目的探讨在血管平滑肌细胞(VSMC)中,同型半胱氨酸诱导活性氧物质(ROS)产生的途径.方法用[3H]-胸腺嘧啶脱氧核苷掺入法测定细胞DNA合成率;用DCF—DA荧光探针测定细胞内ROS;CHOP基因表达升高测定采用RT—RCR.结果(1)同型半胱氨酸诱导VSMC的DNA合成增加,这与和ROS水平升高有关;(2)同型半胱氨酸激活的NADPH氧化酶;抑制该酶活性可以显著地抑制细胞内ROS产生;(3)同型半胱氨酸上调VSMC的CHOP基因表达;(4)在没有同型半胱氨酸存在下,通过转染过表达CHOP基因可诱导细胞内ROS水平升高.结论NADPH氧化酶激活和CHOP基因表达上调都参与了同型半胱氨酸诱导的ROS产牛.
Objective To investigate the pathways of homocysteine-induced production of reactive oxygen species (ROS) in vascular smooth muscle cells (VSMC). Methods DNA synthesis in cells was determined by using [3H] thymidine incorporation. The levels of ROS in cells were measured using DCF-DA as a fluorescence probe. The expression of CHOP was detected by RT-PCR. Results (1) Homocysteine promoted DNA synthesis, which was associated with an increase in the level of ROS in VSMC. (2) Homocysteine activated NADPH oxidases. Inhibition of NADPH oxidases significantly suppressed the formation of ROS. (3) Homocysteine up-regulated the expression of CHOP. (4) Overexpression of CHOP by transfection markedly induced an increase in ROS levels in VSMC in the absence of homocysteine. Conclusion Both the activation of NADPH oxidases and up-regulation of CHOP by homocysteine are involved in the production of ROS.