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DICER1 syndrome is a tumor predisposition syndrome caused by familial genetic mutations in DICER1. Pathogenic variants of DICER1 have been discovered in many rare cancers, including cystic liver tumors. However, the molecular mechanisms underlying liver lesions induced by these variants remain unclear. In the present study, we sought to gain a better understanding of the pathogenesis of these variants by generating a mouse model of liver-specific DICER1 syndrome. The mouse model developed bile duct hyperplasia with fibrosis, similar to congenital hepatic fibrosis, as well as cystic liver tumors resembling those in Caroli's syndrome, intrahepatic cholangiocarcinoma, and hepatocellular carcinoma. Interestingly, the mouse model of DICER1 syndrome showed abnormal formation of primary cilia in the bile duct epithelium, which is a known cause of bile duct hyperplasia and cyst formation. These results indicated that DICER1 mutations contribute to cystic liver tumors by inducing defective primary cilia. The mouse model generated in this study will be useful for elucidating the potential mechanisms of tumorigenesis induced by DICER1 variants and for obtaining a comprehensive understanding of DICER1 syndrome. © 2024 The Pathological Society of Great Britain and Ireland.
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Cilios , ARN Helicasas DEAD-box , Modelos Animales de Enfermedad , Neoplasias Hepáticas , Ribonucleasa III , Animales , Ribonucleasa III/genética , Ribonucleasa III/metabolismo , ARN Helicasas DEAD-box/genética , ARN Helicasas DEAD-box/deficiencia , Cilios/patología , Cilios/metabolismo , Ratones , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/patología , Mutación , Hígado/patología , Hígado/metabolismo , Conductos Biliares/patologíaRESUMEN
Reducing recurrence following radical resection of colon cancer without over- or under-treatment remains a challenge. Postoperative adjuvant chemotherapy (Adj) is currently administered based solely on pathological tumor, node, and metastasis (TNM) stage. However, prognosis can vary significantly among patients with the same disease stage. Therefore, novel classification systems in addition to the TNM are necessary to inform decision-making regarding postoperative treatment strategies, especially stage II and III disease, and to minimize overtreatment and undertreatment with Adj. We developed a prognostic prediction system for colorectal cancer by using a combined convolutional neural network (CNN) and support vector machine (SVM) approach to extract features from hematoxyling and eosin staining (HE) images. We combined the TNM and our AI-based classification system into a TNM-AI (mTNM-AI) classification system with high discriminative power for recurrence-free survival (RFS). Furthermore, the cancer cell population recognized by this system as low risk of recurrence exhibited the mutational signature SBS87 as a genetic phenotype. The novel AI-based classification system developed here is expected to play an important role in prognostic prediction and personalized treatment selection in oncology.
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Risk factors for hepatocarcinogenesis include chronic inflammation due to viral infection, liver fibrosis, and aging. In this study, we separated carcinogenic and non-carcinogenic cases due to hepatitis C virus (HCV) infection, aiming to comprehensively analyze miRNA expression in liver tissues by age, and identify factors that contribute to carcinogenesis. Total RNA was extracted from 360 chronic hepatitis C (CH), 43 HCV infected hepatocellular carcinoma (HCC), and surrounding non-tumor (SNT) tissues. MicroRNA (miRNA) expression patterns were analyzed using microarray. Using machine learning, we extracted characteristic miRNA expression patterns for each disease and age. There were no age-dependent changes in miRNA expression in the disease-specific comparisons; however, miRNA expression differed among the age groups of 50, 60, and 70 years of age between CH and SNT. The expression of miRNA was different between SNT and HCC only in patients in their 70s. Of the 55 miRNAs with significant differences in expression between CH and SNT, 34 miRNAs showed significant differences in expression even in the degree of liver fibrosis. The observation that miRNAs involved in hepatocarcinogenesis differ at different ages suggests that the mechanisms of carcinogenesis differ by age group as well. We also found that many miRNAs whose expression did not affect liver fibrosis were involved in carcinogenesis. These findings are expected to define biomarkers for detection of HCC at early stage, and develop novel therapeutic targets for HCC.
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Carcinoma Hepatocelular , Hepatitis C Crónica , Neoplasias Hepáticas , MicroARNs , Humanos , MicroARNs/genética , MicroARNs/metabolismo , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/virología , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/virología , Persona de Mediana Edad , Masculino , Femenino , Anciano , Hepatitis C Crónica/genética , Hepatitis C Crónica/complicaciones , Hepatitis C Crónica/patología , Cirrosis Hepática/genética , Cirrosis Hepática/virología , Cirrosis Hepática/patología , Hígado/metabolismo , Hígado/patología , Hígado/virología , Perfilación de la Expresión Génica , Carcinogénesis/genética , Adulto , Regulación Neoplásica de la Expresión GénicaRESUMEN
Cell transfer therapy using mesenchymal stem cells (MSCs) has pronounced therapeutic potential, but concerns remain about immune rejection, emboli formation, and promotion of tumor progression. Because the mode of action of MSCs highly relies on their paracrine effects through secretion of bioactive molecules, cell-free therapy using the conditioned medium (CM) of MSCs is an attractive option. However, the effects of MSC-CM on tumor progression have not been fully elucidated. Herein, we addressed this issue and investigated the possible underlying molecular mechanisms. The CM of MSCs derived from human bone marrow greatly inhibited the in vitro growth of several human tumor cell lines and the in vivo growth of the SCCVII murine squamous cell carcinoma cell line with reduced neovascularization. Exosomes in the MSC-CM were only partially involved in the inhibitory effects. The CM contained a variety of cytokines including insulin-like growth factor binding proteins (IGFBPs). Among them, IGFBP-4 greatly inhibited the in vitro growth of these tumors and angiogenesis, and immunodepletion of IGFBP-4 from the CM significantly reversed these effects. Of note, the CM greatly reduced the phosphorylation of AKT, ERK, IGF-1 receptor beta, and p38 MAPK in a partly IGFBP4-dependent manner, possibly through its binding to IGF-1/2 and blocking the signaling. The CM depleted of IGFBP-4 also reversed the inhibitory effects on in vivo tumor growth and neovascularization. Thus, MSC-CM has potent inhibitory effects on tumor growth and neovascularization in an IGFBP4-dependent manner, suggesting that cell-free therapy using MSC-CM could be a safer promising alternative for even cancer patients.
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Proteína 4 de Unión a Factor de Crecimiento Similar a la Insulina , Células Madre Mesenquimatosas , Humanos , Ratones , Animales , Proteína 4 de Unión a Factor de Crecimiento Similar a la Insulina/metabolismo , Proteína 4 de Unión a Factor de Crecimiento Similar a la Insulina/farmacología , Medios de Cultivo Condicionados/farmacología , Medios de Cultivo Condicionados/metabolismo , Médula Ósea/metabolismo , Células Madre Mesenquimatosas/metabolismo , Proteínas de Unión a Factor de Crecimiento Similar a la Insulina/metabolismo , Neovascularización Patológica/metabolismoRESUMEN
Clinical autopsies are performed to reveal the process of the disease that caused patient death and validate the diagnosis and treatment decisions. In pediatric clinical autopsy, the feedback provided to bereaved families has a considerable social impact; however, pediatric diseases are diverse, which makes it difficult to elucidate them. Therefore, it is necessary to employ molecular biology techniques in addition to conventional methods. Formalin-fixed, paraffin-embedded (FFPE) tissues are routinely prepared. However, clinical autopsy FFPE tissue processing is not standardized, and it is unclear whether DNA from such tissues can be used for comprehensive genomic analysis. In this study, we evaluated the DNA quality of FFPE tissues from 15 recent autopsy cases at a single-center children's hospital using quantitative polymerase chain reaction [PCR (Q129/Q41)] and nanoelectrophoresis (DNA integrity number (DIN)). Good quality DNA was obtained from every organ type excluding bone marrow within 6 days of formalin fixation. Prolonged proteinase K digestion (48 h > 24 h > 1 h) and thicker tissue sections (10 µm > 1 µm) improved Q129/Q41; however, 24 h fixed FFPE tissues showed better DNA quality. We propose an optimal and feasible workflow for storing short-term fixed FFPE tissues as DNA-preserved FFPE tissues for future comprehensive genomic searches.
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ADN , Formaldehído , Niño , Humanos , Fijación del Tejido/métodos , Autopsia , Adhesión en Parafina/métodos , ADN/análisis , Pruebas GenéticasRESUMEN
The mouse vagina consists of stratified squamous epithelium and stroma and is regulated by ovarian hormones. Vaginal epithelial cells do not stratify, but rather form a monolayer and show an inconsistent responsiveness to ovarian hormones when cultured on plastic dish or matrix. To address the discrepancy between in vivo and in vitro observations, three-dimensional (3D) co-culture models are developed with clonal vaginal epithelial and stromal cell lines; stromal cells are embedded in collagen gel and epithelial cells are seeded on the gel. In the 3D models, epithelial cells express Transformation related protein 63 (Trp63) and begin to stratify when they are co-cultured with two out of three stromal cell lines, but not with the other stromal cell line. Stroma may consist of various types of cells with distinct functions.
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Vagina/anatomía & histología , Vagina/crecimiento & desarrollo , Animales , Membrana Basal/metabolismo , Línea Celular , Células Clonales/citología , Células Clonales/metabolismo , Técnicas de Cocultivo , Células Epiteliales/citología , Células Epiteliales/efectos de los fármacos , Células Epiteliales/metabolismo , Epitelio/efectos de los fármacos , Epitelio/crecimiento & desarrollo , Epitelio/metabolismo , Estradiol/farmacología , Femenino , Ratones , Ratones Noqueados , Modelos Anatómicos , Fosfoproteínas/deficiencia , Fosfoproteínas/genética , Fosfoproteínas/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Células del Estroma/citología , Células del Estroma/efectos de los fármacos , Células del Estroma/metabolismo , Transactivadores/deficiencia , Transactivadores/genética , Transactivadores/metabolismo , Vagina/metabolismoRESUMEN
Stratification of the vaginal epithelium is regulated by stromal factors. To analyze the mechanisms of stratification in vitro, 3 dimensional (3D) co-culture models were established with clonal cell lines. In the models, stromal cells were embedded in collagen gel and epithelial cells were seeded on the gel. In the 3D co-culture, stromal SV-6c4a1b cells induced epithelial stratification but stromal MV-1e6g1a cells did not, suggesting that SV-6c4a1b cells secrete molecules to induce stratification. Microarray analyses of these stromal cell lines identified chordin-like 1 (Chrd1) and WNT1 inducible signaling pathway protein 2 (Wisp2) as candidate genes inducing stratification. Chrdl1 variant1 and variant2 mRNAs were expressed not only in stromal SV-6c4a1b and MV-1e6g1a cells but also in epithelial SV-4b6b cells. Wisp2-overexpressing MV-1e6g1a cells, secreting WISP2 as much as SV-6c4a1b cells, induced stratification of epithelial cells. In addition, Wisp2-knockdowned SV-6c4a1b cells were unable to induce epithelial stratification. These results suggest that WISP2 is one of the stromal factors inducing stratification of the mouse vaginal epithelium.
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Vagina/anatomía & histología , Vagina/crecimiento & desarrollo , Animales , Células Clonales/citología , Células Clonales/metabolismo , Técnicas de Cocultivo , Células Epiteliales/citología , Células Epiteliales/metabolismo , Epitelio/anatomía & histología , Epitelio/crecimiento & desarrollo , Epitelio/metabolismo , Estradiol/farmacología , Proteínas del Ojo/genética , Femenino , Regulación del Desarrollo de la Expresión Génica/efectos de los fármacos , Técnicas de Silenciamiento del Gen , Variación Genética , Péptidos y Proteínas de Señalización Intracelular/antagonistas & inhibidores , Péptidos y Proteínas de Señalización Intracelular/genética , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Ratones , Proteínas del Tejido Nervioso/genética , Progesterona/farmacología , ARN Mensajero/genética , ARN Mensajero/metabolismo , Transducción de Señal/genética , Células del Estroma/citología , Células del Estroma/metabolismo , Vagina/metabolismoRESUMEN
Recent studies have demonstrated that exosomal microRNAs (miRNAs) have the potential of facilitating molecular diagnosis. Currently, little is known about the underlying mechanism behind late-onset acute graft-versus-host disease (LA GVHD). Identifying differentially expressed miRNAs in exosomes should be useful for understanding the role of miRNAs in this disease. This study was established to investigate the relevance of miRNAs in exosomes derived from patients developing LA GVHD after allogeneic hematopoietic stem cell transplantation (HSCT). Plasma samples were collected from patients with LA GVHD (n = 5), non-GVHD (n = 5), and controls (n = 8) for exosomal miRNA expression profiling using a TaqMan low-density array; the results were validated by quantitative reverse transcription polymerase chain reaction (RT-PCR). We analyzed exosomal miRNAs differentially expressed among these three groups. MirTarBase was employed to predict potential target genes of the miRNAs specific for LA GVHD. We detected 55 miRNAs that were differentially expressed with a significant change >2.0-fold between LA GVHD and non-GVHD. Of these, we selected the 10 miRNAs (miR-423-5p, miR-19a, miR-142-3p, miR-128, miR-193b, miR-30c, miR-193a, miR-191, miR-125b, and miR-574-3p) with the most significant differential expression. Using quantitative RT-PCR, we further identified that miR-128 was significantly upregulated at the onset of LA GVHD compared with that in normal controls and is a promising diagnostic marker of LA GVHD, with an area under the curve (AUC) value of 0.975. MirTarBase analysis revealed that the predicted target genes of miR-128 are involved in the immune system and inflammation. Increased expression of miR-128 may serve as a novel, noninvasive biomarker for early LA GVHD diagnosis.
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Biomarcadores de Tumor/genética , Exosomas/química , Enfermedad Injerto contra Huésped/genética , Neoplasias Hematológicas/genética , Trasplante de Células Madre Hematopoyéticas/efectos adversos , MicroARNs/genética , Enfermedad Aguda , Adulto , Anciano , Área Bajo la Curva , Biomarcadores de Tumor/sangre , Inhibidores de la Calcineurina/uso terapéutico , Estudios de Casos y Controles , Ciclosporina/uso terapéutico , Exosomas/inmunología , Femenino , Perfilación de la Expresión Génica , Enfermedad Injerto contra Huésped/etiología , Enfermedad Injerto contra Huésped/inmunología , Enfermedad Injerto contra Huésped/mortalidad , Neoplasias Hematológicas/inmunología , Neoplasias Hematológicas/mortalidad , Neoplasias Hematológicas/terapia , Humanos , Masculino , Metotrexato/uso terapéutico , MicroARNs/sangre , Persona de Mediana Edad , Agonistas Mieloablativos/uso terapéutico , Curva ROC , Análisis de Supervivencia , Tacrolimus/uso terapéutico , Trasplante HomólogoRESUMEN
PURPOSE OF REVIEW: To review the data on angiogenesis related to exosomes secreted by tumor cells in hematologic neoplasia and to elucidate the role of exosomes and exosomal miRNA in the bone marrow microenvironment, especially under hypoxic conditions. RECENT FINDINGS: Cross-talk between bone marrow tumor cells and surrounding cells, including endothelial cells, is important for tumor growth in hematologic neoplasia. In addition to conventional signaling pathways, exosomes, which are small endosome-derived vesicles containing miRNAs, can help to modulate the microenvironment without directly contacting nontumorous cells. The human myeloid leukemia cell line K562 secretes exosomes containing a large amount of miR-92a that enhances angiogenesis under normoxic and hypoxic conditions. With chronic hypoxia, exosomes secreted by multiple myeloma cells also enhance angiogenesis by targeting factor-inhibiting hypoxia-inducible factor-1 via miR-135b. SUMMARY: Intercellular communication between tumor cells and a heterogeneous population of bone marrow stromal cells is mediated by exosomes containing various functional proteins, mRNA, and miRNA. Hypoxia is a major regulator of exosomal content and affects angiogenesis in various types of hematologic neoplasia. Functional analysis of exosomes and exosome-mediated cell-cell interactions not only clarifies molecular pathogenesis but also suggests new treatment strategies for hematologic neoplasia through targeting exosomes.
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Médula Ósea/irrigación sanguínea , Médula Ósea/patología , Exosomas/metabolismo , Neoplasias Hematológicas/metabolismo , Neoplasias Hematológicas/patología , Neovascularización Patológica/patología , Hipoxia Tumoral , Humanos , Células Madre Mesenquimatosas/metabolismo , Células Madre Mesenquimatosas/patología , Neovascularización Patológica/metabolismoRESUMEN
Exosomes are small endosome-derived vesicles containing a wide range of functional proteins, mRNA, and miRNA. Exosomal miRNA from cancer cells helps modulate the microenvironment. In multiple myeloma (MM), the massive proliferation of malignant plasma cells causes hypoxia. To date, the majority of in vitro hypoxia studies of cancer cells have used acute hypoxic exposure (3-24 hours). Thus, we attempted to clarify the role of MM-derived exosomes in hypoxic bone marrow by using MM cells grown continuously in vitro under chronic hypoxia (hypoxia-resistant MM [HR-MM] cells). The HR-MM cells produced more exosomes than the parental cells under normoxia or acute hypoxia conditions, and miR-135b was significantly upregulated in exosomes from HR-MM cells. Exosomal miR-135b directly suppressed its target factor-inhibiting hypoxia-inducible factor 1 (FIH-1) in endothelial cells. Finally, exosomal miR-135b from HR-MM cells enhanced endothelial tube formation under hypoxia via the HIF-FIH signaling pathway. This in vitro HR myeloma cell model will be useful for investigating MM cell-endothelial cell interactions under hypoxic conditions, which may mimic the in vivo bone marrow microenvironment. Although tumor angiogenesis is regulated by various factors, exosomal miR-135b may be a target for controlling MM angiogenesis.
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Exosomas/genética , MicroARNs/genética , Oxigenasas de Función Mixta/genética , Neovascularización Patológica/genética , Proteínas Represoras/genética , Regiones no Traducidas 3'/genética , Vasos Sanguíneos/crecimiento & desarrollo , Vasos Sanguíneos/metabolismo , Western Blotting , Hipoxia de la Célula , Línea Celular Tumoral , Células Cultivadas , Exosomas/ultraestructura , Perfilación de la Expresión Génica , Regulación Neoplásica de la Expresión Génica , Técnicas de Silenciamiento del Gen , Células Endoteliales de la Vena Umbilical Humana/metabolismo , Células Endoteliales de la Vena Umbilical Humana/fisiología , Humanos , Luciferasas/genética , Luciferasas/metabolismo , Microscopía Electrónica de Transmisión , Oxigenasas de Función Mixta/metabolismo , Mieloma Múltiple/irrigación sanguínea , Mieloma Múltiple/genética , Mieloma Múltiple/patología , Análisis de Secuencia por Matrices de Oligonucleótidos , Proteínas Represoras/metabolismoRESUMEN
Approximately 40% of chronic myeloid leukemia (CML) patients who discontinue imatinib (IM) therapy maintain undetectable minimal residual disease (UMRD) for more than one year (stopping IM (STOP-IM)). To determine a possible biomarker for STOP-IM CML, we examined plasma miRNA expression in CML patients who were able to discontinue IM. We first screened candidate miRNAs in unselected STOP-IM patients, who had sustained UMRD after discontinuing IM for more than six months, in comparison with healthy volunteers, by using a TaqMan low-density array for plasma or exosomes. Exosomal miR-215 and plasma miR-215 were downregulated in the STOP-IM group compared to the control, indicating that the biological relevance of the plasma miR-215 level is equivalent to that of the exosomal level. Next, we performed real-time quantitative RT-PCR in 20 STOP-IM patients, 32 patients with UMRD on continued IM therapy (IM group) and 28 healthy volunteers. The plasma miRNA-215 level was significantly downregulated in the STOP-IM group (p < 0.0001); we determined the cut-off level and divided the IM group patients into two groups according to whether the plasma miR-215 was downregulated or not. The IM group patients with a low plasma miR-215 level had a significantly higher total IM intake, compared to the patients with elevated miR-215 levels (p = 0.0229). Functional annotation of miR-215 target genes estimated by the Database for Annotation, Visualization and Integrated Discovery (DAVID) bioinformatic tools involved cell cycle, mitosis, DNA repair and cell cycle checkpoint. Our study suggests a possible role of miR-215 in successful IM discontinuation.
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Antineoplásicos/uso terapéutico , Mesilato de Imatinib/uso terapéutico , Leucemia Mielógena Crónica BCR-ABL Positiva/tratamiento farmacológico , Leucemia Mielógena Crónica BCR-ABL Positiva/genética , MicroARNs/genética , Regulación hacia Abajo/efectos de los fármacos , Perfilación de la Expresión Génica , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Leucemia Mielógena Crónica BCR-ABL Positiva/sangre , MicroARNs/sangreRESUMEN
A 51-year-old man with chronic myeloid leukemia (CML) was treated with imatinib (IM). After 24 months of treatment, he achieved a complete molecular response (CMR), which he sustained for 3 years. However, 4 months after discontinuing IM treatment, the CML relapsed. The patient was treated again with IM and achieved CMR. A combination of IM and interferon-α (IFNα) was administered for the following year, and then discontinued. The patient has since sustained CMR without therapy for 24 months, to date. This patient was found to have a BCL2L11 (BIM) deletion polymorphism. CML patients with a BIM deletion polymorphism show a low response to IM, and we infer that the BIM deletion polymorphism is a negative factor for discontinuation of IM. IFNα treatment is expected to prevent relapse during immunological surveillance. Therefore, the combination of IM and IFNα might be a feasible approach for CML patients who experience difficulty with IM discontinuation.
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Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Proteínas Reguladoras de la Apoptosis/genética , Leucemia Mielógena Crónica BCR-ABL Positiva/tratamiento farmacológico , Proteínas de la Membrana/genética , Polimorfismo Genético , Proteínas Proto-Oncogénicas/genética , Proteína 11 Similar a Bcl2 , Benzamidas/administración & dosificación , Terapia Combinada , Eliminación de Gen , Humanos , Mesilato de Imatinib , Interferón-alfa/administración & dosificación , Leucemia Mielógena Crónica BCR-ABL Positiva/diagnóstico , Masculino , Persona de Mediana Edad , Piperazinas/administración & dosificación , Pirimidinas/administración & dosificación , Recurrencia , Inducción de Remisión/métodosRESUMEN
Hypoxia plays an important role during the evolution of cancer cells and their microenvironment. Emerging evidence suggests communication between cancer cells and their microenvironment occurs via exosomes. This study aimed to clarify whether hypoxia affects angiogenic function through exosomes secreted from leukemia cells. We used the human leukemia cell line K562 for exosome-generating cells and human umbilical vein endothelial cells (HUVECs) for exosome target cells. Exosomes derived from K562 cells cultured under normoxic (20%) or hypoxic (1%) conditions for 24 h were isolated and quantitated by nanoparticle tracking analysis. These exosomes were then cocultured with HUVECs to evaluate angiogenic activity. The exosomes secreted from K562 cells in hypoxic conditions significantly enhanced tube formation by HUVECs compared with exosomes produced in normoxic conditions. Using a TaqMan low-density miRNA array, we found a subset of miRNAs, including miR-210, were significantly increased in exosomes secreted from hypoxic K562 cells. We demonstrated that cancer cells and their exosomes have altered miRNA profiles under hypoxic conditions. Although exosomes contain various molecular constituents such as proteins and mRNAs, altered exosomal compartments under hypoxic conditions, including miR-210, affected the behavior of endothelial cells. Our results suggest that exosomal miRNA derived from cancer cells under hypoxic conditions may partly affect angiogenic activity in endothelial cells.
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Hipoxia de la Célula , Células Endoteliales/metabolismo , Exosomas/metabolismo , Leucemia/metabolismo , Técnicas de Cocultivo , Células Endoteliales/citología , Regulación Neoplásica de la Expresión Génica , Células Endoteliales de la Vena Umbilical Humana , Humanos , Células K562 , Leucemia/patología , MicroARNs/genética , MicroARNs/metabolismoRESUMEN
Ovarian cancer is a malignant gynecologic disease rarely diagnosed in the early stages. Among the various types of ovarian cancer, clear cell carcinoma has a poor prognosis due to its malignant potential. MicroRNAs (miRNAs/miRs) regulate gene expression in cells by suppressing the translation of target genes or by degrading the target mRNA. miRNAs are also secreted from the cells in the blood, binding to proteins or lipids and assisting in cell-cell communication. Therefore, serum miRNAs may be considered potential diagnostic biomarkers for ovarian cancer. The present study investigated and identified specific miRNAs associated with ovarian clear cell carcinoma and compared them to those in ovarian endometrioma samples and healthy controls. CA125, an ovarian tumor marker, did not differ between patients with ovarian clear cell carcinoma, endometriosis or healthy controls. Subsequently, four miRNAs (miR-146a-5p, miR-191-5p, miR-484 and miR-574-3p) were analyzed. The expression levels of miR-146a-5p and miR-191-5p were significantly increased in the serum samples from patients with ovarian clear cell carcinoma compared with those in the healthy controls, but there was no significant difference compared with in patients with endometriosis. Furthermore, the bioinformatics analysis showed that CCND2 and NOTCH2 were the candidate target genes of miR-146a-5p and miR-191-5p. In conclusion, the results of the present study demonstrated that miR-146a-5p and miR-191-5p may be useful as early and non-invasive diagnostic tools in ovarian clear cell carcinoma. These miRNAs can help in distinguishing between ovarian clear cell carcinoma and ovarian endometrioma. To the best of our knowledge, no previous studies have screened any candidates specifically for ovarian clear cell carcinoma.
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BACKGROUND AND AIM: Circulating micro RNAs (miRNAs) indicate clinical pathologies such as inflammation and carcinogenesis. In this study, we aimed to investigate whether miRNA expression level patterns in could be used to diagnose hepatocellular carcinoma (HCC) and biliary tract cancer (BTC), and the relationship miRNA expression patterns and cancer etiology. METHODS: Patients with HCC and BTC with indications for surgery were selected for the study. Total RNA was extracted from the extracellular vesicle (EV)-rich fraction of the serum and analyzed using Toray miRNA microarray. Samples were divided into two cohorts in order of collection, the first 85 HCC were analyzed using a microarray based on miRBase ver.2.0 (hereafter v20 cohort), and the second 177 HCC and 43 BTC were analyzed using a microarray based on miRBase ver.21 (hereafter v21 cohort). RESULTS: Using miRNA expression patterns, we found that HCC and BTC could be identified with an area under curve (AUC) 0.754 (v21 cohort). Patients with anti-hepatitis C virus (HCV) treatment (SVR-HCC) and without antiviral treatment (HCV-HCC) could be distinguished by an AUC 0.811 (v20 cohort) and AUC 0.798 (v21 cohort), respectively. CONCLUSIONS: In this study, we could diagnose primary hepatic malignant tumor using miRNA expression patterns. Moreover, the difference of miRNA expression in SVR-HCC and HCV-HCC can be important information for enclosing cases that are prone to carcinogenesis after being cured with antiviral agents, but also for uncovering the mechanism for some carcinogenic potential remains even after persistent virus infection has disappeared.
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Carcinoma Hepatocelular , Hepatitis C , Neoplasias Hepáticas , MicroARNs , Humanos , Neoplasias Hepáticas/diagnóstico , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/terapia , Carcinoma Hepatocelular/diagnóstico , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/terapia , MicroARNs/genética , Hepacivirus/genética , CarcinogénesisRESUMEN
There is an unmet need for antifibrotic therapies to prevent the progression of liver cirrhosis. Previously, we conducted an exploratory trial to assess the safety and antifibrotic efficacy of PRI-724, a selective CBP/ß-catenin inhibitor, in patients with liver cirrhosis. PRI-724 was well tolerated and exerted a potential antifibrotic effect. Here, we investigated whether the profiles of circulating microRNAs packaged in extracellular vesicles (EV-miRNAs) are associated with responses to liver fibrosis treatments. Eighteen patients who received PRI-724 for 12 weeks in a phase 1/2a study were classified as responders (n = 10) or non-responders (n = 8) based on changes in liver stiffness. Plasma samples were obtained before and after PRI-724 administration and the levels of EV-miRNAs were analyzed. Three miRNAs (miR-6510-5p, miR-6772-5p, and miR-4261) were identified as predictors of response or non-response to PRI-724, and the levels of three other miRNAs (miR-939-3p, miR-887-3p, and miR-7112-5p) correlated with the efficacy of treatment. Expression of miR-887-3p was detected in hepatocytes and was decreased significantly in liver tissue following PRI-724 treatment. In addition, transfection of a miR-887-3p mimic activated hepatic stellate cells. Thus, decreases in the miR-887-3p level in blood may reflect recovery from liver fibroses in patients with liver cirrhosis treated with PRI-724, although further validation studies are warranted to confirm this.
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Vesículas Extracelulares , MicroARNs , Pirimidinonas , Humanos , MicroARNs/metabolismo , beta Catenina/genética , beta Catenina/metabolismo , Cirrosis Hepática/tratamiento farmacológico , Cirrosis Hepática/genética , Compuestos Bicíclicos Heterocíclicos con Puentes/uso terapéutico , Vesículas Extracelulares/metabolismoRESUMEN
Mesenchyme is generally believed to play critical roles in "secondary induction" during organogenesis. Because of the complexity of tissue interactions in secondary inductions, however, little is known about the precise mechanisms at the cellular and molecular levels. We have demonstrated that, in mouse oviductal development, the mesenchyme determines the fate of undetermined epithelial cells to become secretory or cilial cells. We have established a model for studying secondary induction by establishing clonal epithelial and mesenchymal cell lines from perinatal p53(-/-) mouse oviducts. The signal sequence trap method collected candidate molecules secreted from mesenchymal cell lines. Naive epithelial cells exposed to Follistatin-like-1 (Fstl1), one of the candidates, became irreversibly committed to expressing a cilial epithelial marker and differentiated into ciliated cells. We concluded that Fstl1 is one of the mesenchymal factors determining oviductal epithelial cell fate. This is a unique demonstration that the determination of epithelial cell fate is induced by a single diffusible factor.
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Diferenciación Celular/fisiología , Células Epiteliales/metabolismo , Proteínas Relacionadas con la Folistatina/fisiología , Mesodermo/metabolismo , Animales , Diferenciación Celular/genética , Línea Celular , Técnicas de Cocultivo , Células Epiteliales/citología , Trompas Uterinas/citología , Femenino , Proteínas Relacionadas con la Folistatina/genética , Proteínas Relacionadas con la Folistatina/metabolismo , Factores de Transcripción Forkhead/genética , Factores de Transcripción Forkhead/metabolismo , Expresión Génica , Glicoproteínas/genética , Glicoproteínas/metabolismo , Inmunohistoquímica , Hibridación in Situ , Queratina-18/metabolismo , Mesodermo/citología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Células 3T3 NIH , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Factores de TiempoRESUMEN
BACKGROUND: Polycythemia vera (PV), essential thrombocythemia (ET), and primary myelofibrosis (PMF) are myeloproliferative neoplasms (MPNs) characterized in most cases by a unique somatic mutation, JAK2 V617F. Recent studies revealed that JAK2 V617F occurs more frequently in a specific JAK2 haplotype, named JAK2 46/1 or GGCC haplotype, which is tagged by rs10974944 (C/G) and/or rs12343867 (T/C). This study examined the impact of single nucleotide polymorphisms (SNPs) of the JAK2 locus on MPNs in a Japanese population. METHODS: We sequenced 24 JAK2 SNPs in Japanese patients with PV. We then genotyped 138 MPN patients (33 PV, 96 ET, and 9 PMF) with known JAK2 mutational status and 107 controls for a novel SNP, in addition to two SNPs known to be part of the 46/1 haplotype (rs10974944 and rs12343867). Associations with risk of MPN were estimated by odds ratios and their 95% confidence intervals using logistic regression. RESULTS: A novel locus, rs4495487 (T/C), with a mutated T allele was significantly associated with PV. Similar to rs10974944 and rs12343867, rs4495487 in the JAK2 locus is significantly associated with JAK2-positive MPN. Based on the results of SNP analysis of the three JAK2 locus, we defined the "GCC genotype" as having at least one minor allele in each SNP (G allele in rs10974944, C allele in rs4495487, and C allele in rs12343867). The GCC genotype was associated with increased risk of both JAK2 V617F-positive and JAK2 V617F-negative MPN. In ET patients, leukocyte count and hemoglobin were significantly associated with JAK2 V617F, rather than the GCC genotype. In contrast, none of the JAK2 V617F-negative ET patients without the GCC genotype had thrombosis, and splenomegaly was frequently seen in this subset of ET patients. PV patients without the GCC genotype were significantly associated with high platelet count. CONCLUSIONS: Our results indicate that the C allele of JAK2 rs4495487, in addition to the 46/1 haplotype, contributes significantly to the occurrence of JAK2 V617F-positive and JAK2 V617F-negative MPNs in the Japanese population. Because lack of the GCC genotype represents a distinct clinical-hematological subset of MPN, analyzing JAK2 SNPs and quantifying JAK2 V617F mutations will provide further insights into the molecular pathogenesis of MPN.
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Janus Quinasa 2/genética , Trastornos Mieloproliferativos/genética , Polimorfismo de Nucleótido Simple , Adulto , Anciano , Alelos , Pueblo Asiatico/genética , Femenino , Genotipo , Haplotipos , Humanos , Masculino , Persona de Mediana Edad , Trastornos Mieloproliferativos/diagnóstico , Mutación Puntual , Factores de RiesgoRESUMEN
A method for quantifying global DNA methylation using fluorescence correlation spectroscopy (FCS) has been established. The single-molecule methylation assay (SMMA) is based on two methodologies. One methodology, FCS, estimates the translational diffusion coefficient of molecules in solution, whereas the other methodology uses the high affinity of methyl-CpG-binding domain protein 2 (MBD2) to bind specifically to methylated DNA. We studied the specific binding rates of fluorescence-labeled MBD2 and methylated DNA from biological samples using the automated FCS system. Using a standard curve with methylated control DNA, we developed the SMMA index to assess the global DNA methylation level of the biological samples. A marked decrease in the SMMA index was observed when human leukemia cell lines (U937 and K562) were cultured with DNA demethylating agents. Our findings clearly indicate the applicability of SMMA as a simple and rapid tool for quantifying global DNA methylation. SMMA may prove useful for genome-wide comparative methylation analyses of malignancies and as an indicator of the demethylation effects of epigenetic drugs.