TLR-9 agonist CpG activates macrophages and modulates expression of microRNAs involved in cancer drug resistance and various signaling pathways

Abstract

Abstract TLR-9 agonist, CpG induces pro-inflammatory immune response. We studied the immune response elicited in C57Bl/6 female mice by repeated stimulation of TLR-9 through multiple doses of CpG administration. Complete blood count analysis after CpG treatment revealed peripheral pancytopenia and the spleen showed splenomegaly. Flow cytometry analyses of spleen and liver mononuclear cells disclosed macrophage activation, evidenced by substantial increase in F4/80+ cells in the treatment group compared to control. Analysis of serum by ELISA demonstrated enhanced levels of pro-inflammatory cytokines, IL12/23, IL6 and TNFα in the CpG-treated group whereas no significant change was observed in the amount of anti-inflammatory cytokine, IL10. The spleen cells were further subjected to microarray analysis in order to examine the modulation of microRNA expression pattern following CpG treatment. Investigation of miRNA-mRNA relationships using the ingenuity pathway analysis (IPA) revealed differential expression of miRNAs associated with cancer drug resistance, HOTAIR and BAG signaling pathways along with microRNAs involved in immunomodulation. The major microRNAs altered with CpG treatment included mir27a, mir181, mir130a, mir487a, mir29b1, mir130a, mir3473b, mir21a, mir142 and mir144 targeting genes involved in drug resistance, apoptosis, cell cycle, angiogenesis and major histocompatibility complex-related proteins. Together, our studies demonstrate that chronic macrophage activation may lead to uncontrolled activation and proliferation of immune cells. (Supported by NIH grants P01AT003961, P20GM103641, R01AI129788, R01ES030144, R01AI160896 and R01AI123947). Supported by NIH grants P01AT003961, P20GM103641, R01AI129788, R01ES030144, R01AI160896 and R01AI123947