RESUMEN

Binge Drinking (BD) corresponds to episodes of ingestion of large amounts of ethanol in a short time, typically ≤2 h. BD occurs across all populations, but young and sports-related people are especially vulnerable. However, the short- and long-term effects of episodic BD on skeletal muscle function have been poorly explored. Young rats were randomized into two groups: control and episodic Binge-Like ethanol protocol (BEP) (ethanol 3 g/kg IP, 4 episodes of 2-days ON-2-days OFF paradigm). Muscle function was evaluated two weeks after the last BEP episode. We found that rats exposed to BEP presented decreased muscle strength and increased fatigability, compared with control animals. Furthermore, we observed that skeletal muscle from rats exposed to BEP presented muscle atrophy, evidenced by reduced fiber size and increased expression of atrophic genes. We also observed that BEP induced fibrotic and inflammation markers, accompanied by mislocalization of nNOSµ and high levels of protein nitration. Our findings suggest that episodic binge-like ethanol exposure alters contractile capacity and increases fatigue by mechanisms involving atrophy, fibrosis, and inflammation, which remain for at least two weeks after ethanol clearance. These pathological features are common to several neuromuscular diseases and might affect muscle performance and health in the long term.

Asunto(s)

Consumo Excesivo de Bebidas Alcohólicas , Etanol , Ratas , Animales , Etanol/efectos adversos , Etanol/metabolismo , Músculo Esquelético/metabolismo , Inflamación/metabolismo , Atrofia Muscular/metabolismo , Fuerza Muscular , Fibrosis , Consumo Excesivo de Bebidas Alcohólicas/metabolismo

RESUMEN

Tibetan pigs are characterized by significant phenotypic differences relative to lowland pigs. Our previous study demonstrated that the genes CRYAB and CTGF were differentially expressed in heart tissues between Tibetan (highland breed) and Yorkshire (lowland breed) pigs, indicating that they might participate in hypoxia adaptation. CRYAB (ɑB-crystallin) and CTGF (connective tissue growth factor) have also been reported to be associated with lung development. However, the expression patterns of CRYAB and CTGF in lung tissues at different altitudes and their genetic characterization are not well understood. In this study, qRT-PCR and western blot of lung tissue revealed higher CRYAB expression levels in highland and middle-highland Tibetan and Yorkshire pigs than in their lowland counterparts. With an increase in altitude, the expression level of CTGF increased in Tibetan pigs, whereas it decreased in Yorkshire pigs. Furthermore, two novel single-nucleotide polymorphism were identified in the 5' flanking region of CRYAB (g.39644482C>T and g.39644132T>C) and CTGF (g.31671748A>G and g.31671773T>G). The polymorphism may partially contribute to the differences in expression levels between groups at the same altitude. These findings provide novel insights into the high-altitude hypoxia adaptations of Tibetan pigs.

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Porcos tibetanos são caracterizados por diferenças fenotípicas significativas em relação aos porcos de planície. Nosso estudo anterior demonstrou que os genes CRYAB e CTGF eram expressos diferentemente nos tecidos do coração entre os porcos tibetanos (raça das terras altas) e Yorkshire (raça das terras baixas), indicando que eles poderiam participar da adaptação à hipoxia. CRYAB (ɑB-crystallin) e CTGF (fator de crescimento do tecido conjuntivo) também foram relatados como estando associados ao desenvolvimento pulmonar. Entretanto, os padrões de expressão do CRYAB e CTGF nos tecidos pulmonares em diferentes altitudes e sua caracterização genética não são bem compreendidos. Neste estudo, o qRT-PCR e a mancha ocidental de tecido pulmonar revelou níveis de expressão de CRYAB mais elevados em porcos tibetanos e Yorkshire de altitude e média altitude do que em seus pares de planície. Com um aumento na altitude, o nível de expressão do CTGF aumentou nos porcos tibetanos, enquanto diminuiu nos porcos Yorkshire. Além disso, foram identificados dois novos polimorfismos de um único nucleotídeo na região flanqueadora de CRYAB (g.39644482C>T e g.39644132T>C) e CTGF (g.31671748A>G e g.31671773T>G). O polimorfismo pode contribuir parcialmente com as diferenças nos níveis de expressão entre grupos a uma mesma altitude. Estas descobertas proporcionam novos conhecimentos sobre as adaptações de hipoxia a alta altitude dos porcos tibetanos.

Asunto(s)

Animales , Polimorfismo Genético , Adaptación Biológica/genética , Expresión Génica , Sus scrofa , Mal de Altura/veterinaria , Hipoxia/veterinaria , Tibet

RESUMEN

ABSTRACT Background: Coronavirus disease-2019 (COVID-19) results in acute lung injury. This study examined the usefulness of serum transforming growth factor-beta 1 (TGF-β1) and connective tissue growth factor (CTGF) levels in predicting disease severity in COVID-19 patients with pulmonary involvement. Methods: Fifty patients with confirmed COVID-19 and pulmonary involvement between September 2020, and February 2021 (Group 1) and 45 healthy controls (Group 2) were classified into three subgroups based on clinical severity: moderate, severe, and critical pneumonia. Serum TGF-β1 and CTGF concentrations were measured on days 1 and 7 of admission in Group 1 using an enzyme-linked immunosorbent assay. These concentrations were also measured in control cases. The mean serum TGF-β1 and CTGF levels were then compared among COVID-19 patients, based on clinical severity. Results: Significantly higher mean serum TGF-β1 and CTGF levels were observed on both days in Group 1 than in the control group. The mean serum TGF-β1 and CTGF levels on day 7 were also significantly higher than those on day 1 in Group 1. The critical patient group had the highest serum TGF-β1 and CTGF levels on both days, and the difference between this group and the moderate and severe pneumonia groups was significant. Cutoff values of 5.36 ng/mL for TGF-β1 and 626.2 pg/mL for CTGF emerged as predictors of COVID-19 with pulmonary involvement in receiver-operating characteristic curve analysis. Conclusions: TGF-β1 and CTGF are potential markers that can distinguish COVID-19 patients with pulmonary involvement and indicate disease severity. These findings may be useful for initiating treatment for early-stage COVID-19.

RESUMEN

Connective tissue growth factor or cellular communication network 2 (CCN2/CTGF) is a matricellular protein member of the CCN family involved in several crucial biological processes. In skeletal muscle, CCN2/CTGF abundance is elevated in human muscle biopsies and/or animal models for diverse neuromuscular pathologies, including muscular dystrophies, neurodegenerative disorders, muscle denervation, and muscle overuse. In this context, CCN2/CTGF is deeply involved in extracellular matrix (ECM) modulation, acting as a strong pro-fibrotic factor that promotes excessive ECM accumulation. Reducing CCN2/CTGF levels or biological activity in pathological conditions can decrease fibrosis, improve muscle architecture and function. In this work, we summarize information about the role of CCN2/CTGF in fibrosis associated with neuromuscular pathologies and the mechanisms and signaling pathways that regulate their expression in skeletal muscle.

RESUMEN

The Cellular Communication Network (CCN) family of matricellular proteins comprises six proteins that share conserved structural features and play numerous biological roles. These proteins can interact with several receptors or soluble proteins, regulating cell signaling pathways in various tissues under physiological and pathological conditions. In the skeletal muscle of mammals, most of the six CCN family members are expressed during embryonic development or in adulthood. Their roles during the adult stage are related to the regulation of muscle mass and regeneration, maintaining vascularization, and the modulation of skeletal muscle fibrosis. This work reviews the CCNs proteins' role in skeletal muscle physiology and disease, focusing on skeletal muscle fibrosis and its regulation by Connective Tissue Growth factor (CCN2/CTGF). Furthermore, we review evidence on the modulation of fibrosis and CCN2/CTGF by the renin-angiotensin system and the kallikrein-kinin system of vasoactive peptides.

Asunto(s)

Factor de Crecimiento del Tejido Conjuntivo/metabolismo , Músculo Esquelético/fisiología , Péptidos/metabolismo , Animales , Regulación del Desarrollo de la Expresión Génica , Humanos , Cininas/metabolismo , Familia de Multigenes , Proteínas Musculares/metabolismo , Músculo Esquelético/irrigación sanguínea , Músculo Esquelético/patología , Atrofia Muscular/metabolismo , Atrofia Muscular/patología , Enfermedades Musculares/metabolismo , Enfermedades Musculares/patología , Regeneración , Sistema Renina-Angiotensina

RESUMEN

During ischemia/reperfusion (I/R), cardiomyocytes activate pathways that regulate cell survival and death and release factors that modulate fibroblast-to-myofibroblast differentiation. The mechanisms underlying these effects are not fully understood. Polycystin-1 (PC1) is a mechanosensor crucial for cardiac function. This work aims to assess the role of PC1 in cardiomyocyte survival, its role in profibrotic factor expression in cardiomyocytes, and its paracrine effects on I/R-induced cardiac fibroblast function. In vivo and ex vivo I/R and simulated in vitro I/R (sI/R) were induced in wild-type and PC1-knockout (PC1 KO) mice and PC1-knockdown (siPC1) neonatal rat ventricular myocytes (NRVM), respectively. Neonatal rat cardiac fibroblasts (NRCF) were stimulated with conditioned medium (CM) derived from NRVM or siPC1-NRVM supernatant after reperfusion and fibroblast-to-myofibroblast differentiation evaluated. Infarcts were larger in PC1-KO mice subjected to in vivo and ex vivo I/R, and necrosis rates were higher in siPC1-NRVM than control after sI/R. PC1 activated the pro-survival AKT protein during sI/R and induced PC1-AKT-pathway-dependent CTGF expression. Furthermore, conditioned media from sI/R-NRVM induced PC1-dependent fibroblast-to-myofibroblast differentiation in NRCF. This novel evidence shows that PC1 mitigates cardiac damage during I/R, likely through AKT activation, and regulates CTGF expression in cardiomyocytes via AKT. Moreover, PC1-NRVM regulates fibroblast-to-myofibroblast differentiation during sI/R. PC1, therefore, may emerge as a new key regulator of I/R injury-induced cardiac remodeling.

Asunto(s)

Factor de Crecimiento del Tejido Conjuntivo/biosíntesis , Regulación de la Expresión Génica , Daño por Reperfusión Miocárdica/metabolismo , Miocitos Cardíacos/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Canales Catiónicos TRPP/metabolismo , Animales , Factor de Crecimiento del Tejido Conjuntivo/genética , Masculino , Ratones , Ratones Noqueados , Daño por Reperfusión Miocárdica/genética , Daño por Reperfusión Miocárdica/patología , Miocitos Cardíacos/patología , Proteínas Proto-Oncogénicas c-akt/genética , Ratas , Ratas Sprague-Dawley , Canales Catiónicos TRPP/genética

RESUMEN

Tendon injuries are common and have a high incidence of re-rupture that can cause loss of functionality. Therapies with adipose-derived stem cells (ASC) and the microcurrent (low-intensity electrical stimulation) application present promising effects on the tissue repair. We analyzed the expression of genes and the participation of some molecules potentially involved in the structural recovery of the Achilles tendon of rats, in response to the application of both therapies, isolated and combined. The tendons were distributed in five groups: normal (N), transected (T), transected and ASC (C) or microcurrent (M) or with ASC, and microcurrent (MC). Microcurrent therapy was beneficial for tendon repair, as it was observed a statistically significant increase in the organization of the collagen fibers, with involvement of the TNC, CTGF, FN, FMDO, and COL3A1 genes as well as PCNA, IL-10, and TNF-α. ASC therapy significantly increased the TNC and FMDO genes expression with no changes in the molecular organization of collagen. With the association of therapies, a significant greater collagen fibers organization was observed with involvement of the FMOD gene. The therapies did not affect the expression of COL1A1, SMAD2, SMAD3, MKX, and EGR1 genes, nor did they influence the amount of collagen I and III, caspase-3, tenomodulin (Tnmd), and hydroxyproline. In conclusion, the application of the microcurrent isolated or associated with ASC increased the organization of the collagen fibers, which can result in a greater biomechanical resistance in relation to the tendons treated only with ASC. Future studies will be needed to demonstrate the biological effects of these therapies on the functional recovery of injured tendons.

Asunto(s)

Biomarcadores/análisis , Estimulación Eléctrica/métodos , Regulación de la Expresión Génica , Células Madre Mesenquimatosas/citología , Trasplante de Células Madre/métodos , Traumatismos de los Tendones/terapia , Cicatrización de Heridas , Animales , Diferenciación Celular , Movimiento Celular , Perfilación de la Expresión Génica , Masculino , Células Madre Mesenquimatosas/metabolismo , Ratas , Ratas Wistar , Regeneración , Traumatismos de los Tendones/genética , Traumatismos de los Tendones/metabolismo , Traumatismos de los Tendones/patología

RESUMEN

The objective of this study was to evaluate the effects of red Light Emiting Diode (red LED) irradiation on fibroblasts in adipose-derived mesenchymal stem cells (ASC) co-culture on the scratch assay. We hypothesized that red LED irradiation could stimulate paracrine secretion of ASC, contributing to the activation of genes and molecules involved in cell migration and tissue repair. ASC were co-cultured with NIH/3T3 fibroblasts through direct contact and subjected to red LED irradiation (1.45 J/cm2/5min6s) after the scratch assay, during 4 days. Four groups were established: fibroblasts (F), fibroblasts + LED (FL), fibroblasts + ASC (FC) and fibroblasts + LED + ASC (FLC). The analyzes were based on Ctgf and Reck expression, quantification of collagen types I and III, tenomodulin, VEGF, TGF-ß1, MMP-2 and MMP-9, as well as viability analysis and cell migration. Higher Ctgf expression was observed in FC compared to F. Group FC presented higher amount of tenomodulin and VEGF in relation to the other groups. In the cell migration analysis, a higher number of cells was observed in the scratched area of the FC group on the 4th day. There were no differences between groups considering cell viability, Reck expression, amount of collagen types I and III, MMP-2 and TGF-ß1, whereas TGF-ß1 was not detected in the FC group and the MMP-9 in none of the groups. Our hypothesis was not supported by the results because the red LED irradiation decreased the healing response of ASC. An inhibitory effect of the LED irradiation associated with ASC co-culture was observed with reduction of the amount of TGF-ß1, VEGF and tenomodulin, possibly involved in the reduced cell migration. In turn, the ASC alone seem to have modulated fibroblast behavior by increasing Ctgf, VEGF and tenomodulin, leading to greater cell migration. In conclusion, red LED and ASC therapy can have independent effects on fibroblast wound healing, but the combination of both does not have a synergistic effect. Therefore, future studies with other parameters of red LED associated with ASC should be tested aiming clinical application for tissue repair.

RESUMEN

Hypoxia refers to the decrease in oxygen tension in the tissues, and the central effector of the hypoxic response is the transcription factor Hypoxia-Inducible Factor α (HIF1-α). Transient hypoxia in acute events, such as exercising or regeneration after damage, play an important role in skeletal muscle physiology and homeostasis. However, sustained activation of hypoxic signaling is a feature of skeletal muscle injury and disease, which can be a consequence of chronic damage but can also increase the severity of the pathology and worsen its outcome. Here, we review evidence that supports the idea that hypoxia and HIF-1α can contribute to the establishment of fibrosis in skeletal muscle through its crosstalk with other profibrotic factors, such as Transforming growth factor ß (TGF-ß), the induction of profibrotic cytokines expression, as is the case of Connective Tissue Growth Factor (CTGF/CCN2), or being the target of the Renin-angiotensin system (RAS).

RESUMEN

Several skeletal muscle diseases are characterized by fibrosis, the excessive accumulation of extracellular matrix. Transforming growth factor-ß (TGF-ß) and connective tissue growth factor (CCN2/CTGF) are two profibrotic factors augmented in fibrotic skeletal muscle, together with signs of reduced vasculature that implies a decrease in oxygen supply. We observed that fibrotic muscles are characterized by the presence of positive nuclei for hypoxia-inducible factor-1α (HIF-1α), a key mediator of the hypoxia response. However, it is not clear how a hypoxic environment could contribute to the fibrotic phenotype in skeletal muscle. We evaluated the role of hypoxia and TGF-ß on CCN2 expression in vitro. Fibroblasts, myoblasts and differentiated myotubes were incubated with TGF-ß1 under hypoxic conditions. Hypoxia and TGF-ß1 induced CCN2 expression synergistically in myotubes but not in fibroblasts or undifferentiated muscle progenitors. This induction requires HIF-1α and the Smad-independent TGF-ß signaling pathway. We performed in vivo experiments using pharmacological stabilization of HIF-1α or hypoxia-induced via hindlimb ischemia together with intramuscular injections of TGF-ß1, and we found increased CCN2 expression. These observations suggest that hypoxic signaling together with TGF-ß signaling, which are both characteristics of a fibrotic skeletal muscle environment, induce the expression of CCN2 in skeletal muscle fibers and myotubes.

Asunto(s)

Factor de Crecimiento del Tejido Conjuntivo/metabolismo , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Músculo Esquelético/patología , Factor de Crecimiento Transformador beta1/administración & dosificación , Regulación hacia Arriba , Animales , Diferenciación Celular , Hipoxia de la Célula , Línea Celular , Modelos Animales de Enfermedad , Fibroblastos/citología , Fibroblastos/efectos de los fármacos , Fibroblastos/metabolismo , Fibrosis , Inyecciones Intramusculares , Isquemia/etiología , Ratones , Fibras Musculares Esqueléticas/citología , Fibras Musculares Esqueléticas/efectos de los fármacos , Fibras Musculares Esqueléticas/metabolismo , Músculo Esquelético/citología , Músculo Esquelético/metabolismo , Mioblastos/citología , Mioblastos/efectos de los fármacos , Mioblastos/metabolismo , Células 3T3 NIH , Transducción de Señal , Factor de Crecimiento Transformador beta1/farmacología

RESUMEN

The etiology of polycystic ovary syndrome (PCOS) is complex and the pathogenesis is not fully understood. Some studies have shown that dysregulation of ovarian granulosa cells may be related to abnormal follicles and excessive androgen in women with PCOS. Our team has also confirmed the high expression status of H19 in PCOS patients in the early stage. However, the relationship between H19 and miR-19b in the development of PCOS is still unknown. Therefore, we used bioinformatics to predict the binding sites of human H19 and miR-19b, and of miR-19b and CTGF genes. After the silencing and overexpression of H19, real-time polymerase chain reaction (PCR) was used to detect the expressions of H19, miR-19b, and CTGF. Western blotting was used to detect CTGF protein. Proliferation of KGN cells after H19 silencing was detected by CCK8. Flow cytometry was used to detect the apoptosis of KGN cells after H19 silencing. After the overexpression of H19, it was found that the expression of miR-19b gene decreased and the expression of CTGF increased, whereas silencing of H19 did the opposite. In addition, H19 could promote cell proliferation and decrease cell apoptosis. Finally, luciferase reporter assays showed that the 3′-end sequences of lncRNA H19 and CTGF contained the binding site of miR-19b. In conclusion, our study indicated that lncRNA H19 acted as a ceRNA to bind to miR-19b via a "sponge" to regulate the effect of CTGF on KGN cells, which may play a vital role in PCOS.

Asunto(s)

Humanos , Femenino , Síndrome del Ovario Poliquístico/genética , Apoptosis , MicroARNs/genética , Proliferación Celular , Factor de Crecimiento del Tejido Conjuntivo , ARN Largo no Codificante/genética

RESUMEN

Connective tissue growth factor (CTGF/CCN2) is a matricellular protein that belongs to the CCN family of proteins. Since its discovery, it has been linked to cellular processes such as cell proliferation, differentiation, adhesion, migration, and synthesis of extracellular matrix (ECM) components, among others. The pro-fibrotic role of CTGF/CCN2 has been well-studied in several pathologies characterized by the development of fibrosis. Reduction of CTGF/CCN2 levels in mdx mice, a murine model for Duchenne muscular dystrophy (DMD), decreases fibrosis and improves skeletal muscle phenotype and function. Recently, it has been shown that skeletal muscle of symptomatic hSOD1G93A mice, a model for Amyotrophic lateral sclerosis (ALS), shows up-regulation of CTGF/CCN2 accompanied by excessive deposition ECM molecules. Elevated levels of CTGF/CCN2 in spinal cord from ALS patients have been previously reported. However, there is no evidence regarding the role of CTGF/CCN2 in neurodegenerative diseases such as ALS, in which alterations in skeletal muscle seem to be the consequence of early pathological denervation. In this regard, the emerging evidence shows that CTGF/CCN2 also exerts non-fibrotic roles in the central nervous system (CNS), specifically impairing oligodendrocyte maturation and regeneration, and inhibiting axon myelination. Despite these striking observations, there is no evidence showing the role of CTGF/CCN2 in peripheral nerves. Therefore, even though more studies are needed to elucidate its precise role, CTGF/CCN2 is starting to emerge as a novel therapeutic target for the treatment of neurodegenerative diseases where demyelination and axonal degeneration occurs.

Asunto(s)

Factor de Crecimiento del Tejido Conjuntivo/metabolismo , Músculo Esquelético/metabolismo , Sistema Nervioso/metabolismo , Enfermedades Neurodegenerativas/metabolismo , Animales , Humanos , Modelos Biológicos

RESUMEN

Fibrosis is a common feature of several chronic diseases and is characterized by exacerbated accumulation of ECM. An understanding of the cellular and molecular mechanisms involved in the development of this condition is crucial for designing efficient treatments for those pathologies. Connective tissue growth factor (CTGF/CCN2) is a pleiotropic protein with strong profibrotic activity. In this report, we present experimental evidence showing that ECM stimulates the synthesis of CTGF in response to lysophosphatidic acid (LPA).The integrin/focal adhesion kinase (FAK) signaling pathway mediates this effect, since CTGF expression is abolished by the use of the Arg-Gly-Asp-Ser peptide and also by an inhibitor of FAK autophosphorylation at tyrosine 397. Cilengitide, a specific inhibitor of αv integrins, inhibits the expression of CTGF mediated by LPA or transforming growth factor ß1. We show that ECM obtained from decellularized myofibroblast cultures or derived from activated fibroblasts from muscles of the Duchenne muscular dystrophy mouse model ( mdx) induces the expression of CTGF. This effect is dependent on FAK phosphorylation in response to its activation by integrin. We also found that the fibrotic ECM inhibits skeletal muscle differentiation. This novel regulatory mechanism of CTGF expression could be acting as a positive profibrotic feedback between the ECM and CTGF, revealing a novel concept in the control of fibrosis under chronic damage.

Asunto(s)

Diferenciación Celular/efectos de los fármacos , Factor de Crecimiento del Tejido Conjuntivo/metabolismo , Matriz Extracelular/efectos de los fármacos , Fibroblastos/efectos de los fármacos , Quinasa 1 de Adhesión Focal/metabolismo , Integrina alfaV/metabolismo , Lisofosfolípidos/farmacología , Músculo Esquelético/efectos de los fármacos , Distrofia Muscular de Duchenne/enzimología , Mioblastos/efectos de los fármacos , Animales , Línea Celular , Factor de Crecimiento del Tejido Conjuntivo/genética , Modelos Animales de Enfermedad , Matriz Extracelular/enzimología , Matriz Extracelular/patología , Fibroblastos/enzimología , Fibroblastos/patología , Fibrosis , Integrina alfaV/genética , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos mdx , Músculo Esquelético/enzimología , Músculo Esquelético/patología , Distrofia Muscular de Duchenne/genética , Distrofia Muscular de Duchenne/patología , Mioblastos/enzimología , Mioblastos/patología , Fosforilación , Transducción de Señal/efectos de los fármacos

RESUMEN

Connective tissue growth factor (CTGF/CCN2) has strong inflammatory and profibrotic activities. Its expression is enhanced in skeletal muscular dystrophies such as Duchenne muscular dystrophy (DMD), a myopathy characterized by exacerbated inflammation and fibrosis. In dystrophic tissue, necrotic-regenerative foci, myofibroblasts, newly-regenerated muscle fibers and necrosis all occur simultaneously. To determine if CCN2 is involved in the appearance of the foci, we studied their presence and characteristics in mdx mice (DMD mouse model) compared to mdx mice hemizygous for CCN2 (mdx-Ccn2+/-). We used laser capture microdissection followed by gene expression and immunofluorescence analyses to investigate fibrotic, inflammation and regeneration markers in damaged and non-damaged areas in mdx and mdx-Ccn2+/- skeletal muscle. Mdx mice foci express elevated mRNAs levels of transforming growth factor type beta, collagen, fibronectin, the myofribroblast marker α-SMA, and the myogenic transcription factor myogenin. Mdx foci also show elevated levels of MCP-1 and CD-68 positive cells, indicating that CCN2 could be inducing an inflammatory response. We found a significant reduction in the number of foci in mdx-Ccn2+/- mice muscle. Fibrotic and inflammatory markers were also decreased in these foci. We did not observe any difference in Pax7 mRNA levels, a marker for satellite cells, in mdx mice compared to mdx-Ccn2+/- mice. Thus, CCN2 appears to be involved in the fibrotic response as well as in the inflammatory response in the dystrophic skeletal muscle.

RESUMEN

Investigations demonstrated that oxidative stress plays an important role in injury promotion in cholestatic liver disease. We hypothesized that coffee attenuates cholestasis-induced hepatic necrosis and fibrosis via its antioxidant, anti-inflammatory, and antifibrotic properties. The major aim of this study was to evaluate the hepatoprotective properties of coffee and caffeine in a model of chronic bile duct ligation (BDL) in male Wistar rats. Liver injury was induced by 28-day BDL, and conventional coffee, decaffeinated coffee, or caffeine was administered daily. After treatment, the hepatic oxidative status was estimated by measuring lipid peroxidation, the reduced to oxidized glutathione ratio, and glutathione peroxidase. Fibrosis was assessed by measuring the liver hydroxyproline content. The transforming growth factor-ß, connective tissue growth factor, α-smooth muscle actin, collagen 1, and interleukin-10 proteins and mRNAs were measured by Western blot and polymerase chain reaction, respectively. Conventional coffee suppressed most of the changes produced by BDL; however, caffeine showed better antifibrotic effects. Coffee demonstrated antioxidant properties by restoring the redox equilibrium, and it also prevented the elevation of liver enzymes as well as hepatic glycogen depletion. Interestingly, coffee and caffeine administration prevented collagen increases. Western blot assays showed decreased expression levels of transforming growth factor-ß, connective tissue growth factor, α-smooth muscle actin, and collagen 1 in the coffee- and caffeine-treated BDL groups. Similarly, coffee decreased the mRNA levels of these proteins. We conclude that coffee prevents liver cirrhosis induced by BDL by attenuating the oxidant processes, blocking hepatic stellate cell activation, and downregulating the main profibrotic molecules involved in extracellular matrix deposition.

Asunto(s)

Cafeína/farmacología , Café/química , Cirrosis Hepática Biliar/prevención & control , Actinas/metabolismo , Animales , Antioxidantes/farmacología , Conductos Biliares/metabolismo , Factor de Crecimiento del Tejido Conjuntivo/metabolismo , Modelos Animales de Enfermedad , Fibrosis , Glutatión Peroxidasa/metabolismo , Células Estrelladas Hepáticas/metabolismo , Hidroxiprolina/metabolismo , Peroxidación de Lípido/fisiología , Hígado/metabolismo , Hígado/patología , Masculino , Estrés Oxidativo , Ratas , Ratas Wistar , Especies Reactivas de Oxígeno/metabolismo , Factor de Crecimiento Transformador beta/metabolismo

RESUMEN

CCN2 or connective tissue growth factor (CTGF) is a matricellular protein that regulates several cellular processes. In skeletal muscle, CTGF is a key modulator of fibrogenesis, is increased in pathological conditions such as muscular dystrophies, and plays a major role in the pathology outcome. Overexpression of CTGF in skeletal muscle of wild-type mice results in muscle damage, fibrosis, and reduction of strength. In contrast, a decrease in CTGF in dystrophic mice increases strength and reduces damage and fibrosis. Thus, CTGF is a relevant target to study in skeletal muscle pathology and its possible modulation by different treatments or potential new drugs to develop new strategies for the treatment of muscular dystrophies. We summarize the techniques used to detect CTGF in the skeletal muscle of dystrophic mdx mice.

Asunto(s)

Factor de Crecimiento del Tejido Conjuntivo/genética , Factor de Crecimiento del Tejido Conjuntivo/metabolismo , Distrofias Musculares/genética , Distrofias Musculares/metabolismo , Animales , Modelos Animales de Enfermedad , Inmunohistoquímica , Ratones , Ratones Endogámicos mdx , Músculo Esquelético/metabolismo , Músculo Esquelético/patología , Distrofias Musculares/patología , Reacción en Cadena en Tiempo Real de la Polimerasa

RESUMEN

Fibrotic disorders are characterized by an increase in extracellular matrix protein expression and deposition, Duchene Muscular Dystrophy being one of them. Among the factors that induce fibrosis are Transforming Growth Factor type ß (TGF-ß) and the matricellular protein Connective Tissue Growth Factor (CTGF/CCN2), the latter being a target of the TGF-ß/SMAD signaling pathway and is the responsible for the profibrotic effects of TGF-ß. Both CTGF and TGF are increased in tissues affected by fibrosis but little is known about the regulation of the expression of CTGF mediated by TGF-ß in muscle cells. By using luciferase reporter assays, site directed mutagenesis and specific inhibitors in C2C12 cells; we described a novel SMAD Binding Element (SBE) located in the 5' UTR region of the CTGF gene important for the TGF-ß-mediated expression of CTGF in myoblasts. In addition, our results suggest that additional transcription factor binding sites (TFBS) present in the 5' UTR of the CTGF gene are important for this expression and that SP1/SP3 factors are involved in TGF-ß-mediated CTGF expression.

Asunto(s)

Factor de Crecimiento del Tejido Conjuntivo/genética , Factor de Crecimiento del Tejido Conjuntivo/metabolismo , Mioblastos/efectos de los fármacos , Factor de Transcripción Sp1/metabolismo , Factor de Crecimiento Transformador beta/farmacología , Regiones no Traducidas 5' , Animales , Sitios de Unión , Línea Celular , Factor de Crecimiento del Tejido Conjuntivo/química , Regulación de la Expresión Génica , Ratones , Mutagénesis Sitio-Dirigida , Mioblastos/metabolismo , Mioblastos/fisiología , Transducción de Señal/efectos de los fármacos , Proteína smad3/metabolismo , Factor de Transcripción Sp3/metabolismo

RESUMEN

Connective tissue growth factor (CTGF)/CCN family member 2 (CCN2) is a CCN family member of matricellular signaling modulators. It has been shown that CCN2/CTGF mediates cell adhesion, aggregation and migration in a large variety of cell types, including vascular endothelial cells, fibroblasts, epithelial cells, aortic smooth muscle and also pluripotent stem cells. Others matricellular proteins are capable of interacting with CCN2/CTGF to mediate its function. Cell migration is a key feature for tumor cell invasion and metastasis. CCN2/CTGF seems to be a prognostic marker for cancer. In addition, here we intend to discuss recent discoveries and a new strategy to develop therapies against CCN2/CTGF, in order to treat cancer metastasis.

RESUMEN

Myogenesis consists of a highly organized and regulated sequence of cellular processes aimed at forming or repairing muscle tissue. Several processes occur during myogenesis, including cell proliferation, migration, and differentiation. Cytokines, proteinases, cell adhesion molecules and growth factors are involved, either activating or inhibiting these events, and are modulated by a group of molecules called proteoglycans (PGs), which play critical roles in skeletal muscle physiology. Particularly interesting are some of the factors responsible for the fibrotic response associated with skeletal muscular dystrophies. Transforming growth factor-ß and connective tissue growth factor have gained great attention as factors participating in the fibrotic response in skeletal muscle. This review is focused on the advances achieved in understanding the roles of proteoglycans as modulators of profibrotic growth factors in fibrosis associated with diseases such as skeletal muscle dystrophies.

Asunto(s)

Factor de Crecimiento del Tejido Conjuntivo/metabolismo , Fibrosis/etiología , Desarrollo de Músculos/fisiología , Fibras Musculares Esqueléticas/citología , Proteoglicanos/metabolismo , Factor de Crecimiento Transformador beta/metabolismo , Animales , Fibrosis/metabolismo , Humanos , Fibras Musculares Esqueléticas/fisiología

RESUMEN

Connective tissue growth factor (CCN2/CTGF) is a matricellular-secreted protein involved in extracellular matrix remodeling. The P19 cell line is an embryonic carcinoma line widely used as a cellular model for differentiation and migration studies. In the present study, we employed an exogenous source of CCN2 and small interference RNA to address the role of CCN2 in the P19 cell aggregation phenomenon. Our data showed that increasing CCN2 protein concentrations from 0.1 to 20 nM decreased the number of cell clusters and dramatically increased cluster size without changing proliferation or cell survival, suggesting that CCN2 induced aggregation. In addition, CCN2 specific silencing inhibited typical P19 cell aggregation, which could be partially rescued by 20 nM CCN2. The present study demonstrates that CCN2 is a key molecule for cell aggregation of embryonic P19 cells.

Asunto(s)

Humanos , Agregación Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Factor de Crecimiento del Tejido Conjuntivo/farmacología , Células Madre de Carcinoma Embrionario/efectos de los fármacos , Adhesión Celular , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/fisiología