RESUMO
OBJECTIVE: Medication-related osteonecrosis of the jaw (MRONJ) is the main adverse side effect of bisphosphonates (BPs), mainly owing to the inhibitory effect of BPs on osteoclastogenesis. CircRNAs were identified to be an important factor in regulating cellular processes. The aim of this study was to explore the effect of mmu_circ_0001066 on BP-inhibited osteoclastogenesis. MATERIALS AND METHODS: The expression of MRONJ-related miRNA in RANKL-induced RAW264.7 cells treated with BP was analyzed using qRT-PCR analysis. Bioinformatics techniques were applied to screen potential circRNAs. Tartrate-resistant acid phosphatase (TRAP) staining and bone resorption assays were used to examine the effect of mmu_circ_0001066 on osteoclastogenesis. Bioinformatics analysis, luciferase reporter assays, and Western blotting assays were performed to investigate the underlying mechanism. RESULTS: Four MRONJ-related miRNAs were upregulated in BP-treated RAW264.7 cells, and the expression of mmu_circ_0001066 was negatively correlated with those of MRONJ-related miRNAs. Furthermore, the upregulation of mmu_circ_0001066 partially attenuated the inhibitory effect of BP on osteoclastogenesis in RAW264.7 cells. Mechanistically, upregulated miR-16 suppressed osteoclastogenesis and miR-16 inhibitor increased osteoclastogenesis. Furthermore, we have identified that miR-16 is a downstream effector of mmu_circ_0001066. CONCLUSION: Our results suggest that mmu_circ_0001066 played an important role in the BP-mediated suppression of osteoclastogenesis, which lays a foundation for identifying mmu_circ_0001066 as a potential biomarker for MRONJ.
Assuntos
MicroRNAs , RNA Circular , Difosfonatos/farmacologia , MicroRNAs/genética , MicroRNAs/metabolismo , Osteogênese/genética , RNA Circular/genética , Regulação para CimaRESUMO
PURPOSE: Recent ï¬ndings have shown that circRNA dysregulation was involved in the development of many types of cancer. However, our knowledge of circRNA in oral squamous cell carcinoma (OSCC) remains elusive. METHODS: Here, we explored whether ciRS-7 could function as a ceRNA in promoting metastasis of OSCC via regulating miR-7 activity. The expression levels of ciRS-7 and miR-7 were examined in clinical samples and cell lines by qRT-PCR, and the effects of ectopic expression of ciRS-7 and miR-7 on cell proliferation, migration and invasion were assessed in vitro and in vivo. The effects of ciRS-7 on miR-7 activity were investigated by means of luciferase reporter assay, qRT-PCR and Western blot. In addition, the effects of miR-7 mediated ciRS-7 on the levels of MAPK/AKT signaling proteins were evaluated by Western blot. RESULTS: We found that ciRS-7 was highly expressed in OSCC tissues and cell lines compared with normal counterparts. Ectopic expression of ciRS-7 significantly promoted OSCC cell proliferation, migration and invasion through in vitro and in vivo. Based on bioinformatics analysis, qRT-PCR, Western blot and luciferase reporter assays, we determined that ciRS-7 functioned as a sponge for miR-7, resulting in attenuation of miR-7 targets RAF-1 and PIK3CD, which are core components of the MAPK/AKT signaling pathways. Moreover, miR-7 correlated with perineural and lymphovascular invasion in OSCC patients. Further experiments demonstrated that ciRS-7 overexpression could attenuate the anti-tumor effects of miR-7 on OSCC cells. CONCLUSIONS: Our results suggested that ciRS-7 can interact directly with miR-7, resulting in upregulation of RAF-1/PIK3CD expression and enhancing metastatic progression of OSCC.
Assuntos
Carcinoma de Células Escamosas/genética , Classe I de Fosfatidilinositol 3-Quinases/metabolismo , Progressão da Doença , Sistema de Sinalização das MAP Quinases , Neoplasias Bucais/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteínas Proto-Oncogênicas c-raf/metabolismo , RNA Longo não Codificante/metabolismo , Animais , Sequência de Bases , Carcinogênese/patologia , Carcinoma de Células Escamosas/patologia , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , Camundongos Endogâmicos BALB C , Camundongos Nus , MicroRNAs/genética , MicroRNAs/metabolismo , Neoplasias Bucais/patologia , Invasividade Neoplásica , Metástase Neoplásica , RNA Longo não Codificante/genética , Regulação para Cima/genéticaRESUMO
OBJECTIVE: Increasing evidence points toward the key function of circular RNAs (circRNAs) in various carcinomas. This study aimed to identify aberrant expression of hsa_circ_0072387 in oral squamous cell carcinoma and probe its clinical significance. MATERIALS AND METHODS: Real-time quantitative reverse transcription-polymerase chain reaction (qRT-PCR) was used to assess hsa_circ_0072387 expression levels in 63 paired OSCC tissues and three OSCC cell lines. The area under the receiver operator characteristic (ROC) curve was plotted to assess any potential clinical significance. RESULTS: Our data showed that hsa_circ_0072387 expression in OSCC was significantly downregulated compared with adjacent normal tissues (p < 0.001). Compared to human normal oral keratinocyte cell, the levels of hsa_circ_0072387 were lower in three OSCC cell lines (SCC25, SCC15, CAL27). More significantly, hsa_circ_0072387 expression was associated with the TNM stage in OSCC (p = 0.050). The area under the ROC curve reached up to 0.746. Based on bioinformatics, hsa-miR-129-3p, hsa-miR-141-3p, and hsa-miR-29-3p were predicted to be potential miRNAs binding with hsa_circ_0072387. Furthermore, hsa-miR-129-3p, hsa-miR-141-3p, and hsa-miR-29-3p were involved in multiple tumor-related signaling pathways. CONCLUSION: Our finding suggested that lower expression of has_circ_0072387 could be a key circRNA in OSCC and serve as a potential biomarker in OSCC diagnosis and therapeutic targets.
Assuntos
Biomarcadores/metabolismo , Carcinoma de Células Escamosas/diagnóstico , MicroRNAs , Neoplasias Bucais/diagnóstico , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/metabolismo , Biologia Computacional , Humanos , Neoplasias Bucais/genética , Neoplasias Bucais/metabolismo , RNA , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
PTENp1, non-coding RNA (ncRNA) pseudogene, is involved in oral squamous cell carcinoma (OSCC). The precise effects mediated by PTENp1 transcripts within intricate regulatory networks involving molecular interactions with ancestral gene PTEN and tumorigenicity in OSCC remain unclear. Here, we found that PTENp1 was aberrantly expressed in OSCC. There was a positive correlation between the expression levels of PTENp1 and PTEN. Further, we showed that PTENp1 acted as a competing endogenous RNA that protects PTEN transcripts from being inhibited by miR-21, and consequently inhibited proliferation and colony formation and triggered S-G2/M cell cycle arrest through the AKT pathway. Also, the homogeneous relationship between expression of PTENp1 and PTEN was confirmed in OSCC tumor xenografts. Finally, low expression of PTENp1 and PTEN was negatively associated with histological differentiation and OSCC prognosis. The present work provided the first evidence for the extraordinary crosstalk among PTENp1, PTEN, and miR-21, and rendered a new light on the treatment of OSCC. © 2016 Wiley Periodicals, Inc.
Assuntos
Carcinoma de Células Escamosas/genética , Regulação Neoplásica da Expressão Gênica , MicroRNAs/genética , Neoplasias Bucais/genética , PTEN Fosfo-Hidrolase/genética , Pseudogenes/genética , RNA não Traduzido/genética , Animais , Carcinoma de Células Escamosas/diagnóstico , Carcinoma de Células Escamosas/metabolismo , Carcinoma de Células Escamosas/patologia , Ciclo Celular , Linhagem Celular Tumoral , Proliferação de Células , Feminino , Humanos , Masculino , Camundongos Endogâmicos BALB C , Pessoa de Meia-Idade , Boca/metabolismo , Boca/patologia , Neoplasias Bucais/diagnóstico , Neoplasias Bucais/metabolismo , Neoplasias Bucais/patologia , Prognóstico , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de SinaisRESUMO
BACKGROUND: High mobility group AT-hook 2 (HMGA2) and pleomorphic adenoma gene 1(PLAG1) have been demonstrated to be elevated in many malignant tumors. However, the aim of this study was to evaluate HMGA2 and PLAG1 levels in blood as a non-invasive biomarker for oral squamous cell carcinoma (OSCC) diagnosis. METHODS: qRT-PCR was performed to measure circulating HMGA2 and PLAG1 levels in OSCC patients (n=43) and matched cancer-free blood control group (n=21). Clinical data of all patients were recorded. RESULTS: Circulating HMGA2 and PLAG1 in the 43 OSCC patients was significantly higher than in control group (P<.001, P=.038, respectively). Furthermore, HMGA2 expression in OSCC patients with poor-moderate differentiation was increased compared with well-differentiated group. However, no significant differences in PLAG1 expression were detected when differentiation was considered. In addition, the receiver operating characteristic (ROC) curve analysis for circulating HMGA2 revealed an area under the ROC curve of 0.876 (95% confidence interval, 0.793-0.959; P<.001) with 65.1% sensitivity and 100% specificity in discriminating OSCC from controls at a cutoff value of 14.380, demonstrating significant diagnostic value for OSCC. CONCLUSION: Circulating HMGA2 levels are increased in OSCC patients and may potentially serve as a significant index to evaluate OSCC diagnosis.
Assuntos
Carcinoma de Células Escamosas/sangue , Carcinoma de Células Escamosas/diagnóstico , Proteínas de Ciclo Celular/sangue , Proteína HMGA2/sangue , Neoplasias Bucais/sangue , Neoplasias Bucais/diagnóstico , Fatores de Transcrição/sangue , Proteínas Supressoras de Tumor/sangue , Biomarcadores Tumorais/sangue , Feminino , Humanos , Masculino , Pessoa de Meia-IdadeRESUMO
Rescuing the function of mutant p53 protein is an attractive cancer therapeutic strategy. Small molecule CP-31398 was shown to restore mutant p53 tumor suppressor functions in cancer cells. Here, we determined the effects of CP-31398 on the growth of p53-mutated colorectal cancer (CRC) cells in vitro and in vivo. CRC cells which carry p53 mutation in codon 273 were treated with CP-31398 and the control, and the effects of CP-31398 on cell cycle, cell apoptosis, and proliferation were determined. The expression of p53-responsive downstream genes was evaluated by quantitative reverse transcriptase PCR (RT-PCR) and Western blot. CP-31398 was administrated into xenograft tumors created by the inoculation of HT-29 cells, and then the effect of CP-31398 on the growth of xenograft tumors was examined. CP-31398 induced p53 downstream target molecules in cultured HT-29 cells, which resulted in the inhibition of CRC cell growth assessed by the determination of cell cycle, apoptosis, and cell proliferation. In xenograft tumors, CP-31398 modulated the expression of Bax, Bcl-2, caspase 3, cyclin D, and Mdm2 and then blocked the growth of xenograft tumors. CP-31398 would be developed as a therapeutic candidate for p53-mutated CRC due to the restoration of mutant p53 tumor suppressor functions.
Assuntos
Neoplasias Colorretais/tratamento farmacológico , Pirimidinas/farmacologia , Proteína Supressora de Tumor p53/genética , Animais , Apoptose/efeitos dos fármacos , Apoptose/genética , Caspase 3/genética , Ciclo Celular/efeitos dos fármacos , Ciclo Celular/genética , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/genética , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Ciclina D/genética , Células HT29 , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Proteínas Proto-Oncogênicas c-bcl-2/genética , Proteínas Proto-Oncogênicas c-mdm2/genética , Ensaios Antitumorais Modelo de Xenoenxerto/métodos , Proteína X Associada a bcl-2/genéticaRESUMO
INTRODUCTION: The diagnosis of malignant ascites is a challenging problem in clinical practice, non-invasive techniques should be developed to improve diagnostic accuracy. The diagnostic performances of tumor markers in malignant ascites remained unsettled. Our aim was to evaluate diagnostic performance of tumor markers in differential diagnosis of benign and malignant ascites. MATERIAL AND METHODS: A total of 437 patients were enrolled, and the relevant parameters of the patients were analyzed for the differentiation of benign ascites from malignant ascites. RESULTS: At the predetermined cutoff values of tumor makers, tumor markers in ascitic fluid showed better diagnostic performance than those in serum. Combined use of tumor markers and the cytology increased the diagnostic yield of the latter by 37%. In cytologically negative malignant ascites, tumor markers provided assistance in differentiating malignant ascites from benign ascites, and the combination of ascitic tumor markers yielded 86% sensitivity, 97% specificity. CONCLUSION: Use of a panel of tumor markers exhibited excellent diagnostic performance in diagnosing malignant ascites, which indicated the detection of tumor markers may represent a beneficial adjunct to cytology, thus guiding the selection of patients who might benefit from further invasive procedures.
Assuntos
Ascite/diagnóstico , Líquido Ascítico/química , Biomarcadores Tumorais/análise , Hipertensão Portal/diagnóstico , Cirrose Hepática/diagnóstico , Neoplasias/diagnóstico , Ascite/etiologia , Ascite/metabolismo , Líquido Ascítico/citologia , Antígeno Ca-125/metabolismo , Antígeno CA-19-9/metabolismo , Antígeno Carcinoembrionário/metabolismo , Estudos de Coortes , Diagnóstico Diferencial , Feminino , Humanos , Hipertensão Portal/complicações , Cirrose Hepática/complicações , Masculino , Mucina-1/metabolismo , Neoplasias/complicações , Antígeno Prostático Específico/metabolismo , Sensibilidade e Especificidade , alfa-Fetoproteínas/metabolismoRESUMO
AIMS: To investigate the clinical value and performance of [18F]AlF-NOTA-FAPI-04 PET/CT in assessing early-stage liver fibrosis in liver transplantation (LT) recipients. METHODS: A prospective study including 17 LT recipients and 12 chronic Hepatitis B (CHB) patients was conducted. All patients received liver biopsy, transient elastography (TE), and [18F]AlF-NOTA-FAPI-04 PET/CT. On [18F]AlF-NOTA-FAPI-04 PET/CT scans, the liver parenchyma's maximum standardized uptake values (SUVmax) were measured. The receiver operating characteristic (ROC) curve analysis was applied to determine the diagnostic efficacy of [18F]AlF-NOTA-FAPI-04 PET/CT in early-stage liver fibrosis (S1-S2) compared with the diagnostic performance of TE. RESULTS: Among those 29 patients enrolled in this study, 15(51.7%) had fibrosis S0, 10(34.5%) had S1, and 4(13.8%) had S2, respectively. The SUVmax of patients with early-stage liver fibrosis was significantly higher than those without liver fibrosis in LT recipients and CHB patients (P = 0.004, P = 0.02). In LT recipients, a SUVmax cut-off value of 2.0 detected early-stage liver fibrosis with an AUROC of 0.92 (P = 0.006), and a liver stiffness measurements (LSM) score cut-off value of 8.2 kPa diagnosed early-stage liver fibrosis with an AUROC of 0.80 (P = 0.012). In CHB patients, a SUVmax cut-off value of 2.7 detected early-stage liver fibrosis with an AUROC of 0.94 (P < 0.001) and an LSM scores cut-off value of 8.4 kPa diagnosed early-stage liver fibrosis with an AUROC of 0.91 (P < 0.001). CONCLUSION: [18F]AlF-NOTA-FAPI-04 PET/CT could be applied to evaluate early-stage liver fibrosis in LT recipients and CHB patients properly, with the potential additional advantages in monitoring and predicting complications after LT.
Assuntos
Hepatite B Crônica , Cirrose Hepática , Transplante de Fígado , Tomografia por Emissão de Pósitrons combinada à Tomografia Computadorizada , Compostos Radiofarmacêuticos , Humanos , Masculino , Feminino , Cirrose Hepática/diagnóstico por imagem , Estudos Prospectivos , Pessoa de Meia-Idade , Tomografia por Emissão de Pósitrons combinada à Tomografia Computadorizada/métodos , Hepatite B Crônica/diagnóstico por imagem , Hepatite B Crônica/complicações , Adulto , Técnicas de Imagem por Elasticidade/métodos , Idoso , Fígado/diagnóstico por imagem , Fígado/patologiaRESUMO
OBJECTIVE: To investigate the association of gastric emptying with ghrelin, obestatin and GHSR, GPR-39 in hypothalamus of diabetic rats. METHODS: Sixty Wistar rats were randomly divided into three groups: a normal control group (NC, n = 20), a diabetes mellitus group (DM, n = 20) induced by intraperitoneal injection of streptozotocin (STZ) and an insulin treated group (INS, n = 20). After two and six weeks of STZ injection, gastric emptying was measured by intragastric administration of phenol red, ghrelin and obestatin in hypothalamus measured by ELISA (enzyme-linked immunosorbent assay) and GHSR and GPR-39 by RT-PCR (reverse transcription-polymerase chain reaction). RESULTS: After two weeks of STZ injection, gastric emptying (%) (74 +/- 8, 40 +/- 5), ghrelin level(ng/g) (52 +/- 9, 51 +/- 7) and ratio of ghrelin/obestatin (3.8 +/- 1.0, 2.8 +/- 1.0) increased significantly in DM and INS groups compared to those in NC group [32% +/- 7%, (39 +/- 11) ng/g, 2.1 +/- 0.8, all P < 0.05]. Obestatin level(ng/g) (14.2 +/- 2.0) of hypothesis decreased significantly in DM group as compared to those in NC group (21.7 +/- 4.7) while GHSR/beta-actin increased significantly (1.26 +/- 0.46 vs 0.77 +/- 0.21, P < 0.05). Gastric emptying was positively correlated with ghrelin, ghrelin/obestatin and GHSR/beta-actin of hypothalamus (r = 0.49; r = 0.63; r = 0.73; P < 0.01). But there was a negative correlation with obestatin of hypothalamus (r = -0.74, P < 0.01). After six weeks of STZ injection, gastric emptying (78.97% +/- 8.13% vs 44.06% +/- 5.06%) increased significantly in DM and INS groups as compared to those in NC group (35.06% +/- 3.91%, P < 0.01). Gastric emptying was positively correlated with ghrelin/obestatin of hypothalamus (r = 0.40, P < 0.05). There was no detection of GPR-39 in hypothalamus. CONCLUSION: The rapid gastric emptying may be due to the rising levels of ghrelin and GHSR in hypothalamus during early hyperglycemia. And the duration of hyperglycemia is affected by the rising ratio of ghrelin/obestatin.
Assuntos
Diabetes Mellitus Experimental/metabolismo , Esvaziamento Gástrico , Grelina/metabolismo , Hipotálamo/metabolismo , Receptores de Grelina/metabolismo , Animais , Diabetes Mellitus Experimental/fisiopatologia , Masculino , Ratos , Ratos WistarRESUMO
OBJECTIVE: This study aimed to investigate the specific regulatory roles of microRNA-219-5p (miR-219-5p) on ulcerative colitis (UC), and reveal the potential mechanisms relating with the differentiation of Treg/Th17 cells. METHODS: The mouse model of chronic UC was established by oral administration of 3% dextran sodium sulfate for three cycles. After intravenous injected with lentivirus (LV)-miR-219-5p for 24 h, the disease activity index (DAI), colon length, as well as the serum levels of Interleukin (IL)-6, -17A, -21, and -23 were measured. In addition, the histopathological changes in colon tissues were observed by Hematoxylin-eosin staining. The differentiation of Treg/Th17 cells was detected by Flow cytometry, and the expression of retinoic acid-related orphan receptor (RORrt), signal transducer and activator of transcription 3 (STAT3), and forkhead box p3 (Foxp3) were detected by quantitative real-time PCR and Western blot. RESULTS: MiR-219-5p was downregulated in colonic mucosal tissues of UC mice (P < 0.05). UC mice injected with LV-miR-219-5p exhibited significantly relieved histopathological changes of colon tissues, increased colon length, decreased DAI, as well as decreased serum levels of IL-6, -17A, -21, and -23 (P < 0.05). In addition, the injection of LV-miR-219-5p significantly increased the percentage of Treg cells via upregulating Foxp3, and decreased the percentage of Th17 cells via downregulating RORrt and STAT3 in UC mice (P < 0.05). CONCLUSION: The upregulation of miR-219-5p relieved the colonic damage and inflammation of UC through balancing the differentiation of Treg/Th17 cells.
Assuntos
Colite Ulcerativa , MicroRNAs , Animais , Diferenciação Celular , Colite Ulcerativa/induzido quimicamente , Colite Ulcerativa/genética , Humanos , Camundongos , MicroRNAs/genética , Linfócitos T Reguladores , Células Th17 , Regulação para CimaRESUMO
Retinol-binding protein 1 (RBP1) is involved in several physiological functions, including the regulation of the metabolism and retinol transport. Studies have shown that it plays an important role in the pathogenesis of several types of cancer. However, the role of RBP1 and its correlation with autophagy in oral squamous cell carcinoma (OSCC) pathogenesis remain unknown. In this study, RBP1 was identified as the most significantly upregulated DEPs with a >2-fold change in OSCC samples when compared to normal tissues through iTRAQ-based proteomics analysis coupled with 2D LC-MS/MS. RBP1 overexpression was significantly associated with malignant phenotypes (differentiation, TNM stage, and lymphatic metastasis) of OSCC. In vitro experiments demonstrated that RBP1 was significantly increased in OSCC tissues and cell lines compared with control group. RBP1 overexpression promoted cell growth, migration, and invasion of OSCC cells. Silencing of RBP1 suppressed tumor formation in xenografted mice. We further demonstrated that the RBP1-CKAP4 axis was a critical regulator of the autophagic machinery in OSCC, inactivation of autophagy rescued the RBP1-CKAP4-mediated malignant biological behaviors of OSCC cells. Overall, a mechanistic link was provided by RBP1-CKAP4 between primary oncogenic features and the induction of autophagy, which may provide a potential therapeutic target that warrants further investigation for treatment of OSCC.
Assuntos
Autofagia , Carcinogênese/metabolismo , Carcinoma de Células Escamosas/patologia , Progressão da Doença , Proteínas de Membrana/metabolismo , Neoplasias Bucais/patologia , Proteínas Celulares de Ligação ao Retinol/metabolismo , Transdução de Sinais , Animais , Proteína 5 Relacionada à Autofagia/metabolismo , Carcinogênese/patologia , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/ultraestrutura , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Regulação para Baixo/genética , Regulação Neoplásica da Expressão Gênica , Humanos , Camundongos Endogâmicos BALB C , Camundongos Nus , Neoplasias Bucais/genética , Neoplasias Bucais/ultraestrutura , Invasividade Neoplásica , Ligação Proteica , Vacúolos/metabolismo , Vacúolos/ultraestruturaRESUMO
BACKGROUND: Globally, colorectal cancer (CRC) affects more than 1 million people each year. In addition to non-modifiable and other environmental risk factors, Fusobacterium nucleatum infection has been linked to CRC recently. In this study, we explored mechanisms underlying the role of Fusobacterium nucleatum infection in the progression of CRC in a mouse model. METHODS: C57BL/6 J-Adenomatous polyposis coli (APC) Min/J mice [APC (Min/+)] were treated with Fusobacterium nucleatum (109 cfu/mL, 0.2 mL/time/day, i.g., 12 weeks), saline, or FadA knockout (FadA-/-) Fusobacterium nucleatum. The number, size, and weight of CRC tumors were determined in isolated tumor masses. The human CRC cell lines HCT29 and HT116 were treated with lentiviral vectors overexpressing chk2 or silencing ß-catenin. DNA damage was determined by Comet assay and γH2AX immunofluorescence assay and flow cytometry. The mRNA expression of chk2 was determined by RT-qPCR. Protein expression of FadA, E-cadherin, ß-catenin, and chk2 were determined by Western blot analysis. RESULTS: Fusobacterium nucleatum treatment promoted DNA damage in CRC in APC (Min/+) mice. Fusobacterium nucleatum also increased the number of CRC cells that were in the S phase of the cell cycle. FadA-/- reduced tumor number, size, and burden in vivo. FadA-/- also reduced DNA damage, cell proliferation, expression of E-cadherin and chk2, and cells in the S phase. Chk2 overexpression elevated DNA damage and tumor growth in APC (Min/+) mice. CONCLUSIONS: In conclusion, this study provided evidence that Fusobacterium nucleatum induced DNA damage and cell growth in CRC through FadA-dependent activation of the E-cadherin/ß-catenin pathway, leading to up-regulation of chk2.
Assuntos
Carcinogênese/genética , Quinase do Ponto de Checagem 2/genética , Neoplasias Colorretais/genética , Fusobacterium nucleatum/genética , Animais , Proliferação de Células/genética , Neoplasias Colorretais/microbiologia , Neoplasias Colorretais/patologia , Dano ao DNA/genética , Modelos Animais de Doenças , Progressão da Doença , Fusobacterium nucleatum/patogenicidade , Células HCT116 , Células HT29 , Humanos , Camundongos , Camundongos Endogâmicos C57BLRESUMO
AIM: Previous studies indicated that cerebral ischemia/reperfusion injury (CI/RI) could induce behavioral deficits. Single treatment of vagus nerve stimulation (VNS) or curcumin is reported to restore CI/RI-induced behavioral deficits. However, the synergic effect remains unclear. MATERIALS AND METHODS: Rats were divided into 6 groups: sham, CI/RI, VNS, CI/RIâ¯+â¯VNS, VNSâ¯+â¯curcumin and CI/RIâ¯+â¯VNSâ¯+â¯curcumin groups. Each group was further divided into three or four subgroups for further assessments. In specific, Morris water maze task and shuttle box test were used to evaluate cognitive capacity. Rota-rod test, neurological deficits scores, 2,3,5-triphenyltetrazolium chloride staining, TUNEL staining were performed to estimate motor capacity, neurological deficits, the size of infarct volume and neural apoptosis, respectively. Finally, the expressions of apoptosis-associated proteins and key kinases in the AKT/extracellular signal-regulated kinase-2 (ERK2) pathway were measured by Western blot analysis. RESULTS: Combination of curcumin and VNS significantly restored the CI/RI-induced cognitive and motor impairments compared with the CI/RIâ¯+â¯VNS group (Pâ¯<â¯0.05 and Pâ¯<â¯0.01). Moreover, combination of curcumin and VNS significantly lowered CI/RI-induced neurological deficits, infract volume, neural apoptosis (all Pâ¯<â¯0.05) and inflammatory cytokines release (Pâ¯<â¯0.05 and Pâ¯<â¯0.01) when compared to the CI/RIâ¯+â¯VNS group. Additionally, the phosphorylation levels of AKT and ERK2 were both increased by combination of curcumin and VNS compared with the CI/RIâ¯+â¯VNS group. CONCLUSION: Combination of curcumin and VNS restored CI/RI-induced behavioral deficits by inhibiting apoptosis and inflammatory response. Besides, the AKT/ERK2 pathway might be implicated.
Assuntos
Anti-Inflamatórios não Esteroides/administração & dosagem , Isquemia Encefálica/terapia , Curcumina/administração & dosagem , Aprendizagem em Labirinto/efeitos dos fármacos , Traumatismo por Reperfusão/terapia , Estimulação do Nervo Vago/métodos , Animais , Isquemia Encefálica/metabolismo , Isquemia Encefálica/psicologia , Terapia Combinada/métodos , Mediadores da Inflamação/antagonistas & inibidores , Mediadores da Inflamação/metabolismo , Aprendizagem em Labirinto/fisiologia , Distribuição Aleatória , Ratos , Ratos Sprague-Dawley , Traumatismo por Reperfusão/metabolismo , Traumatismo por Reperfusão/psicologiaRESUMO
The miR-200 family suppresses epithelial-mesenchymal transition by inhibiting ZEB1 and ZEB2 mRNA translation in several types of cancers. Kindlin-2 is a target gene of miR-200b and its expression level correlates positively to ZEB2 in oral squamous cell carcinoma (OSCC). Whether Kindlin-2 and ZEB2 share a competitive endogenous RNAs regulatory network in OSCC remains unclear. Here, we studied the expression levels of miR-200b, Kindlin-2, and ZEB2 and found direct interaction between miR-200b, ZEB2, and Kindlin-2 mRNA in OSCC. A series of experiments was performed to elucidate the role of miR-200b and Kindlin-2 in OSCC cells. To further investigate whether Kindlin-2 regulates ZEB2 as a "ceRNA", we utilized pools of siRNAs to deplete Kindlin-2 or ZEB2 in Tca-8113 cells. Significantly elevated expression levels of Kindlin-2 and ZEB2, down-regulated mRNA levels of miR-200b, and a positive correlation between Kindlin-2 and ZEB2 were found in OSCC cells. Additional results suggest that miR-200b directly targets ZEB2 and that Kindlin-2 3'UTR miR-200b repressed both the migration and invasive functionality of Tca-8113. Kindlin-2 and ZEB2 are involved in accelerated migration and invasion of Tca-113 cells in vitro and Kindlin-2 controlled ZEB2 expression. However, Kindlin-2-mediated ZEB2 regulation did not depend on miRNAs. These results indicate that Kindlin-2 does not act as ZEB2 ceRNA and modify the migration of Tca-8113 cells. Our results improve our understanding of the underlying molecular and cellular mechanisms of oral cancer metastasis.
RESUMO
OBJECTIVE: To study the inhibition of maxizyme (Mz) directed against the mutant-type p53 gene (mtp53) at codon 249 in exon 7 (AGG --> AGT) both in cell-free system and in MHCC97 cell lines. METHODS: Maxizyme and control mutant maxizyme (G5 --> A5) were designed by computer and cloned into the eukaryotic expression vector pBSKneoU6 (pU6Mz, pU6asMz). Mz was driven by T7 RNA polymerase promoter in vitro. In the cell lines, U6 promoter was driven by RNA PolIII. The mutant type p53 gene fragment was cloned into the pGEM-T vector under the T7 promoter control. The 32P-labeled mtp53 transcript was the target RNA. Cold maxizyme transcripts were incubated with 32P-labeled target RNA in vitro. pU6Mz was introduced into MHCC97 cells by Lipofectamine2000 and mtp53 expression was analyzed by RT-PCR and Western blot. RESULTS: In vitro cleavage showed that pU6Mz was very active with cleavage efficiency of 42% while pU6asMz was not. The wild type p53 was not cleaved. Partial down-regulation of mtp53 mRNA and mtp53 protein were observed in MHCC97 cells transfected with pU6Mz but not those with pU6asMz. The proliferation of MHCC cells was inhibited by MTT analysis. CONCLUSION: Our findings suggest that the chimeric U6 maxizyme against the mtp53 is a new promising gene therapeutic agent in treating hepatocellular carcinoma.
Assuntos
Carcinoma Hepatocelular/genética , Neoplasias Hepáticas/genética , Proteína Supressora de Tumor p53/genética , Linhagem Celular Tumoral , Terapia Genética/métodos , Vetores Genéticos , Humanos , Conformação de Ácido Nucleico , Mutação Puntual , Conformação Proteica , RNA Catalítico , RNA Mensageiro/síntese química , RNA Mensageiro/metabolismo , Proteínas Recombinantes de Fusão , Ribonuclease T1/farmacologiaRESUMO
BACKGROUND: The nutritional risk screening (NRS 2002) has been applied increasingly in patients who underwent abdominal surgery for nutritional risk assessment. However, the usefulness of the NRS 2002 for predicting is controversial. This meta-analysis was to examine whether a preoperative evaluation of nutritional risk by NRS 2002 provided prediction of postoperative outcomes in patients undergoing abdominal surgery. METHODS: A systematic literature search for published papers was conducted using the following online databases: MEDLINE, EMBASE, the Cochrane library, EBSCO, CRD databases, Cinahl, PsycInfo and BIOSIS previews. The pooled odds ratio (OR) or weight mean difference (WMD) was calculated using a random-effect model or a fix-effect model. RESULTS: Eleven studies with a total of 3527 patients included in this study. Postoperative overall complications were more frequent in nutritional risk patients versus patients without nutritional risk (the pooled OR 3.13 [2.51, 3.90] p<0.00001). The pooled OR of mortality for the nutritional risk group and non-nutritional risk group was 3.61 [1.38, 9.47] (p = 0.009). Furthermore, the postoperative hospital stay was significant longer in the preoperative nutritional risk group than in the nutritional normal group (WMD 5.58 [4.21, 6.95] p<0.00001). CONCLUSIONS: The present study has demonstrated that patients at preoperative nutritional risk have increased complication rates, high mortality and prolonged hospital stay after surgery. However, NRS 2002 needs to be validated in larger samples of patients undergoing abdominal surgery by better reference method.
Assuntos
Estado Nutricional/fisiologia , Complicações Pós-Operatórias/etiologia , Complicações Pós-Operatórias/fisiopatologia , Abdome/cirurgia , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Tempo de Internação , Masculino , Pessoa de Meia-Idade , Avaliação Nutricional , Período Pós-Operatório , Cuidados Pré-Operatórios , Estudos Prospectivos , Medição de Risco , Adulto JovemRESUMO
AIM: To evaluate the specific inhibition of maxizyme directing against mutant-type p53 gene (mtp53) at codon 249 in exon 7 (AGG-AGT) in vitro. METHODS: Two different monomers of anti-mtp53 maxizyme (maxizyme right MzR, maxizyme left MzL) and control mutant maxizyme (G(5)-A(5)) were designed by computer and cloned into vector pBSKU6 (pBSKU6MzR, pBSKU6MzL). After being sequenced, the restrictive endonuclease site in pBSKU6MzR was changed by PCR and then U6MzR was inserted into pBSKU6MzL, the recombinant vector was named pU6Mz and pU6asMz (mutant maxizyme). Mtp53 and wild-type p53 (wtp53) gene fragments were cloned into pGEM-T vector under the T7 promoter control. The (32)p-labeled mtp53 transcript was the target mRNA. Cold maxizyme transcripts were incubated with (32)p-labeled target RNA in vitro and radioautographed after denaturing polyacrylamide gel electrophoresis. RESULTS: In cell-free systems, pU6Mz showed a specific cleavage activity against target mRNA at 37 degrees and 25 mM MgCL(2). The cleavage efficiency of pU6Mz was 42 %, while pU6asMz had no inhibitory effect. Wtp53 was not cleaved by pU6Mz either. CONCLUSION: pU6Mz had a specific catalytic activity against mtp53 in cell-free system. These lay a good foundation for studying the effects of anti-mtp53 maxizyme in HCC cell lines. The results suggest that maxizyme may be a promising alternative approach for treating hepatocellular carcinoma containing mtp53.
Assuntos
Mutação Puntual , RNA Catalítico/genética , RNA Catalítico/metabolismo , Proteína Supressora de Tumor p53/genética , Carcinoma Hepatocelular/terapia , Sistema Livre de Células , Clonagem Molecular , Terapia Genética/métodos , Vetores Genéticos , Humanos , Técnicas In Vitro , Neoplasias Hepáticas/terapia , Conformação de Ácido Nucleico , RNA Catalítico/química , RNA Mensageiro/metabolismoRESUMO
AIM: To study the preparation and cleavage activity of HpRz directed against the transcript of HBV core gene in vitro. METHODS: HpRz gene designed by computer targeting the transcript of HBV core gene was cloned into the vector p1.5 between 5'-cis-Rz and 3'-cis-Rz. 32p-labeled HpRz transcript proved whether the vector fit for the preparation of hairpin ribozyme in vitro. 32p-labeled pKC transcript containing HBV core region as target-RNA was transcribed using T7 RNA polymerase and purified by denaturing PAGE. Cold HpRz transcript was incubated with 32p-labeled target-RNAs under different conditions and radio autographed after denaturing polyacrylamide gel electrophoresis. RESULTS: HpRz has the specific ability of cleavage of target RNA at 37 degrees and 12 mM MgCl2. Km=26.31 nmol/L, Kcat=0.18/min. These results revealed that the design of HpRz was correct. CONCLUSION: HpRz prepared in this study possesses specific catalytic activity from the identification of cleavage activity. These results indicate that hairpin ribozyme may intracellularly inhibit the replication of HBV, therefore it may become a novel potent weapon for the treatment of hepatitis B.
Assuntos
Vírus da Hepatite B/genética , RNA Catalítico/metabolismo , RNA Viral/metabolismo , Sequência de Bases , Ativação Enzimática , Terapia Genética , Hepatite B/terapia , Hepatite B/virologia , Antígenos do Núcleo do Vírus da Hepatite B/genética , Humanos , Técnicas In Vitro , Cinética , Dados de Sequência Molecular , Plasmídeos , Transcrição GênicaRESUMO
AIM: To study the preparation and cleavage activity of anti-transforming growth factor (TGF)beta1 U1 small nuclear (sn) RNA chimeric hammerhead ribozymes in vitro. METHODS: TGFbeta1 partial gene fragment was cloned into T-vector at the downstream of T7 promoter. (32)p-labeled TGFbeta1 partial transcripts as target RNA were transcribed in vitro and purified by denaturing polyacrylamide gel electrophoresis (PAGE). Anti-TGFbeta1 ribozymes were designed by computer, then synthetic ribozyme fragments were cloned into the U1 ribozyme vector pZeoU1EcoSpe containing U1 snRNA promoter/enhancer and terminator. (32)p-labeled U1 snRNA chimeric ribozyme transcripts were gel-purified, incubated with target-RNAs at different conditions and autoradiographed after running denaturing PAGE. RESULTS: Active U1snRNA chimeric ribozyme (U1Rz803) had the best cleavage activity at 50 degrees; at 37 degrees, it was active, K(m)=34.48 nmol/L, K(cat)=0.14 min(-1); while the point mutant ribozyme U1Rz803(m) had no cleavage activity, so these indicated the design of U1Rz803 was correct. CONCLUSION: U1Rz803 prepared in this study possessed the perfect specific catalytic cleavage activity. These results indicate U1 snRNA chimeric ribozyme U1Rz803 may suppress the expression of TGFbeta1 in vivo, therefore it may provide a new avenue for the treatment of liver fibrosis in the future.
Assuntos
RNA Catalítico/farmacologia , RNA Nuclear Pequeno/farmacologia , RNA/efeitos dos fármacos , Proteínas Recombinantes de Fusão/farmacologia , Fator de Crescimento Transformador beta/antagonistas & inibidores , Fator de Crescimento Transformador beta/genética , Animais , Linhagem Celular Transformada , Clonagem Molecular , Ratos , Fator de Crescimento Transformador beta1RESUMO
OBJECTIVE: To evaluate the mutations of lamivudine-resistance using oligonucleotide microarray in hepatitis B virus (HBV) infected patients. METHODS: A randomized clinical trial was conducted on 20 lamivudine-treated patients for 18 months and 10 patients as controls. The serum HBV DNA was amplified by PCR and the lamivudine-resistance mutations in YMDD region were assayed by 4 sites microarray developed before. RESULTS: This microarray could clearly differentiate the wide-type from mutated-type HBV with lamivudine-resistance mutations. The rate of mutations in YMDD region increased with the time of lamivudine treatment (chi2=6.69, P<0.01). The most common mutated type was M539V+L515M and next M539I. Continuous administration of lamivudine was no benefit for inhibiting the replication of HBV with YMDD mutation but helpful for wide-type HBV. CONCLUSION: The routine serum HBV DNA assay by PCR may introduce prejudice in monitoring HBV inhibitory effect by lamivudine, while the microarray technique can avoid this and is one of the best ways to monitor the lamivudine-resistance mutations in HBV. There is no effect of lamivudine on HBV with YMDD mutation in clinical practice.