Objective To investigate the relationship between the methylation of CpG island of RUNX3 gene promoter and its expression in a human cutaneous malignant melanoma cell line A375, and to assess the role of RUNX3 gene methylation in the pathogenesis of human cutaneous malignant melanoma. Methods Cultured A375 cells were treated with various concentrations (0, 1, 5, 10, 20 μmol/l) of 5-azacyti-dine for 24 or 72 hours followed by another 5 days of culture. Then, methylation-specific PCR (MSP) was performed to evaluate the methylation status of RUNX3 promoter region, and Western-blot analysis to detect the protein expression of RUNX3 in A375 cells. Results The RUNX3 gene promoter region was hypermethylated in untreated A375 cells, along with the absence of protein expression of RUNX3. However, after the treatment with 5-azacytidine, the promoter region of RUNX3 gene was demethylated partly, and the expression of RUNX3 protein was restored in A375 cells. Further, the expression intensity was directly correlated with the concentration of 5-azacytidine. Conclusions The promoter hypermethylation of RUNX3 gene may be related to the silencing of RUNX3 gene expression in A375 cells, whereas 5-azacytidine can cause the demethylation of RUNX3 gene, reactivate the gene which has been inactivated by the promoter hypermethylation, and finally induce the re-expression of RUNX3 protein.

Objective To investigate the relationship between the expression of the survivin gene and CpC methylation in exon 1 of the survivin gene in CA tissue, and to study the expression of survivin protein in CA tissue and its modulation mechanism. Methods Tissue samples were obtained from the CA lesions of 30 patients, normal cervix of 10 female controls, and normal foreskin of 10 male controls. Immunohistochemistry was carried out to detect the expression of survivin protein in these specimens, RT-PCR to measure the mRNA expression of survivin, and methylation specific PCR (MSP) to analyze the methylation status of CpG island in the survivin gene exon 1. Results The positivity rate of survivin protein and mRNA was 90% (27/30) and 93.3% (28/30) in CA tissue specimens, respectively, 5% (1/20) and 5% (1/20) in control tissue specimens, respectively; there was a significant difference between the two groups of specimens in both the parameters (x2 = 35.187, 38.437, both P < 0.01). The demethylation of CpG island in the survivin gene exon 1 was observed in 86.7% (26/30) of the CA tissue specimens and in 15% (3/20) of the control tissue specimens (x2 = 10.865, P < 0.01). There was a positive correlation between the demethylation status of CpG island in exon 1 and the mRNA expression of survivin gene (x2 = 13.929, P = 0.014). Conclusions The expression of survivin protein in CA tissues might be associated with the demethylation of CpG island in exon 1 of the survivin gene, and may play a certain role in the development of CA.

Objective To investigate the efficacy of combined high-dose intravenous immunoglobulin (IVIG) pulse therapy in patients with severe systemic lupus erythematosus (SLE). Methods Thirty-six patients were enrolled into this study, and randomly classified into WIG group (n=17) and methylprednisolone (MP) group (n=19). The treatment of patients in MG group began with a 3-day intravenous MP followed by intravenous WIG 400 mg per kilogram of body weight per day for 3-5 days, then was switched to oral prcdnisone and cyclophosphamide at routine dose. Intravenous MG was given repeatedly with an interval of 1 month for 2-5 sessions. Patients in MP group were treated with the same corticosteroids and immunosuppressants as used in WIG group but without IVIG. Patients were followed up for 3-12 months.The clinical efficacy, related serum parameters, and systemic lupus activity measurement (SLAM) were evaluated and compared between the two groups. Results Most patients in both groups showed a remission of symptoms and reduction in disease activity after treatment. The decrease in SLAM, positivity rates of antinuclear antibodies and anti-double-stranded DNA (anti-dsDNA) antibodies as well as the increase in platelets were faster in IVIG group than those in MP group (all P<0.05), but the long-term efficacy of the two groups was similar (P>0.05). Infections occurred in 11.8% of patients in WIG group and 36.8% of patients in MP group. Conclusions High-dose intravenous immunoglobulin may serve as an effective aid in the treatment of severe SLE, and is particularly beneficial to patients resistant to corticosteroids and immunosuppressants of routine dose and those accompanied by severe infections and intolerable to high dose of corticosteroids and immunosuppressants.

Objective To investigate the changes of blood oxygen saturation and heart rate after urgently going to high‐alti‐tude area ,so as to provide a reference for medical rescue in high‐altitude area .Methods Subjects left from the plain area with an al‐titude of 400 m .Blood oxygen saturation and heart rate were measured before departure and after reaching 4 300 m altitude region . Then the subjects were taken to the destination with an altitude of 3 200 m ,at which they received a dynamic continuous monitoring of blood oxygen saturation /heart rate at the 1st day ,2nd day ,3rd day ,4th day ,5th day ,6th day ,7th day after arrival .After adapting to the environment in 3 200 m altitude area for 1 week ,subjects were taken to the 4 300 m altitude region ,at which they were re‐measured blood oxygen saturation and heart rate .Results After entering the areas of 4 300 m altitude and 3 200 m altitude ,the blood oxygen saturation was significantly decreased compared with that in plain area (P< 0 .05) .The blood oxygen saturation at the 6th and 7th day after entering 3 200 m altitude area was statistically different when compared with that at the 1st day(P< 0 .05) . The blood oxygen saturation had statistical difference between reaching at 4 300 m altitude area for the first time and re‐entering 4 300 m altitude area ,while the heart rate had no statistical difference (P> 0 .05) .Conclusion The arterial oxygen saturation was de‐creased with the increase of altitude ;the people living in plain areas can preliminarily adapt to the environment at 6th day after reaching 3 200 m altitude regions ;people can better adapt to the high‐altitude environment by shortly living in lower‐altitude areas before re‐entering high‐altitude areas .

Objective　To investigate the relationship betwee n tricuspid insufficiency (TI) and the function of the right ventricle, degree o f pulmonary vascular pathological change in patients with rheumatic heart diseas e so as to provide the indication of operation for treating TI. Methods 　41 cases of rheumatic heart disease with TI accompanying with bicuspid pathological changes were reviewed. Correlation analysis was done between the s everity of TI and the right ventricular size(RVS), pulmonary artery pressure(PAP ), pulmonary artery resistance(PAR) etc. Results　It was found t hat the size of the right ventricle, PAP, PAR were positively correlated to th e degree of TI. Tricuspid annuloplasty should be carried out in patients with RV D>40 mm, PAR >48 kPa*s/L, PAP>8 kPa. Conclusion　TI resulted ma inly from insufficient function of the right ventricle and marked patholog ical changes of the pulmonary blood vessels. Doppler echocardiography evaluation for the function of right ventricle and pathological condition of pulmonary blo od vessels might be of significant in deciding whether tricuspid annuloplasty sh ould be performed simutaneously in patients of bicuspid valve replacement.

Objective:This work aims to investigate diallyl disulfide (DADS) inhibition of cell migration and invasion in human colon cancer SW480 cells through the Rac1-ADF/cofilin1 pathway. Methods:The potential of cell migration and invasion was examined by scratch healing assay and transwell membrane assay. The expression of Rac1-ADF/cofilin1 pathway was detected by RT-PCR and Western blot. Results:After the SW480 cells were treated with 40 and 50 mg·L-1 of DADS for 24 h, the number of transmembrane cells through the Matrigel obviously decreased by 57.12%and 64.59%, respectively (P<0.05). After cell treatment for 48 h, the cell migration rates were 23.23%and 12.87%, which were significantly lower compared with the control group (75.86%;P<0.05). After the cells were treated with 45 mg·L-1 of DADS for 24 and 48 h, the expression of Rac1, Rock1, PAK1, LIMK1, and destrin mRNA respectively decreased compared with the control group (P<0.05). However, no significant difference was observed in the expression of cofilin1 mRNA (P>0.05). After the treatment with 45 mg·L-1 of DADS for 6, 12, 24, and 48 h, the expression of Rac1, Rock1, PAK1, LIMK1, and Destrin proteins respectively decreased in a time-dependent manner compared with the control group (P<0.05). However, no significant differences were observed in the expression of the cofilin1 protein (P>0.05). Moreover, the expression of p-LIMK1 and p-cofilin1 notably decreased in a time-dependent manner (P<0.05). Conclusion:DADS inhibits cell migration and invasion, which is related to the down-regulation of Rac1, Rock1, PAK1, LIMK1, p-LIMK1, p-cofilin1, and destrin through the Rac1-ADF/cofilin1 pathway.

Objective To investigate the effect of trichostatin A (TSA) and 5-azacitidine (5-AzaC) on pancreatic β-cells impaired by cytokine,via measuring the proliferation,apoptosis,and function of pancreatic β-cells.Methods RIN-m5f was impaired by interleukin-1β and interferon-γin vitro,and treated with TSA and 5-AzaC.Experiment groups included blank control group,cytokine induction group,0.05/0.10 μmoL/L TSA group,0.63/1.25 μmoL/L 5-AzaC group,and0.10 μmol/L TSA plus 1.25 μmol/L 5-AzaC group.The viability of RIN-m5f cells was detected by MTT assay.Apoptotic rate was determined by Annexin V-fluorescein isothiocyanate (FITC) /propidium iodide flow cytometry.Insulin secretion was measured by enzyme-linked immunosorbent assay.Results The viability of RIN-m5f cells in 0.05/0.10 μmoL/L TSA group,0.63/1.25 μmol/L 5-AzaC group,and 5-AzaC plus TSA group was 70.1％/79.2 ％,67.3 ％/82.9 ％,and 89.1％ respectively,being higher than that in the cytokine group (33.9％,P＜0.05) ; the apoptosis rate was 10.3％/10.5％,7.9％/9.6％,and 8.2％,being lower than that in the cytokine group (16.6％,P＜0.05) ; the capacity of glucose-stimulated insulin secretion of all the treated groups was higher than that in the cytokine group (P＜0.05).Conclusion TSA and 5-AzaC might promote the proliferation of pancreatic β-cells impaired by cytokines,inhibit its apoptosis and recover its insulin secretion.

Aim To explore the effects of DADS in the cell cycle of HT-29 cells. Methods HT-29 cells growth inhibition were measured by MTT assay and cell counting. Phase distribution of cell cycle was analyzed by flow cytometry. Expression of p21~(Waf1),C-myc,Cyclin E was determined by immunohisto- chemistry analysis. Results MTT assay showed that DADS significantly inhibited HT-29 cells and exhibited a time, dose- dependent model. Adding 60 ?mol?L~(-1) and 120 ?mol?L~(-1) DADS for 24 h suppressed HT-29 growth by 23.1%,45.6% respectively. Cell counting showed that average doubling time retarded from 22.58 h in normal cultured HT-29 cells to 31.2 h in 60umol?L~(-1) DADS experimented HT-29 cells(P

AIM To study the inhibitory effect of bioactive compounds from entomogenous fungi(BCEF0083) on monoamine oxidase. METHODS Spectrofluorometer was used to detect the activity of MAO in mouse and rat brain mitochondria; Dose effect and time effect relationship of BCEF in inhibition of MAO were studied in vivo and in vitro in mice and rats; Method of Lineweaver Burk was used to assay the Km of MAO. RESULTS The antagoning action of BCEF0083 on MAO( BCEF 500 mg?kg -1 , 0 5, 1, 2, 4, 8, 16, 24, 48, 72 h after ig; 500, 400, 200, 100, 50, 25 mg?kg -1 , 2 h after ig)showed some extent of dose effect and time effect relationship. BCEF0083 in vitro inhibited the activity of MAO A,B in a dose dependent manner with IC 50 (95% of confidence limits)of 128 88(82 70～200 86),184 14(156 17～217 11) ?g?ml -1 in rats. The antagonism type of BCEF0083 on MAOA, B were both mixed type, their Km were 11 97, 8 13 ?mol?L -1 . CONCLUSION The results suggest that BCEF0083 could obviously inhibite the activity of MAO on brain remitochondria in mice and rats.

Objective To investigate the mRNA expression and methylation status of IL-13 receptor(IL-13R)α1 gene in peripheral T lymphocytes of patients with systemic lupus erythematosus(SLE).Methods Venous blood samples were obtained from 10 SLE patients(5 in active phase,5 in inactive phase)and 6 normal human controls.CD4+ and CD8+ T cells were isolated from these samples via magnetic activated cell sorting(MACS).Real-time quantitative PCR was used to test the mRNA expression of IL-13Rα1 gene,and methylation specific PCR to detect the methylation status.Results The expression level of IL-13Rα1 mRNA was 2.224±0.251,1.712±0.132.and 1.104±0.044 in CD4+ T cells of active SLE patients,inactive SLE patients and controls,respectively;the difference between the three groups was statistically significant(all P＜0.05).The expression level of IL-13Rα1 mRNA in CD8+T cells was significantly higher in active SLE patients than that in the normal controls(1.672±0.142 vs 1.238±0.106,P＜0.05),while no difference was noted between inactive and active SLE patients or normal controls.The methylation index of IL-13Rα1 gene was 0.454±0.023.0.635±0.065.0.844±0.097 in CD4+T cells of active SLE patients,inactive SLE patients and normal controls,respectively,and the difference between the three groups was significant(all P＜0.05),while no significant difference was observed in the methylation index in CD8+T cells among these groups(P＞0.05).The IL-13Rα1 mRNA expression in CD4+T and CD8+T cells was positively correlated with SLE disease activity index(SLEDAI)score(r=0.79,0.76,P=0.007,0.02 respectively).A negative correlation was found between the methylation level Of IL-13Rα1 in CD4+T cells and SLEDAI score(r=-0.89.P＜0.0 1).as well as between the IL-13Rα1 mRNA expression and its methylation level(r=-0.84,P＜0.0 1).Conclusion The development of SLE may be related to the overexpression of IL-13Rα1 gene induced by DNA hypomethylation in T cells.