Project description:We found that co-culturing BNL CL.2 liver cells with RAW 264.7 macrophages increased IRP binding in the first. To further investigate this modulation we investigated the gene expression profile in BNL CL.2 cells cultured alone, with iron, with RAW 264.7 macrophages or in the presence of both iron and macrophages. This novel reconstituted liver cell-macrophage communication pathway with the present gene expression data provides a platform for addressing how macrophages participate in the iron homeostasis of liver cells and, ultimately, in systemic iron homeostasis.
Project description:We found that co-culturing BNL CL.2 liver cells with RAW 264.7 macrophages increased IRP binding in the first. To further investigate this modulation we investigated the gene expression profile in BNL CL.2 cells cultured alone, with iron, with RAW 264.7 macrophages or in the presence of both iron and macrophages. This novel reconstituted liver cell-macrophage communication pathway with the present gene expression data provides a platform for addressing how macrophages participate in the iron homeostasis of liver cells and, ultimately, in systemic iron homeostasis. We used microarrays to determine the gene expression modulation in BNL CL.2 cells in response to 24h culture with 100 micromolar ferric ammonium citrate (FAC), co-culture with RAW 264.7 macrophages or both
Project description:We have searched genome-wide binding sites of mouse group V secretory phospholipase A2 (gV sPLA2) in Raw 264.7 cells. Since we previously had technical problems in immunoblotting analysis and immunoprecipitation of the gV sPLA2 protein due in part to unavailability of a specific antibody against mouse gV sPLA2 and other issues, we used a monoclonal antibody against the Myc-tag to analyze gV sPLA2 knockdown (KD) RAW 264.7 cells with re-constitutive expression of Myc-tagged gV sPLA2 or Myc-tagged gV sPLA2-H48Q, lacking catalytic activity, as well as gV sPLA2 KD RAW 264.7 cells transfected with empty vector. Among the significant peaks, the region corresponding to -375 ~ +6 from the transcriptional start site of the Pgk1 (phosphoglycerate kinase 1) gene was the strongest binding peak in the nucleus of both gV sPLA2 KD cells expressing Myc-tagged gV sPLA2 and gV sPLA2 KD cells expressing Myc-tagged gV sPLA2-H48Q. Meanwhile, the Pgk1 gene was not identified as a significant peak in the nucleus of gV sPLA2 KD cells transfected with empty vector.
