Aberrant fibroblast function plays a key role in the pathogenesis of idiopathic pulmonary fibrosis, a devastating disease of unrelenting extracellular matrix deposition in response to lung injury. Platelet-derived growth factor α-positive (Pdgfra+) lipofibroblasts (LipoFBs) are essential for lung injury response and maintenance of a functional alveolar stem cell niche. Little is known about the effects of lung injury on LipoFB function. Here, we used single-cell RNA-Seq (scRNA-Seq) technology and PdgfraGFP lineage tracing to generate a transcriptomic profile of Pdgfra+ fibroblasts in normal and injured mouse lungs 14 days after bleomycin exposure, generating 11 unique transcriptomic clusters that segregated according to treatment. While normal and injured LipoFBs shared a common gene signature, injured LipoFBs acquired fibrogenic pathway activity with an attenuation of lipogenic pathways. In a 3D organoid model, injured Pdgfra+ fibroblast-supported organoids were morphologically distinct from those cultured with normal fibroblasts, and scRNA-Seq analysis suggested distinct transcriptomic changes in alveolar epithelia supported by injured Pdgfra+ fibroblasts. In summary, while LipoFBs in injured lung have not migrated from their niche and retain their lipogenic identity, they acquire a potentially reversible fibrogenic profile, which may alter the kinetics of epithelial regeneration and potentially contribute to dysregulated repair, leading to fibrosis.

Cancer progresses through distinct stages, and mouse models recapitulating traits of this progression are frequently used to explore genetic, morphological, and pharmacological aspects of tumor development. To complement genomic investigations of this process, we here quantify phosphoproteomic changes in skin cancer development using the SILAC mouse technology coupled to high-resolution mass spectrometry. We distill protein expression signatures from our data that distinguish between skin cancer stages. A distinct phosphoproteome of the two stages of cancer progression is identified that correlates with perturbed cell growth and implicates cell adhesion as a major driver of malignancy. Importantly, integrated analysis of phosphoproteomic data and prediction of kinase activity revealed PAK4-PKC/SRC network to be highly deregulated in SCC but not in papilloma. This detailed molecular picture, both at the proteome and phosphoproteome level, will prove useful for the study of mechanisms of tumor progression.

Plasmacytoid dendritic cells (pDC) are the major producers of type I interferons (IFNs) in humans and rapidly produce IFN-α in response to virus exposure. Although HIV infection is associated with pDC activation, it is unclear why the innate immune response is unable to effectively control viral replication. We systematically compared the effect of HIV, Influenza, Sendai, and HSV-2 at similar target cell multiplicity of infection (M.O.I.) on human pDC function. We found that Influenza, Sendai, HSV-2 and imiquimod are able to rapidly induce IFN-α production within 4 hours to maximal levels, whereas HIV had a delayed induction that was maximal only after 24 hours. In addition, maximal IFN-α induction by HIV was at least 10 fold less than that of the other viruses in the panel. HIV also induced less TNF-α and MIP-1β but similar levels of IP-10 compared to other viruses, which was also mirrored by delayed upregulation of pDC activation markers CD83 and CD86. BDCA-2 has been identified as an inhibitory receptor on pDC, signaling through a pathway that involves SYK phosphorylation. We find that compared to Influenza, HIV induces the activation of the SYK pathway. Thus, HIV delays pDC IFN-α production and pDC activation via SYK phosphorylation, allowing establishment of viral populations.

Lung disease causes significant morbidity and mortality, and is exacerbated by environmental injury, for example through lipopolysaccharide (LPS) or ozone (O3). Toll-like receptors (TLRs) orchestrate immune responses to injury by recognizing pathogen- or danger-associated molecular patterns. TLR4, the prototypic receptor for LPS, also mediates inflammation after O3, triggered by endogenous hyaluronan. Regulation of TLR4 signaling is incompletely understood. TLR5, the flagellin receptor, is expressed in alveolar macrophages, and regulates immune responses to environmental injury. Using in vivo animal models of TLR4-mediated inflammations (LPS, O3, hyaluronan), we show that TLR5 impacts the in vivo response to LPS, hyaluronan and O3. We demonstrate that immune cells of human carriers of a dominant negative TLR5 allele have decreased inflammatory response to O3 exposure ex vivo and LPS exposure in vitro. Using primary murine macrophages, we find that TLR5 physically associates with TLR4 and biases TLR4 signaling towards the MyD88 pathway. Our results suggest an updated paradigm for TLR4/TLR5 signaling.

The protein-tyrosine phosphatase Shp1 is expressed ubiquitously in hematopoietic cells and is generally viewed as a negative regulatory molecule. Mutations in Ptpn6, which encodes Shp1, result in widespread inflammation and premature death, known as the motheaten (me) phenotype. Previous studies identified Shp1 as a negative regulator of TCR signaling, but the severe systemic inflammation in me mice may have confounded our understanding of Shp1 function in T cell biology. To define the T cell–intrinsic role of Shp1, we characterized mice with a T cell–specific Shp1 deletion (Shp1fl/fl CD4-cre). Surprisingly, thymocyte selection and peripheral TCR sensitivity were unaltered in the absence of Shp1. Instead, Shp1(fl/fl) CD4-cre mice had increased frequencies of memory phenotype T cells that expressed elevated levels of CD44. Activation of Shp1-deficient CD4⁺ T cells also resulted in skewing to the Th2 lineage and increased IL-4 production. After IL-4 stimulation of Shp1- deficient T cells, Stat 6 activation was sustained, leading to enhanced Th2 skewing. Accordingly, we observed elevated serum IgE in the steady state. Blocking or genetic deletion of IL-4 in the absence of Shp1 resulted in a marked reduction of the CD44hi population. Therefore, Shp1 is an essential negative regulator of IL-4 signaling in T lymphocytes.