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Copy number alterations are crucial for the development of gastric cancer (GC). Here, we identified Transmembrane Protein 65 (TMEM65) amplification by genomic hybridization microarray to profile copy-number variations in GC. TMEM65 mRNA level was significantly up-regulated in GC compared to adjacent normal tissues, and was positively associated with TMEM65 amplification. High TMEM65 expression or DNA copy number predicts poor prognosis (P < 0.05) in GC. Furtherly, GC patients with TMEM65 amplification (n = 129) or overexpression (n = 78) significantly associated with shortened survival. Ectopic expression of TMEM65 significantly promoted cell proliferation, cell cycle progression and cell migration/invasion ability, but inhibited apoptosis (all P < 0.05). Conversely, silencing of TMEM65 in GC cells showed opposite abilities on cell function in vitro and suppressed tumor growth and lung metastasis in vivo (all P < 0.01). Moreover, TMEM65 depletion by VNP-encapsulated TMEM65-siRNA significantly suppressed tumor growth in subcutaneous xenograft model. Mechanistically, TMEM65 exerted oncogenic effects through activating PI3K-Akt-mTOR signaling pathway, as evidenced of increased expression of key regulators (p-Akt, p-GSK-3ß, p-mTOR) by Western blot. YWHAZ (Tyrosine 3-Monooxygenase/Tryptophan 5-Monooxygenase) was identified as a direct downstream effector of TMEM65. Direct binding of TMEM65 with YWHAZ in the cytoplasm inhibited ubiquitin-mediated degradation of YWHAZ. Moreover, oncogenic effect of TMEM65 was partly dependent on YWHAZ. In conclusion, TMEM65 promotes gastric tumorigenesis by activating PI3K-Akt-mTOR signaling via cooperating with YWHAZ. TMEM65 overexpression may serve as an independent new biomarker and is a therapeutic target in GC.
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Proteínas Proto-Oncogênicas c-akt , Neoplasias Gástricas , Humanos , Proteínas 14-3-3/metabolismo , Carcinogênese/genética , Linhagem Celular Tumoral , Proliferação de Células/genética , Transformação Celular Neoplásica , Glicogênio Sintase Quinase 3 beta/metabolismo , Fosfatidilinositol 3-Quinases/genética , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Neoplasias Gástricas/genética , Neoplasias Gástricas/patologia , Serina-Treonina Quinases TOR/genética , Serina-Treonina Quinases TOR/metabolismo , Fatores de Transcrição/metabolismoRESUMO
Roundabout guidance receptor 2 (Robo2) is closely related to malignant tumors such as pancreatic cancer and liver fibrosis, but there is no relevant research on the role of Robo2 in HCC. The study will further explore the function and mechanism of Robo2 and its downstream target genes in HCC. Firstly, Robo2 protein levels in human HCC tissues and paired adjacent normal liver tissues were detected. Then we established HepG2 and Huh7 hepatoma cell lines with knock-down Robo2 by transfection with lentiviral vectors, and examined the occurrence of EMT, proliferation and apoptosis abilities in HCC cells by western blot, flow cytometry, wound healing assay and TUNEL staining. Then we verified the interaction between Robo2 and its target gene by Co-IP and immunofluorescence co-staining, and further explored the mechanism of Robo2 and YB-1 by rescue study. The protein expression level of Robo2 in HCC was considerably higher than that in the normal liver tissues. After successfully constructing hepatoma cells with knock-down Robo2, it was confirmed that down-regulated Robo2 suppressed EMT and proliferation of hepatoma cells, and accelerated the cell apoptosis. High-throughput sequencing and validation experiments verified that YB-1 was the downstream target gene of Robo2, and over-expression of YB-1 could reverse the apoptosis induced by Robo2 down-regulation and its inhibitory effect on EMT and proliferation. Robo2 deficiency inhibits EMT and proliferation of hepatoma cells and augments the cell apoptosis by regulating YB-1, thus inhibits the occurrence of HCC and provides a new strategy for the treatment of HCC.
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Carcinoma Hepatocelular , Neoplasias Hepáticas , Humanos , Carcinoma Hepatocelular/patologia , Neoplasias Hepáticas/patologia , Regulação para Baixo , Proliferação de Células , Imunofluorescência , Regulação Neoplásica da Expressão Gênica , Linhagem Celular Tumoral , Movimento Celular/genética , Apoptose/genéticaRESUMO
MicroRNA (miRNA) can affect tumor progression by regulating cell proliferation, apoptosis and metastasis. A significant upregulation of miR-17-5p expression was found in colorectal cancer (CRC) tissues by miRNA microarray chip analysis. However, the underlying mechanism of miR-17-5p in CRC is still unclear. The mRNA expression of miR-17-5p was significantly higher in CRC tissues than in adjacent normal tissues. In CRC group, the expression of miR-17-5p in cancer tissues with lymph node metastasis was higher compared with those without lymph node metastasis. The biological function of miR-17-5p was demonstrated through CCK-8, colony formation, flow cytometry and transwell assays. Overexpression of miR-17-5p inhibited CRC cell apoptosis, as well as promoting proliferation, migration and invasion. Transcriptome sequencing and miRNA target prediction software suggested that HSPB2 might be a target gene of miR-17-5p and luciferase reporter detection validated for the first time that miR-17-5p binds directly to the 3'-UTR of HSPB2. In the rescue experiment, the tumor suppressive effect of HSPB2 was detected and miR-17-5p could promote cell proliferation, migration and invasion by targeting HSPB2. Taken together, miR-17-5p promotes invasion and migration by inhibiting HSPB2 in CRC, thereby implicating its potential as a novel diagnostic biomarker and therapeutic target for CRC.
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PURPOSE: To develop new and effective biomarkers for the diagnosis of colorectal cancer (CRC). EXPERIMENTAL DESIGN: The serum expression of ITGB4 (49 CRC and 367 HC) was detected by enzyme-linked immunosorbent assay (ELISA), and its diagnostic value was analyzed using the receiver operating characteristic (ROC) curve. The sensitivity and specificity of ITGB4 in CRC diagnosis were calculated through statistical analysis. The optimal clinical cutoff value was calculated using the Youden index, and diagnostic efficacy was analyzed in a larger serum sample (98 CRC and 1631 non-CRC). The expression of ITGB4 was measured by CyTOF (cell experimental technology) at the single-cell level, and characteristics were analyzed using viSNE and SPADE TREE. RESULTS: Serum ITGB4 and CEA levels were significantly higher in CRC patients than in HC and non-CRC patients. The use of serum ITGB4 levels for the diagnosis of CRC has a high sensitivity (79%) but not high specificity when the clinical cutoff value was 0.70 ng/mL. However, the optimal cutoff value was 1.6 ng/mL with 86.2% specificity and 52.0% sensitivity, and the diagnostic efficacy was greatly improved with high specificity (82.0%) and sensitivity (71.4%) when combined with CEA. ITGB4 expression characteristics were measured and related to the expression of EpCAM, Ck8/18, and perforin at the single-cell level. Single-cell analysis showed that cell clusters with low expression of CK8/18 and ITGB4 were more sensitive to 5FU and radiotherapy (RT). CONCLUSIONS: ITGB4 is an effective diagnostic serum biomarker and a potential therapeutic target for CRC.
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Biomarcadores Tumorais/metabolismo , Neoplasias Colorretais/genética , Integrina beta4/metabolismo , Estudos de Casos e Controles , Neoplasias Colorretais/mortalidade , Feminino , Humanos , Masculino , Estudos Retrospectivos , Análise de Sobrevida , TransfecçãoRESUMO
OBJECTIVE: Presently, the prone position is necessary for popliteal vein puncture access, but it makes the patients uncomfortable and does not allow traditional femoral or jugular access. To address these deficiencies, this study introduces two new methods, anterior and medial access carried out in the supine position. METHODS: Venous interventions with punctures in the popliteal vein of 120 limbs in 97 patients were performed during the period from February 2017 to April 2019. After puncture, venographic guidance was achieved by dorsal vein injection of contrast medium. Interventional therapy was performed after puncture and insertion of the introducer sheath. RESULTS: In all, 120 limbs were punctured in the popliteal vein, with technical success in 118 (98.3% in total) cases: 100%, 96.1%, and 100% successful punctures in, respectively, 32 anterior, 49 medial, and 37 posterior access cases. A comparison of the three groups revealed that the fluoroscopy time and duration of puncture were longer in the medial and anterior access groups than in the posterior access group. The rate of intra-operative and post-operative complications was 7.5% (9/120), with no statistically significant difference between the three access groups. Compared with the pre-operative median score of 2.5, the post-operative SVS (Society of Vascular Surgery) score of the popliteal vein was reduced to 1.5 in the anterior and 0.5 in the medial groups. CONCLUSION: Medial and anterior puncture of the popliteal vein in the supine position can be used as a safe alternative in venous endovascular therapy. The two new methods can mitigate frailty or respiratory problems resulting from the prone position and facilitate traditional femoral and jugular access.
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Cateterismo Periférico , Posicionamento do Paciente , Flebografia , Veia Poplítea/diagnóstico por imagem , Radiografia Intervencionista , Decúbito Dorsal , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Cateterismo Periférico/efeitos adversos , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Punções , Estudos Retrospectivos , Adulto JovemRESUMO
Purpose: We recently reported that tripartite motif-containing 67 (TRIM67) activates p53 to suppress colorectal cancer (CRC). However, the function and mechanism of TRIM67 in the inhibition of CRC cell proliferation and metastasis remains to be further elucidated. Methods: We detected the expression of TRIM67 in CRC tissues compared with normal tissues and confirmed its relationship with clinicopathological features. DNA methylation of TRIM67 was analyzed to determine its significantly hypermethylated sites in CRC tissues. CCK-8, colony formation, transwell migration, and Matrigel invasion assays were performed to evaluate the effects of TRIM67 on cell proliferation and metastasis in CRC cells. RNA sequencing of TRIM67 and TRIM67 rescue experiments were performed to reveal its mechanisms in CRC cell proliferation and metastasis. Results: TRIM67 expression was significantly downregulated in CRC tissues and its expression was associated with clinical stage, invasive depth, tumor size, lymph node metastasis, and Dukes' stage. Three methylation sites were significantly hypermethylated and negatively correlated with TRIM67 expression in CRC tissues. TRIM67 suppressed proliferation, migration, and invasion in CRC cells. RNA sequencing revealed that protein mitogen-activated protein kinase 11 (MAPK11) was a potential downstream negative regulatory gene of TRIM67. Reversing MAPK11 expression could rescue the effects of TRIM67 on the proliferation and metastasis of CRC cells. Conclusion: TRIM67 inhibited cell proliferation and metastasis by mediating MAPK11 in CRC, and may be a potential target to inhibit CRC metastasis.
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MicroRNA (miR)-424-5p is overexpressed in colorectal cancer (CRC); however, its role, clinical significance and underlying molecular mechanism have remained to be fully elucidated. The aim of the present study was to investigate the roles of miR-424-5p in CRC and the underlying mechanisms. It was demonstrated that miR-424-5p is overexpressed in CRC, based on bioinformatics analysis using The Cancer Genome Atlas TCGA and analysis of tissue samples from patients with CRC from The First Hospital of Hebei Medical University, and the expression of miR-424-5p was associated with the depth of invasion and Dukes' staging. In CRC cells, the oncogenic roles of miR-424-5p were also verified by Cell Counting Kit-8, wound healing and Transwell assays. To identify target genes, all transcripts were compared between miR-424-5p mimic-transfected SW480 cells and mimic control cells by transcriptome sequencing. Subsequently, the differentially expressed genes (DEGs) were subjected to Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses. The DEGs were revealed to be significantly enriched in the GO terms 'serine hydrolase activity,' 'serine-type peptidase activity' and 'serine-type endopeptidase activity'. KEGG signaling pathway analysis indicated that the DEGs were significantly enriched in 'endocytosis', 'regulation of actin cytoskeleton', 'Wnt signaling pathway' and 'ubiquitin-mediated proteolysis signaling pathway'. These results suggested that miR-424-5p is a potential target in the treatment of CRC.
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Gastric cancer (GC) causes high morbidity and mortality in patients largely due to its invasion and metastasis. Kiss1 has been shown to be a metastasis suppressor in various malignancies. However, its clinical significance and biological functions in GC have not been thoroughly investigated. The present study investigated the association between Kiss1 expression and its methylation status and clinicopathological features in GC. Kiss1 expression was reduced in GC and its low expression was associated with poor histological grade, lymph node metastasis and TNM III+IV stage. Kiss1 overexpression in AGS GC cells significantly inhibited cell proliferation, migration and invasion in vitro. Kiss1 knockdown promoted the proliferation, migration and invasion of HGC27 cells. In summary, the data demonstrated that a low expression of Kiss1 played a suppressive role for the proliferation, migration and invasion of GC cells. Its expression and methylation levels were associated with the clinical progression of GC. Thus, Kiss1 is a potential diagnostic and prognostic marker as well as a new target for the treatment of GC.
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Biomarcadores Tumorais/genética , Mucosa Gástrica/patologia , Kisspeptinas/genética , Metástase Linfática/genética , Neoplasias Gástricas/genética , Idoso , Biomarcadores Tumorais/análise , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Metilação de DNA , Progressão da Doença , Epigênese Genética , Feminino , Gastrectomia , Mucosa Gástrica/cirurgia , Regulação Neoplásica da Expressão Gênica , Técnicas de Silenciamento de Genes , Humanos , Kisspeptinas/análise , Masculino , Pessoa de Meia-Idade , Invasividade Neoplásica/patologia , Estadiamento de Neoplasias , Prognóstico , Regiões Promotoras Genéticas/genética , Neoplasias Gástricas/diagnóstico , Neoplasias Gástricas/patologia , Neoplasias Gástricas/cirurgiaRESUMO
Competing endogenous RNAs (ceRNAs) are a newly proposed RNA interaction mechanism that has been associated with the tumorigenesis, metastasis, diagnosis, and predicting survival of various cancers. In this study, we constructed a ceRNA network in colorectal cancer (CRC). Then, we sought to develop and validate a composite clinicopathologic-genomic nomogram using The Cancer Genome Atlas (TCGA) database. To construct the ceRNA network in CRC, we analyzed the mRNAseq, miRNAseq data, and clinical information from TCGA database. LncRNA, miRNA, and mRNA signatures were identified to construct risk score as independent indicators of the prognostic value in CRC patients. A composite clinicopathologic-genomic nomogram was developed to predict the overall survival (OS). One hundred sixty-one CRC-specific lncRNAs, 97 miRNAs, and 161 mRNAs were identified to construct the ceRNA network. Multivariate Cox proportional hazards regression analysis indicated that nine-lncRNA signatures, eight-miRNA signatures, and five-mRNA signatures showed a significant prognostic value for CRC. Furthermore, a clinicopathologic-genomic nomogram was constructed in the primary cohort, which performed well in both the primary and validation sets. This study presents a nomogram that incorporates the CRC-specific ceRNA expression profile, clinical features, and pathological factors, which demonstrate its excellent differentiation and risk stratification in predicting OS in CRC patients.
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Colorectal cancer (CRC) is the third most common tumor in the world; however, the role and mechanism of endoplasmic reticulum (ER) stress in CRC metastasis remains largely unclear. Metastasisassociated lung adenocarcinoma transcript 1 (MALAT1) is a long noncoding RNA (lncRNA), which has previously been associated with CRC metastasis. It has been suggested that ER stress pathways regulate lncRNA expression; however, the effect of ER stress on MALAT1 expression in cancer is unknown. The present study aimed to investigate the relationship between ER stress pathways, MALAT1 expression and cell migration in CRC cells. ER stress was induced by thapsigargin (TG); low dose TG induced the migration of HT29 and HCT116 cells, but not SW1116 and SW620 cells. This effect was associated with increased expression levels of MALAT1, as the knockdown of MALAT1 prevented TGinduced cell migration. TGinduced MALAT1 expression was associated with inositolrequiring enzyme 1 (IRE1) expression and activation of the protein kinase R (PKR)like ER kinase (PERK) signaling pathway. Xboxbinding protein 1 (XBP1) and activating transcription factor 4 (ATF4) binding sites were predicted to be located in the MALAT1 gene promoter regions and the expression of MALAT1 was positively associated with XBP1 and ATF4 expression levels in CRC tissue samples. Thus, these findings indicated that ER stress may promote the migration of CRC cells and contribute to the progression of CRC through the activation of the IRE1/XBP1 and PERK/eIF2α/ATF4 signaling pathways. In conclusion, to the best of our knowledge, this study is the first report that lncRNA MALAT1 expression is regulated by the IRE1/XBP1 pathway in CRC.
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Movimento Celular , Neoplasias Colorretais/patologia , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , RNA Longo não Codificante/genética , Tapsigargina/farmacologia , Fator 4 Ativador da Transcrição/metabolismo , Linhagem Celular Tumoral , Neoplasias Colorretais/genética , Neoplasias Colorretais/metabolismo , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Ativação Transcricional , Proteína 1 de Ligação a X-Box/metabolismoRESUMO
BACKGROUND: Colorectal cancer (CRC) is the most common gastrointestinal malignancy. And probiotics may have the function of preventing colon cancer. The aim of this study was to investigate the inhibitory effects of a probiotic mixture on colorectal cancer and its potential mechanisms. METHODS: The effects of the probiotic mixture on proliferation and metastasis of mouse colon cancer CT26 cells were assessed by probiotics and cells co-culture assay, Cell Counting Kit-8 assay, colony formation assay, wound-healing assay, as well as migration and invasion assays. And CT26 cells were also transplanted into BALB/c mice to construct transplanted tumor animal model. The mice were randomly divided into two groups, control group and probiotic mixture intragastric administration group, after injection 21 days the tumor size and infiltration of immune cells in the tumor or spleen tissues were analyzed by hematoxylin and eosin (HE) and immunohistochemistry (IHC) staining. RESULTS: The probiotic mixture significantly inhibited the proliferation, invasion, and migration ability of CT26 cells compare to the control cells (P<0.05). In the animal experiments, the tumor volume of mice that had been fed the probiotic mixture was significantly smaller than that of the control group (P<0.05). Compared with control mice, more apoptotic cells and infiltration of immune cells were showed in the tumor tissues of the mice treated with the probiotic mixture, and an increased number of CD8+ cells in the tumor and spleen tissues but no significant change in tissues. CONCLUSIONS: These results suggested that the probiotic mixture could inhibit the growth of CT26 tumors and induce an immune response in vivo. The probiotic mixture also inhibited the invasion, migration, and proliferation ability of CT26 cells in vitro.
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Integrin ß4 (ITGB4) has been reported to be involved in carcinomas. Currently, ITGB4 has been characterized in colon cancer, however, its clinical significance is not very clear. In the present study, we utilized the large public datasets from NCBI Gene Expression Omnibus (GEO) and The Cancer Genome Atlas (TCGA) databases and collected clinical samples in our center to investigate the transcriptional expressions of ITGB4 in colon cancer, and then explored the associations of ITGB4 with clinicopathological features and overall survival. The statistical analyses suggested that ITGB4 mRNA expressions were up-regulated significantly in colon cancer. High ITGB4 expression was observed to be associated with elder onset age, proximal tumor location, and high microsatellite instability (MSH) status. Further, Kaplan-Meier curves and univariate analysis demonstrated high ITGB4 expression was significantly associated with unfavorable overall survival in colon cancer (HR=1.292, 95%CI=1.084-1.540, P=0.004). And significant association was also found after adjusting the confounding factors including age, gender, and stage (adjusted HR=1.254, 95%CI=1.050-1.497, P=0.012). The annotation of ITGB4 co-expressed genes suggested the pathways including cell growth, positive regulation of cell migration, and apoptotic signaling might be involved in the potential mechanisms of ITGB4 in colon cancer development. The molecular regulation mechanism of ITGB4 ectopic expression in colon cancer was also explored and the results indicated that ITGB4 might be up-regulated by the transcription factor FOSL1 (FOS like 1, AP-1 Transcription Factor Subunit) and its promoter hypomethylation. Our results revealed that ITGB4 might be a therapeutic target and prognosis marker for individual therapy of colon cancer.
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Chemotherapy and radiotherapy are main adjuvant therapies for the treatment of gastric cancer, the treatment effects are individual difference, but the specific mechanism is unknown. CyTOF 2 mass cytometer (CyTOF) enables the detecting up to 135 parameters on single cell, the emergence of which is an opportunity for proteomics research. We first tried to apply CyTOF technique to gastric cancer cells. We verified applicability of CyTOF in gastric cancer cells, and analyzed the responses of seventeen proteins to chemoradiotherapy in human gastric cancer AGS cells. To analyze the high dimensional CyTOF data, we used two statistical and visualization tools including viSNE and Citrus. Two specific clusters were found which had differences in protein expression profiles. CyTOF technology is proved feasibility and value at single cell level of gastric cancer.
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Citometria de Fluxo/métodos , Análise de Célula Única/métodos , Neoplasias Gástricas/patologia , Linhagem Celular Tumoral , Feminino , Humanos , Pessoa de Meia-Idade , Proteômica/métodos , Neoplasias Gástricas/metabolismoRESUMO
Dapper homolog 1 alpha (DACT1α) is a member of DACT family and an important regulator in the planar cell polarity pathway. We aim to clarify its functional role in metastasis ability of gastric cancer. DACT1α was silenced in all gastric cancer cell lines (8/8), but expressed in normal gastric tissue. Ectopic expression of DACT1α in silenced gastric cancer cell lines (AGS, BGC823 and MGC803) by stable transfection significantly suppressed cancer cell spreading (P < 0.05), migration (P < 0.01) and invasion (P < 0.01). These effects were associated with downregulation of planar cell polarity pathway related genes involved in cell proliferation (PDGFB, VEGFA), adhesion (ITGA1, ITGA2, ITGA3, ITGB3) and migration/invasion (PLAU, MMP9, MCAM, Dvl-2 and JNK). DACT1α promoter methylation was detected in 205 gastric cancers and 20 normal controls by direct bisulfite genomic sequencing. DACT1α methylation was detected in 29.3% (60/205) of gastric cancer patients, but not in normal tissues. DACT1α methylation was associated with poor survival of gastric cancer patients. In conclusion, DACT1α plays a pivotal role as a potential tumor suppressor in migration and invasion of gastric cancer. DACT1α methylation may serve as a biomarker for the prognosis of gastric cancer.
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Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Movimento Celular , Plasticidade Celular , Proteínas Nucleares/metabolismo , Neoplasias Gástricas/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Linhagem Celular Tumoral , Proliferação de Células , Forma Celular , Metilação de DNA , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Estimativa de Kaplan-Meier , Masculino , Invasividade Neoplásica , Proteínas Nucleares/genética , Fenótipo , Modelos de Riscos Proporcionais , Transdução de Sinais , Neoplasias Gástricas/genética , Neoplasias Gástricas/mortalidade , Neoplasias Gástricas/patologia , Fatores de Tempo , Transfecção , Proteínas Supressoras de Tumor/genéticaRESUMO
OBJECTIVE: Bi-allelic function-inactivating ENPP1 mutations cause artery media calcification (AMC) with associated severe myointimal hyperplasia in generalized arterial calcification of infancy (GACI), whereas mono-allelic ENPP1 deficiency is phenotypically normal. Here, we tested if ENPP1 deficiency promotes abnormal vascular smooth muscle cell (VSMC)-driven responses to injury, with or without calcification. The ER stress mediator C/EBP homologous protein (CHOP) affects neointimal hyperplasia and atherosclerosis, and has paradoxical effects on bone formation. Hence, we assessed relationships between ENPP1 and CHOP in VSMCs. METHODS: We studied ENPP1-deficient mice and control littermates subjected to left carotid artery ligation, and isolated and studied VSMCs from these and Chop-/- mice, or with CHOP siRNA treatment. RESULTS: Normal Enpp1-/+ mice, in addition to Enpp1-/- mice prior to AMC development, had accelerated neointimal hyperplasia in response to carotid artery ligation at 7-8 weeks age. Neointimal hyperplasia was linked with robust artery media CHOP expression in situ, but with marked AMC only in injured Enpp1-/- arteries. Cultured, ENPP1-deficient and CHOP-deficient VSMCs had increased migration and proliferation to PDGF. Cultured Chop-/- VSMCs demonstrated increased Pi donor-induced calcification. CHOP was significantly increased in Pi donor treated Enpp1-/- and Enpp1-/+ cultured VSMCs. CHOP siRNA treatment of Enpp1-/- VSMCs increased calcification, associated with elevated expression of tissue nonspecific alkaline phosphatase and the master osteoblastic transcription factor RUNX2. CONCLUSIONS: Both mono-allelic and bi-allelic ENPP1 deficiency promote dysregulated VSMC function, with robust lesion CHOP expression and enhanced neointimal hyperplasia after injury in vivo, but marked post-injury calcification limited to Enpp1-/- mice. Intimal hyperplasia in GACI appears regulated by biologic effects of ENPP1 deficiency other than calcification, including ER stress. VSMC CHOP excess in ENPP1 deficiency may primarily function to limit VSMC calcification.
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Estenose das Carótidas/patologia , Miócitos de Músculo Liso/patologia , Neointima/fisiopatologia , Diester Fosfórico Hidrolases/fisiologia , Pirofosfatases/fisiologia , Fator de Transcrição CHOP/biossíntese , Túnica Íntima/lesões , Fosfatase Alcalina/biossíntese , Fosfatase Alcalina/genética , Alelos , Animais , Estenose das Carótidas/metabolismo , Diferenciação Celular , Subunidade alfa 1 de Fator de Ligação ao Core/biossíntese , Subunidade alfa 1 de Fator de Ligação ao Core/genética , Modelos Animais de Doenças , Genótipo , Hiperplasia , Ligadura , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Miócitos de Músculo Liso/metabolismo , Diester Fosfórico Hidrolases/deficiência , Diester Fosfórico Hidrolases/genética , Pirofosfatases/deficiência , Pirofosfatases/genética , Fator de Transcrição CHOP/deficiência , Fator de Transcrição CHOP/genética , Túnica Íntima/patologia , Túnica Média/patologia , Resposta a Proteínas não Dobradas/fisiologia , Calcificação Vascular/etiologia , Calcificação Vascular/genética , Calcificação Vascular/metabolismo , Calcificação Vascular/patologia , Calcificação Vascular/fisiopatologiaRESUMO
OBJECTIVE: Intima formation involves smooth muscle cell (SMC) proliferation and migration that ultimately drives arterial stenosis, thrombosis, and ischemia in atherosclerosis, hypertension, and arterial revascularization. Receptor for advanced glycation endproducts (RAGE) is a transmembrane signaling receptor implicated in diabetic renal and vascular complications, and post-injury intima formation, partly via Signal transducer and activator of transcription 3 (STAT3) activation. The metabolic super-regulator Adenosine monophosphate kinase (AMPK) inhibits SMC proliferation and intima formation. AMPK activation is promoted by liver kinase B1 (LKB1), and LKB1 inhibits STAT3 activation. Here, we tested the hypothesis that RAGE promotes arterial intima formation by modulating both LKB1 and AMPK. METHODS AND RESULTS: RAGE ligands (the calgranulin S100A11, and glycated albumin) suppressed AMPK activation in conjunction with increased proliferation and migration of cultured SMCs. These effects were inhibited both by RAGE deficiency and by prior AMPK activation. In SMCs, RAGE ligands decreased LKB1 activity. Moreover, knockdown of both LKB1 and AMPK were associated with increased STAT3 phosphorylation levels. In response to murine carotid artery ligation, expression of RAGE and S100A11 increased, whereas AMPK and LKB1 activity decreased in situ. Conversely, LKB1 and AMPK activity increased in situ, and neointima formation was attenuated in Rage(-/-) mice. CONCLUSION: The linkage of decreased LKB1 and AMPK activity with increased STAT3 in SMCs mediates the capacity of RAGE ligand-induced signaling to promote neointima formation in response to arterial injury.
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Proteínas Quinases Ativadas por AMP/metabolismo , Lesões das Artérias Carótidas/enzimologia , Fígado/enzimologia , Músculo Liso Vascular/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Receptores Imunológicos/metabolismo , Proteínas Quinases Ativadas por AMP/genética , Animais , Artérias Carótidas/metabolismo , Artérias Carótidas/patologia , Lesões das Artérias Carótidas/genética , Lesões das Artérias Carótidas/patologia , Bovinos , Movimento Celular , Proliferação de Células , Células Cultivadas , Modelos Animais de Doenças , Regulação para Baixo , Ativação Enzimática , Técnicas de Silenciamento de Genes , Produtos Finais de Glicação Avançada , Ligantes , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Músculo Liso Vascular/patologia , Neointima , Fosforilação , Proteína Fosfatase 2/metabolismo , Proteínas Serina-Treonina Quinases/genética , Receptor para Produtos Finais de Glicação Avançada , Receptores Imunológicos/deficiência , Receptores Imunológicos/genética , Proteínas S100/metabolismo , Fator de Transcrição STAT3/metabolismo , Albumina Sérica/metabolismo , Transdução de Sinais , Albumina Sérica GlicadaRESUMO
BACKGROUND: Cysteine protease cathepsins are important in extracellular matrix protein degradation, cell apoptosis, and angiogenesis. Mice lacking cathepsins are protected from tumor progression in several animal models, suggesting that the regulation of cathepsin activities controls the growth of various malignant tumors. METHODS AND RESULTS: We tested the role of cathepsins using a mouse model of multistage epithelial carcinogenesis, in which the human keratin-14 promoter/enhancer drove the expression of human papillomavirus type 16 (HPV16) early region E6/E7 transgenes. During the progression of premalignant dysplasia, we observed increased expression of cysteine protease cathepsin S, but concomitantly reduced expression of cathepsin endogenous inhibitor cystatin C in the skin tissue extract. Absence of cystatin C in these transgenic mice resulted in more progression of dysplasia to carcinoma in situ on the face, ear, chest, and tail. Chest and ear skin extract real time PCR and immunoblot analysis, mouse serum sample ELISA, tissue immunohistological analysis, and tissue extract-mediated in vitro elastinolysis and collagenolysis assays demonstrated that cystatin C deficiency significantly increased cathepsin expression and activity. In skin from both the chest and ear, we found that the absence of cystatin C reduced epithelial cell apoptosis but increased proliferation. From the same tissue preparations, we detected significantly higher levels of pro-angiogenic laminin 5-derived γ2 peptides and concurrently increased neovascularization in cystatin C-deficient mice, compared to those from wild-type control mice. CONCLUSION: Enhanced cathepsin expression and activity in cystatin C-deficient mice contributed to the progression of dysplasia by altering premalignant tissue epithelial proliferation, apoptosis, and neovascularization.
Assuntos
Cistatina C/deficiência , Epiderme/metabolismo , Queratina-14/genética , Proteínas Oncogênicas Virais/genética , Animais , Apoptose , Catepsinas/genética , Catepsinas/metabolismo , Proliferação de Células , Cistatina C/genética , Cistatina C/metabolismo , Progressão da Doença , Epiderme/patologia , Feminino , Papillomavirus Humano 16/genética , Humanos , Hiperplasia , Immunoblotting , Masculino , Camundongos , Camundongos da Linhagem 129 , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Proteínas E7 de Papillomavirus/genética , Regiões Promotoras Genéticas/genética , Proteínas Repressoras/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Neoplasias Cutâneas/genética , Neoplasias Cutâneas/metabolismo , Neoplasias Cutâneas/patologiaRESUMO
Polymorphisms in cytokine genes may contribute to increased susceptibility to different cancers. The aim of this paper is to investigate the association of IL-8-251A/T polymorphism and Helicobacter pylori (H. pylori) infection with the risk of developing gastric cardiac adenocarcinoma (GCA) in the south of Taihang Mountain, a high-incidence area of esophageal cancer in China. The IL-8-251 A/T polymorphism was genotyped in 519 cases of GCA and 504 healthy controls. The H. pylori infection in GCA patients and controls was detected by rapid urease test (RUT), histopathology or (14)C-urea breath test ((14)C-UBT). The results showed that family history of upper gastrointestinal cancer (UGIC) and H. pylori infection significantly increased the risk of developing GCA. The overall genotype and allelotype distributions of IL-8 promoter SNPs in GCA patients were significantly different from those in healthy controls. Compared with TT genotype, AA genotype significantly elevated the risk of developing GCA. The stratification analysis revealed that, compared with the TT genotype, the AA genotype significantly elevated the risk of developing GCA in both positive family history of UGIC and H. pylori infection subgroups. This study provides evidence to support a relationship of increased susceptibility to GCA in individuals of the south Taihang Mountain region with IL-8 251 AA genotype, especially for those individuals who have family history of UGIC or H. pylori infection.
Assuntos
Adenocarcinoma/genética , Predisposição Genética para Doença , Infecções por Helicobacter/complicações , Interleucina-8/genética , Polimorfismo de Nucleotídeo Único/genética , Neoplasias Gástricas/epidemiologia , Neoplasias Gástricas/genética , Adenocarcinoma/complicações , Adenocarcinoma/epidemiologia , Idoso , Sequência de Bases , China/epidemiologia , Demografia , Feminino , Infecções por Helicobacter/genética , Helicobacter pylori/fisiologia , Humanos , Incidência , Masculino , Pessoa de Meia-Idade , Dados de Sequência Molecular , Reação em Cadeia da Polimerase , Lesões Pré-Cancerosas/epidemiologia , Lesões Pré-Cancerosas/genética , Fatores de Risco , Análise de Sequência de DNA , Neoplasias Gástricas/complicaçõesRESUMO
DNMT3B is an important enzyme to modulate the methylation status in mammalian cells. The aim of this study is to investigate the correlation of the DNMT3B G39179T polymorphism with the susceptibilities of colorectal adenomatous polyps and adenocarcinoma. This case-control study included 146 colorectal adenomatous polyps, 170 colorectal adenocarcinoma patients, and 157 normal controls. DNMT3B polymorphism was analyzed by polymerase chain reaction-restriction fragment length polymorphism analysis. Family history of colorectal cancer significantly increases the risk of developing colorectal adenomatous polyps and adenocarcinoma. The genotype frequency of DNMT3B polymorphism (T/T and G/T + G/G) in adenocarcinoma patients was significantly different from that in controls (P value = 0.01). Compared with DNMT3B T/T genotype, the G allelotype (G/T + G/G genotype) had lower risk to develop colorectal adenocarcinoma (OR = 0.50, 95% CI = 0.29-0.87); while there was no significant difference between the colorectal adenomatous polyps patients and controls (OR = 0.63, 95% CI = 0.37-1.09), although descending tendency could be found in this polyps group. In the stratification analysis, a significant association was confined to subgroups of age < 55 (OR = 0.31, 95% CI = 0.12-0.84) and males (OR = 0.35, 95% CI = 0.17-0.71). Meanwhile, combined G/T + G/G genotypes were found to have a lower risk in non-drinkers to develop both colorectal adenomatous polyps and adenocarcinoma (OR = 0.54, 95% CI = 0.31-0.96 and OR = 0.48, 95% CI = 0.27-0.84, respectively). This study also showed a distinct difference in the distribution of DNMT3B G39179T SNP in different ethnics. DNMT3B G39179T SNP may be a potential genetic susceptibility factor for adenocarcinoma of the colon, especially in younger Chinese Han non-drinker men.
Assuntos
Adenocarcinoma/genética , Pólipos Adenomatosos/genética , Neoplasias Colorretais/enzimologia , Neoplasias Colorretais/genética , DNA (Citosina-5-)-Metiltransferases/genética , Predisposição Genética para Doença , Polimorfismo de Nucleotídeo Único/genética , Adenocarcinoma/enzimologia , Pólipos Adenomatosos/enzimologia , Estudos de Casos e Controles , Demografia , Etnicidade/genética , Feminino , Frequência do Gene/genética , Humanos , Masculino , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase , DNA Metiltransferase 3BRESUMO
OBJECTIVES: To investigate lipoprotein(a) (Lp(a)) serum levels in patients with aortic dissection and the influence of smoking on the level of Lp(a) in aortic dissection patients. METHODS: An age-and sex-matched case-control study was conducted. Lp(a) levels in patients with aortic dissection (n = 52) and healthy subjects (n = 104) were studied. The strength of associations between Lp(a) serum levels and aortic dissection was assessed by means of multivariate logistic regression analysis. RESULTS: Patients with aortic dissection had significantly higher Lp(a) serum levels (median, 17.6 mg/dl; range, 6.4-88.7 mg/dl) compared to healthy individuals (median, 12.4 mg/dl; range, 4.9-26.4 mg/dl) (p = 0.005). The Lp(a) concentration in non-smoking patients with aortic dissection (median, 19.1 mg/dl, range, 10.5-88.7 mg/dl) significantly surpassed that of the smoking patients with aortic dissection of comparable age (median, 10.7 mg/dl; range, 6.4-22.1 mg/dl) (p < 0.0001). Multivariate analysis confirmed an independent association between Lp(a) and aortic dissection in the non-smoking population (p = 0.001). CONCLUSIONS: Serum Lp(a) level is significantly elevated in non-smoking patients with aortic dissection independently of other cardiovascular risk factors. Therefore, determination of Lp(a) levels may be important in identifying subjects at risk of aortic dissection among nonsmokers.