Centro de Estudios Farmacológicos y Botánicos (CEFYBO)-CONICET, Facultad de Medicina, Universidad de Buenos Aires, Paraguay 2155 piso 16, Buenos Aires, CP 1121, Argentina.
Inflamm Res. 2010 Mar;59(3):227-38. doi: 10.1007/s00011-009-0097-4. Epub 2009 Oct 13.
Fibroblasts are sentinel cells that could serve as intermediaries in the immune reaction in the inflammatory process. In this work, we investigate the action of the muscarinic agonist carbachol (CARB) on the expression and function of nitric oxide synthase (NOS) and cyclooxygenase (COX) in fibroblasts under normal or inflammatory conditions.
The normal fibroblast cell line, 3T3, from NIH swiss mouse embryo, was used. The inflammatory milieu was mimicked with lipopolysaccharide (LPS) (10 ng/ml) plus interferon gamma (IFNgamma) (0.5 ng/ml). Nitric oxide (NO) and prostaglandin E(2) (PGE(2)) production were measured by Griess reagent and radioimmunoassay, respectively. NOS, COX, and nuclear transcription factor kappa B (NF-kappaB) were studied by Western blot.
CARB increased NO synthesis by 57 +/- 5%, while a 150 +/- 10% increase in NO liberation was triggered by LPS plus IFNgamma treatment. CARB added to LPS plus IFNgamma potentiated NO synthesis by 227 +/- 19%. CARB also upregulated NOS1 protein expression via NF-kappaB activation. In addition CARB and LPS plus IFNgamma stimulated PGE(2) synthesis by 72 +/- 9 and 42 +/- 4%, respectively, while CARB added to LPS plus IFNgamma treated cells produced a synergism in PGE(2) liberation (130 +/- 12%) via COX-2.
Activation of muscarinic acetylcholine receptors can mimic mild inflammatory conditions or can deepen pre-existing inflammation, establishing a fine-tuned set-up on fibroblasts that in turn could be alerting the immune system.
成纤维细胞是哨兵细胞，可作为炎症过程中免疫反应的中介。在这项工作中，我们研究了毒蕈碱激动剂卡巴胆碱 (CARB) 在正常或炎症条件下对成纤维细胞中一氧化氮合酶 (NOS) 和环氧化酶 (COX) 的表达和功能的作用。
使用来自 NIH 瑞士小鼠胚胎的正常成纤维细胞系 3T3。用脂多糖 (LPS) (10 ng/ml) 加干扰素 γ (IFNγ) (0.5 ng/ml) 模拟炎症环境。通过格里斯试剂和放射免疫测定分别测量一氧化氮 (NO) 和前列腺素 E(2) (PGE(2)) 的产生。通过 Western blot 研究 NOS、COX 和核转录因子 kappa B (NF-kappaB)。
CARB 使 NO 合成增加了 57 +/- 5%，而 LPS 加 IFNγ 处理则引发了 150 +/- 10%的 NO 释放增加。CARB 加入 LPS 加 IFNγ 可使 NO 合成增强 227 +/- 19%。CARB 还通过 NF-kappaB 激活上调 NOS1 蛋白表达。此外，CARB 和 LPS 加 IFNγ 分别刺激 PGE(2) 合成增加 72 +/- 9%和 42 +/- 4%，而 CARB 加入 LPS 加 IFNγ 处理的细胞则通过 COX-2 产生协同作用使 PGE(2) 释放增加 130 +/- 12%。
毒蕈碱乙酰胆碱受体的激活可以模拟轻度炎症状态，或加深原有的炎症，使成纤维细胞建立一个精细的调节机制，反过来又可以向免疫系统发出警报。
