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Objective:To investigate the effect and mechanism of miR-195 regulating FOXK1 gene and PI3K/Akt pathway on stomach adenocarcinoma proliferation, invasion and migration ability.Methods:Public database samples were employed to analyze the expression differences and prognostic significance of miR-195 in stomach adenocarcinoma. After overexpression of mir-195-5p in two cell lines, MGC803 and AGS, altered cell proliferation, invasion, and migration abilities were detected by Alamar Blue, Wound healing, and Transwell assays. The potential target genes and binding sites of miR-195 were predicted by the starBase. Western blot was used to detect the expression levels of foxk1 and phosphorylation sites in the PI3K/Akt pathway of target genes after overexpression of mir-195-5p. A Dual-luciferase reporter assay was used to verify the relationship between mir-195-5p and foxk1. Statistical analyses were performed with IBM SPSS 22 software and R 4.0.3.Results:Our results showed a significant over-expression of miR-195 in the tumor tissues, compared with the paired normal tissues ( P<0.001) , which could inhibit the proliferation and invasion of stomach carcinoma cells and significantly correlated with survival ( P=0.011) . Moreover, our study indicated that miR-195 depressed the expression of FOXK1 and significantly reduced the activation of the PI3K/Akt pathway, which had a negative effect on the proliferation and invasion of stomach carcinoma cells. The phosphorylated Akt (s473 site) expression in the PI3K/Akt pathway was significantly decreased after overexpression of miR-195. Conclusion:Overall, our studies clarify the important function of the miR-195 in the diagnosis and therapy of patients with stomach carcinoma and reveal the FOXK1 and PI3K/Akt pathway regulation by the miR-195, which are of important clinical significance in the differential diagnosis.
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Objective@#The aim of the study was to identify the personality types of middle school students and its association with life satisfaction,and to provide a reference for improving personality and mental health of middle school students.@*Methods@#A total of 1 409 students randomly selected form 4 middle schools of a rural area in Wuhan city by cluster stratified sampling method were taken as the object of the research. Adolescents completed the Chinese version of NEO Five-Factor Inventory (NEO-FFI) and the Satisfaction with Life Scale. Latent profile analysis (LPA) was used to identify personality types. ANOVA analyses were used to explore the associations between personality types and life satisfaction.@*Results@#Three profiles were identified: resilient (5.32%), overcontrolled (32.86%) and undercontrolled (61.82%),the differences were of statistical significance(P<0.05). Resilient adolescents had the highest scores (M=5.10, SD=1.34) whereas undercontrolled had the lowest scores (M=3.66, SD=1.09) on life satisfaction.@*Conclusion@#Schools, families and society should pay attention to the high proportion of overcontrolled and undercontrolled in middle school students and take actions to promote the positive development of personality and mental health.
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<p><b>OBJECTIVE</b>To evaluate the feasibility of monitoring clopidogrel resistance with flow cytometric analysis of platelet vasodilator stimulated phosphoprotein (VASP) phosphorylation.</p><p><b>METHODS</b>Twenty patients with coronary heart disease (CHD) and 17 healthy volunteers were examined for platelet aggregation rate and phosphorylation of VASP (calculated as platelet reactivity index, PRI) using traditional optical nephelometry and flow cytometry before and after concurrent therapy of clopidogrel and aspirin.</p><p><b>RESULTS</b>No significant differences were found in PRI between CHD group and healthy control group [(89.45∓5.22)% vs (86.58∓4.35)%] before treatment. The PRI in CHD group was significantly lowered after treatment to (67.66∓19.77)% (P<0.05). Clopidogrel resistance was found in 6 (30%) cases in CHD group by flow cytometric analysis, which showed a higher sensitivity than optical nephelometry (10%).</p><p><b>CONCLUSION</b>Flow cytometric analysis of VASP phosphorylation is a more reliable test to specifically evaluate clopidogrel resistance.</p>
Subject(s)
Aged , Aged, 80 and over , Female , Humans , Male , Middle Aged , Case-Control Studies , Cell Adhesion Molecules , Chemistry , Drug Resistance , Flow Cytometry , Microfilament Proteins , Chemistry , Phosphoproteins , Chemistry , Phosphorylation , Platelet Function Tests , Ticlopidine , PharmacologyABSTRACT
Objective:To study the effects of the overexpression of hsa-miR-202 on the proliferation and molecular mechanism of lung cancer A549 cells. Methods:A sequence of hsa-miR-202 with ppG/miR/eGFP/Blasitidin pasmid was directionally connected and a eukaryotic expression vector pmiR-202 of the target hsa-miR-202 gene was constructed. pmiR-202 was transtected to A549 cell with Lipofectamine 2000. The WST assay was used to detect the cell proliferation rate, and RT-PCR was used to detect the relative gene expression levels. Western blot analysis was used to detect the IL-10 protein expression levels. The interaction between miR-202 and IL-10 was examined using a luciferase reporter assay. Results:The design from the DNA sequencing results shows that a eukaryot-ic expression vector of miR-202 was successfully constructed. The proliferation inhibition rates of A549 cells by Pmir-202 were 12%, 38%, and 52%. The differences in the treatment group compared with the blank control and negative control groups were statistically significant. The RT-PCR results showed that the relative expression levels of miR-202 increased after transfection with pmiR-202. Over-expression of miR-202 can downregulate the relative gene and protein expression levels of IL-10, and the relative levels were 25%and 0.75, respectively. Compared with the blank control and the negative control groups, the difference was statistically significant (P<0.05). The fluorescent activity was reduced when transfection was performed with miR-202 mimics, and IL-10-3'UTR plasmid was cloned. Conclusion: pmiR-202 effectively inhibited the proliferation of A549 cells and exhibited a time-effect relationship with miR-202 by targeted combination with IL-10 3'UTR to downregulate IL-10 expression in A549 cells.
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ObjectiveTo investigate the role of CD4+ CD25+ regulatory T cells(Treg) in peripheral blood and intestinal endotoxin (ET) in liver injury of rat severe acute pancreatitis (SAP).MethodsSixty male Wistar rats were randomly allocated into sham operation group(SO group) and severe acute pancreatitis group(SAP group).The rats in SAP group recevied the injection of Sodium Taurocholate(NaTc) into their far-end of bile-pancreatic duct and were sacrificed in 3-,6-and 12-hour,serum amylase(AMY) and alanine aminotransferase(ALT) determined by full-automatic biochemistry instrument,limulus reagent method is used to determine ET content in plasma,the proportion of Treg among peripheral lymphocytes was determined by Flow Cytometry(FCM),HE stain is used to observe pathological changes in liver and pancreas,the protein of Foxp3 activity was evaluated by immunohistochemistry staining,and the relationships between these indicators were analyzed using Pearson correlation analysis test.ResultsHistopathologic examination and the level of ALT revealed the occurrence of pancreatic inflammation and pathological changes of liver in SAP group.The percentage of Treg in SAP groups significantly increased at 3 h(2.26％ ±0.32％),6 h(2.36 ％ ±0.48％)and 12 h(2.80％ ±0.35％) comparing to the SO groups(P＜0.01) ; plasma ET levels compared with the SO group was statistically significant (P＜ 0,01 ),and 12 h (0.85 ± 0.11) compared to,3 h (0.74±0.11) and 6 h (0.78-±-0.07) was no significant difference (P＞0.05).The expression of Foxp3 protein on the livers were upregulated markedly.Pearson correlation analysis teat showed that quantities of Treg were positively proportional to the levels of ET(r=0.89,P＜0.01) after liver injury of SAP.ConclusionsSAP may lead to liver injury and the high plasma levels of ET and Treg in peripheral blood may play an important role in liver injury of SAP.
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Objective To identify the gene mutations of an inherited coagulation factor Ⅶ deficiency pedigree.Methods PCR and DNA sequencing were used to identify the FⅦ gene mutations in the proband.The identified mutations were validated by PCR followed by restriction fragment length polymorphism technique or DNA sequencing.100 healthy volunteers were chosen randomly as controls. Results R1S2Q and IVS6+1G→T double heterozygous mutations were discovered in the Droband.The pedigree analysis showed that R152Q missense mutation inherited from his father,and IVS6+1G→Twas from his mother. The R1S2Q missense mutation in exon 6 was not found in 100 healthy volunteers. Conclusion The congenital deficiency of F Ⅶ in the proband might be caused by the coinheritance of the R152Q missense mutation in exon 6 and the splicing donor site mutation ( ⅣS6+1G→T)in intron 6.
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Objective To investigate the effect of hypoxia on the expression of vascular endothelial growth factor (VEGF) in villous explants, and the effect of hypoxia-inducible factor 2? (HIF-2?) on the VEGF expression. Methods The villous were obtained from women who underwent suction and curettage for termination at 6-8 gestational weeks. Four groups were divided: normoxia group; hypoxia group; sense HIF-2? oligonucleotide group and anti-sense HIF-2? oligonucleotide group. All villous were cultured in vitro. In anti-sense and sense group, villous were incubated under hypoxia with HIF-2? oligonucleotide. The villous explants were collected after 48 h of cultivation. The protein level of HIF-2? was detected by Western blot and the mRNA of HIF-2? and VEGF were examined by real-time PCR. Results The expression of HIF-2? mRNA and VEGF mRNA as well as the protein level of HIF-2? were increased in hypoxic group comparing with the normoxia group. Compared with the sense group, the expression of HIF-2? protein and HIF-2?, VEGF mRNA showed significant decrease in anti-sense group. Conclusions Hypoxia may result in the increased expression of VEGF in villous explants during the early trimester, which might be mediated by HIF-2?.
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2-Keto-L-gulonic acid (2-KLG), the precurcor of L-ascorbic acid synthesis, was prepared directly from D-glucose by tandem fermentation. In the first step fermentation Erwinia sp. SCB 247 translated D-glucose to 2,5-diketo-D-gluconate (2,5-DKG), which accumenlated 180 mg 2,5-DKG per ml in the broth. In the second step fermentation Corynebacterium sp. SCB 3058 reduced 2,5-DKG to 2-KLG, which accumulated 35 mg 2-KLG per ml in the broth. This reductive fermentation was obtained under aerobic conditions by adding a hydrogen donor such as glucose.The average yield of five batches fermentation was 56.3 mol%, from D-glucose to 2-KLG in 10 L fermentor.