RESUMO
Some studies suggest that the trace element selenium protects against colorectal cancer (CRC). However, the contribution of selenoprotein P (SELENOP), a unique selenocysteine-containing protein, to sporadic colorectal carcinogenesis challenges this paradigm. SELENOP is predominately secreted by the liver but is also expressed in various cells of the small intestine and colon in mice and humans. In this issue of the JCI, Pilat et al. demonstrate that increased SELENOP expression promoted the progression of conventional adenomas to carcinoma. SELENOP functioned as a modulator of canonical WNT signaling activity through interactions with WNT3A and its coreceptor LDL receptor-related protein 5/6 (LRP5/6). Secreted SELENOP formed a concentration gradient along the gut crypt axis, which might amplify WNT signaling activity by binding to LRPL5/6. The mechanism for WNT control via SELENOP may affect colorectal tumorigenesis and provide therapeutic targets for CRC.
Assuntos
Neoplasias Colorretais , Selênio , Humanos , Camundongos , Animais , Selenoproteína P/genética , Selenoproteína P/metabolismo , Carcinogênese/genética , Carcinogênese/metabolismo , Fígado/metabolismo , Transformação Celular Neoplásica/metabolismo , Selênio/metabolismo , Neoplasias Colorretais/genética , Neoplasias Colorretais/metabolismo , Proteína-5 Relacionada a Receptor de Lipoproteína de Baixa Densidade/genética , Proteína-5 Relacionada a Receptor de Lipoproteína de Baixa Densidade/metabolismo , Proteína Wnt3A/genética , Proteína Wnt3A/metabolismoRESUMO
Although selenium deficiency correlates with colorectal cancer (CRC) risk, the roles of the selenium-rich antioxidant selenoprotein P (SELENOP) in CRC remain unclear. In this study, we defined SELENOP's contributions to sporadic CRC. In human single-cell cRNA-Seq (scRNA-Seq) data sets, we discovered that SELENOP expression rose as normal colon stem cells transformed into adenomas that progressed into carcinomas. We next examined the effects of Selenop KO in a mouse adenoma model that involved conditional, intestinal epithelium-specific deletion of the tumor suppressor adenomatous polyposis coli (Apc) and found that Selenop KO decreased colon tumor incidence and size. We mechanistically interrogated SELENOP-driven phenotypes in tumor organoids as well as in CRC and noncancer cell lines. Selenop-KO tumor organoids demonstrated defects in organoid formation and decreases in WNT target gene expression, which could be reversed by SELENOP restoration. Moreover, SELENOP increased canonical WNT signaling activity in noncancer and CRC cell lines. In defining the mechanism of action of SELENOP, we mapped protein-protein interactions between SELENOP and the WNT coreceptors low-density lipoprotein receptor-related proteins 5 and 6 (LRP5/6). Last, we confirmed that SELENOP-LRP5/6 interactions contributed to the effects of SELENOP on WNT activity. Overall, our results position SELENOP as a modulator of the WNT signaling pathway in sporadic CRC.
Assuntos
Adenoma , Neoplasias Colorretais , Selênio , Camundongos , Animais , Humanos , Via de Sinalização Wnt , Selenoproteína P/genética , Selenoproteína P/metabolismo , Neoplasias Colorretais/patologia , Selênio/metabolismo , Carcinogênese/genética , Adenoma/metabolismo , Regulação Neoplásica da Expressão Gênica , Proteína-5 Relacionada a Receptor de Lipoproteína de Baixa Densidade/genética , Proteína-5 Relacionada a Receptor de Lipoproteína de Baixa Densidade/metabolismoRESUMO
Bone erosion is a debilitating pathological process of osteopathic disorder like rheumatoid arthritis (RA). Current treatment strategies render low disease activity but with disease recurrence. To find an alternative, we designed this study with an aim to explore the underlying therapeutic effect of PEGylated liposomal BBR (PEG-BBR) against Wnt1/ß-catenin mediated bone erosion in adjuvant-induced arthritic (AA) rat model and fibroblast-like synoviocytes (FLS) with reference to microRNA-23a (miR-23a) activity. Our initial studies using confocal microscopy and Near-Infrared Imaging (NIR) showed successful internalization of PEG-BBR and PEG-miR-23a in vitro and in vivo respectively and was retained till 48 h. The preferential internalization of PEG-BBR into the inflamed joint region significantly reduced the gene and protein level expression of major Wnt1 signaling mediators and reduced bone erosion in rats. Moreover, PEG-BBR treatment in FLS cells attenuated the gene and protein expression levels of FZD4, LRP5, ß-catenin, and Dvl-1 through the induction of CYLD. Furthermore, inhibition of these factors resulted in reduced bone loss and increased calcium retainability by altering the RANKL/OPG axis. PEG-BBR treatment markedly inhibited the expression of LRP5 protein on par with the DKK-1 (LRP5/Wnt signaling inhibitor) and suppressed the transcriptional activation of ß-catenin inside the cells. We further witnessed that miR-23a altered the expression levels of LRP5 through RNA interference. Overall, our findings endorsed that miR-23a possesses a multifaceted therapeutic efficiency like berberine in RA pathogenesis and can be considered as a potential candidate for therapeutic targeting of Wnt1/ß-catenin signaling in RA disease condition.
Assuntos
Artrite Experimental/tratamento farmacológico , Artrite Reumatoide/tratamento farmacológico , Berberina/farmacologia , MicroRNAs/genética , Animais , Berberina/administração & dosagem , Feminino , Lipossomos , Proteína-5 Relacionada a Receptor de Lipoproteína de Baixa Densidade/genética , Masculino , Polietilenoglicóis/química , Ratos , Ratos Wistar , Via de Sinalização Wnt/efeitos dos fármacos , Proteína Wnt1/metabolismo , beta Catenina/metabolismoRESUMO
Fluoride (F) is capable of promoting abnormal proliferation and differentiation in primary cultured mouse osteoblasts (OB cells), although the underlying mechanism responsible remains rare. This study aimed to explore the roles of wingless and INT-1 (Wnt) signaling pathways and screen appropriate doses of calcium (Ca2+) to alleviate the sodium fluoride (NaF)-induced OB cell toxicity. For this, we evaluated the effect of dickkopf-related protein 1 (DKK1) and Ca2+ on mRNA levels of wingless/integrated 3a (Wnt3a), low-density lipoprotein receptor-related protein 5 (LRP5), dishevelled 1 (Dv1), glycogen synthase kinase 3ß (GSK3ß), ß-catenin, lymphoid enhancer binding factor 1 (LEF1), and cellular myelocytomatosis oncogene (cMYC), as well as Ccnd1 (Cyclin D1) in OB cells challenged with 10-6 mol/L NaF for 24 h. The demonstrated data showed that F significantly increased the OB cell proliferation rate. Ectogenic 0.5 mg/L DKK1 significantly inhibited the proliferation of OB cells induced by F. The mRNA expression levels of Wnt3a, LRP5, Dv1, LEF1, ß-catenin, cMYC, and Ccnd1 were significantly increased in the F group, while significantly decreased in the 10-6 mol/L NaF + 0.5 mg/L DKK1 (FY) group. The mRNA expression levels of Wnt3a, LRP5, ß-catenin, and cMYC were significantly decreased in the 10-6 mol/L NaF + 2 mmol/L CaCl2 (F+CaII) group. The protein expression levels of Wnt3a, Cyclin D1, cMYC, and ß-catenin were significantly increased in the F group, whereas they were decreased in the F+CaII group. However, the mRNA and protein expression levels of GSK3ß were significantly decreased in the F group while significantly increased in the F+CaII group. In summary, F activated the canonical Wnt/ß-catenin pathway and changed the related gene expression and ß-catenin protein location in OB cells, promoting cell proliferation. Ca2+ supplementation (2 mmol/L) reversed the expression levels of genes and proteins related to the canonical Wnt/ß-catenin pathway.
Assuntos
Cálcio/metabolismo , Fluoretos/efeitos adversos , Osteoblastos/efeitos dos fármacos , Proteínas Wnt/metabolismo , beta Catenina/metabolismo , Animais , Diferenciação Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Suplementos Nutricionais/análise , Glicogênio Sintase Quinase 3 beta/genética , Glicogênio Sintase Quinase 3 beta/metabolismo , Peptídeos e Proteínas de Sinalização Intercelular/genética , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Proteína-5 Relacionada a Receptor de Lipoproteína de Baixa Densidade/genética , Proteína-5 Relacionada a Receptor de Lipoproteína de Baixa Densidade/metabolismo , Masculino , Camundongos , Osteoblastos/classificação , Osteoblastos/metabolismo , Proteínas Wnt/genética , Via de Sinalização Wnt/efeitos dos fármacos , beta Catenina/genéticaRESUMO
BACKGROUND: Drug therapy plays an important role in the treatment of cervical cancer, which is one of the most common solid tumors in women. Therefore, it is important to seek more effective and less toxic therapies. PURPOSE: The aim of this study is to investigate the therapeutic potential of HMQ-T-F5 (1-(4-(2-aminoquinazolin-7-yl)phenyl)-3-(2bromo5-(trifluoromethoxy) phenyl)thiourea) (F5) for cervical cancer and explore the related mechanism. METHODS: By performing MTT assay, colony formation assay, flow cytometry, wound-healing assay, transwell assay, immunofluorescent staining and siRNA assay, we study the effect of F5 on human cervical HeLa cells. The mechanism of F5 was also investigated. RESULTS: We found that F5 significantly inhibited HeLa cell proliferation, led to accumulation of cells in the S phase, and induced apoptosis and inhibited migration. Mechanistically, F5 inhibited HeLa cell growth and migration through repressing the expression and nuclear translocation of ß-catenin, enhancing Axin expression, inhibiting the phosphorylation of LRP5/6 and GSK3ß, as well as downregulating the Wnt downstream targeted proteins. Knockdown of a checkpoint ß-catenin by siRNA significantly attenuated HeLa cell proliferation. Furthermore, XAV939, an inhibitor of ß-catenin, was used to treat HeLa cells and the results demonstrated that F5 inhibited proliferation and migration via the inhibition of the Wnt/ß-catenin pathway. CONCLUSION: Our findings demonstrated that F5 can target ß-catenin potentially and is useful in the treatment of cervical cancer.
Assuntos
Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Tioureia/farmacologia , Via de Sinalização Wnt/efeitos dos fármacos , Apoptose/efeitos dos fármacos , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Regulação para Baixo , Feminino , Glicogênio Sintase Quinase 3 beta/metabolismo , Células HeLa , Humanos , Proteína-5 Relacionada a Receptor de Lipoproteína de Baixa Densidade/metabolismo , Proteína-6 Relacionada a Receptor de Lipoproteína de Baixa Densidade/metabolismo , RNA Interferente Pequeno/farmacologia , Tioureia/análogos & derivados , beta Catenina/metabolismoRESUMO
AIM: The aim of this study was to evaluate the therapeutic effect of a traditional Chinese medicine (TCM), Bushen-Jianpi-Huoxue decoction (BJHD), on diabetic osteoporosis (DOP) and the action mechanisms likely mediated by nuclear factor-kappa B (NF-κB) and Wnt signaling pathways. METHODS: Fifty-five male Wistar rats were used in this study; they were divided into normal control (n = 10) and established DOP model (n = 45) groups. The DOP model was induced using a combination high carbohydrate - high fat diet and intraperitoneal injections of streptozotocin (STZ). The successfully induced animals were randomized to the model, Western medicine, TCM and control groups. Levels of fasting blood glucose; insulin; serum Ca, P and alkaline phosphatase, and the femoral bone mineral density (BMD) were measured. Furthermore, messenger RNA (mRNA) levels of cytokines in the Wnt and NF-κB signaling pathways were measured using reverse transcription-polymerase chain reaction (RT-PCR). RESULTS: Thirty rats were successfully established as the DOP model (10/group). After treatment, the levels of fasting blood glucose, insulin resistance and alkaline phosphatase in the TCM group rats were lower, while P and BMD were higher than those in the model groups. The mRNA levels of cytokines in the Wnt signaling pathway were higher in the TCM group than those in the model group. Moreover, the expressions of factors in the NF-κB pathway were markedly lower in the TCM group than they were in the model group. CONCLUSION: Bushen-Jianpi-Huoxue decoction relieved DOP by activating the Wnt signaling pathway while inhibiting NF-κB signaling.
Assuntos
Conservadores da Densidade Óssea/farmacologia , Densidade Óssea/efeitos dos fármacos , Diabetes Mellitus Experimental/tratamento farmacológico , Medicamentos de Ervas Chinesas/farmacologia , Fêmur/efeitos dos fármacos , Hipoglicemiantes/farmacologia , NF-kappa B/metabolismo , Osteoporose/prevenção & controle , Via de Sinalização Wnt/efeitos dos fármacos , Absorciometria de Fóton , Animais , Biomarcadores/sangue , Glicemia/efeitos dos fármacos , Glicemia/metabolismo , Subunidade alfa 1 de Fator de Ligação ao Core/genética , Subunidade alfa 1 de Fator de Ligação ao Core/metabolismo , Diabetes Mellitus Experimental/sangue , Diabetes Mellitus Experimental/etiologia , Dieta Hiperlipídica , Carboidratos da Dieta , Fêmur/diagnóstico por imagem , Fêmur/metabolismo , Regulação da Expressão Gênica , Insulina/sangue , Resistência à Insulina , Proteína-5 Relacionada a Receptor de Lipoproteína de Baixa Densidade/genética , Proteína-5 Relacionada a Receptor de Lipoproteína de Baixa Densidade/metabolismo , Masculino , NF-kappa B/genética , Osteoporose/sangue , Osteoporose/diagnóstico por imagem , Osteoporose/etiologia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos Wistar , Estreptozocina , Fator 6 Associado a Receptor de TNF/genética , Fator 6 Associado a Receptor de TNF/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , beta Catenina/genética , beta Catenina/metabolismoRESUMO
Failure to achieve optimal bone mass in childhood is the primary cause of decreased adult bone mineral density (BMD) and increased bone fragility in later life. Activating and inactivating LRP5 gene mutations has been associated with extreme bone-related phenotypes. Our aim was to investigate the role of LRP5 polymorphism on BMD, mineral biochemical parameters, and body composition in Iranian children. This cross-sectional study was performed on 9-18 years old children (125 boys, 137 girls). The serum level of calcium, phosphorous, alkaline phosphatase, and vitamin D parameters were checked. The body composition and BMD variables were measured by the Hologic system DXA. The rs566442 (V1119V) coding polymorphism in exon 15 of LRP5 was performed using PCR-RFLP method. Linear regression analysis, with adjustment for age, gender, body size parameters, and pubertal status was used to determine the association between LRP5 polymorphism (rs556442) and bone and body composition parameters. The allele frequency of the rs566442 gene was 35.5 % A and 63.9 % G. Our study revealed that LRP5 (rs556442) has not any significant influence on serum calcium, phosphorus, 25OHvitD, and serum alkaline phosphatase (P > 0.05). Total lean mass was greater in GG genotype (P = 0.028). Total body less head area (P = 0.044), spine BMD (P = 0.04), and total femoral BMC (P = 0.049) were lower in AG heterozygote genotype. This study show LRP5 polymorphism may associate with body composition and BMD in Iranian children. However, further investigations should be done to evaluate the role of other polymorphism.
Assuntos
Composição Corporal/genética , Densidade Óssea/genética , Cálcio/sangue , Homeostase/genética , Proteína-5 Relacionada a Receptor de Lipoproteína de Baixa Densidade/genética , Polimorfismo de Nucleotídeo Único/genética , Absorciometria de Fóton , Fosfatase Alcalina/metabolismo , Biomarcadores/sangue , Criança , Feminino , Frequência do Gene , Genótipo , Humanos , Irã (Geográfico) , Masculino , Minerais/sangue , Fósforo/sangueRESUMO
OBJECTIVE: To investigate the effects of Huogu I formula on regulation of lipid metabolism in steroid-induced osteonecrosis of the femoral head (SONFH) rats and verify our hypothesis that Huogu I formula regulates lipid metabolism by down-regulating peroxisome proliferator-activated receptor gamma (PPARgamma) expression and activating Wnt signaling pathways. METHODS: Eighty-five rats were divided into four groups: control, model, Huogu 15 g/kg and Huogu 30 g/kg. Six weeks later, animals were anaesthetized, femora was dissected for histopathological examination of the osteonecrotic changes and repair processes, micro computed tomography (Micro-CT)-based micro-angiography was performed to assess vascularization. Serum lipid levels were detected by haematological examination. The expressions of PPARy, Wnt3a, low density lipoprotein receptor-related protein 5 (LRP5) and beta-catenin were evaluated by immunohistochemistry, Western blot and quantitative real-time polymerase chain reaction analyses. RESULTS: The incidence of osteonecrosis, ratio of empty lacuna, adipose tissue area and adipocyte perimeter in the bone marrow were dramatically lower in the Huogu I formula treatment groups. By micro-CT quantification, Huogu I formula treatment dose-dependently increased vessel volume, vessel surface, percentage of vessel volume and vessel thickness of the femoral heads of SONFH rats. Levels of serum lipid in Huogu 15 g/kg and Huogu 30 g/kg groups reduced significantly. Huogu I formula treatment could suppress the expression of PPARy and increase the expressions of Wnt3a, LRP5 and beta-catenin at both protein and mRNA levels. CONCLUSION: The results of our present study highlight the lipid-lowering potential of Huogu I formula, and provide further evidence of the involvement of the PPARgamma inhibition and Wnt/LRP5/ beta-catenin signaling activation in the effects of Huogu I formula.
Assuntos
Medicamentos de Ervas Chinesas/administração & dosagem , Proteína-5 Relacionada a Receptor de Lipoproteína de Baixa Densidade/metabolismo , Osteonecrose/tratamento farmacológico , PPAR gama/metabolismo , Transdução de Sinais , Esteroides/efeitos adversos , Proteínas Wnt/metabolismo , beta Catenina/metabolismo , Animais , Regulação para Baixo/efeitos dos fármacos , Humanos , Proteína-5 Relacionada a Receptor de Lipoproteína de Baixa Densidade/genética , Masculino , Osteonecrose/induzido quimicamente , Osteonecrose/genética , Osteonecrose/metabolismo , PPAR gama/genética , Ratos , Ratos Wistar , Proteínas Wnt/genética , beta Catenina/genéticaRESUMO
OBJECTIVE: Macrophages are key players in atherosclerotic lesion formation and progression. We have recently demonstrated that lipid-loaded macrophages show activation of the canonical Wnt signaling pathway. METHODS: To test the in vivo role of the canonical Wnt pathway in atherosclerosis we used mice deficient in the Wnt signaling receptor LRP5 (LRP5(-/-)) fed a hypercholesterolemic diet (HC) to induce atherosclerosis. These dietary groups were further subdivided into two subgroups receiving their respective diets supplemented with 2% plant sterol esters (PSE). All mice remained on their assigned diets until age 18 weeks. RESULTS: HC WT mice had mildly increased non-HDL cholesterol levels, developed aortic atherosclerotic lesions and showed upregulated expression levels of aortic Lrp5. HC LRP5(-/-) mice develop larger aortic atherosclerotic lesions than WT mice indicating that LRP5 has a protective function in atherosclerosis progression. The oral administration of PSE, a dietary cholesterol-lowering agent, had an effect in the expression levels of the Wnt signaling receptor and in atherosclerosis progression. We found that PSE reduced serum total cholesterol levels, abolished HC-induced LRP5 overexpression and reduced aortic atherosclerotic plaques. CONCLUSION: The proatherogenic effects of the excess of plasma lipids are in part mediated by modulation of LRP5 in the aorta. LRP5 and canonical Wnt signaling exert a protective defense mechanism against hyperlipidemia and atherosclerosis lesion progression.
Assuntos
Colesterol na Dieta/farmacologia , Proteína-5 Relacionada a Receptor de Lipoproteína de Baixa Densidade/biossíntese , Fitosteróis/farmacologia , Animais , Aorta/efeitos dos fármacos , Aorta/metabolismo , Aorta/patologia , Aterosclerose/metabolismo , Colesterol/sangue , Suplementos Nutricionais , Hiperlipidemias/prevenção & controle , Proteína-5 Relacionada a Receptor de Lipoproteína de Baixa Densidade/genética , Camundongos Endogâmicos C57BL , Placa Aterosclerótica/prevenção & controle , Via de Sinalização Wnt/fisiologiaRESUMO
Recent studies have shown that Er-Zhi-Wan (EZW), a traditional Chinese medicine consisting of Herba Ecliptae (HE) and Fructus Ligustri Lucidi (FLL), had a definite antiosteoporotic effect on osteoporotic femur, but its effect on osteoporosis of alveolar bone remains unknown. In the present study, we investigated the effects of Er-Zhi-Wan (EZW) on the microarchitecture and the regulation of Wnt/ß-catenin signaling pathway in the alveolar bone of ovariectomized rats. Thirty Sprague-Dawley rats were randomly divided into three groups: sham operation group (sham, n=10), ovariectomy (OVX) group (n=10), and OVX with EZW treatment group (EZW group, n=10). From one week after ovariectomy, EZW (100 mg/mL) or vehicle (distilled water) was fed (1 mL/100 g) once per day for 12 weeks until the sacrifice of the rats. The body weights were measured weekly. After sacrifice, the sera and mandible were collected and routinely prepared for the measurement of alveolar trabecular microarchitecture, serum levels of E2, bone-specific alkaline phosphatase (BALP) and tartrate-resistant acid phosphatase 5b (TRAP5b), as well as mandibular mRNA expression of Wnt/ß-catenin signaling pathway molecules wnt3a, low-density lipoprotein receptor-related protein 5 (LRP5), ß-catenin and dickkopf homolog 1 (DKK1). The results showed that EZW treatment significantly prevented the body weight gain, degradation of alveolar trabecular microarchitecture and alveolar bone loss in the OVX rats. Furthermore, we observed that EZW could increase the serum levels of E2 and BALP, and decrease levels of serum TRAP5b in EZW group compared with vehicle group. In addition, RT-PCR results revealed that EZW upregulated the expression levels of wnt3a, LRP5 and ß-catenin, and reduced the expression of DKK1 in OVX rats. Taken together, our results suggested that EZW may have potential anti-osteoporotic effects on osteoporotic alveolar bone by stimulating Wnt/LRP5/ß-catenin signaling pathway.
Assuntos
Processo Alveolar/efeitos dos fármacos , Medicamentos de Ervas Chinesas/farmacologia , Ovariectomia , Via de Sinalização Wnt/efeitos dos fármacos , Fosfatase Ácida/sangue , Fosfatase Alcalina/sangue , Processo Alveolar/metabolismo , Animais , Peso Corporal/efeitos dos fármacos , Estradiol/sangue , Feminino , Expressão Gênica/efeitos dos fármacos , Peptídeos e Proteínas de Sinalização Intercelular/genética , Isoenzimas/sangue , Proteína-5 Relacionada a Receptor de Lipoproteína de Baixa Densidade/genética , Mandíbula/efeitos dos fármacos , Mandíbula/metabolismo , Medicina Tradicional Chinesa/métodos , Tamanho do Órgão/efeitos dos fármacos , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fosfatase Ácida Resistente a Tartarato , Fatores de Tempo , Regulação para Cima/efeitos dos fármacos , Útero/efeitos dos fármacos , Útero/crescimento & desenvolvimento , Via de Sinalização Wnt/genética , Proteína Wnt3A/genética , beta Catenina/genéticaRESUMO
INTRODUCTION: Wnt ligands bind to low-density lipoprotein receptor-related protein (LRP) 5 or 6, triggering a cascade of downstream events that include ß-catenin signaling. Here we explored the roles of LRP5 in interleukin 1ß (IL-1ß)- or Wnt-mediated osteoarthritic (OA) cartilage destruction in mice. METHODS: The expression levels of LRP5, type II collagen, and catabolic factors were determined in mouse articular chondrocytes, human OA cartilage, and mouse experimental OA cartilage. Experimental OA in wild-type, Lrp5 total knockout (Lrp5â»/â») and chondrocyte-specific knockout (Lrp5fl/fl;Col2a1-cre) mice was caused by aging, destabilization of the medial meniscus (DMM), or intra-articular injection of collagenase. The role of LRP5 was confirmed in vitro by small interfering RNA-mediated knockdown of Lrp5 or in Lrp5â»/â» cells treated with IL-1ß or Wnt proteins. RESULTS: IL-1ß treatment increased the expression of LRP5 (but not LRP6) via JNK and NF-κB signaling. LRP5 was upregulated in human and mouse OA cartilage, and Lrp5 deficiency in mice inhibited cartilage destruction. Treatment with IL-1ß or Wnt decreased the level of Col2a1 and increased those of Mmp3 or Mmp13, whereas Lrp5 knockdown ameliorated these effects. In addition, we found that the functions of LRP5 in arthritic cartilage were subject to transcriptional activation by ß-catenin. Moreover, Lrp5â»/â» and Lrp5fl/fl;Col2a1-cre mice exhibited decreased cartilage destruction (and related changes in gene expression) in response to experimental OA. CONCLUSIONS: Our findings indicate that LRP5 (but not LRP6) plays an essential role in Wnt/ß-catenin-signaling-mediated OA cartilage destruction in part by regulating the expression levels of type II collagen, MMP3, and MMP13.
Assuntos
Artrite Experimental/metabolismo , Cartilagem Articular/patologia , Proteína-5 Relacionada a Receptor de Lipoproteína de Baixa Densidade/metabolismo , Osteoartrite/metabolismo , Via de Sinalização Wnt/fisiologia , Animais , Artrite Experimental/patologia , Western Blotting , Cartilagem Articular/metabolismo , Células Cultivadas , Condrócitos/metabolismo , Condrócitos/patologia , Humanos , Imuno-Histoquímica , Marcação In Situ das Extremidades Cortadas , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Microscopia de Fluorescência , Osteoartrite/patologia , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transfecção , Regulação para Cima , Proteínas Wnt/metabolismoRESUMO
Recent studies have shown that Er-Zhi-Wan (EZW), a traditional Chinese medicine consisting of Herba Ecliptae (HE) and Fructus Ligustri Lucidi (FLL), had a definite antiosteoporotic effect on osteoporotic femur, but its effect on osteoporosis of alveolar bone remains unknown. In the present study, we investigated the effects of Er-Zhi-Wan (EZW) on the microarchitecture and the regulation of Wnt/β-catenin signaling pathway in the alveolar bone of ovariectomized rats. Thirty Sprague-Dawley rats were randomly divided into three groups: sham operation group (sham, n=10), ovariectomy (OVX) group (n=10), and OVX with EZW treatment group (EZW group, n=10). From one week after ovariectomy, EZW (100 mg/mL) or vehicle (distilled water) was fed (1 mL/100 g) once per day for 12 weeks until the sacrifice of the rats. The body weights were measured weekly. After sacrifice, the sera and mandible were collected and routinely prepared for the measurement of alveolar trabecular microarchitecture, serum levels of E2, bone-specific alkaline phosphatase (BALP) and tartrate-resistant acid phosphatase 5b (TRAP5b), as well as mandibular mRNA expression of Wnt/β-catenin signaling pathway molecules wnt3a, low-density lipoprotein receptor-related protein 5 (LRP5), β-catenin and dickkopf homolog 1 (DKK1). The results showed that EZW treatment significantly prevented the body weight gain, degradation of alveolar trabecular microarchitecture and alveolar bone loss in the OVX rats. Furthermore, we observed that EZW could increase the serum levels of E2 and BALP, and decrease levels of serum TRAP5b in EZW group compared with vehicle group. In addition, RT-PCR results revealed that EZW upregulated the expression levels of wnt3a, LRP5 and β-catenin, and reduced the expression of DKK1 in OVX rats. Taken together, our results suggested that EZW may have potential anti-osteoporotic effects on osteoporotic alveolar bone by stimulating Wnt/LRP5/β-catenin signaling pathway.
Assuntos
Animais , Feminino , Fosfatase Ácida , Sangue , Fosfatase Alcalina , Sangue , Processo Alveolar , Metabolismo , Peso Corporal , Medicamentos de Ervas Chinesas , Farmacologia , Estradiol , Sangue , Expressão Gênica , Peptídeos e Proteínas de Sinalização Intercelular , Genética , Isoenzimas , Sangue , Proteína-5 Relacionada a Receptor de Lipoproteína de Baixa Densidade , Genética , Mandíbula , Metabolismo , Medicina Tradicional Chinesa , Métodos , Tamanho do Órgão , Ovariectomia , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fosfatase Ácida Resistente a Tartarato , Fatores de Tempo , Regulação para Cima , Útero , Via de Sinalização Wnt , Genética , Proteína Wnt3A , Genética , beta Catenina , GenéticaRESUMO
BACKGROUND AND AIMS: The therapeutic effects of pulsed electromagnetic fields (PEMFs) on osteoporosis have been documented. However, the precise mechanisms by which PEMFs elicit these favorable biological responses are still not fully understood. This study aimed to systematically investigate the effects of PEMFs on bone mass and Wnt/ß-catenin signaling pathway in ovariectomized rats. METHODS: Thirty 3-month-old female Sprague Dawley rats were randomly assigned to one of three groups: sham-operated control (sham), ovariectomy (OVX), and ovariectomy with PEMFs treatment (PEMFs). One week following ovariectomy surgery, rats in the PEMFs group were exposed to PEMFs for 40 min/day, 5 days/week, for 12 weeks. RESULTS: After 12-week interventions, serum 17ß-estradiol and bone-specific alkaline phosphatase levels increased in the PEMFs group. Bone mineral density of the femur and the fifth lumbar vertebral body also increased in the PEMFs group. Histomorphometrical studies showed that PEMFs improved trabecular area, trabecular width, and trabecular number by 77.50%, 17.38% and 51.06%, respectively, and reduced trabecular separation by 44.28% compared with the OVX group. Biomechanical studies showed that PEMFs increased maximum load and energy to failure in the fifth lumbar vertebral body. Quantitative real-time RT-PCR analysis showed that PEMFs increased the mRNA expressions of Wnt3a, low-density lipoprotein receptor-related protein 5(LRP5), ß-catenin, c-myc and runt-related gene 2 (Runx2), and reduced dickkopf1 (DKK1) in ovariectomized rats. However, mRNA expression of Axin2 was not affected by PEMFs. CONCLUSIONS: PEMFs can prevent ovariectomy-induced bone loss and deterioration of bone microarchitecture and strength, at least partly, through activation of Wnt/ß-catenin signaling pathway.
Assuntos
Densidade Óssea/efeitos da radiação , Terapia por Estimulação Elétrica , Magnetoterapia , Transdução de Sinais/efeitos da radiação , Via de Sinalização Wnt/efeitos da radiação , Absorciometria de Fóton , Fosfatase Alcalina/sangue , Animais , Subunidade alfa 1 de Fator de Ligação ao Core/biossíntese , Subunidade alfa 1 de Fator de Ligação ao Core/genética , Modelos Animais de Doenças , Estradiol/sangue , Feminino , Fêmur/efeitos da radiação , Fêmur/ultraestrutura , Regulação da Expressão Gênica/efeitos da radiação , Peptídeos e Proteínas de Sinalização Intercelular/biossíntese , Peptídeos e Proteínas de Sinalização Intercelular/genética , Proteína-5 Relacionada a Receptor de Lipoproteína de Baixa Densidade/biossíntese , Proteína-5 Relacionada a Receptor de Lipoproteína de Baixa Densidade/genética , Vértebras Lombares/efeitos da radiação , Vértebras Lombares/ultraestrutura , Ovariectomia , Distribuição Aleatória , Ratos , Ratos Sprague-Dawley , Proteína Wnt3A/biossíntese , Proteína Wnt3A/genéticaRESUMO
BACKGROUND: Electroacupuncture (EA) treatment has been shown to increase bone mineral density (BMD) in ovariectomised (OVX) rats; however, the underlying mechanisms remain unclear. OBJECTIVE: To systematically evaluate the effects of EA on OVX rats and the Wnt/ß-catenin signalling pathway. METHODS: Three-month-old female Sprague-Dawley rats were randomly divided into three different groups (n=10 each): sham operated control (sham operated), ovariectomy (OVX) and ovariectomy with EA treatment (OVX+EA). Rats in the OVX+EA group received 12-week EA treatments. RESULTS: Serum bone-specific alkaline phosphatase level (p<0.01), BMD of the proximal femoral metaphysis and the fifth lumbar (L5) vertebral body (both, p<0.05) and maximum load and energy to failure of L5 vertebral body (both p<0.01) were significantly higher in the OVX+EA group than in the OVX group. Trabecular area, trabecular width and trabecular number were significantly higher in the OVX+EA group by 66.9%, 29.2% and 30.3%, respectively, than in the OVX group (all, p<0.01). Trabecular separation was 31.9% lower in the OVX+EA group than in the OVX group (p<0.01). Quantitative real-time reverse transcription polymerised chain reaction indicated that the expressions of mRNAs for low-density lipoprotein receptor-related protein 5 and ß-catenin were significantly increased in the OVX+EA group, as compared with the OVX group (p<0.01 and p<0.05, respectively). CONCLUSION: This study demonstrates that EA can prevent OVX-induced bone loss and deterioration of bone architecture and strength by stimulating the Wnt/ß-catenin signalling pathway. These findings suggest that EA may bet a promising adjunct method for inhibiting OVX-induced osteoporosis in clinical settings.
Assuntos
Fosfatase Alcalina/sangue , Densidade Óssea , Osso e Ossos , Eletroacupuntura , Proteína-5 Relacionada a Receptor de Lipoproteína de Baixa Densidade/metabolismo , Osteoporose Pós-Menopausa/prevenção & controle , beta Catenina/metabolismo , Animais , Osso e Ossos/anatomia & histologia , Osso e Ossos/metabolismo , Modelos Animais de Doenças , Feminino , Fêmur/anatomia & histologia , Fêmur/metabolismo , Humanos , Proteína-5 Relacionada a Receptor de Lipoproteína de Baixa Densidade/genética , Vértebras Lombares/anatomia & histologia , Vértebras Lombares/metabolismo , Osteoporose Pós-Menopausa/genética , Osteoporose Pós-Menopausa/metabolismo , Ovariectomia , RNA Mensageiro/metabolismo , Distribuição Aleatória , Ratos , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase Via Transcriptase Reversa , beta Catenina/genéticaRESUMO
OBJECTIVE: To reveal the mechanism of Zuogui Pill (see text) in treatment of glucocorticoid-induced osteoporosis from the angle of the Wnt signal transduction pathway and to provide further experimental evidence for expounding the scientific connotation of "the kidney dominating the bones" in TCM. METHODS: Forty-two male Wistar rats were selected and randomly divided into three groups, control group (n = 12), model group (n = 15) and Zuogui Pill group (n = 15). Form the beginning, The rats were injected dexamethasone for eight weeks to make the model of osteoporosis, and the Zuogui Pill were administered intragastrically to the rats of Zuogui Pill group for eight weeks. The relative morphological parameters were measured in the undecalcified tibial slices. And the protein expression levels of Wnt1, LRP-5 and beta-catenin in rat tibial osteoblasts (OB) and bone marrow stromal cells (BMC) were detected by immunohistochemistry. RESULTS: Compared with the control group, TBV% and TFS% decreased significantly, while TRS% increased significantly, and the protein expression of Wnt1, LRP-5 and beta-catenin in OB and BMC decreased significantly in the model group. And compared with the model group, TBV% and TFS% increased significantly, and expression levels of Wnt1, LRP-5 and beta-catenin proteins increased significantly in the Zuogui pill group. CONCLUSION: Zuogui Pill can prevent and treat glucocorticoid-induced osteoporosis in rats by up-regulating the expression of the key signal molecules Wnt1, LRP-5 and beta-catenin in Wnt signal transduction pathway.
Assuntos
Medicamentos de Ervas Chinesas/uso terapêutico , Glucocorticoides/farmacologia , Osteoporose/tratamento farmacológico , Transdução de Sinais/efeitos dos fármacos , Proteínas Wnt/fisiologia , Animais , Feminino , Proteínas Relacionadas a Receptor de LDL/fisiologia , Proteína-5 Relacionada a Receptor de Lipoproteína de Baixa Densidade , Medicina Tradicional Chinesa , Ratos , Ratos Wistar , beta Catenina/fisiologiaRESUMO
Tissue transglutaminase (TG2) is a ubiquitously expressed member of the transglutaminase family of Ca(2+)-dependent crosslinking enzymes. Unlike other family members, TG2 is a multifunctional protein, which has several other well documented enzymatic and non-enzymatic functions. A significant body of evidence accumulated over the last decade reveals multiple and complex activities of this protein on the cell surface and in the extracellular matrix (ECM), including its role in the regulation of cell-ECM interactions and outside-in signaling by several types of transmembrane receptors. Moreover, recent findings indicate a dynamic regulation of the levels and functions of extracellular TG2 by several complementary mechanisms. This review summarizes and assesses recent research into the emerging functions and regulation of extracellular TG2.
Assuntos
Matriz Extracelular/metabolismo , Proteínas de Ligação ao GTP/fisiologia , Transglutaminases/fisiologia , Adesão Celular/fisiologia , Reagentes de Ligações Cruzadas/metabolismo , Cisteína/metabolismo , Ativação Enzimática , Fibronectinas/metabolismo , Proteínas de Ligação ao GTP/metabolismo , Humanos , Integrinas/metabolismo , Proteína-5 Relacionada a Receptor de Lipoproteína de Baixa Densidade/fisiologia , Proteína-6 Relacionada a Receptor de Lipoproteína de Baixa Densidade/fisiologia , Macrófagos/fisiologia , Metaloproteinases da Matriz Associadas à Membrana/metabolismo , Transplante de Células-Tronco Mesenquimais , Conformação Proteica/efeitos dos fármacos , Proteína 2 Glutamina gama-Glutamiltransferase , Transporte Proteico/fisiologia , Receptores de Fatores de Crescimento/fisiologia , Transdução de Sinais/fisiologia , Sindecana-4/fisiologia , Transglutaminases/metabolismo , beta Catenina/fisiologiaRESUMO
BACKGROUND: There is a need to develop new bone anabolic agents because current bone regeneration regimens have limitations. The Wingless-type MMTV integration site (Wnt) pathway has emerged as a regulator of bone formation and regeneration. OBJECTIVE: To review the molecular basis for Wnt pathway modulation and discuss strategies that target it and improve bone mass. METHODS: Data in peer-reviewed reports and meeting abstracts are discussed. RESULTS/CONCLUSIONS: Neutralizing inhibitors of Wnt signaling have emerged as promising strategies. Small-molecule inhibitors of glycogen synthase kinase 3beta increase bone mass, lower adiposity and reduce fracture risk. Neutralizing antibodies to Dickkopf 1, secreted Frizzled-related protein 1 and sclerostin produce similar outcomes in animal models. These drugs are exciting breakthroughs but are not without risks. The challenges include tissue-specific targeting and consequently, long-term safety.
Assuntos
Doenças Ósseas/tratamento farmacológico , Osteoblastos/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Proteínas Wnt/antagonistas & inibidores , Proteínas Adaptadoras de Transdução de Sinal , Animais , Doenças Ósseas/fisiopatologia , Proteínas Morfogenéticas Ósseas/antagonistas & inibidores , Proteínas Morfogenéticas Ósseas/fisiologia , Reabsorção Óssea/tratamento farmacológico , Avaliação Pré-Clínica de Medicamentos , Fraturas Ósseas/tratamento farmacológico , Regulação da Expressão Gênica/efeitos dos fármacos , Marcadores Genéticos/fisiologia , Quinase 3 da Glicogênio Sintase/antagonistas & inibidores , Glicogênio Sintase Quinase 3 beta , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/deficiência , Peptídeos e Proteínas de Sinalização Intercelular/genética , Peptídeos e Proteínas de Sinalização Intercelular/fisiologia , Proteínas Relacionadas a Receptor de LDL/deficiência , Proteínas Relacionadas a Receptor de LDL/genética , Proteínas Relacionadas a Receptor de LDL/fisiologia , Proteína-5 Relacionada a Receptor de Lipoproteína de Baixa Densidade , Proteínas de Membrana/antagonistas & inibidores , Proteínas de Membrana/deficiência , Proteínas de Membrana/genética , Proteínas de Membrana/fisiologia , Camundongos , Camundongos Knockout , Especificidade de Órgãos , Osteoblastos/fisiologia , Osteogênese/efeitos dos fármacos , Osteoporose/tratamento farmacológico , Inibidores de Proteínas Quinases/farmacologia , Inibidores de Proteínas Quinases/uso terapêutico , Regeneração/fisiologia , Proteínas Wnt/fisiologia , beta Catenina/biossíntese , beta Catenina/genéticaRESUMO
BACKGROUND: LRP5 is known to have an important relationship with bone density and a variety of other biological processes. Mapping to human chromosome 11q13.2, LRP5 shows considerable evolutionary conservation. Orthologs of this gene exist in many species, although comparison of human LRP5 with other non-human primates has not been performed until now. METHODS: We reported the complementary DNA (cDNA) sequence and deduced amino acid sequence for baboon LRP5, and compared the baboon and human sequences. cDNA sequences for 21 baboons were examined to identify single-nucleotide polymorphisms (SNPs). RESULTS: Sequences of coding regions in human and baboon LRP5 showed 97- 99% homology. Twenty-five SNPs were identified in the coding region of baboon LRP5. CONCLUSION: The observed degree of coding sequence homology in LRP5 led us to expect that the baboon may serve as a useful model for future research into the role(s) of this gene in primate metabolic diseases.
Assuntos
Proteínas Relacionadas a Receptor de LDL/genética , Papio anubis/genética , Papio cynocephalus/genética , Polimorfismo de Nucleotídeo Único , Sequência de Aminoácidos , Animais , Densidade Óssea , Feminino , Humanos , Proteínas Relacionadas a Receptor de LDL/química , Proteína-5 Relacionada a Receptor de Lipoproteína de Baixa Densidade , Masculino , Dados de Sequência Molecular , Alinhamento de Sequência , Análise de Sequência de DNA , Homologia de Sequência de AminoácidosRESUMO
The low density lipoprotein receptor-related protein 5 (LRP5) is a key determinant of bone mass, via the Wnt signaling pathway control of osteoblast function. This study examined human LRP5 signaling and the effects of an intracellular domain single nucleotide polymorphism (SNP: p.V1525A) on osteoblast differentiation and mineralization. Constitutively active LRP5 was constructed by deletion of the extracellular domain of LRP5 (LRP5DeltaN). Expression of LRP5DeltaN-V, which carries the allele p.1525V, induced higher beta-catenin/TCF-LEF activity compared to LRP5DeltaN-A, which carries the allele p.1525A. In a yeast two-hybrid assay, LRP5DeltaN-V also demonstrated a stronger interaction with AXIN than LRP5DeltaN-A. Expression of either of the alleles did not change cell proliferation. However, cells expressing LRP5DeltaN-V showed increased alkaline phosphatase activity and bone nodule formation compared to cells transfected with empty vector or LRP5DeltaN-A after osteogenic supplement (OS: beta-glycerophosphate and l-ascorbic acid) treatment. Cells expressing LRP5DeltaN-V revealed significantly increased bone sialoprotein (BSP) expression after 7 days of OS treatment and maintained elevated expression until day 21. Osteocalcin (OCN) mRNA levels were increased after 14-21 days of OS treatment in LRP5DeltaN-V expressing cells. LRP5DeltaN-V expressing cells demonstrated positive interaction with BMP-2 signaling of transcription at the SBE-luc promoter. LRP5 signaling is affected by the cytoplasmic SNP, p.V1525A. mRNA levels of Runx2 and Osterix were not affected by this SNP.
Assuntos
Diferenciação Celular/genética , Proteínas Relacionadas a Receptor de LDL/genética , Osteoblastos/citologia , Polimorfismo de Nucleotídeo Único , Proteínas Wnt/metabolismo , Fosfatase Alcalina/análise , Fosfatase Alcalina/metabolismo , Alelos , Proteína Axina , Proteína Morfogenética Óssea 2 , Proteínas Morfogenéticas Ósseas/metabolismo , Células Cultivadas , Subunidade alfa 1 de Fator de Ligação ao Core/genética , Subunidade alfa 1 de Fator de Ligação ao Core/metabolismo , Citoplasma/metabolismo , Humanos , Proteínas Relacionadas a Receptor de LDL/metabolismo , Proteína-5 Relacionada a Receptor de Lipoproteína de Baixa Densidade , Osteoblastos/metabolismo , Osteocalcina/genética , Osteocalcina/metabolismo , Estrutura Terciária de Proteína , RNA Mensageiro/análise , RNA Mensageiro/metabolismo , Proteínas Repressoras/metabolismo , Deleção de Sequência , Transdução de Sinais , Fator de Transcrição Sp7 , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Técnicas do Sistema de Duplo-Híbrido , beta Catenina/genética , beta Catenina/metabolismoRESUMO
In humans, low peak bone mass is a significant risk factor for osteoporosis. We report that LRP5, encoding the low-density lipoprotein receptor-related protein 5, affects bone mass accrual during growth. Mutations in LRP5 cause the autosomal recessive disorder osteoporosis-pseudoglioma syndrome (OPPG). We find that OPPG carriers have reduced bone mass when compared to age- and gender-matched controls. We demonstrate LRP5 expression by osteoblasts in situ and show that LRP5 can transduce Wnt signaling in vitro via the canonical pathway. We further show that a mutant-secreted form of LRP5 can reduce bone thickness in mouse calvarial explant cultures. These data indicate that Wnt-mediated signaling via LRP5 affects bone accrual during growth and is important for the establishment of peak bone mass.