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Targeting activated fibroblasts, including myofibroblast differentiation, has emerged as a key therapeutic strategy in patients with idiopathic pulmonary fibrosis (IPF). However, there is no available therapy capable of selectively eradicating myofibroblasts or limiting their genesis. Through an integrative analysis of the regulator genes that are responsible for the activation of IPF fibroblasts, we noticed the phosphatidylinositol 4,5-bisphosphate (PIP2)-binding protein, myristoylated alanine-rich C-kinase substrate (MARCKS), as a potential target molecule for IPF. Herein, we have employed a 25-mer novel peptide, MARCKS phosphorylation site domain sequence (MPS), to determine if MARCKS inhibition reduces pulmonary fibrosis through the inactivation of PI3K/protein kinase B (AKT) signaling in fibroblast cells. We first observed that higher levels of MARCKS phosphorylation and the myofibroblast marker α-smooth muscle actin (α-SMA) were notably overexpressed in all tested IPF lung tissues and fibroblast cells. Treatment with the MPS peptide suppressed levels of MARCKS phosphorylation in primary IPF fibroblasts. A kinetic assay confirmed that this peptide binds to phospholipids, particularly PIP2, with a dissociation constant of 17.64 nM. As expected, a decrease of phosphatidylinositol (3,4,5)-trisphosphate pools and AKT activity occurred in MPS-treated IPF fibroblast cells. MPS peptide was demonstrated to impair cell proliferation, invasion, and migration in multiple IPF fibroblast cells in vitro as well as to reduce pulmonary fibrosis in bleomycin-treated mice in vivo. Surprisingly, we found that MPS peptide decreases α-SMA expression and synergistically interacts with nintedanib treatment in IPF fibroblasts. Our data suggest MARCKS as a druggable target in pulmonary fibrosis and also provide a promising antifibrotic agent that may lead to effective IPF treatments.-Yang, D. C., Li, J.-M., Xu, J., Oldham, J., Phan, S. H., Last, J. A., Wu, R., Chen, C.-H. Tackling MARCKS-PIP3 circuit attenuates fibroblast activation and fibrosis progression.

Although stromal-derived factor-1 (SDF-1) via its cognate receptor CXCR4 is assumed to play a critical role in migration of endothelial cells during new vessel formation after tissue injury, CXCR4 expression on endothelial cells is strictly regulated. Erythromycin (EM), a 14-membered ring macrolide, has an anti-inflammatory effect that may account for its clinical benefit in the treatment of chronic inflammatory diseases. However, the effects of EM on endothelial cells and especially their expression of CXCR4 have not been fully evaluated. In this study, we demonstrated that EM markedly induced CXCR4 surface expression on microvascular endothelial cells in vitro and lung capillary endothelial cells in vivo. This ability to induce CXCR4 surface expression on endothelial cells was restricted to 14-membered ring macrolides and was not observed in other antibiotics including a 16-membered ring macrolide, josamycin. Furthermore, this EM-induced expression of CXCR4 on endothelial cells was functionally significant as demonstrated by chemotaxis assays in vitro. These findings suggest that EM-induced CXCR4 surface expression on endothelial cells may promote migration of CXCR4-expressing endothelial cells into sites of tissue injury, which may be associated with the known anti-inflammatory activity of this macrolide. stromal-derived factor-1; angiogenesis

Pulmonary fibrosis is characterized by alterations in fibroblast phenotypes resulting in excessive extracellular matrix accumulation and anatomic remodeling. Current therapies for this condition are largely ineffective. Peroxisome proliferator-activated receptor-[gamma] (PPAR-[gamma]) is a member of the nuclear hormone receptor superfamily, the activation of which produces a number of biological effects, including alterations in metabolic and inflammatory responses. The role of PPAR-[gamma] as a potential therapeutic target for fibrotic lung diseases remains undefined. In the present study, we show expression of PPAR-[gamma] in fibroblasts obtained from normal human lungs and lungs of patients with idiopathic interstitial pneumonias. Treatment of lung fibroblasts and myofibroblasts with PPAR-[gamma] agonists results in inhibition of proliferative responses and induces cell cycle arrest. In addition, PPAR-[gamma] agonists, including a constitutively active PPAR-[gamma] construct (VP16-PPAR-[gamma]), inhibit the ability of transforming growth factor-[beta]1 to induce myofibroblast differentiation and collagen secretion. PPAR-[gamma] agonists also inhibit fibrosis in a routine model, even when administration is delayed until after the initial inflammation has largely resolved. These observations indicate that PPAR-[gamma] is an important regulator of fibroblast/myofibroblast activation and suggest a role for PPAR-[gamma] ligands as novel therapeutic agents for fibrotic lung diseases. troglitazone; ciglitazone; transforming growth factor

In addition to its well-known expression in the germline and in cells of certain cancers, telomerase activity is induced in lung fibrosis, although its role in this process is unknown. To identify the pathogenetic importance of telomerase in lung fibrosis, we examined the effects of telomerase reverse transcriptase (TERT) deficiency in a murine model of pulmonary injury. TERT-deficient mice showed significantly reduced lung fibrosis following bleomycin (BLM) insult. This was accompanied by a significant reduction in expression of lung [alpha]-SMA, a marker of myofibroblast differentiation. Furthermore, lung fibroblasts isolated from BLM-treated TERT-deficient mice showed significantly decreased proliferation and increased apoptosis rates compared with cells isolated from control mice. Transplantation of WT BM into TERT-deficient mice restored BLM-induced lung telomerase activity and fibrosis to WT levels. Conversely, transplantation of BM from TERT-deficient mice into WT recipients resulted in reduced telomerase activity and fibrosis. These findings suggest that induction of telomerase in injured lungs may be caused by BM-derived cells, which appear to play an important role in pulmonary fibrosis. Moreover, TERT induction is associated with increased survival of lung fibroblasts, which favors the development of fibrosis instead of injury resolution., Introduction Telomeres consist of tandem hexanucleotide [(TTAGGG).sub.n] repeats that cap the termini of eukaryotic chromosomes and play essential roles in maintenance of chromosomal stability and cell viability (1). The complex [...]

The bone morphogenetic proteins (BMPs) profoundly affect embryonic development, differentiation and disease. BMP signaling is suppressed by cysteine-rich domain proteins, such as chordin, that sequester ligands from the BMP receptor. We describe a novel protein, KCP, with 18 cysteine-rich domains. Unlike chordin, KCP enhances BMP signaling in a paracrine manner. Smad1-dependent transcription and phosphorylated Smad1 (P-Smad1) levels are increased, as KCP binds to BMP7 and enhances binding to the type I receptor. In vivo, Kcp[sup.-/-] mice are viable and fertile. Because BMPs have a pivotal role in renal disease, we examined the phenotype of Kcp[sup.-/-] mice in two different models of renal injury. Kcp[sup.-/-] animals show reduced levels of P-Smad1, are more susceptible to developing renal interstitial fibrosis, are more sensitive to tubular injury and show substantial pathology after recovery. The data indicate an important role for KCP in attenuating the pathology of renal fibrotic disease., Author(s): Jingmei Lin [1]; Sanjeevkumar R Patel [2]; Xu Cheng [3]; Eun Ah Cho [1]; Inna Levitan [1]; Matthew Ullenbruch [1]; Sem H Phan [1]; John M Park [3]; Gregory [...]

The clinical significance of B7H3 (CD276) and its cleavage product soluble B7H3 (sB7H3) in idiopathic pulmonary fibrosis (IPF) is unknown. Mounting evidence suggests the potential utility of peripheral blood myeloid cell enumeration to predict disease outcome and indicate active lung disease. Here we hypothesized that sB7H3 is involved in regulation of circulating myeloid cells in pulmonary fibrosis. In support of this possibility, both plasma sB7H3 and B7H3+ cells were elevated in IPF patient blood samples, which correlated negatively with lung function. To analyze its function, the effects of sB7H3 on naïve or bleomycin‐treated mice were examined. The results revealed that sB7H3 injection induced an influx of myeloid‐derived suppressor cells (MDSCs) and Ccl2 expression in lung tissue of naïve mice, accompanied by enhanced overall inflammation. Additionally, sB7H3 caused accumulation of MDSCs in bone marrow with increased expression of inflammatory cytokines. Notably, in vitro assays revealed chemotaxis of MDSCs to sB7H3, which was dependent on TLT‐2 (TREML2), a putative receptor for sB7H3. Thus, increased circulating sB7H3 and/or B7H3+ cells in IPF patient blood samples correlated with lung function decline and potential immunosuppressive status. The correlation of sB7H3 with deterioration of lung function might be due to its ability to enhance inflammation and recruitment of MDSCs into the lung and their expansion in the bone marrow, and thus potentially contribute to IPF exacerbation. © 2021 The Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

The Wnt/b-catenin pathway initiates a signaling cascade that is critical in cell differentiation and the normal development of multiple organ systems. The reactivation of this pathway has been documented in experimental and human idiopathic pulmonary fibrosis, wherein Wnt/b-catenin activation has been implicated in epithelial-cell repair. Furthermore, the canonical ligand Wnt3a is known to induce myofibroblast differentiation; however, the role of noncanonical Wnt ligands remains unclear. This study showed significantly higher levels of Wnt11 expression in cells from both patients with idiopathic pulmonary fibrosis and bleomycin-treated mice, as well as in TGFb-treated mouse lung fibroblasts. Moreover, Wnt11 induced myofibroblast differentiation as manifested by increased a-SMA (ACTA2) expression, which was similar to that induced by canonical Wnt3a/b-catenin signaling. Further investigation revealed that Wnt11 induction of a-SMA was associated with the activation of JNK (c-Jun N-terminal kinase)/c-Jun signaling and was inhibited by a JNK inhibitor. The potential importance of this signaling pathway was supported by in vivo evidence showing significantly increased levels of Wnt11 and activated JNK in the lungs of mice with bleomycin-induced pulmonary fibrosis. Interestingly, fibroblasts did not express canonical Wnt3a, but treatment of these cells with exogenous Wnt3a induced endogenous Wnt11 and Wnt5a, resulting in repression of the Wnt3a/b-catenin target gene Axin2. These findings suggested that the noncanonical Wnt induction of myofibroblast differentiation mediated by the JNK/c-Jun pathway might play a significant role in pulmonary fibrosis, in addition to or in synergy with canonical Wnt3a/b-catenin signaling. Moreover, Wnt3a activation of noncanonical Wnt signaling might trigger a switch from canonical to noncanonical Wnt signaling to induce myofibroblast differentiation. [ABSTRACT FROM AUTHOR]