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CCN1 expression by fibroblasts is required for bleomycin-induced skin fibrosis.

ABSTRACT: The microenvironment contributes to the excessive connective tissue deposition that characterizes fibrosis. Members of the CCN family of matricellular proteins are secreted by fibroblasts into the fibrotic microenvironment; however, the role of endogenous CCN1 in skin fibrosis is unknown. Mice harboring a fibroblast-specific deletion for CCN1 were used to assess if CCN1 contributes to dermal homeostasis, wound healing, and skin fibrosis. Mice with a fibroblast-specific CCN1 deletion showed progressive skin thinning and reduced accumulation of type I collagen; however, the overall mechanical property of skin (Young's modulus) was not significantly reduced. Real time-polymerase chain reaction analysis revealed that CCN1-deficient skin displayed reduced expression of mRNAs encoding enzymes that promote collagen stability (including prolyl-4-hydroxylase and PLOD2), although expression of COL1A1 mRNA was unaltered. CCN1-deficent skin showed reduced hydroxyproline levels. Electron microscopy revealed that collagen fibers were disorganized in CCN1-deficient skin. CCN1-deficient mice were resistant to bleomycin-induced skin fibrosis, as visualized by reduced collagen accumulation and skin thickness suggesting that deposition/accumulation of collagen is impaired in the absence of CCN1. Conversely, CCN1-deficient mice showed unaltered wound closure kinetics, suggesting de novo collagen production in response to injury did not require CCN1. In response to either wounding or bleomycin, induction of ?-smooth muscle actin-positive myofibroblasts was unaffected by loss of CCN1. CCN1 protein was overexpressed by dermal fibroblasts isolated from lesional (i.e., fibrotic) areas of patients with early onset diffuse scleroderma. Thus, CCN1 expression by fibroblasts, being essential for skin fibrosis, is a viable anti-fibrotic target.

SUBMITTER: Quesnel K

PROVIDER: S-EPMC7852207 | biostudies-literature | 2019 Aug

REPOSITORIES: biostudies-literature

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CCN1 expression by fibroblasts is required for bleomycin-induced skin fibrosis.

Quesnel Katherine K Shi-Wen Xu X Hutchenreuther James J Xiao Yizhi Y Liu Shangxi S Peidl Alexander A Naskar Deboki D Siqueira Walter L WL O'Gorman David B DB Hinz Boris B Stratton Richard J RJ Leask Andrew A

Matrix biology plus 20190706

The microenvironment contributes to the excessive connective tissue deposition that characterizes fibrosis. Members of the CCN family of matricellular proteins are secreted by fibroblasts into the fibrotic microenvironment; however, the role of endogenous CCN1 in skin fibrosis is unknown. Mice harboring a fibroblast-specific deletion for CCN1 were used to assess if CCN1 contributes to dermal homeostasis, wound healing, and skin fibrosis. Mice with a fibroblast-specific CCN1 deletion showed progr ...[more]

PMID: 33543008

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Project description:BackgroundMacrophages are involved in the pathogenesis of idiopathic pulmonary fibrosis, partially by activating lung fibroblasts. However, how macrophages communicate with lung fibroblasts is largely unexplored. Exosomes can mediate intercellular communication, whereas its role in lung fibrogenesis is unclear. Here we aim to investigate whether exosomes can mediate the crosstalk between macrophages and lung fibroblasts and subsequently induce fibrosis.MethodsIn vivo, bleomycin (BLM)-induced lung fibrosis model was established and macrophages infiltration was examined. The effects of GW4869, an exosomes inhibitor, on lung fibrosis were assessed. Moreover, macrophage exosomes were injected into mice to observe its pro-fibrotic effects. In vitro, exosomes derived from angiotensin II (Ang II)-stimulated macrophages were collected. Then, lung fibroblasts were treated with the exosomes. Twenty-four hours later, protein levels of α-collagen I, angiotensin II type 1 receptor (AT1R), transforming growth factor-β (TGF-β), and phospho-Smad2/3 (p-Smad2/3) in lung fibroblasts were examined. The Student's t test or analysis of variance were used for statistical analysis.ResultsIn vivo, BLM-treated mice showed enhanced infiltration of macrophages, increased fibrotic alterations, and higher levels of Ang II and AT1R. GW4869 attenuated BLM-induced pulmonary fibrosis. Mice with exosomes injection showed fibrotic features with higher levels of Ang II and AT1R, which was reversed by irbesartan. In vitro, we found that macrophages secreted a great number of exosomes. The exosomes were taken by fibroblasts and resulted in higher levels of AT1R (0.22 ± 0.02 vs. 0.07 ± 0.02, t = 8.66, P = 0.001), TGF-β (0.54 ± 0.05 vs. 0.09 ± 0.06, t = 10.00, P < 0.001), p-Smad2/3 (0.58 ± 0.06 vs. 0.07 ± 0.03, t = 12.86, P < 0.001) and α-collagen I (0.27 ± 0.02 vs. 0.16 ± 0.01, t = 7.01, P = 0.002), and increased Ang II secretion (62.27 ± 7.32 vs. 9.56 ± 1.68, t = 12.16, P < 0.001). Interestingly, Ang II increased the number of macrophage exosomes, and the protein levels of Alix (1.45 ± 0.15 vs. 1.00 ± 0.10, t = 4.32, P = 0.012), AT1R (4.05 ± 0.64 vs. 1.00 ± 0.09, t = 8.17, P = 0.001), and glyceraldehyde-3-phosphate dehydrogenase (2.13 ± 0.36 vs. 1.00 ± 0.10, t = 5.28, P = 0.006) were increased in exosomes secreted by the same number of macrophages, indicating a positive loop between Ang II and exosomes production.ConclusionsExosomes mediate intercellular communication between macrophages and fibroblasts plays an important role in BLM-induced pulmonary fibrosis.

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CCN1 expression by fibroblasts is required for bleomycin-induced skin fibrosis.

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CCN1 expression by fibroblasts is required for bleomycin-induced skin fibrosis.

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