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Objective:To investigate the effect of miR-425-5p on glucagon-like peptide-1(GLP-1) secretion in intestinal L cells induced by lipopolysaccharide(LPS), and to explore its mechanism.Methods:GLUTag cells of intestinal L cell line were incubated with LPS to determine the levels of miR-425-5p and GLP-1. Cell viability was determined by MTT assay, and cell apoptosis was detected by flow cytometry. Quantitative real time-PCR and western blot were performed to determine the expressions of miR-425-5p, phosphatase and tensin homology(PTEN), proglucagon, and GLP-1. Activity of Wnt/β-catenin signaling pathway was determined by detecting TOP/FOP ratio. Interaction among miR-425-5p, PTEN, and β-catenin was analyzed using luciferase activity assay and chromatin immunoprecipitation(ChIP)assay.Results:In GLUTag cells, with the elevation of LPS concentration, the expression of miR-425-5p and the apoptosis rate were increased, while the level of active GLP-1 and the cell viability were decreased. MiR-425-5p was involved in the regulation of LPS on GLP-1 secretion and intestinal L cell viability. Inhibition of miR-425-5p reduced the mRNA expression of proglucagon and the TOP/FOP ratio, increased PTEN protein level, and inhibited cell viability. In LPS-treated GLUTag cells, miR-425-5p increased the level of β-catenin by targeting PTEN, and β-catenin acted as a cis-acting element to induce the transcription of proglucagon and promote the secretion of GLP-1.Conclusion:In LPS-induced intestinal L cells, miR-425-5p promotes the expression of GLP-1 by targeting PTEN to modulate β-catenin.
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Objective@#To construct the mmu-miR-155 eukaryotic overexpression vector pmR-155 and to investigate its effect on HBV replication and expression of PTEN in vivo.@*Methods@#The mmu-mir-146a precursor gene fragment pre-mmu-mir-146a was amplified by PCR, then connected to the pmR-mCherry plasmid vector after double enzyme digestion, the accuracy of recombinant vector was verified by colony PCR、double enzyme digestion and sequencing; then the recombinant vector was transfected HBV transgene mice(Experimental Group)with hydrodynamics-based injection via vena caudalis, and pmR-mCherry plasmid、PBS were respectively transfected into the mice as Empty plasmid Group、Blank Group. The concentration of IFN-γ in the serum was detected by ELISA. The expression of SOCS1、PTEN mRNA in the liver was detected by qPCR at 30d post-transfectioned. The Western blot was performed to detect the changes in SOCS1、PTEN、HBX in the liver tissue at 30 d post-transfectioned. The results were analyzed with Student’s t-test, or one-way analysis of variance and the least significant difference test.@*Results@#the colony PCR、double enzyme digestion and sequencing verified that the gene was inserted into the pmR-mCherry vector. Compared with Blank Group, the expression of miR-155 in the Experimental Group was significantly increased(t = 8.90, P < 0.01); the concentration of IFN-γ in the Experimental Group was significantly increased(F = 26.58, P < 0.01); the mRNA(FSOCS1 mRNA = 19.72, P < 0.01; FPTEN mRNA = 7.38, P < 0.05) and protein(FSOCS1 = 50.30, P < 0.01; FPTEN = 129.00, P < 0.01) expression of COCS1、PTEN was significantly decreased in the Experimental group and the protein of HBX was also significantly(FHBX = 77.97, P < 0.01).@*Conclusion@#The pmR-155 eukaryotic overexpression vector is successfully constructed, this recombinant vector can express miR-155 stably; miR-155 can down-regulate cocs1、PTEN gene expression and up-regulate the expression of IFN-γ, it can inhibit the replication of HBV and a potential targets to treating hepatocellular carcinoma.
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Objective To investigate the expression of liver kinase B1 (LKB1) and phosphatase and tensin homology deleted on chromosome ten (PTEN) in hepatocellular carcinoma (HCC) and their relationship with pathological characteristics and prognosis of HCC.Methods hnmunohistochemistry was used to detect expression of LKB1 and PTEN in 115 HCC cases.The relationship between clinicopathologic factors and the expressions was analyzed.Results The positive expression rate of LKB1 and PTEN was 10.4％ (12/115) and 28.7％ (33/115) respectively.The coexpression ratio of LKB1 and PTEN was 5.2％ (6/115).LKB1 and PTEN double deletion rate was 66.1％,with the latter most often found in those ≥ 50 years of age group (x2 =7.968,P =0.001),middle low differentiation HCC group (x2 =11.297,P =0.025) and vascular tumor thrombus group (x2 =6.797,P =0.011).The 5 year survival rates of LKB1 and PTEN protein coexpression and double deletion patients were 100％ and 36.5％ (x2 =10.969,P =0.004),respectively.Multivariate COX regression analysis showed that vascular tumor thrombus,PTEN deletion and LKB1/PTEN double deletion were independent risk factors for the prognosis of HCC.Condusions Double deletion of LKB1/PTEN protein is one of the independent factors that affect the survival time of HCC patients.
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Objective To investigate the effects of over-expression of wild-type PTEN gene and its mutant (G129E) on apoptosis and proliferation of activated hepatic stellate cells (HSC) in vitro and its potential mechanisms. Methods The activated HSC cells were cultured in vitro and transfected with PTEN gene and G129E gene via adenoviral vector. The apoptosis of HSC was measured by MTT , and its proliferation was assessed by TUNEL and flow cytometry (FCM) . Western blotting and real-time fluorescent quantitation PCR were used to detect expression of PTEN in HSC. And the changes of Bcl-2 and Bax expression were tested by Western blotting. Results The wild type PTEN gene and G129E gene were successfully transducted into HSC, which resuted in elevated expression of Bax and reduced expression of Bcl-2 (P<0.01). After transduction, HSC proliferation was markedly inhibited with inhibitory rates of 14.03% and 23.12% at 48 and 72 hours in Ad-PTEN ,respectively, as well as 9.52% and 12.63% in Ad-G129E, respectively. Apoptotic rate of HSC exposed to Ad-PTEN or Ad-G129E for 72 hours increased significantly (P<0.01). Furthermore, wild type PTEN was more powerful than G129E for above-mentioned effects. Conclusions Over-expression of wild type PTEN and its mutant G129E can inhibit the proliferation of activated HSC, and induce HSC apoptosis through the Bcl-2/Bax pathway. In addition, the effect of wild type PTEN is more powerful than that of G129E.
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@#ObjectiveTo investigate the expression and relationship between phosphatase and tensin homology deleted on chromosome ten(PTEN) and vascular endothelial growth factor(VEGF) in bladder transitional cell carcinoma(BTCC) and their clinical significance.MethodsExpression of PTEN and VEGF were detected in 60 specimens by immunohistochemistry test(SP method).ResultsThe positive rate of PTEN was 47%(28/60) in BTCC;VEGF was 68%(41/60).With the pathological grade and the clinical stage of tumors being higher,the lower expression level of PTEN showed(P<0.05);while the expression of VEGF increased with no statistical significance(P>0.05),but having a significant difference between specimens with lymph nodes metastasis and without lymph nodes metastasis(P<0.05).The expression of PTEN was negatively correlated with that of VEGF(r=-0.439,P<0.01).ConclusionThe expression of PTEN and VEGF in BTCC plays an important role during the progress of carcinoma and is helpful to evaluate the prognosis of patients.
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Phosphatase and tensin homology deleted on chromosome 10 (PTEN) plays an important role in the proliferation,migration,differentiation,apoptosis and synapse establishment of nervous system.Elucidation of PTEN function is helpful to understand the mechanisms of neural development,and thus may find new therapies for diseases in central nervous system using PTEN as a target.
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Objective: To research the expression of PTEN and its influence on biological ability in breast cancer cell in vivo. Methods: PTEN-shRNA plasmid was transtected into M231 breast cancer cells to knock down the expression of PTEN. The changes of PTEN expression, proliferation, adhesion and metastasis of PTEN knocked down cell were tested by western-blot, colony formation, adhesion and invasion assay. Results: PTEN-shRNA was successfully transfected into M231 cells and it inhibited PTEN expression efficiently.The capabilities of colony formation, migration and invasion of transfected cell were much greater than those of the controlling cell line. But the transfected cells were more difficulty in adhesion than the scrambled ones. Conclusion: PTEN genecan enhance the adhesion, but restrict the proliferation, migration of breast cancer cells in some degree, so that inhibit the development of the breast cancer. PTEN loss can be a prognostic factors for the patients with breast cancer.
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AIM:To explore the potential correlation between the expression of phosphatase and tensin homology deleted on chromosome 10(PTEN) and RIF in IgA nephropathy.METHODS:Forty-seven patients diagnosed as primary IgA nephropathy by renal histology were involved.10 specimens from normal renal tissue of renal carcinoma served as control group.Tubulointerstitial lesion(TIL) was classified by using Katafuchi scale,including no TIL(group I),mild TIL(group II),moderate TIL(group III),severe TIL(group IV).The expressions of PTEN,TGF-?1,?-SMA and ColⅢ in renal tissue were detected by immunohistochemistry.PTEN mRNA was detected by in situ hybridization.RESULTS:Renal tissues from renal biopsy showed abundant expressions of PTEN and PTEN mRNA in endochylema of renal tubular epithelial cells,and negligible expression in glomeruli.With the progress of TIF in IgA nephropathy,the expressions of PTEN and PTEN mRNA decreased gradually(P