RESUMO
Membrane-type metalloproteinases (including MMP-14 and MMP-15) are enzymes involved in the degradation of extracellular matrix components. In cancer, they are involved in processes such as cellular invasion, angiogenesis and metastasis. Therefore, the aim of this study was to evaluate the expression, content and activity of MMP-14 and MMP-15 in human renal cell carcinoma. Samples of healthy kidney tissue (n = 20) and tissue from clear-cell kidney cancer (n = 20) were examined. The presence and contents of the MMPs were assessed using Western blot and ELISA techniques, respectively. Their activity-both actual and specific-was evaluated using fluorimetric analysis. Both control and cancer human kidney tissues contain MMP-14 and MMP-15 enzymes in the form of high-molecular-weight complexes. Moreover, these enzymes occur in both active and latent forms. Their content in cancer tissues is very similar, but with a noteworthy decrease in content with an increase in the kidney cancer grade for both membrane-type metalloproteinases. Even more notable is the highest content of the investigated enzymes represented by MMP-14 in the control tissues. Considering the actual and specific activity outcomes, MMP-14 dominates over MMP-15 in all of the investigated tissues. Nevertheless, we also noted a significant enhancement of the activity of both metalloproteinases with an increase in the grade of renal cancer. The expression and activity of both enzymes were detected in all examined renal cancer tissues. However, our findings suggest that transmembrane metalloproteinase 14 (MMP-14) plays a much more significant and essential role than MMP-15 in the studied renal carcinoma tissues. Therefore, it seems that MMP-14 could be a promising target in the diagnosis, prognosis and therapy of renal cell carcinoma.
Assuntos
Carcinoma de Células Renais , Neoplasias Renais , Metaloproteinase 14 da Matriz , Metaloproteinase 15 da Matriz , Humanos , Metaloproteinase 14 da Matriz/metabolismo , Neoplasias Renais/patologia , Neoplasias Renais/metabolismo , Neoplasias Renais/enzimologia , Carcinoma de Células Renais/metabolismo , Carcinoma de Células Renais/patologia , Carcinoma de Células Renais/enzimologia , Metaloproteinase 15 da Matriz/metabolismo , Metaloproteinase 15 da Matriz/genética , Feminino , Masculino , Pessoa de Meia-Idade , Idoso , AdultoRESUMO
Here, we delineate the phenotype of two siblings with a bi-allelic frameshift variant in MMP15 gene with congenital cardiac defects, cholestasis, and dysmorphism. Genome sequencing analysis revealed a recently reported homozygous frameshift variant (c.1058delC, p.Pro353Glnfs*102) in MMP15 gene that co-segregates with the phenotype in the family in a recessive mode of inheritance. Relative quantification of MMP15 mRNA showed evidence of degradation of the mutated transcript, presumably by nonsense mediated decay. Likewise, MMP15: p.Gly231Arg, a concurrently reported homozygous missense variant in another patient exhibiting a similar phenotype, was predicted to disrupt zinc ion binding to the MMP-15 enzyme catalytic domain, which is essential for substrate proteolysis, by structural modeling. Previous animal models and cellular findings suggested that MMP15 plays a crucial role in the formation of endocardial cushions. These findings confirm that MMP15 is an important gene in human development, particularly cardiac, and that its loss of function is likely to cause a severe disorder phenotype.
Assuntos
Colestase , Cardiopatias Congênitas , Icterícia , Metaloproteinase 15 da Matriz/genética , Animais , Insuficiência de Crescimento/genética , Cardiopatias Congênitas/genética , Homozigoto , Humanos , FenótipoRESUMO
Aim: To first explore the prognostic value of MMP11 and MMP15 in hepatocellular carcinoma. Methods: MMP11/MMP15 expression was immunohistochemically detected and correlated with clinicopathologic variables and survival and confirmed in publicly available databases. An MMP-based risk score (MMPRS) was established. Results: Tumoral MMP11/MMP15 expression was higher and univariately associated with crucial clinicopathologic parameters, overall survival and disease-free survival in all patients and/or many subsets. Multivariately, MMP11/MMP15 expression remained significant. Their overexpression and prognostic value were confirmed in the Ualcan and Kaplan-Meier plotter databases. Critically, the novel MMPRS integrating MMP11, MMP15 and tumor-node-metastasis stage identified subgroups with the best and worst prognoses, with much higher predictive power. Conclusion: MMP11 and MMP15 served as prognosticators in hepatocellular carcinoma. MMPRS might work more accurately.
MMP11 and MMP15, involved in cancer dissemination, were found to have important biological functions in several cancers. However, their prognostic value in hepatocellular carcinoma (HCC) remains unknown. In the present study, it was found that MMP11 and MMP15 were overexpressed and predictive of the outcome of HCC. Moreover, the novel MMP-based risk score integrating MMP11, MMP15 and tumornodemetastasis stage had much higher prognostic power. MMP11, MMP15 and especially the MMP-based risk score were identified as promising indicators of prognosis in HCC.
Assuntos
Carcinoma Hepatocelular , Neoplasias Hepáticas , Biomarcadores Tumorais/metabolismo , Carcinoma Hepatocelular/diagnóstico , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/cirurgia , Humanos , Estimativa de Kaplan-Meier , Neoplasias Hepáticas/diagnóstico , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/cirurgia , Metaloproteinase 11 da Matriz/metabolismo , Metaloproteinase 15 da Matriz , Prognóstico , Fatores de RiscoRESUMO
Adequate anchoring of the placenta in the uterus through invasion of first trimester cytotrophoblasts (CTB) is required for a successful pregnancy. This process is mediated by matrix metalloproteinases (MMPs) and regulated by the maternal environment. Obesity is known to alter the intrauterine milieu and has been related to impaired invasion. We hypothesized that placental MMP15, a novel membrane-type MMP, is involved in CTB invasion and regulated by maternal obesity in early pregnancy. Thus, in this study MMP15 was immunolocalized to invasive extravillous and interstitial CTB. MMP15 silencing in chorionic villous explants using two different siRNAs reduced trophoblast outgrowth length (-35%, P ≤ .001 and -26%, P < .05) and area (-43%, P ≤ .001 and -36%, P ≤ .01) without altering trophoblast proliferation or apoptosis. Short-term treatment of primary first trimester trophoblasts with IL-6 (10 ng/mL), interleukin 10 (IL-10) (50 ng/mL), and tumor necrosis factor α (TNF-α) (10 ng/mL) did not affect MMP15 protein levels. Likewise, MMP15 mRNA and protein levels were unaltered between human first trimester placentas from control pregnancies vs those complicated with maternal obesity. Overall, our results suggest that the role of MMP15 in placental development and function in early pregnancy is limited to CTB invasion without being affected by short- and long-term inflammation.
Assuntos
Movimento Celular/fisiologia , Metaloproteinase 15 da Matriz/metabolismo , Obesidade Materna/metabolismo , Primeiro Trimestre da Gravidez/metabolismo , Trofoblastos/metabolismo , Trofoblastos/fisiologia , Adulto , Apoptose/fisiologia , Proliferação de Células/fisiologia , Células Cultivadas , Feminino , Humanos , Interleucina-10/metabolismo , Interleucina-6/metabolismo , Masculino , Placenta/metabolismo , Placenta/fisiologia , Gravidez , RNA Mensageiro/metabolismo , RNA Interferente Pequeno/metabolismo , Fator de Necrose Tumoral alfa/metabolismoRESUMO
BACKGROUND: Relapses and flares with delayed wound healing are among the main symptoms of systemic lupus erythematosus (SLE), a rheumatic autoimmune disease. The orientation of immune responses in SLE disease depends on the function of the population of macrophages. This study investigated the effect of indole-3-carbinol (I3C) on transcriptional profiling of macrophage-derived monocytes (MDMs) in four stages of the wound-healing process. METHODS: In the first phase of study, MDMs were generated from peripheral blood mononuclear cells of three new SLE cases (unmedicated) and two healthy controls. The cases and controls were then divided into I3C treated and untreated groups after 24 hours of exposure to I3C. Single-end RNA sequencing was performed using an Illumina NextSeq 500 platform. After comprehensive analysis among differentially expressed genes, CDKN1A, FN1 and MMP15 were validated by quantitative real-time polymerase chain reaction as upregulated ranked genes involved in wound-healing stages. RESULTS: The RNA sequencing analysis of treated cases and treated controls versus untreated cases and untreated controls (group 3 vs. group 4) revealed upregulation of various genes, for example: C1S, C1R, IGKV1-5, IGKV4-1, SERPING1, IGLC1 and IGLC2 in coagulation; ADAM19, CEACAM1 and CEACAM8 in M2 reprogramming; IRS1, FN1, THBS1 and LIMS2 in extracellular matrix organization; and STAT1, THBS1 and ATP2A3 in the proliferation stage of wound healing. CONCLUSIONS: The results showed that treatment with I3C could modulate the gene expression involved in wound healing in SLE cases and healthy controls.
Assuntos
Indóis/farmacologia , Lúpus Eritematoso Sistêmico/tratamento farmacológico , Lúpus Eritematoso Sistêmico/genética , Macrófagos/efeitos dos fármacos , Cicatrização/efeitos dos fármacos , Adulto , Estudos de Casos e Controles , Inibidor de Quinase Dependente de Ciclina p21/genética , Feminino , Fibronectinas/genética , Perfilação da Expressão Gênica , Humanos , Indóis/uso terapêutico , Macrófagos/metabolismo , Masculino , Metaloproteinase 15 da Matriz/genética , Pessoa de Meia-Idade , Análise de Sequência de RNA , Transdução de Sinais/efeitos dos fármacos , Regulação para CimaRESUMO
Using medaka, we found that in vitro follicle ovulation, but not germinal vesicle breakdown, was inhibited by three gap junction blockers, carbenoxolone, mefloquine, and flufenamic acid. The blockers specifically inhibited follicular expression of matrix metalloproteinase-15 mRNA and the protein (mmp15/Mmp15), a protease indispensable for medaka ovulation, indicating that gap junctional communication may be required for successful ovulation and mmp15/Mmp15 expression. Further experiments using carbenoxolone as the representative of the gap junction blockers showed that expression of nuclear progestin receptor (Pgr), a transcription factor required for mmp15 expression, was not affected by carbenoxolone treatment, but the formation of phosphorylated Pgr was considerably suppressed. Carbenoxolone treatment caused a decrease in the Pgr binding to the promoter region of mmp15. mRNA expression of cyclin-dependent protein kinase-9 (cdk9) and cyclin I (ccni), whose translation products are demonstrated to be involved in Pgr phosphorylation in the medaka ovulating follicles, was suppressed by carbenoxolone treatment. Transcripts of connexin 34.5 (cx34.5) and connexin 35.4 (cx35.4) were dominantly expressed in the follicle cells of ovulating follicles. The results indicate that gap junctional communication plays an important role in medaka ovulation.
Assuntos
Disruptores Endócrinos/farmacologia , Junções Comunicantes/efeitos dos fármacos , Hormônio Luteinizante/farmacologia , Metaloproteinase 15 da Matriz/genética , Oryzias/fisiologia , Ovulação/efeitos dos fármacos , Animais , Carbenoxolona/farmacologia , Feminino , Ácido Flufenâmico/farmacologia , Junções Comunicantes/metabolismo , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Metaloproteinase 15 da Matriz/efeitos dos fármacos , Metaloproteinase 15 da Matriz/metabolismo , Mefloquina/farmacologia , Folículo Ovariano/efeitos dos fármacos , Folículo Ovariano/metabolismo , Ovulação/genética , Ativação Transcricional/efeitos dos fármacosRESUMO
Cervical cancer is the second most frequent type of gynecologic cancer worldwide. Prokineticin 2 (PROK2) is reported to be involved in tumor progression in some malignant tumors. However, the role of PROK2 in the development of cervical cancer remains unknown. Our results indicate that PROK2 is overexpressed in the human cervical cancer. Cervical cancer patients with high PROK2 expression have a shorter overall survival rate (OS) and disease-free survival rate (DFS). PROK2 acts as a potential biomarker for predicting OS and DFS of cervical cancer patients. We further show that PROK2 is important factor for oncogenic migration and invasion in human cervical cancer cells. Knockdown PROK2 significantly inhibited cell migration, invasion, and MMP15 protein expression in HeLa cells. High expression of MMP15 is confirmed in the human cervical cancer, is significantly associated with the shorter overall survival rate (OS) and is correlated with PROK2 expression. Overexpression of PROK2 using PROK2 plasmid significantly reverses the function of knockdown PROK2, and further upregulates MMP15 expression, migration and invasion of human cervical cancer cells. In conclusion, our findings are the first to demonstrate the role of PROK2 as a novel and potential biomarker for clinical use, and reveal the oncogenic functions of PROK2 as therapeutic target for cervical cancer.
Assuntos
Biomarcadores Tumorais/metabolismo , Hormônios Gastrointestinais/metabolismo , Regulação Neoplásica da Expressão Gênica , Metaloproteinase 15 da Matriz/metabolismo , Neuropeptídeos/metabolismo , Neoplasias do Colo do Útero/patologia , Apoptose , Biomarcadores Tumorais/genética , Ciclo Celular , Movimento Celular , Proliferação de Células , Feminino , Hormônios Gastrointestinais/antagonistas & inibidores , Hormônios Gastrointestinais/genética , Humanos , Metaloproteinase 15 da Matriz/química , Metaloproteinase 15 da Matriz/genética , Invasividade Neoplásica , Neuropeptídeos/antagonistas & inibidores , Neuropeptídeos/genética , Prognóstico , Células Tumorais Cultivadas , Neoplasias do Colo do Útero/genética , Neoplasias do Colo do Útero/metabolismoRESUMO
BACKGROUND: Gingival hyperplasia could occur after the administration of cyclosporine A. Up to 90% of the patients submitted to immunosuppressant drugs have been reported to suffer from this side effect. The role of fibroblasts in gingival hyperplasia has been widely discussed by literature, showing contrasting results. In order to demonstrate the effect of cyclosporine A on the extracellular matrix component of fibroblasts, we investigated the gene expression profile of human fibroblasts after cyclosporine A administration. MATERIALS AND METHODS: Primary gingival fibroblasts were stimulated with 1000 ng/mL cyclosporine A solution for 16 h. Gene expression levels of 57 genes belonging to the "Extracellular Matrix and Adhesion Molecules" pathway were analyzed using real-time PCR in treated cells, compared to untreated cells used as control. RESULTS: Expression levels of different genes were significantly de-regulated. The gene CDH1, which codes for the cell adhesion protein E-cadherin, showed up-regulation. Almost all the extracellular matrix metalloproteases showed down-regulation (MMP8, MMP11, MMP15, MMP16, MMP24, MMP26). The administration of cyclosporine A was followed by down-regulation of other genes: COL7A1, the transmembrane receptors ITGB2 and ITGB4, and the basement membrane constituents LAMA2 and LAMB1. CONCLUSION: Data collected demonstrate that cyclosporine inhibits the secretion of matrix proteases, contributing to the accumulation of extracellular matrix components in the gingival connective tissue, causing gingival overgrowth. Patients affected by gingival overgrowth caused by cyclosporine A need to be further investigated in order to determine the role of this drug on fibroblasts.
Assuntos
Ciclosporina/farmacologia , Ciclosporina/uso terapêutico , Gengiva/efeitos dos fármacos , Hiperplasia Gengival/tratamento farmacológico , Células Cultivadas , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Gengiva/metabolismo , Hiperplasia Gengival/metabolismo , Humanos , Metaloproteinase 11 da Matriz/metabolismo , Metaloproteinase 15 da Matriz/metabolismo , Metaloproteinase 16 da Matriz/metabolismo , Metaloproteinase 8 da Matriz/metabolismo , Metaloproteinases da Matriz Associadas à Membrana/metabolismo , Metaloproteinases da Matriz Secretadas/metabolismoRESUMO
Matrix metalloproteinases (MMPs), a family of zinc-dependent endopeptidases, are involved in a variety of physiological and pathological processes. We analyzed 11 data sets from Gene Expression Omnibus Database and found that MMP7 and MMP15 were highly expressed in multiple carcinomas. GSE13204 showed that MMP7 and MMP15 were overexpressed in acute myeloid leukemia (AML) patients. The Cancer Genome Atlas data set exhibited that high expression of MMP7 or MMP15 in bone marrow (BM) of AML patients predicted poor overall survival. The χ 2 test results indicated that high expression level of MMP7 and MMP15 were correlated with high-risk stratification and high BM blast cell percentage in AML patients. To confirm these findings, we performed reverse-transcription quantitative polymerase chain reaction (RT-qPCR) and found that MMP7 and MMP15 were highly expressed in three AML cell lines. Further study showed that MMP7 and MMP15 were highly expressed both in BM and peripheral blood in collected AML samples compared with healthy individuals. Additionally, long noncoding RNA (lncRNA) microarray of BM samples of AML patients revealed that multiple lncRNAs were correlated with MMP7 and MMP15, suggesting that lncRNAs might be involved in the pathogenesis of AML via modulating MMPs. In conclusion, our study uncovers the potential roles of MMP7 and MMP15 in the prognosis of AML.
Assuntos
Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/mortalidade , Metaloproteinase 15 da Matriz/genética , Metaloproteinase 7 da Matriz/genética , Adolescente , Estudos de Casos e Controles , Linhagem Celular Tumoral , Criança , Feminino , Regulação Enzimológica da Expressão Gênica , Regulação Leucêmica da Expressão Gênica , Humanos , Leucemia Mieloide Aguda/patologia , Masculino , Prognóstico , Modelos de Riscos Proporcionais , Mapas de Interação de Proteínas , RNA Longo não Codificante/genéticaRESUMO
Cadherin-based intercellular adhesions are essential players in epithelial homeostasis, but their dynamic regulation during tissue morphogenesis and remodeling remain largely undefined. Here, we characterize an unexpected role for the membrane-anchored metalloproteinase MT2-MMP in regulating epithelial cell quiescence. Following co-immunoprecipitation and mass spectrometry, the MT2-MMP cytosolic tail was found to interact with the zonula occludens protein-1 (ZO-1) at the apical junctions of polarized epithelial cells. Functionally, MT2-MMP localizes in the apical domain of epithelial cells where it cleaves E-cadherin and promotes epithelial cell accumulation, a phenotype observed in 2D polarized cells as well as 3D cysts. MT2-MMP-mediated cleavage subsequently disrupts apical E-cadherin-mediated cell quiescence resulting in relaxed apical cortical tension favoring cell extrusion and re-sorting of Src kinase activity to junctional complexes, thereby promoting proliferation. Physiologically, MT2-MMP loss of function alters E-cadherin distribution, leading to impaired 3D organoid formation by mouse colonic epithelial cells ex vivo and reduction of cell proliferation within intestinal crypts in vivo Taken together, these studies identify an MT2-MMP-E-cadherin axis that functions as a novel regulator of epithelial cell homeostasis in vivo.
Assuntos
Caderinas/metabolismo , Homeostase/fisiologia , Mucosa Intestinal/metabolismo , Metaloproteinase 15 da Matriz/metabolismo , Junções Aderentes/metabolismo , Caderinas/genética , Movimento Celular/fisiologia , Proteínas do Citoesqueleto/metabolismo , Células Epiteliais/metabolismo , Humanos , Junções Intercelulares/metabolismo , Junções Íntimas/metabolismoRESUMO
During late embryogenesis, mammary epithelial cells initiate migration programs that drive ductal invasion into the surrounding adipose-rich mesenchyme. Currently, branching morphogenesis is thought to depend on the mobilization of the membrane-anchored matrix metalloproteinases MMP14 (MT1-MMP) and MMP15 (MT2-MMP), which drive epithelial cell invasion by remodeling the extracellular matrix and triggering associated signaling cascades. However, the roles that these proteinases play during mammary gland development in vivo remain undefined. Here, we characterize the impact of global Mmp14 and Mmp15 targeting on early postnatal mammary gland development in mice. Unexpectedly, both Mmp14-/- and Mmp15-/- mammary glands retain the ability to generate intact ductal networks. Although neither proteinase is required for branching morphogenesis, transcriptome profiling reveals a key role for MMP14 and MMP15 in regulating mammary gland adipocyte differentiation. Whereas MMP14 promotes the generation of white fat depots crucial for energy storage, MMP15 differentially controls the formation of thermogenic brown fat. Taken together, these data not only indicate that current paradigms relevant to proteinase-dependent morphogenesis need be revisited, but also identify new roles for the enzymes in regulating adipocyte fate determination in the developing mammary gland.
Assuntos
Glândulas Mamárias Animais/crescimento & desenvolvimento , Metaloproteinase 14 da Matriz/fisiologia , Metaloproteinase 15 da Matriz/fisiologia , Morfogênese/genética , Adipócitos/fisiologia , Adipogenia/genética , Animais , Animais Recém-Nascidos , Diferenciação Celular/genética , Metabolismo Energético/genética , Feminino , Metaloproteinase 14 da Matriz/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Termogênese/genéticaRESUMO
The direct role of melatonin in mammary glands of dairy goats has remained obscure. This study aimed to evaluate the expression of melatonin membrane receptors (MT1 and MT2) in the pituitary and mammary glands of dairy goats during lactation, and to investigate the role of melatonin in mammary function. Both MT1 and MT2 were consistently expressed in the pituitary and mammary eight glands throughout the lactation period, and their levels were lower in 9 March (group I), June (group III), and September (group V) than in May (group II) and August (group IV). The expression patterns of pituitary and mammary MT1 and MT2 were consistent with those of blood melatonin during lactation. Furthermore, the mammary prolactin (PRL), and pituitary growth hormone (GH) and PRL mRNA expression showed an inverse trend in relation to blood melatonin levels. In mammary tissues, MT1 and MT2 immunoreactivity was predominantly located in the mammary epithelial cells (MECs). In addition, a dose- and time-dependent inhibition on cell viability was observed in cultured MECs. At the dose of 10 and 100 pg/ml, melatonin decreased mammary ß-casein and PRL expression. Furthermore, the inhibitory effects of melatonin were blocked by luzindole, a nonselective MT1 and MT2 receptor antagonist. In addition, melatonin promoted MT1 and MT2 expression in cultured MECs. In conclusion, the presence of MT1 and MT2 in the pituitary and mammary glands and the inhibitory effects of melatonin on cell viability, ß-casein, and PRL expression in MECs suggest the potential regulation by melatonin in goat mammary function.
Assuntos
Cabras/fisiologia , Lactação/efeitos dos fármacos , Glândulas Mamárias Animais/efeitos dos fármacos , Melatonina/farmacologia , Animais , Caseínas/genética , Caseínas/metabolismo , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular , Feminino , Regulação da Expressão Gênica , Metaloproteinase 14 da Matriz/genética , Metaloproteinase 14 da Matriz/metabolismo , Metaloproteinase 15 da Matriz/genética , Metaloproteinase 15 da Matriz/metabolismo , Prolactina/genética , Prolactina/metabolismoRESUMO
Non-small-cell carcinoma (NSCLC) is the most common cancer along with high mortality rate worldwide. In the present study, our data showed that lncRNA MAF BZIP Transcription Factor G Antisense RNA 1 (MAFG-AS1) was over-expressed in NSCLC tissues and cell lines. Overexpression of MAFG-AS1 promoted the migration, invasion and enhanced epithelial-mesenchymal transition (EMT) of NSCLC cell. In addition, miR-339-5p was predicted to be a target of MAFG-AS1 and the level of miR-339-5p was down-regulated in NSCLC. Over-expression of MAFG-AS1 significantly decreased the level of miR-339-5p in NSCLC cell. Moreover, the matrix metalloproteinase 15 (MMP15) was identified to be a target of miR-339-5p. The level of MMP15 was negatively regulated by miR-339-5p whereas positively controlled by MAFG-AS1. In addition, up-regulation of miR-339-5p neutralized the promoting impact of MAFG-AS1 on the migration, invasion and EMT of NSCLC cell. Finally, the xenograft model suggested that MAFG-AS1 promoted the metastasis of NSCLC cell in vivo. Altogether, we proved that MAFG-AS1-miR-339-5p-MMP15 axis might be a promising therapeutic target for the treatment of patients with NSCLC.
Assuntos
Carcinoma Pulmonar de Células não Pequenas/genética , Neoplasias Pulmonares/genética , Metaloproteinase 15 da Matriz/metabolismo , MicroRNAs/metabolismo , RNA Antissenso/metabolismo , Animais , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Carcinoma Pulmonar de Células não Pequenas/patologia , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Transição Epitelial-Mesenquimal/genética , Xenoenxertos , Humanos , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patologia , Fator de Transcrição MafG/genética , Fator de Transcrição MafG/metabolismo , Metaloproteinase 15 da Matriz/genética , Camundongos , Camundongos Endogâmicos BALB C , MicroRNAs/genética , Invasividade Neoplásica , RNA Antissenso/genética , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismoRESUMO
Embryo development block seriously limits the success of in vitro embryo production and assisted reproductive technology. Although numerous researchers have explored this problem, it remains to be solved. In this study, we found that melatonin supplementation at 10-8 and 10-9 M in M16 significantly reduced two-cell block of mouse embryos. When those melatonin-treated four-cell embryos were transplanted into the oviducts of female recipient mice, the litter sizes were significantly increased compared with those of the controls. Mechanism study discovered that melatonin treatment markedly reduced reactive oxygen species and mitochondrial superoxide. Quantitative polymerase chain reaction revealed that melatonin significantly upregulated the transcription of catalase, superoxide dismutase 2, glutathione peroxidase, and the antiapoptotic factors Bcl-2 and Bcl-x while downregulated the transcription of pro-apoptotic genes p53 and Bax. In addition, we found Dux, an important gene which promotes zygotic genome activation, and zygotic genes (zinc finger and SCAN4B and eukaryotic translation initiation factor 1A) were all increased after melatonin treatment. Melatonin membrane receptors have two isoforms, melatonin receptor 1 and 2 (MT1, MT2). Further studies with luzindole (a nonselective MT1 and MT2 antagonist) demonstrated that the beneficial effects of melatonin on reducing two-cell block were not mediated by the melatonin membrane receptors. This study shows that melatonin can be used for improving the embryo quality and production efficiency cultured in vitro and also identifies the underlying mechanism by which melatonin decreases two-cell block.
Assuntos
Melatonina/farmacologia , Animais , Antioxidantes/metabolismo , Feminino , Glutationa Peroxidase/metabolismo , Metaloproteinase 14 da Matriz/metabolismo , Metaloproteinase 15 da Matriz/metabolismo , Melatonina/antagonistas & inibidores , Camundongos , Superóxido Dismutase/metabolismo , Triptaminas/farmacologiaRESUMO
STUDY QUESTION: Does endothelin-1 (ET-1) regulate matrix metalloproteinase (MMP) 14 and 15 production and invasion of human first trimester trophoblasts? SUMMARY ANSWER: ET-1 in pathophysiological concentrations down-regulates MMP14 and MMP15 expression via endothelin receptor (ETR) type B and decreases trophoblast migration and invasion. WHAT IS KNOWN ALREADY: MMP14 and MMP15 are involved in trophoblast invasion. Impairment of invasion has been linked to pregnancy complications such as pre-eclampsia (PE). ET-1 is up-regulated in PE. STUDY DESIGN, SIZE, DURATION: In vitro study using primary human trophoblasts from 50 first trimester placentas (gestational week 7-12). PARTICIPANTS/MATERIALS, SETTING, METHODS: Trophoblasts were cultured in the absence or presence of 10-100 nM ET-1. MMP14 and MMP15 mRNA and protein were quantified by RT-qPCR and Western blotting, respectively. Selective antagonists for ETRA (BQ-123) or ETRB (BQ-788) were used to identify ETR subtypes involved. Functional ET-1 effects were tested in first trimester chorionic villous explants and transwell invasion assays. The roles of tumor necrosis factor (TNF)-α (25 ng/ml) and oxygen (1%) in ET-1 regulation of MMP14 and 15 expression were assessed by Western blotting. MAIN RESULTS AND THE ROLE OF CHANCE: ET-1 down-regulated MMP14 and MMP15 mRNA (-21% and -26%, respectively, P < 0.05) and protein levels (-18% and -22%, respectively, P < 0.05). This effect was mediated via ETRB. ET-1 decreased trophoblast outgrowth in placental explants (-24%, P < 0.05) and trophoblast invasion (-26%, P ≤ 0.01). TNF-α enhanced ET-1 mediated MMP15 down-regulation (by 10%, P < 0.05), whereas hypoxia abolished the effect of ET-1 on both MMPs. LARGE SCALE DATA: N/A. LIMITATIONS, REASONS FOR CAUTION: Only primary trophoblasts were used in this study. Since trophoblast yield from first trimester placental material is limited, further aspects of MMP14 and 15 regulation could not be characterized. Other anti-invasive factors may be altered by ET-1 in trophoblasts and, thus, contribute to the reduced invasion, but have not been investigated. Oxygen levels similar to those found in the decidua (5-8% O2) were not analyzed in this study. WIDER IMPLICATIONS OF THE FINDINGS: ET-1 modifies placental function already during the first trimester of pregnancy, the time-window when the placental changes implicated in PE occur. Thus, our results improve the understanding of the placental mechanisms underlying trophoblast invasion and PE. STUDY FUNDING/COMPETING INTERESTS: The study was funded by the Oesterreichische Nationalbank (Anniversary Fund, project number: 14796) and the Herzfelder'sche Familienstiftung (to J.P.; number: 00685). AMM received funding from the Austrian Science Fund FWF (W1241) and the Medical University Graz through the PhD Program Molecular Fundamentals of Inflammation (DK-MOLIN). The authors have no conflict of interest.
Assuntos
Regulação para Baixo/efeitos dos fármacos , Endotelina-1/farmacologia , Metaloproteinase 14 da Matriz/metabolismo , Metaloproteinase 15 da Matriz/metabolismo , Receptor de Endotelina B/metabolismo , Trofoblastos/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Relação Dose-Resposta a Droga , Feminino , Humanos , Metaloproteinase 14 da Matriz/genética , Metaloproteinase 15 da Matriz/genética , Placenta/efeitos dos fármacos , Placenta/metabolismo , Gravidez , Primeiro Trimestre da Gravidez/metabolismo , Receptor de Endotelina B/genética , Trofoblastos/metabolismo , Fator de Necrose Tumoral alfa/farmacologiaRESUMO
Melatonin (MLT; N-acetyl-5-methoxytryptamine) exhibits analgesic properties in chronic pain conditions. While researches linking MLT to epigenetic mechanisms have grown exponentially over recent years, very few studies have investigated the contribution of MLT-associated epigenetic modification to pain states. Here, we report that together with behavioral allodynia, spinal nerve ligation (SNL) induced a decrease in the expression of catalytic subunit of phosphatase 2A (PP2Ac) and enhanced histone deacetylase 4 (HDAC4) phosphorylation and cytoplasmic accumulation, which epigenetically alleviated HDAC4-suppressed hmgb1 gene transcription, resulting in increased high-mobility group protein B1 (HMGB1) expression selectively in the ipsilateral dorsal horn of rats. Focal knock-down of spinal PP2Ac expression also resulted in behavioral allodynia in association with similar protein expression as observed with SNL. Notably, intrathecal administration with MLT increased PP2Ac expression, HDAC4 dephosphorylation and nuclear accumulation, restored HDAC4-mediated hmgb1 suppression and relieved SNL-sensitized behavioral pain; these effects were all inhibited by spinal injection of 4P-PDOT (a MT2 receptor antagonist, 30 minutes before MLT) and okadaic acid (OA, a PP2A inhibitor, 3 hr after MLT). Our findings demonstrate a novel mechanism by which MLT ameliorates neuropathic allodynia via epigenetic modification. This MLT-exhibited anti-allodynia is mediated by MT2-enhanced PP2Ac expression that couples PP2Ac with HDAC4 to induce HDAC4 dephosphorylation and nuclear import, herein increases HDAC4 binding to the promoter of hmgb1 gene and upregulates HMGB1 expression in dorsal horn neurons.
Assuntos
Histona Desacetilases/metabolismo , Hiperalgesia/metabolismo , Metaloproteinase 15 da Matriz/metabolismo , Melatonina/farmacologia , Proteína Fosfatase 2/metabolismo , Corno Dorsal da Medula Espinal/metabolismo , Transcrição Gênica/efeitos dos fármacos , Animais , Proteína HMGB1/biossíntese , Hiperalgesia/patologia , Masculino , Ratos , Corno Dorsal da Medula Espinal/patologiaRESUMO
Internal tandem duplication (ITD) of the fms-related tyrosine kinase-3 (FLT3) gene occurs in 30% of acute myeloid leukemias (AMLs) and confers a poor prognosis. Thirteen relapsed or chemo-refractory FLT3-ITD(+) AML patients were treated with sorafenib (200-400 mg twice daily). Twelve patients showed clearance or near clearance of bone marrow myeloblasts after 27 (range 21-84) days with evidence of differentiation of leukemia cells. The sorafenib response was lost in most patients after 72 (range 54-287) days but the FLT3 and downstream effectors remained suppressed. Gene expression profiling showed that leukemia cells that have become sorafenib resistant expressed several genes including ALDH1A1, JAK3, and MMP15, whose functions were unknown in AML. Nonobese diabetic/severe combined immunodeficiency mice transplanted with leukemia cells from patients before and during sorafenib resistance recapitulated the clinical results. Both ITD and tyrosine kinase domain mutations at D835 were identified in leukemia initiating cells (LICs) from samples before sorafenib treatment. LICs bearing the D835 mutant have expanded during sorafenib treatment and dominated during the subsequent clinical resistance. These results suggest that sorafenib have selected more aggressive sorafenib-resistant subclones carrying both FLT3-ITD and D835 mutations, and might provide important leads to further improvement of treatment outcome with FLT3 inhibitors.
Assuntos
Antineoplásicos/administração & dosagem , Benzenossulfonatos/administração & dosagem , Resistencia a Medicamentos Antineoplásicos , Regulação Leucêmica da Expressão Gênica , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/genética , Mutação , Piridinas/administração & dosagem , Tirosina Quinase 3 Semelhante a fms/antagonistas & inibidores , Tirosina Quinase 3 Semelhante a fms/genética , Adulto , Aldeído Desidrogenase/biossíntese , Aldeído Desidrogenase/genética , Família Aldeído Desidrogenase 1 , Animais , Antineoplásicos/efeitos adversos , Benzenossulfonatos/efeitos adversos , Medula Óssea/enzimologia , Medula Óssea/patologia , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/genética , Feminino , Perfilação da Expressão Gênica , Humanos , Janus Quinase 3/biossíntese , Janus Quinase 3/genética , Leucemia Mieloide Aguda/enzimologia , Leucemia Mieloide Aguda/patologia , Masculino , Metaloproteinase 15 da Matriz/biossíntese , Metaloproteinase 15 da Matriz/genética , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Pessoa de Meia-Idade , Transplante de Neoplasias , Niacinamida/análogos & derivados , Compostos de Fenilureia , Estrutura Terciária de Proteína , Piridinas/efeitos adversos , Retinal Desidrogenase , Sorafenibe , Fatores de Tempo , Transplante Heterólogo , Tirosina Quinase 3 Semelhante a fms/metabolismoRESUMO
BACKGROUND: Formation of the epicardium requires several cellular processes including migration, transformation, invasion, and differentiation in order to give rise to fibroblast, smooth muscle, coronary endothelial and myocyte cell lineages within the developing myocardium. Snai1 is a zinc finger transcription factor that plays an important role in regulating cell survival and fate during embryonic development and under pathological conditions. However, its role in avian epicardial development has not been examined. RESULTS: Here we show that Snai1 is highly expressed in epicardial cells from as early as the proepicardial cell stage and its expression is maintained as proepicardial cells migrate and spread over the surface of the myocardium and undergo epicardial-to-mesenchymal transformation in the generation of epicardial-derived cells. Using multiple in vitro assays, we show that Snai1 overexpression in chick explants enhances proepicardial cell migration at Hamburger Hamilton Stage (HH St.) 16, and epicardial-to-mesenchymal transformation, cell migration, and invasion at HH St. 24. Further, we demonstrate that Snai1-mediated cell migration requires matrix metalloproteinase activity, and MMP15 is sufficient for this process. CONCLUSIONS: Together our data provide new insights into the multiple roles that Snai1 has in regulating avian epicardial development.
Assuntos
Regulação da Expressão Gênica no Desenvolvimento , Pericárdio/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/fisiologia , Animais , Diferenciação Celular , Linhagem da Célula , Movimento Celular , Proliferação de Células , Embrião de Galinha , Coração/embriologia , Metaloproteinase 15 da Matriz/metabolismo , Metaloproteinases da Matriz/metabolismo , Camundongos , Miocárdio/metabolismo , Pericárdio/embriologia , Reação em Cadeia da Polimerase em Tempo Real , Fatores de Transcrição da Família SnailRESUMO
Magnolin (MGL), a compound derived from the magnolia plant, has inhibitory effects on tumor cell invasion and growth. His study aims to explore the antitumor effect and underlying molecular mechanism of MGL against human cervical cancer. We found that MGL inhibited the proliferation, migration, and invasiveness of cervical cancer cells in vitro and in vivo. The underlying mechanism was shown to involve MGL-induced inhibition of JNK/Sp1-mediated MMP15 transcription and translation. Overexpression of JNK/Sp1 resulted in significant restoration of MMP15 expression and the migration and invasion capabilities of MGL-treated cervical cancer cells. MGL modulated the cervical cancer microenvironment by inhibiting cell metastasis via targeting IL-10/IL-10 receptor B (IL-10RB) expression, thereby attenuating JNK/Sp1-mediated MMP15 expression. Analysis of the gut microbiota of mice fed MGL revealed a significant augmentation in Lachnospiraceae bacteria, known for their production of sodium butyrate. In vivo experiments also demonstrated synergistic inhibition of cervical cancer cell metastasis by MGL and sodium butyrate co-administration. Our study provides pioneering evidence of a novel mechanism by which MGL inhibits tumor growth and metastasis through the IL-10/IL-10RB targeting of the JNK/Sp1/MMP15 axis in human cervical cancer cells.