BHLHE40 regulates myeloid cell polarization through IL-10-dependent and independent mechanisms

Better understanding of the host responses to Mycobacterium tuberculosis (Mtb) infections is required to prevent tuberculosis and develop new therapeutic interventions. The host transcription factor BHLHE40 is essential for controlling Mtb infection, in part by repressing Il10 expression, where excess IL-10 contributes to the early susceptibility of Bhlhe40-/- mice to Mtb infection. Deletion of Bhlhe40 in lung macrophages and dendritic cells is sufficient to increase the susceptibility of mice to Mtb infection, but how BHLHE40 impacts macrophage and dendritic cell responses to Mtb is unknown. Here we report that BHLHE40 is required in myeloid cells exposed to GM-CSF, an abundant cytokine in the lung, to promote the expression of genes associated with a pro-inflammatory state and better control of Mtb infection. Loss of Bhlhe40 expression in murine bone marrow derived myeloid cells cultured in the presence of GM-CSF results in lower levels of pro-inflammatory associated signaling molecules IL-1b, IL-6, IL-12, TNFα, iNOS, IL-2, KC, and RANTES, as well as higher levels of the anti-inflammatory associated molecules MCP-1 and IL-10 following exposure to heat-killed Mtb. Deletion of Il10 in Bhlhe40-/- myeloid cells restored some, but not all, pro-inflammatory signals, demonstrating that BHLHE40 promotes pro-inflammatory responses via both IL-10-dependent and independent mechanisms. In addition, we show that macrophages and neutrophils within the lungs of Mtb-infected Bhlhe40-/- mice exhibit defects in iNOS production compared to infected WT mice, supporting that BHLHE40 promotes pro-inflammatory responses in innate immune cells, which may contribute to the essential role for BHLHE40 during Mtb infection in vivo.