Effect of hypoxia in skeletal muscle fibrosis; regulation of CTGF/CCN2 expression by HIF-1α and TGF-β

Fibrosis is a recurrent condition of several pathologies associated to the skeletal muscle, among them; Duchenne Muscular Dystrophy (DMD), motor-neuron diseases including Amyotrophic Lateral Sclerosis (ALS) or denervation, and chronic damage. Muscle fibrosis is characterized by the excessive accumul...

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Autor: Valle Tenney, Roger
Tipo de documento: tese
Estado:Versão publicada
Data de publicação:2019
País:Chile
OAI Identifier:oai:repositorio.anid.cl:10533/236575
Acesso em linha:https://hdl.handle.net/10533/236575
Access Level:Acceso aberto
Palavra-chave:Ciencias Naturales
Otras Ciencias Naturales
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dc.title.es_CL.fl_str_mv Effect of hypoxia in skeletal muscle fibrosis; regulation of CTGF/CCN2 expression by HIF-1α and TGF-β
title Effect of hypoxia in skeletal muscle fibrosis; regulation of CTGF/CCN2 expression by HIF-1α and TGF-β
spellingShingle Effect of hypoxia in skeletal muscle fibrosis; regulation of CTGF/CCN2 expression by HIF-1α and TGF-β
Valle Tenney, Roger
Ciencias Naturales
Otras Ciencias Naturales
title_short Effect of hypoxia in skeletal muscle fibrosis; regulation of CTGF/CCN2 expression by HIF-1α and TGF-β
title_full Effect of hypoxia in skeletal muscle fibrosis; regulation of CTGF/CCN2 expression by HIF-1α and TGF-β
title_fullStr Effect of hypoxia in skeletal muscle fibrosis; regulation of CTGF/CCN2 expression by HIF-1α and TGF-β
title_full_unstemmed Effect of hypoxia in skeletal muscle fibrosis; regulation of CTGF/CCN2 expression by HIF-1α and TGF-β
title_sort Effect of hypoxia in skeletal muscle fibrosis; regulation of CTGF/CCN2 expression by HIF-1α and TGF-β
dc.creator.none.fl_str_mv Valle Tenney, Roger
author Valle Tenney, Roger
author_facet Valle Tenney, Roger
author_role author
dc.contributor.advisor.none.fl_str_mv Brandan Siques, Enrique
dc.contributor.institution.es_CL.fl_str_mv PONTIFICIA UNIVERSIDAD CATOLICA DE CHILE
dc.subject.oecd1n.es_CL.fl_str_mv Ciencias Naturales
topic Ciencias Naturales
Otras Ciencias Naturales
dc.subject.oecd2n.es_CL.fl_str_mv Otras Ciencias Naturales
description Fibrosis is a recurrent condition of several pathologies associated to the skeletal muscle, among them; Duchenne Muscular Dystrophy (DMD), motor-neuron diseases including Amyotrophic Lateral Sclerosis (ALS) or denervation, and chronic damage. Muscle fibrosis is characterized by the excessive accumulation of extracellular matrix (ECM) proteins, persistent inflammation, muscle waste, and over-expression of profibrotic factors. Such as transforming growth factor type β (TGF-β) and connective tissue growth factor (CTGF/CCN2). Several reports show that fibrotic muscles are also associated with vascular damage. Preliminary results from our laboratory show that murine models that develop fibrosis are associated with a decrease in the number of capillaries that surrounds each muscle fiber. In consequence, this is linked to a decrease in blood perfusion; and therefore, to a drop in the oxygen tension in the tissue. This phenomenon is called hypoxia and triggers the activation of molecular mechanisms of hypoxic response that allows the survival and adaptation under low oxygen concentrations. However, the relationship between hypoxia and fibrosis in skeletal muscle has not been studied. The transcription factor Hypoxia-Inducible Factor 1α (HIF-1α) is the central regulator of hypoxic response, while CTGF/CCN2 is a key matricellular protein in fibrosis induction. There is evidence that links CTGF/CCN2 expression regulation by hypoxia, and specifically by HIF-1α in opposite ways in different tissues. However, the regulation of CTGF/CCN2 expression by hypoxia in skeletal muscle has not been described. The aim of this thesis is to study the molecular mechanism and the signaling pathways involved in the regulation of expression of CTGF/CCN2 in the different cell types that compose skeletal muscle in vitro and in vivo. For this analysis, we systematically exposed fibroblasts, myoblasts and myotubes to hypoxic conditions in a controlled atmosphere cell culture chamber together with TGF-β stimulation. First, we found that hypoxic treatment at different times under hypoxic atmosphere did not induce the expression of CTGF/CCN2 in none of the cell types mentioned above. Interestingly, we found that specifically myotubes over-express CTGF/CCN2 after combined activation of hypoxic and TGF-β signaling pathways. We classify this phenotype as a synergistic effect and requires the HIF-1α transcription factor and non-canonical TGF-β signaling pathways. Finally, we demonstrate these results in vivo using two different methodologies to activate hypoxic signaling mediated by HIF-1α; using pharmacological HIF-1α stabilizers and by ischemia through femoral artery scission. We combined these methodologies with intramuscular injections of TGF-β, and we were able to resemble the effect observed in vitro. HIF-1α activation, together with an intramuscular injection of TGF-β synergistically induce CTGF/CCN2 expression in skeletal muscle. Moreover, CTGF/CCN2 was localized in muscle fibers in vivo. Based on these results, we hypothesize that muscle fibers are the main source of CTGF/CCN2 in the skeletal muscle in response to hypoxia and TGF-β, contributing to the fibrotic pathology progression.
publishDate 2019
dc.date.accessioned.none.fl_str_mv 2019-10-03T13:47:19Z
2022-08-18T14:12:50Z
dc.date.available.none.fl_str_mv 2019-10-03T13:47:19Z
2022-08-18T14:12:50Z
dc.date.issued.es_CL.fl_str_mv 2019
dc.type.none.fl_str_mv Tesis Doctorado
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spelling PONTIFICIA UNIVERSIDAD CATOLICA DE CHILEValle Tenney, Roger2019https://hdl.handle.net/10533/236575http://purl.org/coar/access_right/c_abf2Otras Ciencias NaturalesCiencias NaturalesEffect of hypoxia in skeletal muscle fibrosis; regulation of CTGF/CCN2 expression by HIF-1α and TGF-βBrandan Siques, EnriquePONTIFICIA UNIVERSIDAD CATOLICA DE CHILEChileValle Tenney, Roger2019-10-03T13:47:19Z2022-08-18T14:12:50Z2019-10-03T13:47:19Z2022-08-18T14:12:50Z2019Fibrosis is a recurrent condition of several pathologies associated to the skeletal muscle, among them; Duchenne Muscular Dystrophy (DMD), motor-neuron diseases including Amyotrophic Lateral Sclerosis (ALS) or denervation, and chronic damage. Muscle fibrosis is characterized by the excessive accumulation of extracellular matrix (ECM) proteins, persistent inflammation, muscle waste, and over-expression of profibrotic factors. Such as transforming growth factor type β (TGF-β) and connective tissue growth factor (CTGF/CCN2). Several reports show that fibrotic muscles are also associated with vascular damage. Preliminary results from our laboratory show that murine models that develop fibrosis are associated with a decrease in the number of capillaries that surrounds each muscle fiber. In consequence, this is linked to a decrease in blood perfusion; and therefore, to a drop in the oxygen tension in the tissue. This phenomenon is called hypoxia and triggers the activation of molecular mechanisms of hypoxic response that allows the survival and adaptation under low oxygen concentrations. However, the relationship between hypoxia and fibrosis in skeletal muscle has not been studied. The transcription factor Hypoxia-Inducible Factor 1α (HIF-1α) is the central regulator of hypoxic response, while CTGF/CCN2 is a key matricellular protein in fibrosis induction. There is evidence that links CTGF/CCN2 expression regulation by hypoxia, and specifically by HIF-1α in opposite ways in different tissues. However, the regulation of CTGF/CCN2 expression by hypoxia in skeletal muscle has not been described. The aim of this thesis is to study the molecular mechanism and the signaling pathways involved in the regulation of expression of CTGF/CCN2 in the different cell types that compose skeletal muscle in vitro and in vivo. For this analysis, we systematically exposed fibroblasts, myoblasts and myotubes to hypoxic conditions in a controlled atmosphere cell culture chamber together with TGF-β stimulation. First, we found that hypoxic treatment at different times under hypoxic atmosphere did not induce the expression of CTGF/CCN2 in none of the cell types mentioned above. Interestingly, we found that specifically myotubes over-express CTGF/CCN2 after combined activation of hypoxic and TGF-β signaling pathways. We classify this phenotype as a synergistic effect and requires the HIF-1α transcription factor and non-canonical TGF-β signaling pathways. Finally, we demonstrate these results in vivo using two different methodologies to activate hypoxic signaling mediated by HIF-1α; using pharmacological HIF-1α stabilizers and by ischemia through femoral artery scission. We combined these methodologies with intramuscular injections of TGF-β, and we were able to resemble the effect observed in vitro. HIF-1α activation, together with an intramuscular injection of TGF-β synergistically induce CTGF/CCN2 expression in skeletal muscle. Moreover, CTGF/CCN2 was localized in muscle fibers in vivo. Based on these results, we hypothesize that muscle fibers are the main source of CTGF/CCN2 in the skeletal muscle in response to hypoxia and TGF-β, contributing to the fibrotic pathology progression.PFCHA-BecasPFCHA-Becas21150188https://hdl.handle.net/10533/236575instname: Conicytreponame: Repositorio Digital RI2.0info:eu-repo/grantAgreement//DOCTORADOinfo:eu-repo/grantAgreement//21150188info:eu-repo/semantics/dataset/hdl.handle.net/10533/93488https://repositorio.uc.cl/handle/11534/26333info:eu-repo/semantics/openAccessCiencias NaturalesOtras Ciencias NaturalesEffect of hypoxia in skeletal muscle fibrosis; regulation of CTGF/CCN2 expression by HIF-1α and TGF-βTesis Doctoradoinfo:eu-repo/semantics/doctoralThesisinfo:eu-repo/semantics/publishedVersionTesisTesishttps://hdl.handle.net/10533/236575PFCHA-Becasc55d1038-7d31-4099-b343-bba9bea70409virtual::53558-1c55d1038-7d31-4099-b343-bba9bea70409virtual::53558-1ORIGINALTesis de doctorado Roger Valle version final RV.pdfapplication/pdf1711641https://repositorio.anid.cl/bitstreams/1330a80c-d4a8-4167-b7f3-4ddfbb29a6b9/downloadffa63c17876b9c5ebd9ac1ada36cdccbMD51LICENSElicense.txttext/plain1779https://repositorio.anid.cl/bitstreams/b809626a-60b9-4789-9013-1cabf39b6db0/download593a6e7305c66c56041a9f9e15a649c1MD52TEXTTesis de doctorado Roger Valle version final RV.pdf.txtExtracted texttext/plain165800https://repositorio.anid.cl/bitstreams/df1728b7-0e91-4e85-b5cf-d641dc7f36e8/downloadeca033c085b7fa4dbd5a7b70b347fc87MD53THUMBNAILTesis de doctorado Roger Valle version final RV.pdf.jpgIM Thumbnailimage/jpeg2149https://repositorio.anid.cl/bitstreams/85de9db5-d4d3-4f9e-a247-035945237e84/downloadeffdc5e90ae9bf35e7d28cb6f9a0d326MD5410533/236575oai:repositorio.anid.cl:10533/2365752023-07-24 03:48:45.113https://repositorio.anid.clRepositorio ANIDaletelier@anid.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