ω-3多不饱和脂肪酸对血管内皮细胞衰老的影响及机制研究

Nutritional studies indicate that ω-3 polyunsaturated fatty acids (ω-3 PUFAs) are beneficial in alleviating the cardiovascular impairments associated with aging. Vascular biology studies show that vascular aging manifested by endothelial cell (EC) senescence is associated with increased oxidative and inflammatory stresses in ECs. Our preliminary experiments show that docosahexaenoic acid (DHA) exerts anti-inflammatory and anti-oxidative effects in ECs, which delays the EC senescence caused by high level of palmitic acid. However, the molecular basis of such beneficial effect of DHA remains to be determined. Activation and/or increased expression of SIRT1, an NAD+-dependent deacetylase, shows a marked effect in anti-cell senescence, which is contributed by SIRT1 attenuation of reactive oxygen species and increase of eNOS-derived NO generation. Thus, our hypothesis is that ω-3 PUFAs activate and/or increase the expression of SIRT1 in ECs. As a result, ECs show an anti-senescence phenotype with increased anti-inflammatory and anti-oxidative capacity. The three Specific Aims are: (1) to investigate the effects of ω-3 PUFAs activation of SIRT1 on palmitic acid-induced EC aging; (2) to study the molecular consequences resulted from ω-3 PUFAs activation of SIRT1 in the context of SIRT1 target genes and their anti-inflammatory and anti-oxidative effects in ECs; and (3) to elucidate EC aging in C57BL/6 wild-type mice fed a high fat diet in the presence or absence ofω-3 PUFAs. The results will then be compared with that in EC-specific SIRT1 knockout (EC-SIRT1-/-) mice with the same feeding protocols. Thus, results from this study will provide new information of the beneficial effects of ω-3 PUFAs on vascular aging and nutritional intervention.

ω-3多不饱和脂肪酸（ω-3 PUFAs）在预防血管衰老相关疾病方面发挥着积极作用。血管衰老始发于内皮细胞衰老，我们的预实验结果发现ω-3 PUFAs中的二十二碳六烯酸（DHA）可以延缓棕榈酸诱导的内皮细胞衰老，而其分子机制尚不清楚。SIRT1是NAD+依赖的脱乙酰基酶，可以通过增加NO产生以及清除活性氧来发挥抗细胞衰老作用。因此我们提出假说：ω-3 PUFAs可以通过抗炎、抗氧化延缓内皮细胞衰老，其机制与增加SIRT1的活性和/或表达有关。本研究将首先在细胞、分子水平探讨ω-3 PUFAs对棕榈酸诱导的内皮细胞衰老的影响，阐明SIRT1在其中的作用及分子机制；其次利用高脂诱导和内皮细胞SIRT1基因缺陷的衰老动物模型在整体动物水平上探讨SIRT1在ω-3 PUFAs抗血管衰老中的作用。本研究将为ω-3 PUFAs对内皮细胞衰老相关慢性疾病的预防提供理论基础和营养干预新策略。

内皮细胞衰老是血管衰老的起始事件，是引发动脉粥样硬化、冠心病、血栓以及阿尔兹海默病等心脑血管疾病的早期病变基础。ω-3多不饱和脂肪酸（ω-3 PUFAs）被证实在预防和治疗衰老相关疾病方面发挥着重要作用。然而，目前尚无研究阐明ω-3 PUFAs对血管内皮衰老的影响以及潜在机制。SIRT1是NAD+依赖的蛋白脱乙酰基酶,在动脉粥样硬化发生发展过程中具有保护内皮细胞功能和延缓细胞衰老的作用。因此为深入研究ω-3 PUFAs对衰老相关疾病防治的分子机制，并在此基础上提出合理的膳食指导，实验室利用高脂饲料喂养的中年雄性小鼠和人脐静脉内皮细胞（HUVECs）为试验材料，通过慢病毒介导的RNA干扰技术下调小鼠主动脉血管内皮SIRT1的表达，利用动物行为学、分子生物学、细胞生物学等实验技术分析各组小鼠在摄食与体重、胰岛素敏感性和葡萄糖耐受性、机体炎症反应以及血管内皮功能方面的变化，探索SIRT1在二十二碳六烯酸（DHA）延缓血管内皮衰老中的作用，并在此基础上提出合理的膳食指导对动脉硬化等内皮衰老相关慢性疾病的防治有重要意义。 ..获得的主要研究结果如下：.（1）成功建立稳定的C57/BL6J品系的SIRT1 KD小鼠；.（2）DHA补充可显著改善高脂饲料喂养导致的机体胰岛素敏感性及葡萄糖耐受性降低，并降低机体炎性水平；.（3）DHA补充以及DHA体外孵育均能可显著改善高脂饲料喂养导致的小鼠主动脉血管内皮的舒张功能减退； .（4）DHA可通过SIRT1改善小鼠血管内皮功能，缓解其衰老。.（5）DHA可在体外降低棕榈酸诱导的HUVECs衰老；.（6）DHA可通过CaMKKβ/AMPK/SIRT1通路增加细胞NO含量，减少细胞炎性反应，从而延缓HUVECs衰老；..以上实验结果证实，DHA在延缓高脂诱导的血管内皮衰老过程中，SIRT1发挥了重要且积极的作用。以上结果为进一步研究ω-3 PUFAs对动脉硬化等内皮衰老相关慢性疾病的防治提供理论基础和营养干预新策略。