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Chinese Medical Journal ; (24): 154-164, 2020.
Article in English | WPRIM | ID: wpr-781627


BACKGROUND@#Gastric cancer (GC) is one of the most common malignancies, and intestinal-type GC is the main histopathologic type of GC in China. We previously reported that casein kinase 2 interacting protein 1 (CKIP-1) acts as a candidate tumor suppressor in intestinal-type GC. CKIP-1 participates in the regulation of multiple signaling pathways, including the Wnt/β-catenin pathway, of which caudal-related homeobox 1 (CDX1) may be a downstream target gene. The purpose of this study was to investigate the relationship between CKIP-1 and CDX1 in intestinal-type GC.@*METHODS@#Sixty-seven gastroscopy biopsy specimens and surgically resected gastric specimens were divided into four groups: gastric mucosa group, intestinal metaplasia (IM) group, dysplasia group, and intestinal-type GC group. The expression levels of CKIP-1 and CDX1 were detected in these groups and GC cell lines, and the correlations between these expression levels were analyzed. SGC7901 and BGC823 cells were divided into CKIP-1 shRNA groups and CKIP-1 over-expression groups, and CDX1 expression was detected. β-Catenin expression was detected in intestinal-type GC tissue samples and CKIP-1 shRNA and CKIP-1 over-expression SGC7901 cells, and its correlation with CKIP-1 expression in intestinal-type GC tissue was analyzed. The Wnt/β-catenin pathway inhibitor DKK-1 and activator LiCl were incubated with SGC7901 cells, BGC823 cells, and CKIP-1 shRNA and CKIP-1 over-expression SGC7901 and BGC823 cells, following which CDX1 and Ki-67 expression were detected.@*RESULTS@#The expression levels of CKIP-1 and CDX1 were lower in patients with intestinal-type GC than in patients with IM and dysplasia (both P < 0.05). CKIP-1 and CDX1 expression levels were positively correlated in IM, dysplasia, and intestinal-type GC tissue and cell lines (r = 0.771, P < 0.01; r = 0.597, P < 0.01; r = 0.654, P < 0.01; r = 0.811, P < 0.01, respectively). CDX1 expression was decreased in the CKIP-1 shRNA groups and increased in the CKIP-1 over-expression groups of SGC7901 and BGC823 cells compared to that in the corresponding control groups (both P < 0.05). CKIP-1 expression was negatively correlated with β-catenin expression in intestinal-type GC patients (r = -0.458, P < 0.01). Compared to the control group, β-catenin expression was increased in the CKIP-1 shRNA SGC7901 cell group and decreased in the CKIP-1 over-expression SGC7901 cell group (P < 0.05). CDX1 expression was increased in SGC7901 and BGC823 cells treated with DKK-1, DKK-1 increased CDX1 expression and decreased Ki-67 expression in the CKIP-1 shRNA group; the opposite result was observed in SGC7901 and BGC823 cells treated with LiCl, and LiCl decreased CDX1 expression and increased Ki-67 expression in the CKIP-1 over-expression group (both P < 0.05).@*CONCLUSIONS@#Through the Wnt/β-catenin signaling pathway, CKIP-1 may positively regulate CDX1 in intestinal-type GC.

Article in Chinese | WPRIM | ID: wpr-701164


AIM:To investigate the effect of miR-483-5p on P3 promoter-driven mRNA(P3 mRNA)expres-sion of human insulin-like growth factor 2(IGF2)gene and its role in the development of hepatocellular carcinoma (HCC).METHODS: The expression levels of miR-483-5p and P3 mRNA were analyzed by real-time PCR in human HCC cell lines Huh7,Hep3B,Bel-7402,HepG2 and SMMC-7721,normal human liver cell line HL-7702,83 cases of hu-man HCC tissues and their matched adjacent nontumorous tissues(MANT), and 22 cases of normal adult liver tissues (NALT).The association between P3 mRNA level and miR-483-5p level was evaluated by Pearson correlation analysis. The full-length sequences of 5'UTR of P3 mRNA containing wild-type and mutant miR-483-5p-binding sequences were cloned into pGL3 promoter vector, which were termed pGL3-P3-5'UTR-WT and pGL3-P3-5'UTR-MUT, respectively. These luciferase reporter constructs were transfected into HeLa,293T and Huh7 cells together with miR-483-5p mimic, miR-483-5p inhibitor or scrambled control,and the luciferase activity was measured using dual-luciferase reporter system. The miR-483-5p mimic,miR-483 inhibitor and scrambled control were also transfected into Huh 7 cells and Hep3B cells, and P3 mRNA level was detected by real-time PCR.The expression levels of miR-483-5p in the nuclear and cytoplasmic fractions of Hep3B cells and Huh7 cells were detected by real-time PCR.The effect of miR-483-5p on P3 mRNA transcrip-tion was evaluated by nuclear run-on assay.The effect of miR-483-5p on the stability of P3 mRNA was analyzed by RNA stability assay.Furthermore,the effects of miR-483-5p on the viability, apoptosis, migration and invasion of Huh7 cells were investigated.RESULTS:Significamtly high levels of miR-483-5p and P3 mRNA were detected in the 5 human HCC cell lines and the human HCC tissues as compared with the human normal liver cell line HL-7702, and the MANT and NALT,respectively.Linear correlation analysis revealed that P 3 mRNA level was positively correlated to miR-483-5p level in the 5 human HCC cell lines and the human HCC tissues.miR-483-5p directly recognized the P3 mRNA 5'UTR to pro-mote gene expression.Overexpression of miR-483-5p resulted in a significant increase in P3 mRNA expression in a dose-dependent manner in the Huh7 cells and Hep3B cells.The mature miR-483-5p was present in both cytoplasm and nucleus of Hep3B cells and Huh7 cells.miR-483-5p induced nascent P3 mRNA transcription in the nucleus of Huh7 cells.miR-483-5p did not alter P3 mRNA stability in Huh7 cells.Furthermore,miR-483-5p led to increased viability,apoptosis inhi-bition,and enhanced migration and invasion abilities in the Huh 7 cells.CONCLUSION:High expression of miR-483-5p promotes the growth,migration and invasion of HCC cells in part through up-regulating P3 mRNA transcription,and is con-sequently involved in the development of HCC.

Article in Chinese | WPRIM | ID: wpr-308669


<p><b>OBJECTIVE</b>To observe the effect of pretreatment with puerarin on activation of LPS -induced RAW264. 7 cells and secretory cytokines, and discuss its anti-inflammatory mechanism.</p><p><b>METHOD</b>Well-grown RAW264. 7 cells in the exponential phase were collected and randomly divided them into the blank control group, the LPS group and the puerarin pretreatment + LPS group. The cellular toxic effect of puerarin on RAW264. 7 cells was examined by CCK-8 assay, cell morphology was detected by Giemsa stain method, the changes in TNF-alpha and MIP-2 were tested by ELISA, and the expression of NF-kappaB p65 mRNA were determined by qRT-PCR.</p><p><b>RESULTS</b>When puerarin was cultured with 1 mg x L(-1) LPS at a concentration of lower than 400 micromol x L(-1), it had not showed the cellular toxic effect (P < 0.05). Compared with the control group, the LPS group could significantly change the morphology of RAW264. 7 cells (increase in cell body, irregular shape, with a large number of pseudopodia extending). After intervention, the puerarin 100 micromol x L(-1) group could significantly inhibit LPS-induced cell morphological changes, while the puerarin 200 micromol x L(-1) and 400 micromol x L(-1) puerarin groups showed more notable inhibitory effects. However, there was no obvious difference between the two groups. The pretreatment with puerarin could inhibit the expression of TNF-alpha and MIP-2 in cell supernatant and NF-kappaB p65 mRNA in cells (P < 0.05). With increase in the puerarin concentration, its inhibitory effect gradually grew (P < 0.05), but did not reach the level of the blank control group.</p><p><b>CONCLUSION</b>As a safe and effective natural anti-inflammatory drug, puerarin can significantly reduce the expression of inflammatory cytokines (TNF-alpha, MIP-2). Its mechanism may be related to the reduction of NF-kappaB p65 mRNA expression.</p>

Animals , Cell Line , Isoflavones , Pharmacology , Lipopolysaccharides , Allergy and Immunology , Macrophage Activation , Macrophages , Allergy and Immunology , Mice , NF-kappa B , Genetics , Allergy and Immunology , Plant Extracts , Pharmacology , Sincalide , Genetics , Allergy and Immunology , Transcription Factor RelA , Genetics , Allergy and Immunology , Tumor Necrosis Factor-alpha , Genetics , Allergy and Immunology
Article in Chinese | WPRIM | ID: wpr-814066


OBJECTIVE@#To investigate the effect of pinggan-qianyang (PGQY), a Chinese medicine, on hypothalamic proteome in the hyperthyroid rats with hyperactivity of liver-yang, and to explore its mechanism.@*METHODS@#The rat model was established by intraperitoneal injection of levo-thyroxine (L-T4) and fuzi decotion. All the quantitative and qualitative changes of the protein expressions were compared among the normal group,the model group and the treatment group by proteomic techniques.@*RESULTS@#The protein spots in the 3 groups were mainly displayed at the isoelectric point (pI) 3 approximately 10, and the molecular weights were 13.8 approximately 98.8 kD.Compared with the normal group, 6 spots of protein expression increased and 10 decreased in the model group. All the changed protein in the model group returned to normal level after PGQY treatment. Mass-spectrometer and bio-informatics indicated that these proteins were Prohibitin, Peroxiredoxin-6, histidine triad nucleotide-binding protein 1, protein-tyrosine-phosphatase, predicted protein, profilin-2, peroxir doxin-II, heat shock protein-27, and annexin-A1.@*CONCLUSION@#There are differences in the expression of hypothalamus proteins in the hyperthyroid rats with hyperactivity of liver-yang after the treatment with PGQY, and the 9 identified protein spots may be associated with the mechanism of PGQY.

Animals , Drugs, Chinese Herbal , Therapeutic Uses , Female , Hyperthyroidism , Drug Therapy , Metabolism , Hypothalamus , Metabolism , Male , Medicine, Chinese Traditional , Nerve Tissue Proteins , Metabolism , Peroxiredoxin VI , Metabolism , Phytotherapy , Proteome , Metabolism , Random Allocation , Rats , Rats, Sprague-Dawley , Repressor Proteins , Metabolism
Article in Chinese | WPRIM | ID: wpr-680332


0,is advantageous to the patient. NNT=3.984,the expression in order to cause 1 patient to be bad avoid the curative effect,must apply outside AO the bracket fixing to treat 4 patients.Conclusion Outside conclusion AO bracket fixing treatment specific weight open shin fibula bone fracture regardless of from statistics significance comparison,the clinical significance analysis, the even curative effect is high,is advantageous to the patient.