可可影响血脂谱的荟萃分析及FOXP3基因甲基化修饰与动脉粥样硬化的相关性研究
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摘要
背景：以往关于黑巧克力、可可粉等可可产物对血脂谱影响的研究结论不一致。
目的：通过Meta分析的方法综合评价随机对照试验中可可对总胆固醇(TC)、低密度脂蛋白胆固醇(LDL)和高密度脂蛋白胆固醇(HDL)的影响,为健康饮食提供可参考的循证学证据。
方法：系统检索PubMed、Cochrane library、Embase电子数据库,并通过阅读相关综述及文章的参考文献进一步获取信息。对目前已发表的,研究食用可可与血脂水平关系的随机对照实验进行荟萃分析。应用固定效应和随机效应模型,以加权平均差计算血脂指标的净变化。以预设的亚组分析探索影响效应的试验因素。
结果：最终纳入8个短期干预的随机对照试验,总共包括215个研究对象。综合分析显示,与对照组相比,食用可可能够显著减低LDL(平均差：-5.87mg/dL,95%CI：-11.13,-0.61,P<0.05),并且对TC有改善的趋势(平均差：-5.82mg/dL,95%CI：-12.39,0.76mg,P=0.08)。但是,敏感性分析中,在排除质量较低的实验后,综合分析高质量实验(仅纳入3项实验)显示,可可对LDL的作用不明显(平均差：-4.98mg/dL,95%CI：-3.18,3.21,P=0.23)。亚组分析显示,食用低剂量可可(多酚摄入量<260mg/天)能够显著降低胆固醇(TC：平均差：-9.92mg/dL；95%CI：-15.74,-4.14；P=0.0008；LDL平均差:-8.07 mg/dL; 95%CI:-15.15,-0.99；P=0.03),而中量或大量摄入可可对血脂谱没有影响。另外,可可改善血脂的作用在有心血管相关疾病组更加明显(TC：平均差：-8.01 mg/dL；95%CI：-13.83,-2.20；P=0.0007；LDL：平均差：7.60 mg/dL；95%CI：-14.70,-0.51：P=0.04)。
结论：短期摄入适量可可能够显著降低血胆固醇,但是可可的这种效应依赖于可可摄入量和受试者的健康状况。本研究仅分析了短期实验的效果,尚需更多的高质量的长期干预试验提供证据。
目的：动物学研究中发现,调节性T细胞在动脉粥样硬化中起保护作用。然而,在人类冠心病的研究中,调节性T细胞的变化水平尚存在争议。实验证明FOXP3基因的去甲基化状态保证其基因能够稳定表达,是调节性T细胞发挥抑制性功能的基础。研究还发现了一项新的调节性T细胞的检测方法,即基于调节性T细胞FOXP3基因特殊区域(TSDR)的去甲基化水平来反应调节性T细胞的水平。所以本研究旨在两方面的研究,一是应用这种新的方法来评估冠心病患者调节性T细胞水平；二是探讨动脉粥样硬化发生发展过程中是否伴有FOXP3基因的甲基化改变。
方法和结果：本研究纳入20位ST段抬高的急性心肌梗死患者,16位非ST段抬高的急性冠脉综合征患者以及20位冠脉造影正常者。分离外周血单个核细胞,分别用流式细胞分析仪和基于PCR的FOXP3基因甲基化分析来测量调节性T细胞水平。实验发现,冠心病患者用基于PCR的FOXP3基因甲基化分析所得到的调节性T细胞水平均是降低的(正常对照4.22%±1.92%,非ST段抬高的急性冠脉综合征3.29%±1.45%；ST段抬高的急性心肌梗死2.86%±1.26%),而流式细胞仪所得到的调节性T细胞水平在冠心病患者中的改变不一致。FOXP3-TSDR的去甲基化水平与冠心病的严重程度呈反比(r=-0.308,P<0.05)。在去除传统危险因素(年龄、吸烟、体重指数、平均动脉压、血脂和血糖)的影响后,这种相关性依然显著存在(β=-0.336,P<0.05)。进一步检测冠心病患者和正常对照者CD4+CD25+T细胞中甲基化转移酶的表达水平发现,冠心病患者CD4+CD25+T细胞中的甲基化转移酶DNMT3b的表达是显著增高的。我们用氧化低密度脂蛋白与分离的健康人的CD4+CD25+T细胞共培养发现,氧化低密度脂蛋白能够明显增加FOXP3-TSDR的甲基化程度达41%,减少FOXP3基因的表达超过3倍,同时伴有氧化低密度脂蛋白诱导DNMT3a和DNMT3b表达显著增加。此外,我们在单个核细胞的培养中还发现甲基转移酶抑制剂EGCG可逆转氧化低密度脂蛋白对调节性T细胞的作用。
结论：本研究显示冠心病患者的去甲基化FOXP3所代表的调节性T细胞水平显著降低,调节性T细胞的减少和动脉粥样硬化的严重程度呈负相关关系。冠心病的危险因素之一,氧化低密度脂蛋白能够增加FOXP3基因的甲基化程度,减少FOXP3基因表达。提示调节性T细胞在冠心病的发病中起重要作用,表观遗传学改变调控动脉粥样硬化进程。
Background:The effect of cocoa products on lipid changes is still controversial. Objectives:We aimed to identify and quantify the effect of cocoa on total cholesterol (TC), low density lipoprotein cholesterol (LDL) and high density lipoprotein cholesterol (HDL).
Design:A comprehensive literature search was conducted for relevant trials of cocoa on lipid profile. Weighted mean differences were calculated for net changes in lipid concentrations by using fixed-effect or random-effect models. Previously defined subgroup analyses were performed to identify the source of heterogeneity.
Results:Eight trials (215 participants) were included and evaluated. Because there was only one relatively longer term study, we focused on the short-term data to evaluate the effects of cocoa on plasma lipid. Cocoa consumption significantly reduced LDL by 5.87 mg/dL (95%CI:-11.13,-0.61; P<0.05) and marginally decreased TC by 5.82 mg/dL (95%CI:-12.39,0.76; P=0.08). However, no significant change was seen in LDL in the high-quality studies (3 studies included,-4.98 mg/dL,95%CI:-13.18,3.21,P=0.23). Subgroup analyses suggested that the cholesterol-lowering effect only was identified in those with low dose of cocoa consumption and with cardiovascular disease risks. No evidence was found in supporting a dose-effect relationship, of any effect in healthy subjects, or of any change in HDL.
Conclusion:Short-term cocoa consumption significantly reduced blood cholesterol, but the effects were dependent on the dose of cocoa consumption and the healthy status of participants. No dose response and no effect were found in healthy participants. Future high quality studies are needed to determine the efficiency of moderate cocoa on lipid profile in long-term intervention and in subjects with other cardiometabolic risk factors.
Objective:Regulatory T (Treg) cells play a protective role in experimental atherosclerosis. However, the role of Treg in the development of human coronary artery disease (CAD) is still inconsistent. Demethylation in the DNA that encodes transcription factor forkhead box P3 (FOXP3) is a prerequisite for the stable maintenance of suppressive properties in Tregs, which is also identified as the most specific marker for human Tregs. We aim to evaluate Treg levels in CAD patients by a novel method based on Treg-specific DNA demethylation within FOXP3 gene and testify DNA methylation modification of FOXP3 in the development of atherosclerosis.
Methods and Results:We included acute ST-elevation myocardial infarction (STEMI) patients, non-ST-elevation acute coronary syndromes (NSTACS) patients, and control subjects with normal coronary artery on angiography (NCA) in our study. Peripheral blood mononuclear cells were collected to determine Treg levels by both flow cytometry and PCR based DNA methylation analysis. We found that Treg levels, quantified by the demethylation rate of Treg-specific demethylated region in FOXP3 gene, but not by flow cytometry, were univocally decreased in NSTACS and in STEMI patients (NCA 4.22%±1.92%; NSTACS 3.29%±1.45%; AMI 2.86%±1.26%). This decline was inversely related to the severity of CAD (γ=-0.308,p<0.05). After adjustment for traditional risk factors, the association remained (/?=-0.336, P<0.05). Furthermore, we detected the expression of DNA methyltransferases in CD4+CD25+Tregs sorted from included subjects. A significant increase of DNA methyltransferase 3b was observed in CAD patients compared with normal controls. Treatment of CD4+CD25+Tregs with ox-LDL induced a 41%increase in the methylation of FOXP3, accompanied with a more than 3-fold reduction of FOXP3 mRNA expression and increases in the expressions of DNA methyltransferase 3a and 3b. Moreover, the effect of ox-LDL on Tregs can be reversed by methyltransferase inhibitor (-)-epigallocatechin-3-gallate.
Conclusion:Our data demonstrate that Treg cells are decreased in CAD patients and its reduction are associated with the severity of atherosclerosis. Ox-LDL decreases the expression of FOXP3 by DNA methylation. Thus, we confirm the important role of Treg cells in human CAD and suggest potential epigenetic regulation in the progression of atherosclerosis.
目的：通过Meta分析的方法综合评价随机对照试验中可可对总胆固醇(TC)、低密度脂蛋白胆固醇(LDL)和高密度脂蛋白胆固醇(HDL)的影响,为健康饮食提供可参考的循证学证据。
方法：系统检索PubMed、Cochrane library、Embase电子数据库,并通过阅读相关综述及文章的参考文献进一步获取信息。对目前已发表的,研究食用可可与血脂水平关系的随机对照实验进行荟萃分析。应用固定效应和随机效应模型,以加权平均差计算血脂指标的净变化。以预设的亚组分析探索影响效应的试验因素。
结果：最终纳入8个短期干预的随机对照试验,总共包括215个研究对象。综合分析显示,与对照组相比,食用可可能够显著减低LDL(平均差：-5.87mg/dL,95%CI：-11.13,-0.61,P<0.05),并且对TC有改善的趋势(平均差：-5.82mg/dL,95%CI：-12.39,0.76mg,P=0.08)。但是,敏感性分析中,在排除质量较低的实验后,综合分析高质量实验(仅纳入3项实验)显示,可可对LDL的作用不明显(平均差：-4.98mg/dL,95%CI：-3.18,3.21,P=0.23)。亚组分析显示,食用低剂量可可(多酚摄入量<260mg/天)能够显著降低胆固醇(TC：平均差：-9.92mg/dL；95%CI：-15.74,-4.14；P=0.0008；LDL平均差:-8.07 mg/dL; 95%CI:-15.15,-0.99；P=0.03),而中量或大量摄入可可对血脂谱没有影响。另外,可可改善血脂的作用在有心血管相关疾病组更加明显(TC：平均差：-8.01 mg/dL；95%CI：-13.83,-2.20；P=0.0007；LDL：平均差：7.60 mg/dL；95%CI：-14.70,-0.51：P=0.04)。
结论：短期摄入适量可可能够显著降低血胆固醇,但是可可的这种效应依赖于可可摄入量和受试者的健康状况。本研究仅分析了短期实验的效果,尚需更多的高质量的长期干预试验提供证据。
目的：动物学研究中发现,调节性T细胞在动脉粥样硬化中起保护作用。然而,在人类冠心病的研究中,调节性T细胞的变化水平尚存在争议。实验证明FOXP3基因的去甲基化状态保证其基因能够稳定表达,是调节性T细胞发挥抑制性功能的基础。研究还发现了一项新的调节性T细胞的检测方法,即基于调节性T细胞FOXP3基因特殊区域(TSDR)的去甲基化水平来反应调节性T细胞的水平。所以本研究旨在两方面的研究,一是应用这种新的方法来评估冠心病患者调节性T细胞水平；二是探讨动脉粥样硬化发生发展过程中是否伴有FOXP3基因的甲基化改变。
方法和结果：本研究纳入20位ST段抬高的急性心肌梗死患者,16位非ST段抬高的急性冠脉综合征患者以及20位冠脉造影正常者。分离外周血单个核细胞,分别用流式细胞分析仪和基于PCR的FOXP3基因甲基化分析来测量调节性T细胞水平。实验发现,冠心病患者用基于PCR的FOXP3基因甲基化分析所得到的调节性T细胞水平均是降低的(正常对照4.22%±1.92%,非ST段抬高的急性冠脉综合征3.29%±1.45%；ST段抬高的急性心肌梗死2.86%±1.26%),而流式细胞仪所得到的调节性T细胞水平在冠心病患者中的改变不一致。FOXP3-TSDR的去甲基化水平与冠心病的严重程度呈反比(r=-0.308,P<0.05)。在去除传统危险因素(年龄、吸烟、体重指数、平均动脉压、血脂和血糖)的影响后,这种相关性依然显著存在(β=-0.336,P<0.05)。进一步检测冠心病患者和正常对照者CD4+CD25+T细胞中甲基化转移酶的表达水平发现,冠心病患者CD4+CD25+T细胞中的甲基化转移酶DNMT3b的表达是显著增高的。我们用氧化低密度脂蛋白与分离的健康人的CD4+CD25+T细胞共培养发现,氧化低密度脂蛋白能够明显增加FOXP3-TSDR的甲基化程度达41%,减少FOXP3基因的表达超过3倍,同时伴有氧化低密度脂蛋白诱导DNMT3a和DNMT3b表达显著增加。此外,我们在单个核细胞的培养中还发现甲基转移酶抑制剂EGCG可逆转氧化低密度脂蛋白对调节性T细胞的作用。
结论：本研究显示冠心病患者的去甲基化FOXP3所代表的调节性T细胞水平显著降低,调节性T细胞的减少和动脉粥样硬化的严重程度呈负相关关系。冠心病的危险因素之一,氧化低密度脂蛋白能够增加FOXP3基因的甲基化程度,减少FOXP3基因表达。提示调节性T细胞在冠心病的发病中起重要作用,表观遗传学改变调控动脉粥样硬化进程。
Background:The effect of cocoa products on lipid changes is still controversial. Objectives:We aimed to identify and quantify the effect of cocoa on total cholesterol (TC), low density lipoprotein cholesterol (LDL) and high density lipoprotein cholesterol (HDL).
Design:A comprehensive literature search was conducted for relevant trials of cocoa on lipid profile. Weighted mean differences were calculated for net changes in lipid concentrations by using fixed-effect or random-effect models. Previously defined subgroup analyses were performed to identify the source of heterogeneity.
Results:Eight trials (215 participants) were included and evaluated. Because there was only one relatively longer term study, we focused on the short-term data to evaluate the effects of cocoa on plasma lipid. Cocoa consumption significantly reduced LDL by 5.87 mg/dL (95%CI:-11.13,-0.61; P<0.05) and marginally decreased TC by 5.82 mg/dL (95%CI:-12.39,0.76; P=0.08). However, no significant change was seen in LDL in the high-quality studies (3 studies included,-4.98 mg/dL,95%CI:-13.18,3.21,P=0.23). Subgroup analyses suggested that the cholesterol-lowering effect only was identified in those with low dose of cocoa consumption and with cardiovascular disease risks. No evidence was found in supporting a dose-effect relationship, of any effect in healthy subjects, or of any change in HDL.
Conclusion:Short-term cocoa consumption significantly reduced blood cholesterol, but the effects were dependent on the dose of cocoa consumption and the healthy status of participants. No dose response and no effect were found in healthy participants. Future high quality studies are needed to determine the efficiency of moderate cocoa on lipid profile in long-term intervention and in subjects with other cardiometabolic risk factors.
Objective:Regulatory T (Treg) cells play a protective role in experimental atherosclerosis. However, the role of Treg in the development of human coronary artery disease (CAD) is still inconsistent. Demethylation in the DNA that encodes transcription factor forkhead box P3 (FOXP3) is a prerequisite for the stable maintenance of suppressive properties in Tregs, which is also identified as the most specific marker for human Tregs. We aim to evaluate Treg levels in CAD patients by a novel method based on Treg-specific DNA demethylation within FOXP3 gene and testify DNA methylation modification of FOXP3 in the development of atherosclerosis.
Methods and Results:We included acute ST-elevation myocardial infarction (STEMI) patients, non-ST-elevation acute coronary syndromes (NSTACS) patients, and control subjects with normal coronary artery on angiography (NCA) in our study. Peripheral blood mononuclear cells were collected to determine Treg levels by both flow cytometry and PCR based DNA methylation analysis. We found that Treg levels, quantified by the demethylation rate of Treg-specific demethylated region in FOXP3 gene, but not by flow cytometry, were univocally decreased in NSTACS and in STEMI patients (NCA 4.22%±1.92%; NSTACS 3.29%±1.45%; AMI 2.86%±1.26%). This decline was inversely related to the severity of CAD (γ=-0.308,p<0.05). After adjustment for traditional risk factors, the association remained (/?=-0.336, P<0.05). Furthermore, we detected the expression of DNA methyltransferases in CD4+CD25+Tregs sorted from included subjects. A significant increase of DNA methyltransferase 3b was observed in CAD patients compared with normal controls. Treatment of CD4+CD25+Tregs with ox-LDL induced a 41%increase in the methylation of FOXP3, accompanied with a more than 3-fold reduction of FOXP3 mRNA expression and increases in the expressions of DNA methyltransferase 3a and 3b. Moreover, the effect of ox-LDL on Tregs can be reversed by methyltransferase inhibitor (-)-epigallocatechin-3-gallate.
Conclusion:Our data demonstrate that Treg cells are decreased in CAD patients and its reduction are associated with the severity of atherosclerosis. Ox-LDL decreases the expression of FOXP3 by DNA methylation. Thus, we confirm the important role of Treg cells in human CAD and suggest potential epigenetic regulation in the progression of atherosclerosis.
引文
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