RESUMEN
Lactoferrin (LF) is known to possess anti-inflammatory activity, although its mechanisms of action are not well-understood. The present study asked whether LF affects the commitment of inducible regulatory T cells (Tregs). LF substantially promoted Foxp3 expression by mouse activated CD4+T cells, and this activity was further enhanced by TGF-ß1. Interestingly, blocking TGF-ß with anti-TGF-ß Ab completely abolished LF-induced Foxp3 expression. However, no significant amount of soluble TGF-ß was released by LF-stimulated T cells, suggesting that membrane TGF-ß (mTGF-ß) is associated. Subsequently, it was found that LF binds to TGF-ß receptor III, which induces reactive oxygen species production and diminishes the expression of mTGF-ß-bound latency-associated peptide, leading to the activation of mTGF-ß. It was followed by phosphorylation of Smad3 and enhanced Foxp3 expression. These results suggest that LF induces Foxp3+ Tregs through TGF-ß receptor III/reactive oxygen species-mediated mTGF-ß activation, triggering canonical Smad3-dependent signaling. Finally, we found that the suppressive activity of LF-induced Tregs is facilitated mainly by CD39/CD73-induced adenosine generation and that this suppressor activity alleviates inflammatory bowel disease.
Asunto(s)
Lactoferrina/metabolismo , Receptores de Factores de Crecimiento Transformadores beta/inmunología , Linfocitos T Reguladores/inmunología , Factor de Crecimiento Transformador beta/inmunología , Animales , Diferenciación Celular/efectos de los fármacos , Diferenciación Celular/inmunología , Colitis/inmunología , Colitis/metabolismo , Lactoferrina/farmacología , Activación de Linfocitos/efectos de los fármacos , Activación de Linfocitos/inmunología , Ratones Endogámicos BALB C , Receptores de Factores de Crecimiento Transformadores beta/efectos de los fármacos , Receptores de Factores de Crecimiento Transformadores beta/metabolismo , Transducción de Señal/efectos de los fármacos , Transducción de Señal/inmunología , Linfocitos T Reguladores/efectos de los fármacos , Linfocitos T Reguladores/metabolismo , Factor de Crecimiento Transformador beta/efectos de los fármacos , Factor de Crecimiento Transformador beta/metabolismoRESUMEN
BACKGROUND: Exposure to endocrine-disrupting chemicals, such as Bisphenol A (BPA) and Bisphenol S (BPS), is widespread and has negative implications on embryonic development. Preliminary evidence revealed that in women undergoing IVF treatment, urinary BPA levels were associated with low serum anti-Mullerian hormone, however a definitive relationship between the two has not yet been characterized. METHODS: This study aimed to evaluate BPA and BPS effects on in vitro oocyte maturation and early preimplantation embryo development through i) analysis of anti-Mullerian hormone (AMH) and anti-Mullerian hormone receptor II (AMHRII), ii) investigation of developmental parameters, such as cleavage, blastocyst rates and developmental arrest, iii) detection of apoptosis and iv) assessment of possible sex ratio skew. An in vitro bovine model was used as a translational model for human early embryonic development. We first assessed AMH and AMHRII levels after bisphenol exposure during oocyte maturation. Zygotes were also analyzed during cleavage and blastocysts stages. Techniques used include in vitro fertilization, quantitative polymerase chain reaction (qPCR), western blotting, TUNEL and immunofluorescence. RESULTS: Our findings show that BPA significantly decreased cleavage (p < 0.001), blastocyst (p < 0.005) and overall developmental rates as well as significantly increased embryonic arrest at the 2-4 cell stage (p < 0.05). Additionally, both BPA and BPS significantly increased DNA fragmentation in 2-4 cells, 8-16 cells and blastocyst embryos (p < 0.05). Furthermore, BPA and BPS alter AMH and AMHRII at the mRNA and protein level in both oocytes and blastocysts. BPA, but not BPS, also significantly skews sex ratios towards female blastocysts (p < 0.05). CONCLUSION: This study shows that BPA affects AMH and AMHRII expression during oocyte maturation and that BPS exerts its effects to a greater extent after fertilization and therefore may not be a safer alternative to BPA. Our data lay the foundation for future functional studies, such as receptor kinetics, downstream effectors, and promoter activation/inhibition to prove a functional relationship between bisphenols and the AMH signalling system.
Asunto(s)
Hormona Antimülleriana/metabolismo , Compuestos de Bencidrilo/toxicidad , Blastocisto/efectos de los fármacos , Desarrollo Embrionario/efectos de los fármacos , Disruptores Endocrinos/toxicidad , Oocitos/efectos de los fármacos , Fenoles/toxicidad , Sulfonas/toxicidad , Animales , Hormona Antimülleriana/genética , Apoptosis/efectos de los fármacos , Bovinos , Femenino , Técnicas de Maduración In Vitro de los Oocitos , ARN Mensajero/efectos de los fármacos , ARN Mensajero/metabolismo , Receptores de Péptidos/efectos de los fármacos , Receptores de Péptidos/genética , Receptores de Péptidos/metabolismo , Receptores de Factores de Crecimiento Transformadores beta/efectos de los fármacos , Receptores de Factores de Crecimiento Transformadores beta/genética , Receptores de Factores de Crecimiento Transformadores beta/metabolismo , Cigoto/efectos de los fármacosRESUMEN
Methamphetamine (MA) abuse is associated with the development of pulmonary arterial hypertension (PAH) and subsequent right ventricular failure. A recent clinical study demonstrated that female sex is a major risk factor for MA-induced PAH. The mechanisms associated with increased prevalence and severity of MA-induced PAH in females are still unclear. We hypothesized that MA may promote changes in gene expression in the right ventricle contributing to the development and/or worsening of PAH in females. Male and female C57BL/6 mice were treated with either MA or vehicle. Right and left ventricular systolic pressures (RVSP and LVSP, respectively) were assessed and tissue samples were collected for gene expression and histology. LVSP and RVSP were not affected by MA in either males or females. Right ventricular hypertrophy was significantly increased by MA in females but it was not affected by MA in males. In the female mice, MA-induced right ventricular hypertrophy was associated with increased expression of brain natriuretic peptide gene and members of the TGF-ß receptor signaling pathway such as TGF-ß receptor-1, smad3 and smad7. In male mice, there were no changes in right ventricular gene expression. Our results suggest that MA caused right ventricular hypertrophy in female mice, but not in males and that this was associated with an increase in hypertrophic genes. The right ventricular hypertrophy was not dependent on increased RVSP suggesting a direct effect of MA on the right ventricle. If this translates to PAH patients, it might explain the poor outcome observed in MA-associated female PAH patients.
Asunto(s)
Estimulantes del Sistema Nervioso Central/farmacología , Hipertrofia Ventricular Derecha/genética , Metanfetamina/farmacología , Trastornos Relacionados con Anfetaminas/genética , Animales , Presión Sanguínea/efectos de los fármacos , Femenino , Regulación de la Expresión Génica/efectos de los fármacos , Masculino , Ratones , Ratones Endogámicos C57BL , Hipertensión Arterial Pulmonar/genética , Receptores de Factores de Crecimiento Transformadores beta/efectos de los fármacos , Caracteres Sexuales , Transducción de Señal/efectos de los fármacos , Función Ventricular Izquierda/efectos de los fármacosRESUMEN
BACKGROUND: Epigenetic reprogramming using DNA demethylating drugs is a promising approach for cancer therapy, but its efficacy is highly dependent on the dosing regimen. Low-dose treatment for a prolonged period shows a remarkable therapeutic efficacy, despite its small demethylating effect. Here, we aimed to explore the mechanisms of how such low-dose treatment shows this remarkable efficacy by focusing on epigenetic reprograming at the single-cell level. METHODS: Expression profiles in HCT116 cells treated with decitabine (DAC) were analyzed by single-cell RNA-sequencing (scRNA-seq). Functional consequences and DNA demethylation at the single-cell level were analyzed using cloned HCT116 cells after DAC treatment. RESULTS: scRNA-seq revealed that DAC-treated cells had highly diverse expression profiles at the single-cell level, and tumor-suppressor genes, endogenous retroviruses, and interferon-stimulated genes were upregulated in random fractions of cells. DNA methylation analysis of cloned HCT116 cells revealed that, while only partial reduction of DNA methylation levels was observed in bulk cells, complete demethylation of specific cancer-related genes, such as cell cycle regulation, WNT pathway, p53 pathway, and TGF-ß pathway, was observed, depending upon clones. Functionally, a clone with complete demethylation of CDKN2A (p16) had a larger fraction of cells with tetraploid than parental cells, indicating induction of cellular senescence due to normalization of cell cycle regulation. CONCLUSIONS: Epigenetic reprogramming of specific cancer-related pathways at the single-cell level is likely to underlie the remarkable efficacy of low-dose DNA demethylating therapy.
Asunto(s)
Reprogramación Celular/genética , Metilación de ADN/efectos de los fármacos , Epigenómica/métodos , Células HCT116/efectos de los fármacos , Neoplasias/tratamiento farmacológico , Análisis de la Célula Individual/métodos , Puntos de Control del Ciclo Celular/efectos de los fármacos , Inhibidor p16 de la Quinasa Dependiente de Ciclina/efectos de los fármacos , Inhibidor p16 de la Quinasa Dependiente de Ciclina/genética , Desmetilación del ADN , Decitabina/farmacología , Relación Dosis-Respuesta a Droga , Inhibidores Enzimáticos/farmacología , Epigénesis Genética/efectos de los fármacos , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Genes p53/efectos de los fármacos , Células HCT116/metabolismo , Humanos , Neoplasias/genética , Receptores de Factores de Crecimiento Transformadores beta/efectos de los fármacos , Resultado del Tratamiento , Regulación hacia Arriba/efectos de los fármacos , Vía de Señalización Wnt/efectos de los fármacosRESUMEN
Anti-Müllerian hormone (Amh) is a peptide factor that is known to regulate sexual differentiation and gonadal function in mammals. Although Amh is also suggested to be associated with cognitive development and function in the postnatal brain, little is known about its expression or direct effects on neuronal activities in the hippocampus. Therefore, we assessed Amh and its receptor expression in the hippocampus of male and female mice using PCR, Western blot, and immunofluorescence staining. While Amh-specific receptor expression was comparable between males and females, mRNA and protein levels of Amh were higher in females than those of males. Electrophysiological recordings on acute hippocampal slices showed that exogenous Amh protein addition increased synaptic transmission and long-term synaptic plasticity at the Cornu Ammonis (CA) 3-CA1 synapses. Amh exposure also increased the excitatory postsynaptic potential at CA1 synapses. Our findings support direct and rapid actions of Amh as a paracrine and/or autocrine factor in regulating hippocampal neuronal activities. Data provide functional evidence of Amh-mediated postsynaptic modulation of synaptic transmission and Amh-regulated long-term synaptic plasticity in the hippocampus. These results suggest a potential role of Amh in learning and memory, and a possible cause of the sex differences in cognitive development and function.
Asunto(s)
Hormona Antimülleriana/farmacología , Potenciales Postsinápticos Excitadores/fisiología , Plasticidad Neuronal/fisiología , Receptores de Péptidos/metabolismo , Receptores de Factores de Crecimiento Transformadores beta/metabolismo , Transmisión Sináptica/fisiología , Animales , Potenciales Postsinápticos Excitadores/efectos de los fármacos , Femenino , Hipocampo/efectos de los fármacos , Hipocampo/fisiología , Aprendizaje/efectos de los fármacos , Aprendizaje/fisiología , Masculino , Memoria/efectos de los fármacos , Memoria/fisiología , Ratones , Plasticidad Neuronal/efectos de los fármacos , Neuronas/metabolismo , Neuronas/fisiología , Receptores de Péptidos/efectos de los fármacos , Receptores de Factores de Crecimiento Transformadores beta/efectos de los fármacos , Caracteres Sexuales , Transmisión Sináptica/efectos de los fármacosRESUMEN
A disintegrin and metalloproteinase (ADAM)12 is considered to promote cardiac dysfunction based on the finding that a small-molecule ADAM12 inhibitor, KB-R7785, ameliorated cardiac function in a transverse aortic constriction (TAC) model by inhibiting the proteolytic activation of heparin-binding-EGF signaling. However, this compound has poor selectivity for ADAM12, and the role of ADAM12 in cardiac dysfunction has not yet been investigated using genetic loss-of-function mice. We revealed that ADAM12 knockout mice showed significantly more advanced cardiac hypertrophy and higher mortality rates than wild-type mice 4 wk after TAC surgery. An ADAM12 deficiency resulted in significantly more expanded cardiac fibrosis accompanied by increased collagen-related gene expression in failing hearts. The results of a genome-wide transcriptional analysis suggested a strongly enhanced focal adhesion- and fibrosis-related signaling pathway in ADAM12 knockout hearts. The loss of ADAM12 increased the abundance of the integrinß1 subunit and transforming growth factor (TGF)-ß receptor types I and III, and this was followed by the phosphorylation of focal adhesion kinase, Akt, mammalian target of rapamycin, ERK, and Smad2/3 in the heart, which resulted in cardiac dysfunction. The present results revealed that the loss of ADAM12 enhanced focal adhesion and canonical TGF-ß signaling by regulating the abundance of the integrinß1 and TGF-ß receptors.NEW & NOTEWORTHY In contrast to a long-believed cardio-damaging role of a disintegrin and metalloproteinase (ADAM)12, cardiac hypertrophy was more severe, cardiac function was lower, and mortality was higher in ADAM12 knockout mice than in wild-type mice after transverse aortic constriction surgery. The loss of ADAM12 enhanced focal adhesion- and fibrosis-related signaling pathways in the heart, which may compromise cardiac function. These results provide insights for the development of novel therapeutics that target ADAM12 to treat heart failure.
Asunto(s)
Proteína ADAM12/genética , Cardiomegalia/prevención & control , Desintegrinas/uso terapéutico , Insuficiencia Cardíaca/prevención & control , Miocardio/patología , Proteína ADAM12/antagonistas & inhibidores , Proteína ADAM12/efectos de los fármacos , Animales , Presión Sanguínea , Fibrosis , Adhesiones Focales/efectos de los fármacos , Integrina beta1/genética , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Receptores de Factores de Crecimiento Transformadores beta/efectos de los fármacos , Receptores de Factores de Crecimiento Transformadores beta/genética , Transducción de Señal/efectos de los fármacosRESUMEN
Rapamycin, the macrolide immunosuppressant and active pharmaceutic in drug-eluting stents (DES), has a well-recognized antiproliferative action that involves inhibition of the mTOR pathway after binding to the cytosolic protein FKBP12. TGF receptor-type I (TGFRI) spontaneous activation is inhibited by the association with FKBP12. We hypothesized that rapamycin, in addition to inhibition of mTOR signaling, activates TGFRI independent of TGFß. Human umbilical vein endothelial cells (HUVECs) were treated with rapamycin (10 nmol/l) and/or TGFß RI kinase inhibitor (TGFRIi, 100 nmol/l) for 24 h. Rapamycin induced SMAD phosphorylation (SMAD1, SMAD2, and SMAD5) and PAI-1 up-regulation, which was specifically abrogated by SMAD2 knockdown. TGFRIi efficiently blocked phosphorylation of SMAD2, but not SMAD1/5. Interestingly, the inhibitor did not alter cell proliferation arrest induced by rapamycin. Active TGFß secretion was not affected by the treatment. Neutralizing TGFß experiments did not influence SMAD2 phosphorylation or PAI-1 expression indicating that activation of this pathway is independent of the ligand. In addition, rapamycin induction of endothelial-to-mesenchymal transition (EndMT) was potentiated by IL-1ß and efficiently blocked by TGFRIi. In vivo, the prothrombogenic effects of rapamycin and up-regulation of PAI-1 in murine carotid arteries were reduced by TGFRIi treatment. In conclusion, we provide evidence that rapamycin activates TGF receptor independent of its ligand TGFß, in concert with promotion of PAI-1 expression and changes in endothelial phenotype. These undesirable effects, the prothrombogenic state, and activation of EndMT are SMAD2-dependent and independent of the therapeutic rapamycin-induced cell proliferation arrest.
Asunto(s)
Células Endoteliales de la Vena Umbilical Humana/efectos de los fármacos , Receptores de Factores de Crecimiento Transformadores beta/efectos de los fármacos , Sirolimus/farmacología , Proteína Smad2/efectos de los fármacos , Proteínas Portadoras/efectos de los fármacos , Humanos , Fosforilación/efectos de los fármacos , Transducción de Señal/efectos de los fármacos , Serina-Treonina Quinasas TOR/efectos de los fármacos , Factor de Crecimiento Transformador beta/efectos de los fármacosRESUMEN
Altered transforming growth factor-beta (TGF-ß) signaling has been implicated in the pathogenesis of leukemia. Although TGF-ß type II receptor (TßRII) isoforms have been isolated from human leukemia cells, their expression patterns and functions of these variants are unclear. In this study, we determined that two TßRII isoforms (TßRII and TßRII-B) are abnormally expressed in leukemic cells, as compared to normal hematopoietic cells. TßRII-B, but not TßRII, was found to promote cell cycle arrest, apoptosis, and differentiation of leukemic cells. TßRII-B also enhanced TGF-ß1 binding and downstream signaling and reduced tumorigenicity in vivo. By contrast, TßRII blocked all-trans retinoic acid-induced differentiation through inhibition of TßRII-B. Overall survival was significantly lower in acute myeloid leukemia (AML) patients with high compared to low TßRII expression. Thus, whereas TßRII-B is a potent inducer of cell cycle arrest, apoptosis, and differentiation, higher TßRII expression correlates with poor clinical prognosis in AML.
Asunto(s)
Leucemia Mieloide Aguda/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Receptores de Factores de Crecimiento Transformadores beta/metabolismo , Animales , Antineoplásicos/farmacología , Apoptosis , Puntos de Control del Ciclo Celular , Diferenciación Celular , Proliferación Celular , Femenino , Regulación Leucémica de la Expresión Génica , Células HL-60 , Humanos , Células K562 , Estimación de Kaplan-Meier , Leucemia Mieloide Aguda/tratamiento farmacológico , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/patología , Masculino , Ratones Endogámicos BALB C , Ratones Desnudos , Fosforilación , Pronóstico , Isoformas de Proteínas , Proteínas Serina-Treonina Quinasas/efectos de los fármacos , Proteínas Serina-Treonina Quinasas/genética , Interferencia de ARN , Receptor Tipo II de Factor de Crecimiento Transformador beta , Receptores de Factores de Crecimiento Transformadores beta/efectos de los fármacos , Receptores de Factores de Crecimiento Transformadores beta/genética , Transducción de Señal , Proteínas Smad/metabolismo , Factores de Tiempo , Transfección , Factor de Crecimiento Transformador beta1/farmacología , Tretinoina/farmacología , Células U937RESUMEN
BACKGROUND/PURPOSE: In order to clarify the role of transforming growth factor beta 1 (TGF-ß1) in pulp repair/regeneration responses, we investigated the differential signaling pathways responsible for the effects of TGF-ß1 on collagen turnover, matrix metalloproteinase-3 (MMP-3), and tissue inhibitor of metalloproteinase-1 (TIMP-1) production in human dental pulp cells. METHODS: Pulp cells were exposed to TGF-ß1 with/without pretreatment and coincubation by 1,4-diamino-2,3-dicyano-1,4-bis(o-aminophenyl mercapto)butadiene (U0126; a mitogen-activated protein kinase kinase [MEK]/extracellular signal-regulated kinase [ERK] inhibitor) and 4-(5-benzol[1,3]dioxol-5-yl-4-pyrldin-2-yl-1H- imidazol-2-yl)-benzamide hydrate (SB431542; an activin receptor-like kinase-5/Smad signaling inhibitor). Sircol collagen assay was used to measure cellular collagen content. Culture medium procollagen I, TIMP-1, and MMP-3 levels were determined by enzyme-linked immunosorbent assay. RESULTS: TGF-ß1 increased the collagen content, procollagen I, and TIMP-1 production, but slightly decreased MMP-3 production of pulp cells. SB431542 and U0126 prevented the TGF-ß1-induced increase of collagen content and TIMP-1 production of dental pulp cells. CONCLUSION: These results indicate that TGF-ß1 may be involved in the healing/regeneration processes of dental pulp in response to injury by stimulation of collagen and TIMP-1 production. These events are associated with activin receptor-like kinase-5/Smad2/3 and MEK/ERK signaling.
Asunto(s)
Colágeno Tipo I/metabolismo , Colágeno/fisiología , Regeneración/fisiología , Inhibidor Tisular de Metaloproteinasa-1/metabolismo , Factor de Crecimiento Transformador beta1/fisiología , Benzamidas/farmacología , Butadienos/farmacología , Células Cultivadas , Pulpa Dental/citología , Dioxoles/farmacología , Humanos , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Sistema de Señalización de MAP Quinasas/fisiología , Nitrilos/farmacología , Proteínas Serina-Treonina Quinasas/efectos de los fármacos , Proteínas Serina-Treonina Quinasas/fisiología , Receptor Tipo I de Factor de Crecimiento Transformador beta , Receptores de Factores de Crecimiento Transformadores beta/efectos de los fármacos , Receptores de Factores de Crecimiento Transformadores beta/fisiología , Regeneración/efectos de los fármacos , Proteínas Smad/efectos de los fármacos , Proteínas Smad/fisiologíaRESUMEN
BACKGROUND: Hypertrophic scars (HPSs) are characterized by excessive fibrosis associated with aberrant function of fibroblasts. Currently no satisfactory treatment has been developed. OBJECTIVES: To investigate the effect of baicalein on HPSs and its underlying mechanisms. METHODS: Baicalein was administered intradermally (10 µmol L(-1) in 100 µL sterile saline plus 10% dimethylsulfoxide) to mechanical-load-induced scars in mice once a day for 14 or 28 days. Histological and immunohistochemical studies were performed to evaluate scar hypertrophy and the function of fibroblasts. Human HPS-derived fibroblasts (HSFs) were determined by immunofluorescence study, collagen gel contraction assay and wound-healing assay. Also, protein expression of the transforming growth factor (TGF)-ß signalling pathway was detected using Western blotting. Lastly, a molecular docking study and kinase binding assay were conducted in search of the potential interaction between baicalein and activin receptor-like kinase (ALK)5. RESULTS: Baicalein treatment significantly attenuated HPS formation and collagen deposition in a mechanical-load-induced mouse model. Baicalein also inhibited the proliferation and activation of fibroblasts in vitro and in vivo. Furthermore, baicalein impaired the contractile and migration ability of HSFs. Protein expression investigation revealed that baicalein had an inhibitory effect on TGF-ß/Smad2/3 signalling. Such bioactivity of baicalein may result from its selective binding to the ATP-binding pocket of ALK5, as suggested by the molecular docking study and kinase binding assay. CONCLUSIONS: Baicalein could serve as a promising agent for treatment of HPSs and a selective ALK5 inhibitor.
Asunto(s)
Antioxidantes/farmacología , Cicatriz Hipertrófica/tratamiento farmacológico , Fármacos Dermatológicos/farmacología , Flavanonas/farmacología , Proteína Smad2/antagonistas & inhibidores , Proteína smad3/antagonistas & inhibidores , Factor de Crecimiento Transformador beta1/antagonistas & inhibidores , Adolescente , Adulto , Animales , Niño , Cicatriz Hipertrófica/metabolismo , Femenino , Fibroblastos/efectos de los fármacos , Humanos , Masculino , Ratones Endogámicos BALB C , Proteínas Serina-Treonina Quinasas/efectos de los fármacos , Receptor Tipo I de Factor de Crecimiento Transformador beta , Receptores de Factores de Crecimiento Transformadores beta/efectos de los fármacos , Transducción de Señal/efectos de los fármacos , Adulto JovenRESUMEN
OBJECTIVE: The aetiology of OA is not fully understood although several adipokines such as leptin are known mediators of disease progression. Since leptin levels were increased in synovial fluid compared to serum in OA patients, it was suggested that joint cells themselves could produce leptin. However, exact mechanisms underlying leptin production by chondrocytes are poorly understood. Nevertheless, prednisolone, although displaying powerful anti-inflammatory properties has been recently reported to be potent stimulator of leptin and its receptor in OA synovial fibroblasts. Therefore, we investigated, in vitro, spontaneous and prednisolone-induced leptin production in OA chondrocytes, focusing on transforming growth factor-ß (TGFß) and Wnt/ß-catenin pathways. DESIGN: We used an in vitro dedifferentiation model, comparing human freshly isolated hip OA chondrocytes cultivated in monolayer during 1 day (type II, COL2A1 +; type X, COL10A1 + and type I collagen, COL1A1 -) or 14 days (COL2A1 -; COL10A1 - and COL1A1+). RESULTS: Leptin expression was not detected in day1 OA chondrocytes whereas day14 OA chondrocytes produced leptin, significantly increased with prednisolone. Activin receptor-like kinase 1 (ALK1)/ALK5 ratio was shifted during dedifferentiation, from high ALK5 and phospho (p)-Smad2 expression at day1 to high ALK1, endoglin and p-Smad1/5 expression at day14. Moreover, inactive glycogen synthase kinase 3 (GSK3) and active ß-catenin were only found in dedifferentiated OA chondrocytes. Smad1 and ß-catenin but not endoglin stable lentiviral silencing led to a significant decrease in leptin production by dedifferentiated OA chondrocytes. CONCLUSIONS: Only dedifferentiated OA chondrocytes produced leptin. Prednisolone markedly enhanced leptin production, which involved Smad1 and ß-catenin activation.
Asunto(s)
Condrocitos/metabolismo , Leptina/metabolismo , Osteoartritis de la Cadera/metabolismo , ARN Mensajero/metabolismo , Receptores de Activinas Tipo II/efectos de los fármacos , Receptores de Activinas Tipo II/genética , Receptores de Activinas Tipo II/metabolismo , Adulto , Anciano , Anciano de 80 o más Años , Cartílago Articular/citología , Cartílago Articular/efectos de los fármacos , Cartílago Articular/metabolismo , Desdiferenciación Celular/efectos de los fármacos , Desdiferenciación Celular/genética , Condrocitos/efectos de los fármacos , Colágeno Tipo X/efectos de los fármacos , Colágeno Tipo X/genética , Colágeno Tipo X/metabolismo , Subunidad alfa 1 del Factor de Unión al Sitio Principal/efectos de los fármacos , Subunidad alfa 1 del Factor de Unión al Sitio Principal/genética , Subunidad alfa 1 del Factor de Unión al Sitio Principal/metabolismo , Femenino , Glucocorticoides/farmacología , Glucógeno Sintasa Quinasa 3/efectos de los fármacos , Glucógeno Sintasa Quinasa 3/genética , Glucógeno Sintasa Quinasa 3/metabolismo , Humanos , Técnicas In Vitro , Linfotoxina-alfa/efectos de los fármacos , Linfotoxina-alfa/genética , Linfotoxina-alfa/metabolismo , Masculino , Metaloproteinasa 13 de la Matriz/efectos de los fármacos , Metaloproteinasa 13 de la Matriz/genética , Metaloproteinasa 13 de la Matriz/metabolismo , Persona de Mediana Edad , Osteoartritis de la Cadera/genética , Prednisolona/farmacología , Proteínas Serina-Treonina Quinasas/efectos de los fármacos , Proteínas Serina-Treonina Quinasas/genética , Proteínas Serina-Treonina Quinasas/metabolismo , ARN Mensajero/efectos de los fármacos , Receptor Tipo I de Factor de Crecimiento Transformador beta , Receptores de Factores de Crecimiento Transformadores beta/efectos de los fármacos , Receptores de Factores de Crecimiento Transformadores beta/genética , Receptores de Factores de Crecimiento Transformadores beta/metabolismo , Factor de Transcripción SOX9/efectos de los fármacos , Factor de Transcripción SOX9/metabolismo , Proteína Smad1/efectos de los fármacos , Proteína Smad1/genética , Proteína Smad1/metabolismo , Proteína Smad2/efectos de los fármacos , Proteína Smad2/genéticaRESUMEN
Growth differentiation factor-8 (GDF-8) has been recently shown to be expressed in human granulosa cells, and the mature form of GDF-8 protein can be detected in the follicular fluid. However, the biological function and significance of this growth factor in the human ovary remains to be determined. Here, we investigated the effects of GDF-8 on steroidogenic enzyme expression and the potential mechanisms of action in luteinized human granulosa cells. We demonstrated that treatment with GDF-8 did not affect the mRNA levels of P450 side-chain cleavage enzyme and 3ß-hydroxysteroid dehydrogenase, whereas it significantly down-regulated steroidogenic acute regulatory protein (StAR) expression and decreased progesterone production. The suppressive effect of GDF-8 on StAR expression was abolished by the inhibition of the TGF-ß type I receptor. In addition, treatment with GDF-8 activated both Smad2/3 and ERK1/2 signaling pathways. Furthermore, knockdown of activin receptor-like kinase 5 reversed the effects of GDF-8 on Smad2/3 phosphorylation and StAR expression. The inhibition of Smad3 or ERK1/2 signaling pathways attenuated the GDF-8-induced down-regulation of StAR and production of progesterone. Interestingly, the concentrations of GDF-8 were negatively correlated with those of progesterone in human follicular fluid. These results indicate a novel autocrine function of GDF-8 to down-regulate StAR expression and decrease progesterone production in luteinized human granulosa cells, most likely through activin receptor-like kinase 5-mediated Smad3 and ERK1/2 signaling pathways. Our findings suggest that granulosa cells might play a critical role in the regulation of progesterone production to prevent premature luteinization during the final stage of folliculogenesis.
Asunto(s)
Células de la Granulosa/efectos de los fármacos , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Miostatina/farmacología , Proteínas Serina-Treonina Quinasas/efectos de los fármacos , ARN Mensajero/efectos de los fármacos , Receptores de Factores de Crecimiento Transformadores beta/efectos de los fármacos , Proteína smad3/efectos de los fármacos , 3-Hidroxiesteroide Deshidrogenasas/efectos de los fármacos , 3-Hidroxiesteroide Deshidrogenasas/genética , Línea Celular , Regulación hacia Abajo , Femenino , Líquido Folicular/química , Células de la Granulosa/metabolismo , Humanos , Luteinización , Fosfoproteínas/efectos de los fármacos , Fosfoproteínas/genética , Fosforilación/efectos de los fármacos , Progesterona/biosíntesis , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Proteínas Serina-Treonina Quinasas/metabolismo , ARN Mensajero/metabolismo , Receptor Tipo I de Factor de Crecimiento Transformador beta , Receptores de Factores de Crecimiento Transformadores beta/antagonistas & inhibidores , Receptores de Factores de Crecimiento Transformadores beta/metabolismo , Transducción de Señal , Proteína Smad2/efectos de los fármacos , Proteína Smad2/metabolismo , Proteína smad3/metabolismoRESUMEN
Transforming growth factor-ß (TGF-ß) has both tumor suppressive and oncogenic activities. Autocrine TGF-ß signaling supports tumor survival and growth in certain types of cancer, and the TGF-ß signaling pathway is a potential therapeutic target for these types of cancer. TGF-ß induces p21 expression, and p21 is considered as an oncogene as well as a tumor suppressor, due to its anti-apoptotic activity. Thus, we hypothesized that autocrine TGF-ß signaling maintains the expression of p21 at levels that can support cell growth. To verify this hypothesis, we sought to examine p21 expression and cell growth in various cancer cells following the inhibition of autocrine TGF-ß signaling using siRNAs targeting TGF-ß signaling components and SB431542, a TGF-ß receptor inhibitor. Results from the present study show that p21 expression and cell growth were reduced by knockdown of TGF-ß signaling components using siRNA in MDA-MB231 and A549 cells. Cell growth was also reduced in p21 siRNA-transfected cells. Downregulation of p21 expression induced cellular senescence in MDA-MB231 cells but did not induce apoptosis in both cells. These data suggest that autocrine TGF-ß signaling is required to sustain p21 levels for positive regulation of cell cycle. On the other hand, treatment with SB431542 up-regulated p21 expression while inhibiting cell growth. The TGF-ß signaling pathway was not associated with the SB431542-mediated induction of p21 expression. Specificity protein 1 (Sp1) was downregulated by treatment with SB431542, and p21 expression was increased by Sp1 knockdown. These findings suggest that downregulation of Sp1 expression is responsible for SB43154-induced p21 expression.
Asunto(s)
Benzamidas/farmacología , Neoplasias de la Mama/tratamiento farmacológico , Inhibidor p21 de las Quinasas Dependientes de la Ciclina/metabolismo , Dioxoles/farmacología , Proteínas Serina-Treonina Quinasas/efectos de los fármacos , Receptores de Factores de Crecimiento Transformadores beta/efectos de los fármacos , Transducción de Señal/efectos de los fármacos , Factor de Crecimiento Transformador beta/farmacología , Animales , Comunicación Autocrina/efectos de los fármacos , Neoplasias de la Mama/genética , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/patología , Células COS , Células CACO-2 , Proliferación Celular/efectos de los fármacos , Senescencia Celular/efectos de los fármacos , Chlorocebus aethiops , Inhibidor p21 de las Quinasas Dependientes de la Ciclina/genética , Femenino , Regulación Neoplásica de la Expresión Génica , Células HCT116 , Humanos , Células MCF-7 , Masculino , Proteínas Serina-Treonina Quinasas/genética , Proteínas Serina-Treonina Quinasas/metabolismo , Interferencia de ARN , Receptor Tipo I de Factor de Crecimiento Transformador beta , Receptores de Factores de Crecimiento Transformadores beta/genética , Receptores de Factores de Crecimiento Transformadores beta/metabolismo , Factor de Transcripción Sp1/genética , Factor de Transcripción Sp1/metabolismo , Factores de Tiempo , TransfecciónRESUMEN
The roles of the extracellular domain of type II TGF-ß receptor (TBRII-ECD) in physiological processes ranging from development to cancer to wound healing render it an attractive target for exploration with chemical tools. For such applications, large amounts of active soluble protein are needed, but the yields of TBRII-ECD we obtained with current folding protocols were variable. To expedite the identification of alternative folding conditions, we developed an on-plate screen. This assay indicated that effective folding additives included the non-detergent sulfobetaine-201 (NDSB-201). Although NDSB-201 can facilitate protein folding, the mode by which it does so is poorly understood. We postulated that specific interactions between NDSB-201 and TBRII-ECD might be responsible. Analysis by X-ray crystallography indicates that the TBRII-ECD possesses a binding pocket for NDSB-201. The pyridinium group of the additive stacks with a phenylalanine side chain in the binding site. The ability of NDSB-201 to occupy a pocket on the protein provides a molecular mechanism for the additive's ability to minimize TBRII-ECD aggregation and stabilize the folded state. NDSB-201 also accelerates TBRII-ECD crystallization, suggesting it may serve as a useful crystallization additive for proteins refolded with it. Our results also suggest there is a site on TBRII-ECD that could be targeted by small-molecule modulators.
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Betaína/análogos & derivados , Chaperonas Moleculares/química , Pliegue de Proteína/efectos de los fármacos , Proteínas Serina-Treonina Quinasas/química , Proteínas Serina-Treonina Quinasas/metabolismo , Receptores de Factores de Crecimiento Transformadores beta/química , Receptores de Factores de Crecimiento Transformadores beta/metabolismo , Betaína/química , Betaína/farmacología , Cristalografía por Rayos X , Humanos , Chaperonas Moleculares/farmacología , Proteínas Serina-Treonina Quinasas/efectos de los fármacos , Receptor Tipo II de Factor de Crecimiento Transformador beta , Receptores de Factores de Crecimiento Transformadores beta/efectos de los fármacosRESUMEN
OBJECTIVE: Pain is the main problem for patients with osteoarthritis (OA). Pain is linked to inflammation, but in OA a subset of patients suffers from pain without inflammation, indicating an alternative source of pain. Nerve Growth Factor (NGF) inhibition is very efficient in blocking pain during OA, but the source of NGF is unclear. We hypothesize that damaged cartilage in OA releases Transforming Growth Factor-ß (TGF-ß), which in turn stimulates chondrocytes to produce NGF. DESIGN: Murine and human chondrocyte cell lines, primary bovine and human chondrocytes, and cartilage explants from bovine metacarpal joints and human OA joints were stimulated with TGF-ß1 and/or Interleukin-1 (IL-1)ß. We analyzed NGF expression on mRNA level with QPCR and stained human OA cartilage for NGF immunohistochemically. Cultures were additionally pre-incubated with inhibitors for TAK1, Smad2/3 or Smad1/5/8 signaling to identify the TGF-ß pathway inducing NGF. RESULTS: NGF expression was consistently induced in higher levels by TGF-ß than IL-1 in all of our experiments: murine, bovine and human origin, in cell lines, primary chondrocytes and explants cultures. TAK1 inhibition consistently reduced TGF-ß-induced NGF whereas it fully blocked IL-1ß-induced NGF expression. In contrast, ALK5-Smad2/3 inhibition fully blocked TGF-ß-induced NGF expression. Despite the large variation in basal NGF in human OA samples (mRNA and histology), TGF-ß exposure led to a consistent high level of NGF induction. CONCLUSION: We show for the first time that TGF-ß induces NGF expression in chondrocytes, in a ALK5-Smad2/3 dependent manner. This reveals a potential alternative non-inflammatory source of pain in OA.
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Cartílago Articular/efectos de los fármacos , Condrocitos/efectos de los fármacos , Interleucina-1beta/farmacología , Factor de Crecimiento Nervioso/efectos de los fármacos , Osteoartritis/metabolismo , Dolor/metabolismo , ARN Mensajero/metabolismo , Factor de Crecimiento Transformador beta1/farmacología , Animales , Cartílago Articular/metabolismo , Bovinos , Línea Celular , Condrocitos/metabolismo , Humanos , Ratones , Factor de Crecimiento Nervioso/genética , Factor de Crecimiento Nervioso/metabolismo , Osteoartritis/complicaciones , Osteoartritis/genética , Dolor/etiología , Dolor/genética , Proteínas Serina-Treonina Quinasas/efectos de los fármacos , Proteínas Serina-Treonina Quinasas/genética , Proteínas Serina-Treonina Quinasas/metabolismo , Reacción en Cadena en Tiempo Real de la Polimerasa , Receptor Tipo I de Factor de Crecimiento Transformador beta , Receptores de Factores de Crecimiento Transformadores beta/efectos de los fármacos , Receptores de Factores de Crecimiento Transformadores beta/genética , Receptores de Factores de Crecimiento Transformadores beta/metabolismo , Transducción de Señal , Proteína Smad2/efectos de los fármacos , Proteína Smad2/genética , Proteína Smad2/metabolismo , Proteína smad3/efectos de los fármacos , Proteína smad3/genética , Proteína smad3/metabolismoRESUMEN
OBJECTIVES: We have previously described the antifibrotic role of the soluble guanylate cyclase (sGC). The mode of action, however, remained elusive. In the present study, we describe a novel link between sGC signalling and transforming growth factor ß (TGFß) signalling that mediates the antifibrotic effects of the sGC. METHODS: Human fibroblasts and murine sGC knockout fibroblasts were treated with the sGC stimulator BAY 41-2272 or the stable cyclic guanosine monophosphate (cGMP) analogue 8-Bromo-cGMP and stimulated with TGFß. sGC knockout fibroblasts were isolated from sGCI(fl/fl) mice, and recombination was induced by Cre-adenovirus. In vivo, we studied the antifibrotic effects of BAY 41-2272 in mice overexpressing a constitutively active TGF-ß1 receptor. RESULTS: sGC stimulation inhibited TGFß-dependent fibroblast activation and collagen release. sGC knockout fibroblasts confirmed that the sGC is essential for the antifibrotic effects of BAY 41-2272. Furthermore, 8-Bromo-cGMP reduced TGFß-dependent collagen release. While nuclear p-SMAD2 and 3 levels, SMAD reporter activity and transcription of classical TGFß target genes remained unchanged, sGC stimulation blocked the phosphorylation of ERK. In vivo, sGC stimulation inhibited TGFß-driven dermal fibrosis but did not change p-SMAD2 and 3 levels and TGFß target gene expression, confirming that non-canonical TGFß pathways mediate the antifibrotic sGC activity. CONCLUSIONS: We elucidated the antifibrotic mode of action of the sGC that increases cGMP levels, blocks non-canonical TGFß signalling and inhibits experimental fibrosis. Since sGC stimulators have shown excellent efficacy and tolerability in phase 3 clinical trials for pulmonary arterial hypertension, they may be further developed for the simultaneous treatment of fibrosis and vascular disease in systemic sclerosis.
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Fibroblastos/patología , Guanilato Ciclasa/metabolismo , Receptores Citoplasmáticos y Nucleares/metabolismo , Esclerodermia Sistémica/patología , Esclerodermia Sistémica/fisiopatología , Transducción de Señal/fisiología , Piel/patología , Factor de Crecimiento Transformador beta/antagonistas & inhibidores , Animales , Estudios de Casos y Controles , Células Cultivadas , Colágeno/metabolismo , GMP Cíclico/metabolismo , Modelos Animales de Enfermedad , Fibroblastos/metabolismo , Fibrosis/metabolismo , Fibrosis/prevención & control , Guanilato Ciclasa/deficiencia , Humanos , Técnicas In Vitro , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Sistema de Señalización de MAP Quinasas/fisiología , Ratones , Ratones Noqueados , Pirazoles/farmacología , Piridinas/farmacología , Receptores Citoplasmáticos y Nucleares/agonistas , Receptores Citoplasmáticos y Nucleares/deficiencia , Receptores de Factores de Crecimiento Transformadores beta/efectos de los fármacos , Receptores de Factores de Crecimiento Transformadores beta/metabolismo , Esclerodermia Sistémica/metabolismo , Transducción de Señal/efectos de los fármacos , Piel/metabolismo , Proteínas Smad/metabolismo , Guanilil Ciclasa Soluble , Factor de Crecimiento Transformador beta/metabolismoRESUMEN
Transforming growth factor beta (TGF-ß) plays different roles in health and disease. TGF-ß has been assumed as a dual factor in tumor growth, since it can repress epithelial tumor development in early stages, while it acts as a tumor promoter in the late stages of tumor progression. The cancer cells, during cancerogenesis, acquire migration and invasion capacities and finally they metastasize. The urokinase type plasminogen activator (uPA) system, comprised of uPA, the cell surface receptor (uPAR) and plasminogen-plasmin, is involved in the proteolytic degradation of the extracellular matrix and it also regulates several critical cellular events by its capacity to trigger the activation of intracellular signaling pathways. This enables the cancer cell survival, its dissemination, and enhancement of cell malignancy during tumor progression. The expression of both uPA and uPAR is finely regulated in normal development, but their expression is deregulated in cancer. TGF-ß regulates uPA expression in cancer cells while uPA, by conversion of plasminogen to active form, plasmin, may release TGF-ß from its latent state. Thus, these pathways cross-regulate each other by mutual feedback contributing to tumor progression. Here, we review the specific roles and the interplay between TGF-ß and uPA system in cancer cells, the current cancer therapies and the novel patents focused mainly on uPA and TGF-beta ligands and their cell surface receptors respectively. Finally, with regard to the mutual activity of uPA and TGF-ß in tumorigenesis, the aim of this review is to expose the potentiality of TGF-ß and uPA systems as becoming combinatorial targets for therapies and patents.
Asunto(s)
Antineoplásicos/farmacología , Antineoplásicos/uso terapéutico , Carcinogénesis/efectos de los fármacos , Neoplasias/tratamiento farmacológico , Factor de Crecimiento Transformador beta/antagonistas & inhibidores , Activador de Plasminógeno de Tipo Uroquinasa/antagonistas & inhibidores , Humanos , Patentes como Asunto , Receptores de Factores de Crecimiento Transformadores beta/efectos de los fármacos , Factor de Crecimiento Transformador beta/fisiología , Activador de Plasminógeno de Tipo Uroquinasa/fisiologíaRESUMEN
Exposure to oxidants results in cellular alterations that are implicated in aging and age-associated diseases. Here, we report that brief, low-level oxidative exposure leads to long-term elevation of cellular reactive oxygen species (ROS) levels and oxidative damage in human skin fibroblasts. Elevated ROS impairs the transforming growth factor-ß (TGF-ß) pathway, through reduction of type II TGF-ß receptor (TßRII) and SMAD3 protein levels. This impairment results in reduced expression of connective tissue growth factor (CTGF/CCN2) and type I collagen, which are regulated by TGF-ß. Restoration of TßRII and SMAD3 together, but not separately, reinstates TGF-ß signaling and increases CTGF/CCN2 and type I collagen levels. Treatment with the anti-oxidant N-acetylcysteine reduces ROS elevation and normalizes TGF-ß signaling and target gene expression. These data reveal a novel linkage between limited oxidant exposure and altered cellular redox homeostasis that results in impairment of TGF-ß signaling. This linkage provides new insights regarding the mechanism by which aberrant redox homeostasis is coupled to decline of collagen production, a hallmark of human skin aging.
Asunto(s)
Fibroblastos/metabolismo , Regulación del Desarrollo de la Expresión Génica , Proteínas Serina-Treonina Quinasas/genética , ARN Mensajero/genética , Receptores de Factores de Crecimiento Transformadores beta/genética , Envejecimiento de la Piel/genética , Piel/metabolismo , Proteína smad3/genética , Acetilcisteína/farmacología , Colágeno Tipo I/metabolismo , Fibroblastos/citología , Fibroblastos/efectos de los fármacos , Depuradores de Radicales Libres/farmacología , Humanos , Proteínas Serina-Treonina Quinasas/biosíntesis , Proteínas Serina-Treonina Quinasas/efectos de los fármacos , Receptor Tipo II de Factor de Crecimiento Transformador beta , Receptores de Factores de Crecimiento Transformadores beta/biosíntesis , Receptores de Factores de Crecimiento Transformadores beta/efectos de los fármacos , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Transducción de Señal , Piel/citología , Piel/efectos de los fármacos , Envejecimiento de la Piel/efectos de los fármacos , Proteína smad3/biosíntesis , Proteína smad3/efectos de los fármacosRESUMEN
The glial cell-line derived neurotrophic factor (GDNF) is crucial for ureteric bud morphogenesis, spermatogenesis, and development of the enteric nervous system and is a potent survival factor for various neuronal populations. However, the impact of GDNF, at least on cell survival, was found to depend strongly on the presence of transforming growth factor ß (TGF-ß). In this study, we investigate the role of TGF-ß in GDNF-induced neuronal differentiation. In a cell culture paradigm of N2aGT cells (neuroblastoma cell line), we show that TGF-ß signaling localizes the GDNF ligand-binding receptor GFRa1 to the cell surface, which is a known mechanism by which TGF-ß is able to facilitate GDNF signaling. TGF-ß-mediated GDNF signaling slightly elevated the phosphorylation state of Ret, the canonical coreceptor for the GPI-linked (glycosyl-phosphatidylinositol) GFRa1. On the basis of morphological as well as immunocytological data, we finally show that GDNF-mediated neuronal differentiation is intensified when GDNF and TGF-ß act in concert.
Asunto(s)
Diferenciación Celular/fisiología , Factor Neurotrófico Derivado de la Línea Celular Glial/farmacología , Neuronas/fisiología , Factor de Crecimiento Transformador beta/farmacología , Animales , Biotinilación , Western Blotting , Línea Celular , Membrana Celular/metabolismo , Receptores del Factor Neurotrófico Derivado de la Línea Celular Glial/biosíntesis , Receptores del Factor Neurotrófico Derivado de la Línea Celular Glial/genética , Inmunohistoquímica , Inmunoprecipitación , Ratones , Neuritas/efectos de los fármacos , Fosforilación , Proteínas Proto-Oncogénicas c-ret/metabolismo , Reacción en Cadena en Tiempo Real de la Polimerasa , Receptores de Factores de Crecimiento Transformadores beta/efectos de los fármacos , Proteína Smad2/metabolismoRESUMEN
UNLABELLED: Transforming growth factor-beta (TGF-ß) is an important regulatory suppressor factor in hepatocytes. However, liver tumor cells develop mechanisms to overcome its suppressor effects and respond to this cytokine by inducing other processes, such as the epithelial-mesenchymal transition (EMT), which contributes to tumor progression and dissemination. Recent studies have placed chemokines and their receptors at the center not only of physiological cell migration but also of pathological processes, such as metastasis in cancer. In particular, CXCR4 and its ligand, stromal cell-derived factor 1α (SDF-1α) / chemokine (C-X-C motif) ligand 12 (CXCL12) have been revealed as regulatory molecules involved in the spreading and progression of a variety of tumors. Here we show that autocrine stimulation of TGF-ß in human liver tumor cells correlates with a mesenchymal-like phenotype, resistance to TGF-ß-induced suppressor effects, and high expression of CXCR4, which is required for TGF-ß-induced cell migration. Silencing of the TGF-ß receptor1 (TGFBR1), or its specific inhibition, recovered the epithelial phenotype and attenuated CXCR4 expression, inhibiting cell migratory capacity. In an experimental mouse model of hepatocarcinogenesis (diethylnitrosamine-induced), tumors showed increased activation of the TGF-ß pathway and enhanced CXCR4 levels. In human hepatocellular carcinoma tumors, high levels of CXCR4 always correlated with activation of the TGF-ß pathway, a less differentiated phenotype, and a cirrhotic background. CXCR4 concentrated at the tumor border and perivascular areas, suggesting its potential involvement in tumor cell dissemination. CONCLUSION: A crosstalk exists among the TGF-ß and CXCR4 pathways in liver tumors, reflecting a novel molecular mechanism that explains the protumorigenic effects of TGF-ß and opens new perspectives for tumor therapy.