Objective To construct recombinant adenoviral vector expressing autocatalysis caspase- 3 driven by human telomerase reverse transcriptase promoter amplified by two-step transcription amplification (hTERTp-TSTA),and investigate its antitumor effect in ovarian cancer iri vitro and in vivo.Methods Recombinant adenoviruses expressing autocatalytic caspase-3(rev-caspase-3)driven by hTERTp-TSTA were prepared,which were named as AdHTVP2G5-rev-casp3.AdHT-rev-casp3,Ad-rev-casp3 and AdHTVP2G5- EGEP,which express rev-easpase-3 driven by hTERTp,cytomegalovirus promoter(CMVp)and enhanced green fluorescent protein(EGFP),respectively,were used as controls.Western blot,cell counting kit (CCK-8),flow cytometry(FCM)and TdT-mediated dUTP-biotin nick end labeling(TUNEL)were used to detect the expression of p17,active subunit of caspase-3,and p85,and to measure cell survival rates, apoptotic rates and cell cycle distribution in ovarian cell line AO and normal human umbilical vein endothelial cell line HUVEC,following treatments of AdHTVP2G5-rev-casp3.subcutaneous tumor models and abdominally spread tumor models of human ovarian carcinoma using AO cells in BALB/e nude mice were established.Following treatments of AdHTVP2G5-rev-easp3,western blot was used to detect the expression of active caspase-3 in abdominally spread tumors and liver tissues,respectively,and the mouse survival rates and the volume of tumor nodules were measured,and the serum level of alanine transaminase (ALT)and aspartate transaminase(AST)were analyzed to monitor liver damages and HE staining was used to detect the histopathological changes of various organs.Results The levels of p17 expression in AdHTVP2G5-rev-casp3-treated AO cells were significantly higher than that in Ad-rev-casp3 or AdHT-rev- casp3 treated AO cells,while no expression was observed in AdHTVP2G5-rev-casp3-treated HUVEC.There was strong cell killing of AdHTVP2G5-rev-casp3 of hTERT positive AO cells,but not of the hTERT-negative HUVEC cells.Cell survival rate and apoptotic rate of AO cells treated with AdHTVP2G5-rev-casp3 were 17.8% and 40.2%,respectively,significantly different from that treated with AdHT-rev-casp3(75.2% and 16.1%)at the multiplicity of infection(MOI)of 70(P0.05) .Significant expressions of active caspase-3 were shown in AdHTVP2G5-rev-casp3-treated tumors,whereas no expression was shown in liver.In contrast,both tumors and liver tissues showed active caspase-3 expression following treatments of Ad-rev-casp3.AdHTVP2G5-rev-casp3 and Ad-rev-casp3 prolonged mouse survival[mean survival time of(259?14)d and(213?16)d],when compared with treatment with AdHT- rev-casp3[(177?12)d]and AdHTVP2G5-EGFP[(109?7)d;P

Objective: To compare tea polysaccharides(TPS) characteristics and their role in scavenging free radicals and reducing blood glucose(BG) in diabetic mice(DM). Methods: TPS was extracted from green,Oolong and black tea which were made from the same fresh leaves from Hubei,Fujian and Yunnan. Then the recovery rate of TPS, contents of neutral sugar, uronic acid and protein were analysed, and scavenging rate of -2Oand 稯H in vitro and hypoglycemic effect were also determined. Results: 1. The yield and contents of neutral sugar, uronic acid and protein of green tea TPS were the highest, and those of black tea TPS were the lowest. Oolong tea TPS acted the best in scavenging-2O and 稯H . 2. The hypoglycemic effect of TPS from Hubei tea was the best . The effect of TPS extracted from semi-fermented Oolong tea and fermented black tea was better than that of non-fermented green tea. 3. There were obvious differences in yield, free radical scavenging rate and effect of reducing BG among TPS extracted from tea in different regions. TPS extracted from Fujian tea had the best effect in reducing BG,but that from Yunnan tea had not. Conclusion: There was remarkable effect of region and process on physico-chemical characteristics,effect of scavenging radical and reducing blood sugar TSP.

Culture environment is the key factor in the construction of dermal skin.It was investigated that the effects of the culture methods,including the static culture and spinner flask culture,and stir speeds on the cells proliferation,metabolism and distribution within collagenchitosan sponges.A higher cell density and specific growth rate was obtained with spinner flask culture versus static culture,especially,the 80 r/min spinner flask culture.The cell distribution in dermal substitutes from stirred culture system was more uniform than static culture,as well as that with increase of stir speeds in spinner flask.In summary,the spinner flasks culture with proper stir speed shows promise for the construction of dermal substitutes in vitro.

Background The sex hormones plays an important role in the incidence of dry eye,especially for the regulation of function.However,the effects of sex hormones on lacrimal gland epithelial cells are below understand.Objective This study was to investgate the effects of estradiol and testosterone on the apoptosis of lacrimal gland cells induced by H2O2.Methods The lacrimal gland tissue was obtained from 2- or 3-month-old clean male New Zealand rabbits and the lacrimal gland epithelial cells were cultured in vitro using esplant culture method.The cells were identified by pan cytokeratin antibodies with immunocytochemistry.lacrimal gland epithelial cells were incubated in the 96 well plate at the density of 5 × l04 cells/ml for 44 hours.Estradiol or testosterone with the concentrations of 1 × 10-5,1 × 10-6,1 × 10-7,1 × 10-8 mol/L were added into the medium for 24 hours respectively and 1× 10-4 mol/L H2O2 treated the cells for 1 hour to induce the apoptosis in experimental groups.The cells treated by only 1 × 10-4 mol/L H2O2 were used as apoptotic control group,and the cells cultured by regular method were used as blank control group.The cell viability in different groups was detected using MTT at 570 nm ( A570 ),and the apoptotic rates of the cells were assayed using Annexin V/PI double staining.This use and maintain of experimental animals followed the Regulation for the Administration of Affairs Concerning Experimental Animals by State Science and Technology Commission.Results The cultured cells showed the irregular polygon in shape,and about 80％ cells was positive response for cytokeratin.MTT assay showed that the lower A570 values were detected in the H2O2-induced group,various concentrations of estradiol or testosterone groups compared with blank control group (P＜0.01 ).The A570 values in 1 × 10-5,1 × 10-6,1 × 10-7 mol/L estradiol groups or 1 × 10-6 mol/L testosterone group were significantly higher than ones of H2 O2-induced group (P＜0.01 ).Compared with corresponding concentrations of testosterone groups,the A570values in various concentrations of estradiol groups were elevated( P＜0.01 ).The apoptosis rates at the early and later phase were significantly declined in both estradiol group and testosterone group in comparison with H2 O2-induced group (P ＜ 0.01,P＜ 0.05 ),and those in estradiol group were lower than the testosterone group( P＜0.01,P＜0.05 ).Conclusions Estradiol and testosterone suppress the apoptosis of lacrimal gland cells induced by H2O2,and the stronger effect is found in estrogen.The inhibition of estrogen on lacrimal gland cell apoptosis show a dose-dependent manner to some extent.

To simulate intervention measures in controlling an outbreak of acute hemorrhagic conjunctivitis on one school campus by using the Susceptible-Infected-Recovered (SIR) model, to provide evidence for preparedness and response to the epidemic. Classical SIR model was used to model the epidemic. Malthusian exponential decline method was employed to estimate the infective coefficient β for interventions. The initial value of parameters was determined based on empirical data. The modeling was implemented using Matlab 7.1 software. Without interventions, the outbreak was expected to experience three phrases: (1)early stage (the first 5 days) in which the epidemic developed slowly and could be intervened easily; (2) rapid growing stage (6-15 days) in which the number of infected cases increased quickly and the epidemic could not be well controlled;and (3) medium and late stage (16 days and later) in which more than 90% of the susceptible persons were infected but the intervention measures failed to prevent the epidemic. With the implementation of interventions, the epidemic was predicted to be controlled in the early stage, under the SIR model. The simulation based on the SIR model kept an acceptable consistency with the actual development of epidemic after the implementation of intervention measures. The SIR model seemed effective in modeling interventions to the epidemic of acute hemorrhagic conjunctivitis in the schools.

Pulmonary embolism is a common and lethal, which accounts for substantial morbidity and mortality. Clinical manifestations of pulmonary embolism are nonspecific during general anesthesia. A 60 years old female received elective operation for left femur fracture under general anesthesia. At the end of operation, she suddenly became hypotensive and developed cyanosis. Immediate cardiopulmonary resuscitation(CPR) was performed without definitive diagnosis. Pulmonary embolism was suspected by clinical signs and echocardiography. So, patient was transferred to intensive care unit and with intensive care and aggressive treatment, patient's vital signs and ventilatory status were progressively improved. However, the endotracheal tube was accidentally extubated by the patient at the second postoperative day, and then cardiac arrest was developed and the patient expired. The primary goal of therapy for pulmonary embolization is to prevent reembolization. In the pulmonary thromboembolization, early diagnosis and intensive care improve outcome.
Anesthesia, General* ; Cyanosis ; Diagnosis ; Early Diagnosis ; Echocardiography ; Embolism ; Female ; Femur* ; Heart Arrest ; Humans ; Critical Care ; Intensive Care Units ; Middle Aged ; Mortality ; Pulmonary Embolism* ; Vital Signs

OBJECTIVETo investigate the degradation of artificial basement membrane (matrigel) co-cultured with oral carcinoma-associated fibroblasts (CAFs) and its possible mechanism.

METHODSCAFs and normal fibroblasts (NFs) were incubated on matrigel for 24, 48, 72 h. Equivalent amounts of conditioned medium were collected and assayed for total protein, hydroxyproline and matrix metalloproteinase-2 (MMP-2), matrix metalloproteinase-9 (MMP-9) activity by gelatin zymography.

RESULTSOral CAFs were superior to oral NFs in total protein and hydroxyproline density, CAFs present more pro-MMP-2 and activated MMP-2.

CONCLUSIONCAFs were superior to NFs in degradation of matrigel. CAFs might play a key role in the reconstitution of extracellular matrix and the progression of tumor.

Basement Membrane ; Coculture Techniques ; Enzyme Precursors ; Fibroblasts ; Gelatinases ; Humans ; Matrix Metalloproteinase 2 ; Matrix Metalloproteinase 9 ; Membranes, Artificial ; Mouth Neoplasms

This study was aimed to investigate the cytogenetic characteristics of hematopoietic cells (HC) and bone marrow mesenchymal stoma cells (BMMSC) isolated from patients with myelodysplastic syndrome (MDS) and healthy individuals as normal controls, and to clarify whether HC and BMMSC are simultaneously involved and participate in pathogenesis and development of MDS. Both marrows of 22 newly diagnosed patients with MDS and 7 healthy individuals were collected; BMMSC were isolated and amplified by using established culture system, as well as were identified according to morphologic features and surface antigens detected by flow cytometry; the characteristics of BMMSC from MDS patients were analyzed; the BMMSC karyotyping analysis of MDS patients was performed by banding of HC and BMMSC with pancreatin-Giemsa technique (GTG) and in accordance with ISCN (2005) requirements; the cytogenetic characteristics of HC and BMMSC were compared. The results showed that in vitro culture system for isolation and amplification was successfully established. The light microscopy and flow cytometry confirmed that BMMSC possessed characteristics showing long and thin spindle form and expressing CD29, CD73, CD90 antigens, and unexpressing CD34, CD45 antigens. In BMMSC of 14 MDS patients (64%), the cytogenetic abnormalities were found usually involving the loss of chromosomal material (92%), among which clonal loss was observed in 7 cases (50%). The detection indicated a random loss of chromosomal material in significant proportion of BMMSC. A high proportion of chromosomal material random loss may be a marker of chromosomal instability in BMMSC of MDS patients, the detection also showed that completely consistent aberration types did not exist between HC and BMMSC. The aberrations were observed in 3 cases of MDS with normal karyotype of HC, its aberration rate (33%) was obviously lower than that in MDS patients with abnormal karyotypes (92%). It is concluded that the cytogenetic abnormalities exist in BMMSC of MDS patients, the unbalanced aberration of chromosomal materials may be a genetic instability marker of BMMSC. The difference of aberration types in BMMSC from HC suggests that genetic susceptibility of BMMSC and HC is similar, but no completely identical, which indicates the potential involvement of BMMSC in pathogenesis of MDS. Studying this potential role of BMMSC can be helpful to further understanding of MDS biological characteristics and provide the new approaches for diagnosis and therapy of MDS.
Adult ; Aged ; Aged, 80 and over ; Bone Marrow Cells ; pathology ; Case-Control Studies ; Chromosome Aberrations ; Female ; Humans ; Male ; Mesenchymal Stromal Cells ; pathology ; Middle Aged ; Myelodysplastic Syndromes ; genetics ; pathology ; Young Adult

OBJECTIVETo investigate the expression and relationship of p27(kip1) and its nuclear export factor Jab1 during proliferation process of lymphoma cell.

METHODSJurkat and Raji cells were treated with serum starvation and then serum release. The protein and mRNA expression of p27(kip1), Jab1 in the cells were detected by Western blot and RT-PCR respectively. LMB were used for stimulating Jurkat cells during their proliferation process, and then the expression changes of p27(kip1) and Jab1 were detected. An eukaryotic expression plasmid(pcDNA3. 1-myc) containing Jab1 was constructed. Jurkat cell were transfected in vitro with or without pcDNA3. 1-myc-Jab1. Double immunolabelling was used to identify the localization of p27(kip1). Immunoprecipitation was used to detect the combination of p27(kip1) and Jab1.

RESULTSThe growth of Jurkat and Raji cells were blocked by serum starvation. The total protein amount of p27(kip1) increased while that of Jab1 decreased. The reverse changes were happened after serum release, but the mRNA expression of p27(kip1) has no significant change. LMB could inhibit the cell proliferation caused by serum release. The expression of p27(kip1) was up-regulated and Jab1 down-regulated when Jurkat cells were treated with LMB. After pcDNA3. 1-myc-Jab1 infected Jurkat cells for 48 h, the distribution of p27(kip1) was translocated from nucleus into cytoplasma. p27(kip1) and Jab1 could form compound in Jurkat and Raji cells detected by Immunoprecipitation.

CONCLUSIONJab1 may influence the location and expression of p27(kip1) through integrating with p27(kip1), and then participates in regulating the growth of NHL cell through interfering with the function of p27(kip1).

COP9 Signalosome Complex ; Cyclin-Dependent Kinase Inhibitor p27 ; metabolism ; Humans ; Intracellular Signaling Peptides and Proteins ; metabolism ; Jurkat Cells ; Peptide Hydrolases ; metabolism ; Plasmids ; RNA, Messenger ; metabolism ; Transfection

OBJECTIVETo investigate the biological difference of clonal cells between myelodysplastic syndromes (MDS) and acute myeloid leukemia (AML).

METHODBone marrow (BM) clonal cells (which had cytogenetic markers detected by FISH assay) and blasts were quantitatively analysed in 51 MDS and 11 AML patients. The clonal cell percentage in orthochromatic normoblasts, granulocytes and megakaryocytes were assayed. The biological functions for phagocytosis and oxidation of MDS peripheral blood (PB) neutrophils were compared with that of normal controls.

RESULTSAlmost all MDS patients BM had a higher clonal cell percentage (mean 48.2%) than blasts percentage (mean 6.7%) (P < 0.01), but with the subtype of MDS advancing this percentage gap was closing up, and in 11 AML patients no such gap was observed. This gap in MDS patients with + 8 abnormality was smaller than in those with 5q -. In MDS BM, clonal cells were detected in segmented granulocytes (mean 45.9%), orthochromatic normoblasts (mean 46.0%) and mature megakaryocytes (mean 38.0%). In Addition, an approximate amount of clonal cells with the same karyotype abnormality in BM were detected in MDS PB (mean 37.3% in blood vs 48.6% in marrow). Functional analysis showed that the neutrophils in MDS PB could exert nearly normal physiological functions (P > 0.05), but those from AML could not as compared to healthy donors (P < 0.01).

CONCLUSIONThere is a significant difference in the biological features between MDS and AML clonal cells.

Adult ; Aged ; Aged, 80 and over ; Bone Marrow Cells ; pathology ; Cell Differentiation ; Clone Cells ; Female ; Humans ; Karyotyping ; Leukemia, Myeloid, Acute ; pathology ; Male ; Middle Aged ; Myelodysplastic Syndromes ; pathology