Heme oxygenase-1 dampens the macrophage sterile inflammasome response and regulates its components in the hypoxic lung.

Exposure to hypoxia causes an inflammatory reaction in the mouse lung, and this response can be modulated by overexpressing the hypoxia-inducible stress-response enzyme, heme oxygenase-1 (HO-1). We hypothesized that the inflammasome activity may be a central pathway by which HO-1 controls pulmonary inflammation following alveolar hypoxia. Therefore, we investigated whether HO-1 controls inflammasome activation by altering its expression in macrophages primed with classic NOD-like receptor containing a pyrin domain 3 (NLRP3) inducers, and in murine lungs lacking HO-1 and exposed to acute hypoxia. We found that lack of HO-1 activated lipopolysaccharide (LPS) and ATP-treated bone marrow-derived macrophages, causing an increase in secreted levels of cleaved interleukin (IL)-1B, IL-18, and caspase-1, markers of increased inflammasome activity, whereas HO-1 overexpression suppressed IL-1B, NLRP3, and IL-18. The production of cleaved IL-1B and the activation of caspase-1 in LPS- and ATP-primed macrophages were inhibited by hemin, an HO-1 inducer, and two HO-1 enzymatic products [bilirubin and carbon monoxide (CO)]. Exposure of mice to hypoxia induced the expression of several inflammasome mRNA components (IL-1B, Nlrp3, and caspase-1), and this was further augmented by HO-1 deficiency. This pronounced inflammasome activation was detected as increased protein levels of apoptosis-associated speck-like protein containing a COOH-terminal caspase recruitment domain, IL-18, procaspase-1, and cleaved caspase-1 in the lungs of hypoxic mice. Systemically, Hmox1-deficient mice showed increased basal levels of IL-18 that were further increased after 48 h of hypoxic exposure. Taken together, these finding point to a pivotal role for HO-1 in the control of baseline and hypoxic inflammasome signaling, perhaps through the antioxidant properties of bilirubin and CO's pleiotropic effects.

Mesenchymal Stromal Cell Exosomes Ameliorate Experimental Bronchopulmonary Dysplasia and Restore Lung Function through Macrophage Immunomodulation.

RATIONALE: Mesenchymal stem/stromal cell (MSC) therapies have shown promise in preclinical models of pathologies relevant to newborn medicine, such as bronchopulmonary dysplasia (BPD). We have reported that the therapeutic capacity of MSCs is comprised in their secretome, and demonstrated that the therapeutic vectors are exosomes produced by MSCs (MSC-exos). OBJECTIVES: To assess efficacy of MSC-exo treatment in a preclinical model of BPD and to investigate mechanisms underlying MSC-exo therapeutic action. METHODS: Exosomes were isolated from media conditioned by human MSC cultures. Newborn mice were exposed to hyperoxia (HYRX; 75% O2), treated with exosomes on Postnatal Day (PN) 4 and returned to room air on PN7. Treated animals and appropriate controls were harvested on PN7, -14, or -42 for assessment of pulmonary parameters. MEASUREMENTS AND MAIN RESULTS: HYRX-exposed mice presented with pronounced alveolar simplification, fibrosis, and pulmonary vascular remodeling, which was effectively ameliorated by MSC-exo treatment. Pulmonary function tests and assessment of pulmonary hypertension showed functional improvements after MSC-exo treatment. Lung mRNA sequencing demonstrated that MSC-exo treatment induced pleiotropic effects on gene expression associated with HYRX-induced inflammation and immune responses. MSC-exos modulate the macrophage phenotype fulcrum, suppressing the proinflammatory "M1" state and augmenting an antiinflammatory "M2-like" state, both in vitro and in vivo. CONCLUSIONS: MSC-exo treatment blunts HYRX-associated inflammation and alters the hyperoxic lung transcriptome. This results in alleviation of HYRX-induced BPD, improvement of lung function, decrease in fibrosis and pulmonary vascular remodeling, and amelioration of pulmonary hypertension. The MSC-exo mechanism of action is associated with modulation of lung macrophage phenotype.