RESUMO
Amyotrophic lateral sclerosis (ALS) is a progressive degenerative disorder of the upper and lower motor neuron systems. The high incidence of ALS in the southern part of the Kii Peninsula of Japan (K-ALS) was reported in the 1960s, but it has gradually decreased to the worldwide average. Although causes of the high incidence of ALS in this area are unknown, our previous studies suggested that environmental factors, including essential mineral deficiency and increased metal-induced oxidative stress, play a role in its development. Recently, it has been reported that microRNAs (miRNA) contribute to the degeneration of nervous system such as ALS. The aim of this study is to explore specific miRNAs in K-ALS and evaluate relationships between oxidative stress. We comprehensively analyzed serum miRNAs and examined urinary 8-hydroxy-2'-deoxyguanosine (8-OHdG), serum Cu/Zn superoxide dismutase (SOD) and serum NÉ-hexanoyl lysin (HEL) as oxidative stress markers in the patients with K-ALS, sporadic ALS (S-ALS), residents in this area (K-residents) and controls from another area. The expression levels of miR-92a-3p and miR-486-5p in the patients with K-ALS were significantly higher than those in controls. The HEL levels were significantly higher in the patients with K-ALS than in those with S-ALS and controls. The expression levels of miR-92a-3p and miR-486-5p were not correlated with the levels of HEL. A set of high levels of miR-92a-3p, miR-486-5p and serum HEL may be a useful biomarker for K-ALS in the Kii Peninsula. The findings should be further studied by a large number of subjects.
Assuntos
Esclerose Lateral Amiotrófica , Biomarcadores/sangue , MicroRNAs/sangue , Idoso , Idoso de 80 Anos ou mais , Esclerose Lateral Amiotrófica/sangue , Esclerose Lateral Amiotrófica/epidemiologia , Esclerose Lateral Amiotrófica/genética , Feminino , Humanos , Incidência , Japão/epidemiologia , Masculino , Pessoa de Meia-Idade , Estresse Oxidativo/fisiologiaRESUMO
PURPOSE: To clarify whether fibulins-5 is associated with primary spontaneous pneumothorax (PSP) in young PSP patients. METHODS: Forty-six surgically resected, fresh lung specimens were used. Patients were divided into 3 groups: younger than 25 years with pneumothorax (group Y), 25 years or older with pneumothorax (group O), and without pneumothorax (group C). Chest X-ray, computed tomography data, height/width ratio (H/W) and anteroposterior/transverse diameter ratio (a/b) were measured. Elastica van Gieson staining and immunofluorescence staining for fibulin-5 were performed. Fibulin-5 mRNA expression and protein levels were measured by real-time PCR and western blotting. Direct sequences of the fibulin-5 gene in PSP patients were performed. RESULTS: The mean H/W ratio in group Y was significantly larger than that in the other groups (p <0.01). The mean a/b ratio in group Y was significantly smaller than that in the other groups (p = 0.02). Fibulin-5 mRNA expression was not significantly different among the groups (p = 0.64). The relative intensity of fibulin-5 protein in group Y was significantly lower than that in group O (p = 0.006), with no significant differences between groups O and C (p = 0.14). CONCLUSIONS: We showed that fibulin-5 is reduced in patients with PSP who are younger than 25 years.
Assuntos
Proteínas da Matriz Extracelular/análise , Pulmão/química , Pneumotórax/metabolismo , Adolescente , Adulto , Idoso , Análise de Variância , Western Blotting , Estudos de Casos e Controles , Distribuição de Qui-Quadrado , Regulação para Baixo , Proteínas da Matriz Extracelular/genética , Feminino , Humanos , Pulmão/diagnóstico por imagem , Pulmão/cirurgia , Masculino , Microscopia de Fluorescência , Pessoa de Meia-Idade , Pneumotórax/diagnóstico por imagem , Pneumotórax/genética , Pneumotórax/cirurgia , RNA Mensageiro/análise , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Análise de Sequência de DNA , Tomografia Computadorizada por Raios X , Adulto JovemRESUMO
The role of microRNAs (miRNAs) in vascular calcification is currently unclear. To examine how miRNAs are involved in vascular smooth muscle cell (VSMC) calcification, we explored the alteration of miRNAs in VSMC calcification in vitro and in vivo. Klotho homozygous mutant mice (kl/kl) display vascular calcification and have perturbations of calcium handling. We therefore hypothesized that the calcium perturbations in VSMCs could be mediated by miRNAs. Using an miRNA array analysis, we demonstrated that miRNAs are aberrantly expressed in the aortic media of 3-week-old kl/kl mice compared with wild-type (WT) mice. The expression levels of miR-135a(*), miR-762, miR-714, and miR-712(*) in the aortic media of kl/kl mice were significantly higher than in WT mice. We used quantitative real-time reverse transcriptase polymerase chain reaction to further confirm that these miRNAs were increased in the aortic media of kl/kl mice and in cultured VSMCs treated with high phosphate and calcium. A search of the miRNA database indicated that the Ca(2+) efflux proteins NCX1, PMCA1, and NCKX4 frequently appeared as potential targets of these miRNAs. The transfection of miRNA mimics into cultured VSMCs reduced the protein levels of each potential target. Conversely, miRNA inhibitors reduced phosphate and calcium-induced VSMC calcification. Furthermore, these inhibitors decreased the intracellular Ca(2+) concentration in cultured VSMCs after treatment with phosphate and calcium. Our results suggest that increased expression of miR-135a(*), miR-762, miR-714, and miR-712(*) in VSMCs may be involved in VSMC calcification by disrupting Ca(2+) efflux proteins.
Assuntos
Aorta/parasitologia , Calcinose/genética , Cálcio/metabolismo , Proteínas de Membrana Transportadoras/genética , MicroRNAs/genética , Músculo Liso Vascular/patologia , Animais , Western Blotting , Cálcio/sangue , Células Cultivadas , Técnicas de Silenciamento de Genes , Camundongos , Fosfatos/sangue , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
Mutation of the human TRPS1 gene leads to trichorhinophalangeal syndrome (TRPS), which is characterized by an abnormal development of various organs including the craniofacial skeleton. Trps1 has recently been shown to be expressed in the jaw joints of zebrafish; however, whether Trps1 is expressed in the mammalian temporomandibular joint (TMJ), or whether it is necessary for TMJ development is unknown. We have analyzed (1) the expression pattern of Trps1 during TMJ development in mice and (2) TMJ development in Trps1 knockout animals. Trps1 is expressed in the maxillo-mandibular junction at embryonic day (E) 11.5. At E15.5, expression is restricted to the developing condylar cartilage and to the surrounding joint disc progenitor cells. In Trps1 knockout mice, the glenoid fossa of the temporal bone forms relatively normally but the condylar process is extremely small and the joint disc and cavities do not develop. The initiation of condyle formation is slightly delayed in the mutants at E14.5; however, at E18.5, the flattened chondrocyte layer is narrowed and most of the condylar chondrocytes exhibit precocious chondrocyte maturation. Expression of Runx2 and its target genes is expanded toward the condylar apex in the mutants. These observations underscore the indispensable role played by Trps1 in normal TMJ development in supporting the differentiation of disc and synoviocyte progenitor cells and in coordinating condylar chondrocyte differentiation.
Assuntos
Fatores de Transcrição GATA/metabolismo , Articulação Temporomandibular/embriologia , Articulação Temporomandibular/metabolismo , Animais , Cartilagem/metabolismo , Diferenciação Celular/genética , Proliferação de Células , Condrócitos/metabolismo , Condrócitos/patologia , Anormalidades Craniofaciais/metabolismo , Anormalidades Craniofaciais/patologia , Fatores de Transcrição GATA/deficiência , Fatores de Transcrição GATA/genética , Regulação da Expressão Gênica no Desenvolvimento , Humanos , Côndilo Mandibular/metabolismo , Côndilo Mandibular/patologia , Camundongos , Camundongos Knockout , Mutação/genética , Proteínas Repressoras , Articulação Temporomandibular/patologiaRESUMO
Smad3 is an intracellular signaling molecule in the transforming growth factor ß (TGF-ß) pathway that serves as a regulator of chondrogenesis and osteogenesis. To investigate the role of the TGF-ß/Smad3 signaling in the process of fracture healing, an open fracture was introduced in mouse tibiae, and the histology of the healing process was compared between wild-type (WT) and Smad3-null (KO) mice. In KO mice, the bone union formed more rapidly with less formation of cartilage in the callus and eventually the fracture was repaired more rapidly than in WT mice. Alkaline phosphatase staining showed that osteoblastic differentiation in the fracture callus was promoted in KO mice. Additionally, TRAP staining and the TUNEL assay revealed that the induction of osteoclasts and apoptotic cells was significantly promoted in the healing callus of KO mice. Sox9 expression clearly decreased at both mRNA and protein levels in the early stage of fracture in KO mice. In contrast, the expression of genes for osteogenesis and osteoclast formation increased from day 5 until day 14 post-fracture in KO mice compared to WT mice. From these results, we concluded that the loss of TGF-ß/Smad3 signaling promoted callus formation by promoting osteogenesis and suppressing chondrogenesis, which resulted in faster fracture healing.
Assuntos
Calo Ósseo/citologia , Consolidação da Fratura/genética , Proteína Smad3/fisiologia , Fosfatase Ácida/metabolismo , Fosfatase Alcalina/metabolismo , Animais , Calo Ósseo/metabolismo , Calo Ósseo/patologia , Diferenciação Celular , Condrogênese/genética , Feminino , Isoenzimas/metabolismo , Camundongos , Camundongos Knockout , Osteoclastos/metabolismo , Osteoclastos/patologia , Osteogênese/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Radiografia , Fatores de Transcrição SOX9/genética , Fatores de Transcrição SOX9/metabolismo , Proteína Smad3/genética , Fosfatase Ácida Resistente a Tartarato , Tíbia/diagnóstico por imagem , Tíbia/metabolismo , Fatores de Tempo , Fator de Crescimento Transformador beta/genética , Fator de Crescimento Transformador beta/metabolismoRESUMO
Tricho-rhino-phalangeal syndrome (TRPS) is an autosomal dominant skeletal disorder caused by mutations of the Trps1 gene, which encodes a GATA type transcriptional repressor. To investigate the genes that act downstream of Trps1, we performed a DNA array using ATDC5 cells. One of the target genes identified from the DNA array was Runx1, which is essential for hematopoiesis and like Runx2 plays a significant role in chondrogenesis. A luciferase promoter assay and a chromosome immunoprecipitation assay showed that Runx1 expression in mouse epiphyseal cartilage was repressed by Trps1 binding to the GATA domain of the P2 promoter; the proximal segment of two promoters of the Runx1 gene. The aberrant expression of P2 transcripts was detected in growth plate chondrocytes from Trps1-null mice by in situ hybridization. In conclusion, Trps1 binds to the P2 promoter of the Runx1 gene and down-regulates Runx1 expression, which is necessary for normal cartilage formation.
Assuntos
Cartilagem/metabolismo , Subunidade alfa 2 de Fator de Ligação ao Core/genética , Epífises/metabolismo , Fatores de Transcrição GATA/deficiência , Regiões Promotoras Genéticas/genética , Animais , Sítios de Ligação , Western Blotting , Cartilagem/patologia , Linhagem Celular , Condrócitos/metabolismo , Condrócitos/patologia , Condrogênese/genética , Imunoprecipitação da Cromatina , Subunidade alfa 2 de Fator de Ligação ao Core/metabolismo , Regulação para Baixo/genética , Epífises/patologia , Fatores de Transcrição GATA/metabolismo , Lâmina de Crescimento/metabolismo , Lâmina de Crescimento/patologia , Luciferases/metabolismo , Camundongos , Análise de Sequência com Séries de Oligonucleotídeos , Ligação Proteica , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Proteínas Repressoras , Transcrição GênicaRESUMO
SNAIL, a potent repressor of E-cadherin expression, plays a key role in inducing epithelial-to-mesenchymal transition (EMT) in epithelial cells. During EMT, epithelial cells lose cell polarity and adhesion, and undergo drastic morphological changes acquiring highly migratory abilities. Although there is increasing evidence that EMT is involved in the progression of some human cancers, its significance in the progression of pancreatic cancer remains elusive. In Panc-1, a well-known human pancreatic cancer cell line in which EMT is triggered by TGF-ß1 treatment, SNAIL and vimentin are highly expressed, whereas E-cadherin expression is scant. In contrast, another human pancreatic cancer cell line, BxPC3, in which SNAIL expression is not detected, has high levels of E-cadherin expression and does not undergo EMT upon TGF-ß1 treatment. After transfecting the SNAIL gene into BxPC3, however, the cells undergo EMT with remarkable alterations in cell morphology and molecular expression patterns without the addition of any growth factors. Furthermore, in an orthotopic transplantation model using SCID mice, SNAIL-transfected BxPC3 displayed highly metastatic and invasive activities. In the immunohistochemical analysis of the tumor derived from the SNAIL-expressing BxPC3, alterations suggestive of EMT were observed in the invasive tumor front. SNAIL enabled BxPC3 to undergo EMT, endowing it with a highly malignant potential in vivo. These results indicate that SNAIL-mediated EMT may be relevant in the progression of pancreatic cancer, and SNAIL could be a molecular target for a pancreatic cancer intervention.
Assuntos
Células Epiteliais/patologia , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patologia , Fatores de Transcrição/fisiologia , Animais , Caderinas/antagonistas & inibidores , Linhagem Celular , Linhagem Celular Tumoral , Progressão da Doença , Feminino , Humanos , Camundongos , Camundongos SCID , Invasividade Neoplásica , Metástase Neoplásica , Transplante de Neoplasias , Neoplasias Pancreáticas/genética , Fatores de Transcrição da Família Snail , Transfecção , Fator de Crescimento Transformador beta1/farmacologia , Vimentina/genética , Vimentina/metabolismoRESUMO
Mutations in TRPS1 cause tricho-rhino-pharyngeal syndrome (TRPS). Trps1 is essential for nephron development, acting downstream of Bmp7. Because Bmp7 counteracts epithelial-to-mesenchymal transition (EMT) and reverses chronic renal injury, we examined the function of Trps1 in renal fibrosis. Immunohistochemistry revealed Trps1 expression in proximal tubular epithelial cells of mice. Unilateral ureteral obstruction reduced mRNA and protein expression of Trps1 in wild-type and heterozygous Trps1-knockout (Trps1(+/-)) mice. Trps1 haploinsufficiency promoted tubulointerstitial fibrosis via increased phosphorylation of Smad3 and decreased Smad7 protein. In primary culture, Trps1 deficiency promoted TGF-beta1-mediated EMT in proximal tubule cells. Trps1(+/-)-derived cells had higher levels of phosphorylated Smad3, and TGF-beta1 induced a time-dependent decrease in Smad7 protein in wild-type and Trps1(+/-) kidneys. In addition, compared with wild-type cells, Trps1(+/-) cells had double the amount of the E3 ubiquitin ligase Arkadia, and TGF-beta1 induced further Arkadia expression. Furthermore, knockdown of Arkadia inhibited TGF-beta1-induced EMT in Trps1(+/-) cells. Collectively, these data suggest that Trps1 haploinsufficiency enhances TGF-beta1-induced EMT and tubulointerstitial fibrosis by modulating the amount of Smad7 through Arkadia/ubiquitin-mediated degradation.
Assuntos
Fatores de Transcrição GATA/fisiologia , Rim/patologia , Ubiquitina-Proteína Ligases/fisiologia , Animais , Células Endoteliais/patologia , Fibrose , Fatores de Transcrição GATA/deficiência , Fatores de Transcrição GATA/genética , Mesoderma/patologia , Camundongos , Proteínas dos Microfilamentos/análise , Proteínas Repressoras , Transdução de Sinais , Proteína Smad3/fisiologia , Proteína Smad7/análise , Fator de Crescimento Transformador beta1/fisiologia , Obstrução Ureteral/patologiaRESUMO
During embryonic development, the mesenchyme of the lungs, gut, kidneys, and other tissues expresses Trps1, an atypical member of the GATA-type family of transcription factors. Our previous work suggested the possibility that Trps1 acts downstream of bone morphogenic protein 7 (Bmp7), which is essential for normal renal development. To examine the role of Trps1 during early renal development, we generated Trps1-deficient mice and examined their renal histology. Compared with wild-type mice, Trps1-deficient newborn mice had fewer tubules and glomeruli, an expanded renal interstitium, and numerous uninduced metanephric mesenchymal cells, which resulted in fewer nephrons. In wild-type kidneys, Trps1 expression was present in ureteric buds, cap mesenchyme, and renal vesicles, whereas Trps1 was virtually absent in Bmp7-deficient kidneys. Furthermore, Trps1-deficient kidneys had low levels of Pax2 and Wt1, which are markers of condensed mesenchymal cells, suggesting that a lack of Trps1 affects the differentiation of cap mesenchyme to renal vesicles. In cultured metanephric mesenchymal cells, Bmp7 induced Trps1 and E-cadherin and downregulated vimentin. Knockdown of Trps1 with small interference RNA inhibited this Bmp7-induced mesenchymal-to-epithelial transition. Last, whole-mount in situ hybridization of Wnt9b and Wnt4 demonstrated prolonged branching of ureteric buds and sparse cap mesenchyme in the kidneys of Trps1-deficient mice. Taken together, these findings suggest that normal formation of nephrons requires Trps1, which mediates mesenchymal-to-epithelial transition and ureteric bud branching during early renal development.
Assuntos
Proteína Morfogenética Óssea 7/fisiologia , Fatores de Transcrição GATA/fisiologia , Rim/crescimento & desenvolvimento , Animais , Animais Recém-Nascidos , Proteína Morfogenética Óssea 7/biossíntese , Células Cultivadas , Fatores de Transcrição GATA/biossíntese , Camundongos , Proteínas RepressorasRESUMO
TNF-alpha and TGF-beta1 have a complementary relationship in fibrogenesis. This study was performed to investigate the role of TNF-alpha in renal tubular interstitial fibrosis. We compared the extent of renal tubular interstitial fibrosis after unilateral ureteral obstruction (UUO) between wild-type and TNF-alpha-deficient mice by using immunohistochemistry, enzyme-linked immunoassay, and the real-time polymerase chain reaction (PCR). In comparison with wild-type mice, there was no significant difference in the extent of renal fibrosis in the TNF-alpha-deficient mice at 2 weeks after UUO. By 4 weeks after UUO, however, fibrosis marked an increase in the TNF-alpha-deficient mice to exceed that in the wild-type mice. Immunohistochemistry, enzyme-linked immunoassay, and real-time PCR demonstrated an increase of extracellular matrix in the kidneys of TNF-alpha-deficient mice that was caused by upregulation of the expression of TGF-beta1 and Snail, which in turn resulted from an increase of infiltrating macrophages. Real-time PCR revealed an increase in expression of the TNF-alpha type 2 receptor at 4 weeks after UUO, which explained the difference in the extent of renal fibrosis between TNF-alpha-deficient and wild-type mice. In the chronic stage of renal fibrosis, TNF-alpha suppresses the infiltration of macrophages by inducing TNF-alpha type 2 receptor expression, resulting in the amelioration of fibrosis.
Assuntos
Túbulos Renais/patologia , Fator de Necrose Tumoral alfa/deficiência , Obstrução Ureteral/complicações , Obstrução Ureteral/patologia , Actinas/genética , Actinas/metabolismo , Animais , Colágeno Tipo I/genética , Colágeno Tipo I/metabolismo , Fibrose , Regulação da Expressão Gênica , Imuno-Histoquímica , Túbulos Renais/metabolismo , Macrófagos/metabolismo , Camundongos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptores Tipo I de Fatores de Necrose Tumoral/genética , Receptores Tipo I de Fatores de Necrose Tumoral/metabolismo , Receptores Tipo II do Fator de Necrose Tumoral/genética , Receptores Tipo II do Fator de Necrose Tumoral/metabolismo , Fatores de Transcrição da Família Snail , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Fator de Crescimento Transformador beta1/genética , Fator de Crescimento Transformador beta1/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Obstrução Ureteral/genéticaRESUMO
We have reported that elongation of the columnar proliferative zone of long bone growth plates in Trps1-/- mice during the late fetal stage in the previous study [1]. Since expression of Trps1 protein was found to overlap with that of mRNAs for Indian hedgehog (Ihh), PTH/PTHrP receptor (PPR), and PTHrP, we hypothesized that Trps1 may inhibit the hypertrophic differentiation of chondrocytes by interacting with the Ihh/PTHrP feedback loop. To investigate whether Trps1 has a role in this Ihh/PTHrP feedback loop, we compared the growth plates of Trps1-/- mice and wild-type (Trps1+/+) mice. Immunohistochemistry showed that Trps1 protein was strongly expressed in the periarticular and prehypertrophic zones of the fetal growth plate in wild-type mice on embryonic day 18.5 (E18.5). On the other hand, Ihh, PPR, and PTHrP mRNAs were predominantly expressed in the prehypertrophic zone at this stage of development. While expression of Ihh and PPR by prehypertrophic chondrocytes was unaffected in the growth plates of Trps1-/- mice, the range of PTHrP expression was expanded toward the proliferating zone in these mice. Quantitative real-time PCR analysis demonstrated upregulation of PTHrP in the epiphyseal growth plates of Trps1-/- mice. Furthermore, promoter analysis combined with the chromatin immunoprecipitation (ChIP) assay demonstrated that direct binding of Trps1 to the PTHrP promoter suppressed the transcription of PTHrP. Finally, organ culture of E14.5 tibiae in the absence or the presence of Pthrp revealed that the proliferative zone of the tibial growth plate was elongated by culture with Pthrp compared to that of control tibiae. Taken together, these data provide the first genetic evidence that lack of Trps1 leads to overexpression of PTHrP, and that Trps1 is required to maintain the normal organization of chondrocytes in the growth plate.
Assuntos
Proliferação de Células , Fatores de Transcrição GATA/fisiologia , Lâmina de Crescimento/citologia , Proteína Relacionada ao Hormônio Paratireóideo/fisiologia , Regulação para Cima/fisiologia , Animais , Sequência de Bases , Imunoprecipitação da Cromatina , Primers do DNA , Fatores de Transcrição GATA/genética , Imuno-Histoquímica , Hibridização In Situ , Camundongos , Camundongos Knockout , Proteína Relacionada ao Hormônio Paratireóideo/genética , RNA Mensageiro/genética , Proteínas Repressoras , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
Tricho-rhino-phalangeal syndrome (TRPS) is an autosomal dominant skeletal disorder caused by mutations of TRPS1. Based on the similar expression patterns of Trps1 and Gdf5, we hypothesized a possible functional interaction between these two molecules. Using a chondrogenic cell line (ATDC5), we investigated the association of Gdf5-mediated signaling pathways with Trps1 and the phenotypic changes of ATDC5 cells due to over-expression or suppression of Trps1. Treatment of cells with Gdf5 enhanced Trps1 protein levels and phosphorylation of p38 mitogen-activated protein kinase (MAPK) in a dose-dependent manner. Nuclear translocation of Trps1 was also induced by Gdf5. These effects were blocked by a dominant negative form of activin-linked kinase 6 (dn-Alk6) and by SB203580, an inhibitor of the p38 MAPK pathway. Conversely, Gdf5 expression was suppressed by the over-expression of Trps1. Trps1-overexpressing ATDC5 (O/E) cells differentiated into chondrocytes more quickly than mock-infected control cells, whereas cells transfected with dn-Alk6 showed slower differentiation. On the other hand, O/E cells showed an increase of apoptosis along with the up-regulation of cleaved caspase 3 and down-regulation of Bcl-2, whereas dn-Alk6 cells showed suppression of apoptosis. In conclusion, Trps1 acts downstream of the Gdf5 signaling pathway and promotes the differentiation and apoptosis of ATDC5 cells.
Assuntos
Apoptose/fisiologia , Proteínas Morfogenéticas Ósseas/metabolismo , Condrogênese/fisiologia , Fatores de Transcrição GATA/metabolismo , Animais , Sequência de Bases , Doenças do Desenvolvimento Ósseo/genética , Receptores de Proteínas Morfogenéticas Ósseas Tipo I/genética , Receptores de Proteínas Morfogenéticas Ósseas Tipo I/metabolismo , Linhagem Celular , Anormalidades Craniofaciais/genética , Primers do DNA/genética , Retroalimentação , Fatores de Transcrição GATA/genética , Expressão Gênica , Fator 5 de Diferenciação de Crescimento , Camundongos , Fenótipo , Proteínas Repressoras , Transdução de Sinais , Transfecção , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismoRESUMO
Mutations in the TRPS1 gene lead to the tricho-rhino-phalangeal syndrome, which is characterized by skeletal defects and abnormal hair development. The TRPS1 gene encodes an atypical member of the GATA-type family of transcription factors. Here we show that mice with a disrupted Trps1 gene develop a chondrodysplasia characterized by diminished chondrocyte proliferation and decreased apoptosis in growth plates. Our analyses revealed that Trps1 is a repressor of Stat3 expression, which in turn controls chondrocyte proliferation and survival by regulating the expression of cyclin D1 and Bcl2. Our conclusion is supported (i) by siRNA-mediated depletion of Stat3 in Trps1-deficient chondrocytes, which normalized the expression of cyclin D1 and Bcl2, (ii) by overexpression of Trps1 in ATDC5 chondrocytes, which diminished Stat3 levels and increased proliferation and apoptosis, and (iii) by mutational analysis of the GATA-binding sites in the Stat3 gene, which revealed that their integrity is critical for the direct association with Trps1 and for Trps1-mediated repression of Stat3. Altogether our findings identify Trps1 as a novel regulator of chondrocytes proliferation and survival through the control of Stat3 expression.