Department of Biochemistry, Medical School and the Research Institute of Medical Science, Chonnam National University, 5 Hak-Dong, Gwangju, 501-746, Republic of Korea.
Mech Ageing Dev. 2010 Feb;131(2):119-23. doi: 10.1016/j.mad.2009.12.010. Epub 2010 Jan 12.
This study was to investigate factors underlying the age-related decrease in NO production in vascular endothelial cells. The age-related changes in NO production, the activity and expression level of eNOS, and eNOS binding proteins, were studied in HUVECs. NO production in HUVECs significantly decreased in an age-dependent manner. The potentiation of NO production by L-Arg was significantly suppressed by L-NIO (eNOS-specific inhibitor) in young HUVECs and was suppressed by 1400W (iNOS-specific inhibitor) in aged HUVECs. The aged HUVECs had lower eNOS protein levels than young cells. eNOS phosphorylation at Ser-1177 (active) decreased gradually from PDL 23 through 40, and eNOS phosphorylation at Thr-495 (inactive) increased in aged cells. Changes of intracellular eNOS binding proteins, such as caveolin-1, pAkt, and Hsp90, as well as interaction between eNOS and eNOS binding proteins, indicated decreasing enzyme activity in aged HUVECs. Aging might decrease the activity as well as expression level of eNOS in HUVECs. And the decrease in eNOS activity probably implicated to the alterations in the regulatory binding proteins. For further study, it needs to be confirmed that the age-related change in the intracellular distribution of eNOS and the relative contribution of eNOS and iNOS on vascular dysfunction in aged endothelial cells.
本研究旨在探讨血管内皮细胞中与年龄相关的 NO 产生减少的相关因素。我们研究了 HUVECs 中 NO 产生、eNOS 的活性和表达水平以及 eNOS 结合蛋白的年龄相关性变化。HUVECs 中 NO 的产生随年龄呈依赖性下降。在年轻的 HUVECs 中，L-Arg 增强 NO 产生的作用被 L-NIO（eNOS 特异性抑制剂）显著抑制，而在衰老的 HUVECs 中，该作用被 1400W（iNOS 特异性抑制剂）抑制。衰老的 HUVECs 的 eNOS 蛋白水平低于年轻细胞。eNOS 在 Ser-1177 处的磷酸化（活性）逐渐从 PDL 23 降低到 40，而在 Thr-495 处的磷酸化（非活性）在衰老细胞中增加。细胞内 eNOS 结合蛋白的变化，如 caveolin-1、pAkt 和 Hsp90，以及 eNOS 与 eNOS 结合蛋白之间的相互作用，表明衰老 HUVECs 中酶活性下降。衰老可能会降低 HUVECs 中 eNOS 的活性和表达水平。而 eNOS 活性的降低可能与调节结合蛋白的改变有关。为了进一步研究，需要证实 eNOS 的细胞内分布以及 eNOS 和 iNOS 在衰老内皮细胞血管功能障碍中的相对贡献随年龄的变化。
