Background and purpose:The Smac/DIABLO(the second mitochondrial derived activator of caspase/direct IAP binding protein with low pI)is a new kind of mitochondria protein,it inhibits IAPs(inhibitors of apoptosis protein),including Survivin,and activates caspase-9 and caspase-3,accordingly promotes apoptosis.In this study,we investigated the effect of over-expressing mitochondrial protein-Smac/DIABLO while silencing inhibitor of apoptosis protein-Survivin on the cell growth,cell cycle and apoptosis of human colon cancer cells through cotransfecting both genes.Methods:Constructed survivin-shRNA-EGFP plasmid was transected with Smac-pcDNA3.1 plasmid into Lovo cells at either half each dose or total dose,respectively.Western blot was used to survey the protein level of Smac and Survivin;Hoechst 33258 staining was used to detect the karyomorphological diversity of apoptotic cells and evaluate apoptotic ratio roughly;PI staining and flow cytometry analysis were used to examine cell cycle;caspase-3 Detection Kits was used to detect the activity of caspase-3.Results:The expression of Smac protein increased but Survivin protein decreased 48 hr after transfection.Karyomorphological diversify of apoptotic cells were obviously observed by hoechst 33258 staining.The apoptosis rate in Smav+Survivin shRNA group was(18.5?1.7)%,which was higher than that in Smac group(9.6?1.8)% and Survivin shRNA group(15.0?0.3)%,and all of three groups were significantly higher than the control groups;The cells of G0/G1 phase increased to(51.0?6.2)% in Smac group,and the cells of S stage increased to(53.3?1.3)% in Survivin shRNA group(P

The development and present situation of high power and luminous flux LED is introduced. The feasibility of high luminous flux LED in the field surgery illumination is analyzed in detail. Then the realizable plans are discussed separately from device, driving circuit, optical system and mechanical system.

Objective To investigate the inducing effects of selenium dioxide(SeO2 ) on the apoptosis in human cervical carcino-ma cell line Hela and its influence on the expression of apoptosis-related proteins caspase-3 and P53 .Methods Hela cells were trea-ted with different concentrations of SeO2 for 24 h in vitro ;the morphological changes of Hela cells were observed by the optical mi-croscope;the influence of SeO2 on the cell proliferation and vitality was examined by the MTT assay ;the flow cytometry was em-ployed to detect the cell apoptosis rate ;the expressions of caspase-3 and P53 proteins in Hela cells were determined by the Western blot analysis .Results Under the optical microscopy ,SeO2 generated the obvious influence on the cell growth morphology ,a large number of cells became rounded and shrunken ,and lost the normal form ,while the adherence cell number was evidently decreased and the proliferation was slowed down ;the MTT results showed that SeO2 markedly inhibited the cell proliferation and viability in a dose-dependent manner ,in which ,the cell apoptosis rates induced by the 0 ,1 .875 ,3 .750 ,7 .500 ,15 .000 and 30 .000 μmol/L con-centrations of SeO2 were 3 .12% ,30 .56% ,33 .42% ,37 .50% ,45 .43% and 69 .38% respectively ,which revealing the obviously in-creasing trend;the Western blot assay revealed that SeO2 could up-regulate the caspase-3 and P53 levels ,and reached the peak value at the concentration of 7 .500μmol/L .Conclusion SeO2 could induce the cervical cancer cell apoptosis possibly by up-regulating the expressions of caspase-3 and p53 in Hela cells .

Objective To investigate the effects of lanthanum chloride on proliferation and migration activity of human cervical cancer cells in vitro which may be a new anti-cervical cancer drug and provide experimental data for cervical cancer treatment. Methods HeLa cells cultured in vitro were divided into two groups: experimental group and control group. In experimental group, the cells were respectively treated with lanthanum chloride at different concentrations, 5, 50 and 100 μmol/L, while the cells in the control group were not treated with lanthanum chloride. The cell growth was observed by inverted microscope and the morphology changes of the cells were observed by the laser scanning confocal microscope (LSCM).Proliferation of HeLa cells in the two groups was detected by methyl thiazolyl tetrazolium (MTT) test;apoptosis rate was analyzed by flow cytometry (FCM). Cell migration test was applied to observe the effect of lanthanum chloride on migration. Reverse transcription (RT)-PCR was employed to evaluate the effects of lanthanum chloride on proliferation gene (cyclinD1), anti-apoptosis gene (zinc finger protein A20) and migration-related gene (matrix metalloproteinase 9, MMP-9). Results The status of cell growth was observed under the inverted microscope: with the increased of the lanthanum chloride concentrations, the cell density of reduced, the granule in cytoplasm increased, color intensifying and intercellular space enlarged; some cells became rounding and dead, floating in the culture media; the exfoliated cells increased gradually in the experimental groups. While In the control group, the cells grew adherently, with clear morphology and plump cytoplasm, and adjacent cell grew in lamellar. Observed with LSCM: the nuclear chromatin condensated and marginated with the volume of nuclear decreased in experimental groups. With the increase of the lanthanum chloride concentrations, nuclei in the experimental groups became pyknotic and then underwent karyorrhexis. However, the nuclear of the cells in control group were inact. The growth inhibition rates of lanthanum chloride groups (5, 50, 100 μmol/L) were 24%, 51% and 78%,respectively, in which each was significantly higher than that of the control group (P < 0. 05); the apoptosis rates of lanthanum chloride group were (4. 91 + 0. 39) %, (7. 30 + 0. 71) % and (13.48 + 0. 92) %,respectively, which were all significantly higher than that of the control group [(0. 89 + 0. 11) %, P <0.01]. The migration ability of the cells was also decreased by the treatment of lanthanum chloride, the number of migrated cells in lanthanum chloride groups were 22.2±4. 3, 12. 0±3.2 and 7. 8±2. 6 respectively, which were all significantly lower than that of the control group (41.2±5.4, P < 0. 01). The expression of genes of cyclinD1, A20 and MMP-9, were all decreased by the treatment of lanthanum chloride in a dose-dependent manner. Conclusion Lanthanum chloride can inhibit the proliferation and migration of cervical cancer cells, and induce apoptosis by down-regulating cyclinD1, A20, and MMP-9 expressions in vitro.

Immuno-fluorescence technique can qualitatively determine certain nuclear translocation, of which NF-κB/ p65 implicates the activation of NF-κB signal pathways. Immuno-fluorescence analysis software with independent property rights is able to quantitatively analyze dynamic location of NF-κB/p65 by computing relative fluorescence units in nuclei and cytoplasm. We verified the quantitative analysis by Western Blot. When we applied the software to analysis of nuclear translocation in lipopolysaccharide (LPS) induced (0. 5 h, 1 h, 2 h, 4 h) primary human umbilical vein endothelial cells (HUVECs) , we found that nuclear translocation peak showed up at 2h as with calculated Western blot verification results, indicating that the inventive immuno-fluorescence analysis software can be applied to the quantitative analysis of immuno-fluorescence.
Active Transport, Cell Nucleus ; Cell Nucleus ; metabolism ; Cytoplasm ; metabolism ; Fluorescent Antibody Technique ; Human Umbilical Vein Endothelial Cells ; Humans ; NF-kappa B p50 Subunit ; metabolism ; Software

Objective To explore the influences of platelet and platelet-derived 5-hydroxytryptamine (5-HT) on liver function, post-operative recurrence and long-term survival of patients with early stage hepatocellular carcinoma (HCC) following curative resection. Methods Prospective cohort study was employed in this research. A total of 297 consecutive patients who met the Milan criteria and received HCC curative resection from January 2009 to December 2009 in our hospital were selected, and their clinical data were collected. Patients' serum samples were stored at -80°C. Serum 5-HT concentration was detected by ELISA kits. Patients were regularly followed-up to observe their health condition after operation. The influences of PLT and 5-HT on liver dysfunction (LD), overall survival (OS) and recurrence-free survival (RFS) of patients with early stage HCC resection were analyzed via Cox' proportional hazard regression model. Results It was found that the pre- and post-operation PLT counts of patients with post-operation LD were significantly lower than those with normal post-operation liver function (P<0.001). Serum 5-HT concentration was positively correlated with PLT count(r=0.712, P<0.001). Low pre-operative PLT (OR=2.952, 95%CI:1.206-7.229, P=0.018)and low preoperative serum 5-HT concentration (OR=4.989, 95%CI: 2.004-12.422, P=0.001) were the independent risk factors of LD after early stage HCC resection. Low pre-operative PLT (OR=1.782, 95%CI:1.086-2.924, P=0.022) and low pre-operative serum 5-HT concentration (OR=1.754, 95%CI:1.014-3.034, P=0.045) were also the independent risk factors of OS; however, the pre-operative PLT and 5-HT were not the independent risk factors of RFS. Conclusion Early stage HCC patients with low pre-operative 5-HT and PLT tend to have poorer perioperative and long-term outcomes after curative hepatic-resection.

Objective: To investigate the effects of perioperative application of intravenous hemostatics on the long-term survival and recurrence of patients with early stage hepatocellular carcinoma (HCC) following curative resection. Methods: A total of 504 patients undergoing hepatectomy during 2005-2007 in our hospital were included in this study. The HCC tumors had a diameter less than 5 cm, at T1-2N0M0, and with pathological negative margins. The liver function of patients was Child-Pugh score A, B grade. Cox model with stepwise regression analysis was used to analyze the factors related to survival of patients and recurrence after surgery, and Kaplan-Meier analysis was used to clarify whether intravenous hemostatic agent is related to the overall survival time (OS) and recurrence-free survival time (RFS). Results: The median follow-up time for the patients was 64 months (7-72 months). Perioperative intravenous hemostatic agents were used in 326 of the 504 patients and the rest did not receive any hemostatic agents during the perioperative period. The 5-year OS rate of patients receiving hemostatic agents was significantly lower than those receiving no hemostatic agents (61.04% vs 75.28%, P=0.002), and the same was also true for the 5-year RFS rate in the two groups (49.08% vs 61.80%, P=0.001). Cox model with stepwise regression analysis showed that perioperative use of intravenous hemostatic agents was an independent risk factor of patients' OS (P=0.001, relative risk 1.872, 95%CI 1.298-2.702) and RFS (P=0.005, relative risk 1.523, 95% CI 1.136-2.043). Conclusion: Application of intravenous hemostatic agents during perioperative period might be associated with poor overall and recurrence-free survival of patients with early stage HCC after curative resection.

Objective To study the telomerase expression in peripheral blood mononuclear cells(PBMCs) in children with acute Kawasaki disease(KD) and its clinical significance.Methods The PBMCs of 64 children with acute KD [25 cases of them with coronary artery lesions(CAL),while the rest without] from 2 months to 6 years old admitted into Jiangxi Children's Hospital from Mar.2005 to Dec.2008 and those of 52 sex-age-matched healthy children (healthy control group) from 5 months to 7 years old were all assayed by Roche telomerase polymerase chain reaction enzymelinked immunosorbent assay(PCR ELISA).WBC,ESR and CRP were also detected.SPSS 11.0 software was used to analyze the data.Results The telomerase expression frequency of PBMCs in children with KD was 32.8%(21/64 cases),while that in healthy control group was only 15.4%(8/52 cases),the difference between the 2 groups was significant (?2= 4.65,P0.05).There were no significant difference of WBC,ESR and CRP between the telomerase of PBMCs positive group and negative group.Conclusions The higher frequency of telomerase expression in peripheral blood lymphocytes might be related to the development and progression of KD.

Objective To explore the significance of nuclear factor-?B(NF-?B)activation in peripheral blood mononuclear cells(PBMC)during acute Kawasaki disease(KD).Methods Peripheral blood was collected from children with acute KD(n=30)and healthy age-matched children(n=20).PBMC were cultured in vitro and divided into 3 groups:naturally cultured blank control group,protein kinase C(PKC)activator stimulated phorbol 12-myristate 13-acetate(PMA)group and PMA plus NF-?B inhibitor treated PMA plus pyrrolidine dithiocarbamate(PDTC)group.Percentages of NF-?B activation were detected by immunohistochemistry.Results Under natural culturing,the percentage of cells with activated NF-?B was significantly higher in acute KD blank control group than that in healthy blank control group.The percentage of cells with activated NF-?B was significantly higher in acute KD PMA group than that in acute KD blank group and that in normal control PMA group,respectively(Pa0.05).Conclusions NF-?B activation in PBMC during acute KD is markedly increased,which suggests that NF-?B activation plays an important role in the formation of vasulitis and CAL in this disease.NF-?B activation in PBMCs in children with KD is regulated by the PKC signaling pathway and PDTC obviously inhibits the activation of NF-?B.J Appl Clin Pediatr,2009,24(1):35-37

Aim To investigate the effect of ribonucleic acidⅡon apoptosis in human leukemia cell lines K562 and KG1 a.Methods Cell counting kit-8(CCK-8)as-say was performed to detect proliferation activity of K562 and KG1 a cells treated with ribonucleic acidⅡ. Apoptosis index was assessed by flow cytometry(FCM) and fluorescent Hoechst 33258 staining was used for observing morphologic changes of apoptosis.Expres-sion levels of p53,Bax,Bcl-2 and cleaved caspase-3 were analyzed by Western blot.Results The prolifera-tion of K562 and KG1 a cells was significantly inhibited by ribonucleic acid Ⅱ treatment for 12 h,24 h,48 h at concentrations of 100~300 mg·L-1 ,which indica-ted the inhibitory effect of ribonucleic acid Ⅱ was in dose-dependent and time-dependent manners.FCM re-sults displayed a dose-dependent increase in cell apop-totic rate.Hoechst 33258 staining showed the typical apoptotic morphology in some leukemic cells treated with ribonucleic acid Ⅱ,including increased nuclear chromatin concentration and edge accumulation.West-ern blot analysis showed the increased expression of p53,Bax,cleaved caspase-3 and decreased expression of Bcl-2 in K562 and KG1 a cells treated with ribonu-cleic acid Ⅱ.Conclusions Ribonucleic acid Ⅱ can induce apoptosis of leukemia K562 and KG1 a cells by up-regulating p53,which mediates Bcl-2/Bax balance and activates caspase-3 .