Department of Pharmacology, Institute of Post Graduate Medical Education and Research, Kolkata, India.
PLoS One. 2011 Apr 5;6(4):e18467. doi: 10.1371/journal.pone.0018467.
A complex interplay between Leishmania and macrophages influences parasite survival and necessitates disruption of signaling molecules, eventually resulting in impairment of macrophage function. In this study, we demonstrate the immunomodulatory activity of Berberine chloride in Leishmania infected macrophages.
The IC(50) of Berberine chloride, a quaternary isoquinoline alkaloid was tested in an amastigote macrophage model and its safety index measured by a cell viability assay. It eliminated intracellular amastigotes, the IC(50) being 2.8 fold lower than its IC(50) in promastigotes (7.10 µM vs. 2.54 µM) and showed a safety index >16. Levels of intracellular and extracellular nitric oxide (NO) as measured by flow cytometry and Griess assay respectively showed that Berberine chloride in Leishmania infected macrophages increased production of NO. Measurement of the mRNA expression of iNOS, IL-12 and IL-10 by RT-PCR along with levels of IL-12p40 and IL-10 by ELISA showed that in infected macrophages, Berberine chloride enhanced expression of iNOS and IL-12p40, concomitant with a downregulation of IL-10. The phosphorylation status of extracellular signal related kinase (ERK1/2) and p38 mitogen activated protein kinase (p38 MAPK) was studied by western blotting. In infected macrophages, Berberine chloride caused a time dependent activation of p38 MAPK along with deactivation of ERK1/2; addition of a p38 MAPK inhibitor SB203580 inhibited the increased generation of NO and IL-12p40 by Berberine chloride as also prevented its decrease of IL-10.
Berberine chloride modulated macrophage effector responses via the mitogen activated protein kinase (MAPK) pathway, highlighting the importance of MAPKs as an antiparasite target.
利什曼原虫和巨噬细胞之间的复杂相互作用影响寄生虫的存活，并需要破坏信号分子，最终导致巨噬细胞功能受损。在这项研究中，我们证明了盐酸黄连素在利什曼原虫感染的巨噬细胞中的免疫调节活性。
盐酸黄连素，一种季铵异喹啉生物碱，在无鞭毛体巨噬细胞模型中的 IC50 进行了测试，并通过细胞活力测定法测量了其安全性指数。它消除了细胞内无鞭毛体，其 IC50 比前鞭毛体低 2.8 倍（7.10 µM 对 2.54 µM），且安全性指数>16。通过流式细胞术和格里斯测定法分别测量细胞内和细胞外一氧化氮（NO）的水平表明，盐酸黄连素在感染巨噬细胞中增加了 NO 的产生。通过 RT-PCR 测量 iNOS、IL-12 和 IL-10 的 mRNA 表达，并通过 ELISA 测量 IL-12p40 和 IL-10 的水平，结果表明，在感染的巨噬细胞中，盐酸黄连素增强了 iNOS 和 IL-12p40 的表达，同时下调了 IL-10。通过 Western blot 研究了细胞外信号相关激酶（ERK1/2）和 p38 有丝分裂原激活蛋白激酶（p38 MAPK）的磷酸化状态。在感染的巨噬细胞中，盐酸黄连素引起 p38 MAPK 的时间依赖性激活，同时 ERK1/2 的失活；添加 p38 MAPK 抑制剂 SB203580 抑制了盐酸黄连素增加的 NO 和 IL-12p40 的产生，也阻止了其减少 IL-10。
盐酸黄连素通过丝裂原激活蛋白激酶（MAPK）途径调节巨噬细胞效应反应，强调了 MAPKs 作为抗寄生虫靶标的重要性。
