Evidence for reprogramming of monocytes into reparative alveolar macrophages in vivo by targeting PDE4b

Abstract

Increased lung vascular permeability and neutrophilic inflammation are hallmarks of acute lung injury. Alveolar macrophages (AMϕ), the predominant sentinel cell type in the airspace, die in massive numbers while fending off pathogens. Recent studies indicate that the AMϕ pool is replenished by airspace-recruited monocytes, but the mechanisms instructing the conversion of recruited monocytes into reparative AMϕ remain elusive. Cyclic AMP (cAMP) is a vascular barrier protective and immunosuppressive second messenger in the lung. Here, we subjected mice expressing GFP under the control of the Lysozyme-M promoter (LysM-GFP mice) to the LPS model of rapidly resolving lung injury to address the impact of mechanisms determining cAMP levels in AMϕ and regulation of mobilization of the reparative AMϕ-pool. RNA-seq analysis of flow-sorted Mϕ identified phosphodiesterase 4b (PDE4b) as the top LPS-responsive cAMP-regulating gene. We observed that PDE4b expression markedly increased at the time of peak injury (4 h) and then decreased to below the basal level during the resolution phase (24 h). Activation of transcription factor NFATc2 was required for transcription of PDE4b in Mϕ. Inhibition of PDE4 activity at the time of peak injury, using i.t. rolipram, increased cAMP levels, augmented the reparative AMϕ pool, and resolved lung injury. This response was not seen following conditional depletion of monocytes, thus establishing airspace-recruited PDE4b-sensitive monocytes as the source of reparative AMϕ. Interestingly, adoptive transfer of rolipram-educated AMϕ into injured mice resolved lung edema. We propose suppression of PDE4b as an effective approach to promote reparative AMϕ generation from monocytes for lung repair.

RGS2 is an innate immune checkpoint for TLR4 and Gαq-mediated IFNγ generation and lung injury

“…LPS induces inflammatory signaling primarily by activating TLR4 (Joshi et al, 2021; Lu et al, 2015; Rochford et al, 2021). Because TLR4 signaling forms complex with Ca 2+ dependent machinery (Park et al, 2016; Tauseef et al, 2012), we surmised that RGS2 limits Gαq access to TLR4 to regulate IFNγ synthesis.…”

“…Femur and tibia were flushed with RPMI-1640 media (Gibco), containing 25 ng/ml M-CSF, 10% FBS and 1% antibiotic/anti-mycotic as described previously (Joshi et al, 2020; Rayees et al, 2022; Rayees et al, 2019; Rochford et al, 2021). On 5 th -day, bone marrow-derived macrophages were incubated in 0.1% FBS containing RPMI for one hour before adding LPS (1.0 µg/ml) with 1% FBS containing media.…”

CLIC4 Regulates Endothelial Barrier Control by Mediating PAR1 Signaling via RhoA

“…Evans blue albumin extravasation was calculated by transendothelial albumin influx in lungs versus plasma. 36,37…”

“…After 24 hours, the lung dry weight was taken and lung wet/dry weight ratio was calculated as described previously. 36,37 Vascular permeability was quantified by centrifuging the plasma and lung homogenate to isolate the supernatant. The optical density was measured spectrophotometrically at 595 nm (Evans blue) and 750 nm (hemoglobin correction).…”

Regulation of cGAS Activity and Downstream Signaling

“…ALI is a devastating inflammatory condition of the lungs that is characterized by disruption of alveolar-capillary junctions leading to an uncontrolled influx of proteins rich fluid and blood cells into the alveolar space [ 102 , 103 , 104 , 105 ]. Neutrophils and tissue-resident macrophages propagate inflammatory responses by releasing various inflammatory cytokines and chemoattractants.…”