Depletion of Collagen IX Alpha1 Impairs Myeloid Cell Function.

The trabecular extracellular matrix (ECM) forms a three-dimensional scaffold to stabilize the bone marrow, provide substrates for cell-matrix interactions and retain, present or release signals to modulate hematopoietic stem and progenitor cell development. However, the impact of trabecular ECM components on hematopoiesis has been poorly studied. Using collagen IX alpha1 - deficient (Col9a1(-/-) ) mice, we revealed that a lack of collagen IX alpha1 results in a disorganized trabecular network enriched in fibronectin, and in a reduction in myeloid cells, which was accompanied by a decrease in colony-stimulating factor 1 receptor expression on monocytes from the bone marrow. In contrast, B-cell numbers in the bone marrow and T-cell numbers in the thymus remained unchanged. Alterations in the bone marrow microenvironment may not only reduce myeloid cell numbers, but also have long-term implications for myeloid cell function. Mice were infected with Listeria moncytogenes to analyze the function of myeloid cells. In this case, an inadequate macrophage-dependent clearance of bacterial infections was observed in Col9a1(-/-) mice in vivo. This was mainly caused by an impaired interferon-gamma/tumor necrosis factor-alpha-mediated activation of macrophages. The loss of collagen IX alpha1 therefore destabilizes the trabecular bone network, impairs myeloid cell differentiation, and affects the innate immune response against Listeria. Stem Cells 2018;36:1752-1763.

NK and NKT Cell Depletion Alters the Outcome of Experimental Pneumococcal Pneumonia: Relationship with Regulation of Interferon-γ Production.

Background. Natural killer (NK) and natural killer T (NKT) cells contribute to the innate host defense but their role in bacterial sepsis remains controversial. Methods. C57BL/6 mice were infected intratracheally with 5 × 10(5) cfu of Streptococcus pneumoniae. Animals were divided into sham group (Sham); pretreated with isotype control antibody (CON) group; pretreated with anti-asialo GM1 antibody (NKd) group; and pretreated with anti-CD1d monoclonal antibody (NKTd) group before bacterial challenge. Serum and tissue samples were analyzed for bacterial load, cytokine levels, splenocyte apoptosis rates, and cell characteristics by flow cytometry. Splenocyte miRNA expression was also analyzed and survival was assessed. Results. NK cell depletion prolonged survival. Upon inhibition of NKT cell activation, spleen NK (CD3-/NK1.1+) cells increased compared to all other groups. Inhibition of NKT cell activation led to higher bacterial loads and increased levels of serum and splenocyte IFN-γ. Splenocyte miRNA analysis showed that miR-200c and miR-29a were downregulated, while miR-125a-5p was upregulated, in anti-CD1d treated animals. These changes were moderate after NK cell depletion. Conclusions. NK cells appear to contribute to mortality in pneumococcal pneumonia. Inhibition of NKT cell activation resulted in an increase in spleen NK (CD3-/NK1.1+) cells and a higher IFN-γ production, while altering splenocyte miRNA expression.

Listeria monocytogenes infection in macrophages induces vacuolar-dependent host miRNA response.

Listeria monocytogenes is a gram-positive facultative intracellular pathogen, causing serious illness in immunocompromised individuals and pregnant women. Upon detection by macrophages, which are key players of the innate immune response against infection, L. monocytogenes induces specific host cell responses which need to be tightly controlled at transcriptional and post-transcriptional levels. Here, we ask whether and how host miRNAs, which represent an important mechanism of post-transcriptional regulation in a wide array of biological processes, are altered by a model pathogen upon live infection of murine bone marrow derived macrophages. We first report that L. monocytogenes subverts the host genome-wide miRNA profile of macrophages in vitro. Specifically, we show that miR-155, miR-146a, miR-125a-3p/5p and miR-149 were amongst the most significantly regulated miRNAs in infected macrophages. Strikingly, these miRNAs were highly upregulated upon infection with the Listeriolysin-deficient L. monocytogenes mutant Δhly, that cannot escape from the phagosome thus representing a vacuolar-contained infection. The vacuolar miRNA response was significantly reduced in macrophages deficient for MyD88. In addition, miR-146a and miR-125a-3p/5p were regulated at transcriptional levels upon infection, and miR-125a-3p/5p were found to be TLR2 responsive. Furthermore, miR-155 transactivation in infection was regulated by NF-κB p65, while miR-146a and miR-125a-3p/5p expression was unaffected in p65-deficient primary macrophages upon L. monocytogenes infection. Our results demonstrate that L. monocytogenes promotes significant changes in the miRNA expression profile in macrophages, and reveal a vacuolar-dependent miRNA signature, listeriolysin-independent and MyD88-dependent. These miRNAs are predicted to target immune genes and are therefore most likely involved in regulation of the macrophage innate immune response against infection at post-transcriptional levels.