules was found when the HDL concentration reached 200mg/L, indicating that HDL had the most significant inhibitory effect on inflammatory response (P0.05). The levels of HMGB-1,TNF- 偽, ICAM-1 and the expression of HMGB-1,RAGE protein in the supernatant of the HDL group were higher than those in the control group, but lower than those in the simulated group and higher than those in the blank group (P0.05). Conclusion: 1.HDL can play an anti-inflammatory effect by inhibiting the HMGB-1/RAGE inflammatory pathway of human umbilical vein endothelial cells. 2. The anti-inflammatory effect of HDL in patients with premature coronary heart disease is weaker than that in healthy subjects compared with HDL, which may be related to the decrease of its inhibitory effect on the inflammatory pathway of HMGB-1/RAGE. The results suggest that the HMGB-1/RAGE inflammatory pathway may be a target for the prevention and treatment of early onset coronary heart disease.
【學位授予單位】：山西醫(yī)科大學
【學位級別】：碩士
【學位授予年份】：2017
【分類號】：R541.4
本文編號：2293513
[Abstract]:Objective: to investigate the effect of high density lipoprotein (High Density Lipoprotein,HDL) on oxidative low density lipoprotein (Oxidized Low-Density Lipoprotein,ox-LDL) -induced inflammation of human umbilical vein endothelial cells (Human Umbilical Vein Endothelial Cells,HUVECs) and its possible mechanism in patients with premature coronary heart disease (CHD) compared with healthy subjects (HDL). To further clarify the pathogenesis of early coronary heart disease and provide a new idea for clinical prevention and treatment of early onset coronary heart disease. Methods: 1. Blood samples were collected from healthy subjects (male 55 years old, female 65 years old) and premature coronary heart disease patients (55 years old for males and 65 years old for women). HDL;2.ox-LDL was separated by density gradient centrifugation to treat HUVECs for different time. The expression of inflammatory factors, such as high mobility group protein B1 (High Mobility Group Protein Box 1 (HMGB-1), tumor necrosis factor- 偽 (Tumor Necrosis Factor- 偽 (TNF- 偽) and intercellular adhesion molecule-1 (Intercellular Cell Adhesion Molecule-1,ICAM-1), was detected by Elisa assay. The expression of HMGB-1 and advanced glycosylation end product receptor (Receptor for Advanced Glycation End Product,RAGE) was detected by Western Blot to determine the most significant time point of inflammatory reaction. 3. After HUVECs was incubated with different concentrations of HDL, the expression of the above inflammatory molecules and inflammatory pathway molecules was detected, and the most significant anti-inflammatory effect of HDL was determined. 4. To compare the anti-inflammatory effects of HDL and HDL in patients with early coronary heart disease (CHD). Results: 1.50mg/L ox-LDL could induce the inflammatory reaction of HUVECs, and the levels of HMGB-1,TNF- 偽 and ICAM-1 in the supernatant of cells increased with time (P0.05) and reached the peak at 48 hours, the expression of HMGB-1,RAGE protein increased, reached the peak at 24 hours, and decreased at 48 hours. However, the expression of RAGE was still higher than that of normal cells, and the expression of RAGE reached its peak at 48 hours (P0.05). 2.HDL could inhibit the inflammatory response of HUVECs induced by ox-LDL. The levels of HMGB-1,TNF- 偽, ICAM-1 and the expression of HMGB-1,RAGE protein in the supernatant decreased with the increase of HDL concentration (P0.05). The lowest expression of the above inflammatory molecules was found when the HDL concentration reached 200mg/L, indicating that HDL had the most significant inhibitory effect on inflammatory response (P0.05). The levels of HMGB-1,TNF- 偽, ICAM-1 and the expression of HMGB-1,RAGE protein in the supernatant of the HDL group were higher than those in the control group, but lower than those in the simulated group and higher than those in the blank group (P0.05). Conclusion: 1.HDL can play an anti-inflammatory effect by inhibiting the HMGB-1/RAGE inflammatory pathway of human umbilical vein endothelial cells. 2. The anti-inflammatory effect of HDL in patients with premature coronary heart disease is weaker than that in healthy subjects compared with HDL, which may be related to the decrease of its inhibitory effect on the inflammatory pathway of HMGB-1/RAGE. The results suggest that the HMGB-1/RAGE inflammatory pathway may be a target for the prevention and treatment of early onset coronary heart disease.
【學位授予單位】：山西醫(yī)科大學
【學位級別】：碩士
【學位授予年份】：2017
【分類號】：R541.4
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