Real-time monitoring of mesangial cell-macrophage cross-talk using SEAP in vitro and ex vivo
Real-time monitoring of mesangial cell-macrophage cross-talk using SEAP in vitro and ex vivo
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DOI：
10.1111/j.1523-1755.2005.00471.x
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发表时间：
2005-08-01
影响因子：
19.6
通讯作者：
Kitamura, M
中科院分区：
文献类型：
--
作者：
Meng, YM;Kasai, A;Kitamura, M
SEAP在体外和离体实时监测肾小球系膜细胞-巨噬细胞串扰。巨噬细胞与系膜细胞的相互作用在肾小球肾炎的发病机制中起着至关重要的作用。我们建立了一个新的系统，用于连续，实时监测巨噬细胞和系膜细胞之间的串扰在体外和离体。通过基因工程改造大鼠肾小球系膜细胞，使其在核因子-κ B（NF-κ B）增强子元件的控制下产生分泌型碱性磷酸酶（SEAP）。将建立的感受器细胞暴露于巨噬细胞或巨噬细胞衍生因子，并评估SEAP的产生水平。在体外，当暴露于活化的巨噬细胞或巨噬细胞产生的细胞因子时，建立的细胞表达并分泌SEAP。培养液中SEAP活性的动力学与SEAP mRNA的表达水平密切相关。传感器细胞也分泌SEAP响应巨噬细胞积累，发炎大鼠肾小球条件培养基。将感受器细胞过继转移到急性抗Thy 1肾炎大鼠肾小球后，分离的含有感受器细胞的肾小球可快速、进行性地分泌SEAP。这些数据表明，建立的系统提供了简单和有用的工具，在体外和离体监测巨噬细胞和系膜细胞之间的串扰。这种方法将是有用的研究的分子机制参与肾小球系膜细胞-巨噬细胞的相互作用，也为筛选有效地干扰浸润性白细胞和驻地肾小球细胞之间的联系的治疗药物。
Real-time monitoring of mesangial cell-macrophage cross-talk using SEAP in vitro and ex vivo.Background. Macrophage-mesangial cell interaction plays a crucial role in the pathogenesis of glomerulonephritis. We established a novel system for continuous, real-time monitoring of cross-talk between macrophages and mesangial cells in vitro and ex vivo.Methods. Rat mesangial cells were genetically engineered to produce secreted alkaline phosphatase (SEAP) under the control of the nuclear factor-kappa B (NF-kappa B) enhancer elements. The established sensor cells were exposed to macrophages or macrophage-derived factors, and the level of SEAP production was evaluated.Results. In vitro, the established cells expressed and secreted SEAP when exposed to activated macrophages or to cytokines produced by macrophages. The kinetics of SEAP activity in culture media was closely correlated with the expression level of SEAP mRNA. The sensor cells also secreted SEAP in response to media conditioned by macrophage-accumulating, inflamed rat glomeruli. When the sensor cells were transferred adoptively into rat glomeruli subjected to acute anti-Thy 1 glomerulonephritis, the isolated glomeruli containing sensor cells secreted SEAP rapidly and progressively.Conclusion. These data suggested that the established system provides simple and useful tools for monitoring of cross-talk between macrophages and mesangial cells in vitro and ex vivo. This approach would be useful for investigation of molecular mechanisms involved in mesangial cell-macrophage interaction and also for screening of therapeutic agents that efficiently interfere with the link between infiltrating leukocytes and resident glomerular cells.