흰쥐 혈관 평활근 세포에서 일산화 질소 합성 효소 유전자의 유전자 발현 조절 기전

Background : The expression of inducible nitric oxide synthase (iNOS) in the vascular smooth muscle cells (VSMCs) is dependent on the multiple regulatory mechanisms of which are largely unknown. In this study, we focused on the role of IFN-r in IL1 b-mediated expression of the VSMCs iNOS gene. Mater...

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Published inKosin Medical Journal (Online) pp. 120 - 131
Main Authors 안호준, 김택상, 강주석, 류현열, 정헌택
Format Journal Article
LanguageEnglish
Published 고신대학교 의과대학 학술지 01.02.2005
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ISSN2005-9531
2586-7024

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Summary:Background : The expression of inducible nitric oxide synthase (iNOS) in the vascular smooth muscle cells (VSMCs) is dependent on the multiple regulatory mechanisms of which are largely unknown. In this study, we focused on the role of IFN-r in IL1 b-mediated expression of the VSMCs iNOS gene. Materials and Methods : VSMCs were isolated from rat thoracic aorta by enzymatic dissociation. Phosphorothioate-modified ODNs were designed to block initiation of target mRNA translation. Nitrite concentrations were measured by mixing 100uL of culture medium with 100uL of Griess reagent. The biological activity of IFN-r was demonstrated by testing the effect of supernatants collected from the IL-1 b-treated VSMCs on the production of NO in RAW 264.7 cells stimulated with LPS. Supernatant levels of IFN-r were measured by Endogen Rat Interferon gamma enzyme-linked immunosorbent assay kit. Total RNA was prepared from cultured VSMCs using RNA zol (Tel-Test) 12 hours after IL1 b treatment. RNA (1ug) from each sample was reverse-transcribed with Oligo-dT as the first-strand cDNA primer and Maloney murine leukemia virus reverse transcriptase. Western blotting and electrophoretic mobility shift assay were performed. Results : In a system of rat VSMCs in primary culture, we found that IL-1b induced the productions of both NO and IFN-r. IFN-r protein and mRNA were expressed in the IL-1b-stimulated VSMCs. Anti-IFN-r antibody or antisense synthesis in the IL 1b-stimulated cells. Furthermore, antisense IFN-r ODN abolished the IL 1b-induced activation of interferon regulatory factor-1 (IRF-1) without affecting NF-kB activity as determined by electrophretic mobility shift assay. The repletion of IFN-r to the system restored NO production and IRF-1 activity. Conclusion : These results suggest that the expression of iNOS gene in the IL1 b-stimulated VSMCs is dependent on the endogenous IFN-r synthesis and regulated through the activations of both IRF-1 and NF-kB. KCI Citation Count: 0
ISSN:2005-9531
2586-7024