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Sepsis is a life-threatening process related to a dysregulated host response to an underlying infection which results in organ dysfunction and poor outcomes. Therapeutic strategies using mesenchymal stromal cells (MSCs) are under investigation for sepsis, with efforts to improve cellular utility. Syndecan (SDC) proteins are transmembrane proteoglycans involved with cellular signaling events including tissue repair and modulating inflammation. Bone marrow-derived human MSCs express syndecan-2 (SDC2) at a level higher than other SDC family members, thus we explored SDC2 in MSC function. Administration of human MSCs silenced for SDC2 in experimental sepsis resulted in decreased bacterial clearance, and increased tissue injury and mortality compared with wild-type MSCs. These findings were associated with a loss of resolution of inflammation in the peritoneal cavity, and higher levels of pro-inflammatory mediators in organs. MSCs silenced for SDC2 had a decreased ability to promote phagocytosis of apoptotic neutrophils by macrophages in the peritoneum, and also a diminished capability to convert macrophages from a pro-inflammatory to a pro-resolution phenotype via cellular or paracrine actions. Extracellular vesicles are a paracrine effector of MSCs that may contribute to resolution of inflammation, and their production was dramatically reduced in SDC2 silenced human MSCs. Collectively, these data demonstrate the importance of SDC2 for cellular and paracrine function of human MSCs during sepsis.

Rationale: CD148/PTRJ (receptor-like protein tyrosine phosphatase η) exerts antifibrotic effects in experimental pulmonary fibrosis via interactions with its ligand syndecan-2; however, the role of CD148 in human pulmonary fibrosis remains incompletely characterized. Objectives: We investigated the role of CD148 in the profibrotic phenotype of fibroblasts in idiopathic pulmonary fibrosis (IPF). Methods: Conditional CD148 fibroblast-specific knockout mice were generated and exposed to bleomycin and then assessed for pulmonary fibrosis. Lung fibroblasts (mouse lung and human IPF lung), and precision-cut lung slices from human patients with IPF were isolated and subjected to experimental treatments. A CD148-activating 18-aa mimetic peptide (SDC2-pep) derived from syndecan-2 was evaluated for its therapeutic potential. Measurements and Main Results: CD148 expression was downregulated in IPF lungs and fibroblasts. In human IPF lung fibroblasts, silencing of CD148 increased extracellular matrix production and resistance to apoptosis, whereas overexpression of CD148 reversed the profibrotic phenotype. CD148 fibroblast-specific knockout mice displayed increased pulmonary fibrosis after bleomycin challenge compared with control mice. CD148-deficient fibroblasts exhibited hyperactivated PI3K/Akt/mTOR signaling, reduced autophagy, and increased p62 accumulation, which induced NF-κB activation and profibrotic gene expression. SDC2-pep reduced pulmonary fibrosis in vivo and inhibited IPF-derived fibroblast activation. In precision-cut lung slices from patients with IPF and control patients, SDC2-pep attenuated profibrotic gene expression in IPF and normal lungs stimulated with profibrotic stimuli. Conclusions: Lung fibroblast CD148 activation reduces p62 accumulation, which exerts antifibrotic effects by inhibiting NF-κB–mediated profibrotic gene expression. Targeting the CD148 phosphatase with activating ligands such as SDC2-pep may represent a potential therapeutic strategy in IPF.

Idiopathic pulmonary fibrosis (IPF) is a lethal disease with an average life expectancy of 3 to 5 years. IPF is characterized by progressive stiffening of the lung parenchyma due to excessive deposition of collagen, leading to gradual failure of gas exchange. Although two therapeutic agents have been approved from the FDA for IPF, they only slow disease progression with little impact on outcome. To develop a more effective therapy, we have exploited the fact that collagen-producing myofibroblasts express a membrane-spanning protein, fibroblast activation protein (FAP), that exhibits limited if any expression on other cell types. Because collagen-producing myofibroblasts are only found in fibrotic tissues, solid tumors, and healing wounds, FAP constitutes an excellent marker for targeted delivery of drugs to tissues undergoing pathologic fibrosis. We demonstrate here that a low-molecular weight FAP ligand can be used to deliver imaging and therapeutic agents selectively to FAP-expressing cells. Because induction of collagen synthesis is associated with phosphatidylinositol 3-kinase (PI3K) activation, we designed a FAP-targeted PI3K inhibitor that selectively targets FAP-expressing human IPF lung fibroblasts and potently inhibited collagen synthesis. Moreover, we showed that administration of the inhibitor in a mouse model of IPF inhibited PI3K activation in fibrotic lungs, suppressed production of hydroxyproline (major building block of collagen), reduced collagen deposition, and increased mouse survival. Collectively, these studies suggest that a FAP-targeted PI3K inhibitor might be promising for treating IPF.

Fibrotic diseases cause organ failure that lead to ~45% of all deaths in the United States. Activated macrophages stimulate fibrosis by secreting cytokines that induce fibroblasts to synthesize collagen and extracellular matrix proteins. Although suppression of macrophage‐derived cytokine production can halt progression of fibrosis, therapeutic agents that prevent release of these cytokines (e.g., TLR7 agonists) have proven too toxic to administer systemically. Based on the expression of folate receptor β solely on activated myeloid cells, we have created a folate‐targeted TLR7 agonist (FA‐TLR7‐54) that selectively accumulates in profibrotic macrophages and suppresses fibrosis‐inducing cytokine production. We demonstrate that FA‐TLR7‐54 reprograms M2‐like fibrosis‐inducing macrophages into fibrosis‐suppressing macrophages, resulting in dramatic declines in profibrotic cytokine release, hydroxyproline biosynthesis, and collagen deposition, with concomitant increases in alveolar airspaces. Although nontargeted TLR7‐54 is lethal at fibrosis‐suppressing doses, FA‐TLR7‐54 halts fibrosis without evidence of toxicity. Taken together, FA‐TLR7‐54 is shown to constitute a novel and potent approach for treating fibrosis without causing dose‐limiting systemic toxicities., A folate‐targeted TLR7 agonist (FA‐TLR7‐54) is shown to alleviate pulmonary fibrosis with no detectable toxicity. Achievement of this outcome is enabled by selective reprogramming of profibrotic to antifibrotic macrophages, suppressing the consequent fibroblast activation and collagen biosynthesis.

Supplemental Digital Content is available in the text., Objectives: Mesenchymal stromal cells are being investigated as a cell-based therapy for a number of disease processes, with promising results in animal models of systemic inflammation and sepsis. Studies are ongoing to determine ways to further improve the therapeutic potential of mesenchymal stromal cells. A gas molecule that improves outcome in experimental sepsis is carbon monoxide. We hypothesized that preconditioning of mesenchymal stromal cells with carbon monoxide ex vivo would promote further therapeutic benefit when cells are administered in vivo after the onset of polymicrobial sepsis in mice. Design: Animal study and primary cell culture. Setting: Laboratory investigation. Subjects: BALB/c mice. Interventions: Polymicrobial sepsis was induced by cecal ligation and puncture. Mesenchymal stromal cells, mesenchymal stromal cells-conditioned with carbon monoxide, fibroblasts, or fibroblasts-conditioned with carbon monoxide were delivered by tail vein injections to septic mice. The mice were assessed for survival, bacterial clearance, and the inflammatory response during sepsis in each of the groups. Mesenchymal stromal cells were also assessed for their ability to promote bacterial phagocytosis by neutrophils, the production of specialized proresolving lipid mediators, and their importance for mesenchymal stromal cells function using gene silencing. Measurements and Main Results: Ex vivo preconditioning with carbon monoxide allowed mesenchymal stromal cells to be administered later after the onset of sepsis (6 hr), and yet maintain their therapeutic effect with increased survival. Carbon monoxide preconditioned mesenchymal stromal cells were also able to alleviate organ injury, improve bacterial clearance, and promote the resolution of inflammation. Mesenchymal stromal cells exposed to carbon monoxide, with docosahexaenoic acid substrate, produced specialized proresolving lipid mediators, particularly D-series resolvins, which promoted survival. Silencing of lipoxygenase pathways (5-lipoxygenase and 12/15-lipoxygenase), which are important enzymes for specialized proresolving lipid mediator biosynthesis, resulted in a loss of therapeutic benefit bestowed on mesenchymal stromal cells by carbon monoxide. Conclusions: Taken together, these data suggest that production of specialized proresolving lipid mediators contribute to improved mesenchymal stromal cell efficacy when exposed to carbon monoxide, resulting in an improved therapeutic response during sepsis.