Kumar V B Sameer, Viji R I, Kiran M S, Sudhakaran P R
Department of Biochemistry, University of Kerala, Thiruvananthapuram, Kerala, India.
J Cell Physiol. 2009 Apr;219(1):123-31. doi: 10.1002/jcp.21659.
Nitric oxide (NO) regulates the vascular tone, and influences survival and apoptosis of endothelial cells (ECs). NO is produced by nitric oxide synthase (NOS) and eNOS is the constitutive enzyme in the endothelium. Though the extracellular matrix (ECM) has been reported to regulate various EC functions, the role of ECM in the regulation of eNOS is not clear. The present study was designed to analyze if laminin-1 (Ln-1), the major glycoprotein of the basement membrane, can regulate eNOS. The activity of eNOS was significantly low in ECs maintained on Ln-1 as compared to those on Col I and polylysine. Reversal of the effect of Ln-1 on treatment with inhibitor of p38 MAPK and changes in Thr and Ser phosphorylation in purified eNOS suggested that eNOS activity in cells maintained on Ln-1 is negatively regulated by post-translational phosphorylation at Ser and Thr residues by recruiting p38 MAPK pathway. Increase in eNOS activity and induction of apoptosis upon inhibition of p38 MAPK and reversal of this on inhibition of NOS by L-NAME suggested that increased NO induced apoptosis in ECs maintained on Ln-1 when p38 MAPK was inhibited. These results suggest that Ln contributes to survival of ECs by negatively modulating eNOS in a p38 MAPK dependent pathway.
一氧化氮（NO）调节血管张力，并影响内皮细胞（ECs）的存活和凋亡。NO由一氧化氮合酶（NOS）产生，而内皮型一氧化氮合酶（eNOS）是内皮中的组成型酶。尽管细胞外基质（ECM）已被报道可调节多种EC功能，但ECM在eNOS调节中的作用尚不清楚。本研究旨在分析基底膜的主要糖蛋白层粘连蛋白-1（Ln-1）是否能调节eNOS。与在I型胶原和聚赖氨酸上培养的ECs相比，在Ln-1上培养的ECs中eNOS的活性显著降低。用p38丝裂原活化蛋白激酶（MAPK）抑制剂处理后Ln-1作用的逆转以及纯化的eNOS中苏氨酸（Thr）和丝氨酸（Ser）磷酸化的变化表明，在Ln-1上培养的细胞中，eNOS活性通过招募p38 MAPK途径在Ser和Thr残基处进行翻译后磷酸化而受到负调控。抑制p38 MAPK后eNOS活性增加并诱导凋亡，而用L-硝基精氨酸甲酯（L-NAME）抑制NOS后这种情况逆转，这表明当p38 MAPK被抑制时，Ln-1上培养的ECs中增加的NO诱导了细胞凋亡。这些结果表明，Ln通过在p38 MAPK依赖性途径中负向调节eNOS来促进ECs的存活。
