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Objective: To investigate the inhibitory effect of T-2 toxin on proliferation of the human hepatocellular carcinoma HepG2 cells and its promotion effect of apoptosis.Methods:The HepG2 cells in the logarithmic phase were selected and divided into control group (without T-2 toxin)and experimental groups (given 0.25,2.50,25.00,250.00 and 2500.00 μg·L-1 T-2 toxin).After 24 h treatment,the morphology of cells was observed under inverted microscope;the inhibitory rate of proliferation of cells was determined by MTT assay;the cell cycle and apoptotic rate of cells were analyzed by flow cytometry;Hochest 33258 staining was used to observe the apoptotic morphology of cells;the activity of caspase-3 in HepG2 cells was detected.Results:After treated for 24 h,the inverted microscope observation results showed that the number of the cells in experimental groups was decreased significantly and the cells shrank and deformed.The MTT results showed that compared with control group,the inhibitory rates of proliferation of the cells in experimental groups were increased (P <0.01).The flow cytometry results showed that compared with control group, the percentage of the cells in SubG1 phase in experimental group was significantly increased,and the apoptotic rates of the cells in experimental groups were significantly increased.The Hoechest 33258 staining results showed that the chromatin condensation was observed in the cells in experimental groups,and the nuclei were dense and stained.Compared with control group,the activities of intracellular caspase-3 of the cells in experimental groups were significantly increased (P < 0.01 ). Conclusion:T-2 can inhibit the proliferation of human hepatocellular carcinoma HepG2 cells,and induce the apoptosis.
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Objective:To modify Ganodermalucidum polysaccharides(GLP) with sulfate and observe the protective effect of Ganodermalucidum polysaccharide sulfate (GLPS) on the cerebral ischemia reperfusion injury in the rats,and to investigate its mechanism.Methods:GLP was modified by sulfation to obtain GLPS.A total of 100 SD rats were randomly divided into sham operation group, model group, GLP group (40 mg·kg-1·d-1), GLPS group (40 mg·kg-1·d-1) and nimodipine group (1 mg·kg-1·d-1).The cerebral ischemia reperfusion models were established by middle cerebral artery occlusion method in the rats.The neurologic deficit score and the content of water in brain tissue of the rats with cerebral ischemia reperfusion injury were detected and the activities of superoxide dismutase(SOD) and the levels of malondialdehyde (MDA) were detected.The levels of nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB),tumor necrosis factor-alpha (TNF-α), interleukin-1 (IL-1) and interleukin-6 (IL-6) in the brain tissue homogenate were detected by ELISA.Western blotting method was used to detect the protein expression levels of HSP70 and p-Akt in the brain tissue of the rats.Results:Compared with model group, the neurological function scores of the rats in GLP group and GLPS group were decreased(P<0.01),the water contents in brain tissue were decreased(P<0.05), the SOD activities were increased and the MDA levels were decreased(P<0.05), and the levels of NF-κB, TNF-α, IL-1 and IL-6 were decreased(P<0.05);the effect in GLPS group was significantly better than that in GLP group(P<0.05).The results of Western blotting method showed that the p-Akt protein expression levels in the brain tissue of the rats in GLP and GLPS groups were increased compared with model group (P<0.05);compared with model group, the HSP-70 protein expression level in the brain tissue of the rats in GLPS group was increased(P<0.01),but the effect in GLP group was not obvious.Conclusion:Sulfation can significantly improve the protective effect of GLP on the cerebral ischemia reperfusion injury in the rats and its mechanism may be related to regulating the HSP70/PI3K/Akt signaling pathway and inhibiting the inflammatory reaction damage to the nerve cells of reperfusion.
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Objective:To prepare the velvet antler polypeptide-collagen/chitosan composite materials,and to investigate its promotive effect on cicatrization of mandibular defect and possible mechanism.Methods:The collagen and chitosan solution were mixed.The composite material was prepared by glutaraldehyde crosslinking method.The microstructure of the composite material was observed by transmission electron microscope (SEM).The unilateral mandibular defect models of 36 rabbits were established.The rabbits were divided into experiment and control groups,and each group was divided into 4-,8-and 12-week subgroups,and there were 6 rabbits in each sub group.The rabbits in experiment group were implanted with velvet antler polypeptide-collagen /chitosan composite materials and the rabbits in control group were treated.4,8 and 12 weeks after operation,the histology of bone defect and peripheral nerve reconstruction of the rabbit models were detected by CT;the expression of vascular endothelial growth factor (VEGF) in bone tissue of the rabbits was detected by immunohistochemistry;the ultrastructure of bone defect was observed by SEM.Results:The structure of composite materials had layered folds and the inner diameter of the stent became larger and mainly dominated by sheet structure,which was the ideal structure of biological materials.4 weeks after operation,the new bone was formatted in experiment group,most of the new bone like-tissue materials were degraded,and the VEGF expression showed an increasing trend;8 weeks after operation,the trabecular bone in the bone defect of the rabbits in experiment group was increased obviously and the expression of VEGF was decreased.12 weeks after operation,the new bone formation and the density in experiment group was consistent with the normal tissue,and the expression level of VEGF returned to normal.At each the point after operation,the degree of bone defect healing and bone formation rate in experiment group were obviously prior to control group.Conclusion:Velvet antler polypeptide-collagen /chitosan composite material has the promotive effect on the fracture healing of mandibular defect of the rabbits and its possible mechanism may be related to promoting the expression of VEGF.
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Objective:To explore the mechanism of promotion effect of juglone combined with cisplatin on the apoptosis of human cervical cancer HeLa cells,and to clarify the effects of its associated signal transduction pathways.Methods:The HeLa cells at logarithmic growth phase were divided into control group,juglone group, cisplatin group and juglone combined with cisplatin group (combined treatment group).The inhibitory rates of proliferation of HeLa cells were detected by MTT assay.The apoptosis was detected by Hoechst 33258 staining. The expressions of Bcl-2, Bax and caspase-3, AKt, and pAKt were detected by Western blotting method. Results:The MTT results showed that the HeLa cell proliferation at 24,48 72 h in each drug group was inhibited;compared with control group,the profileration of HeLa cells in juglone group and cisplatin group was significantly inhibited,especially in combined group. Compared with single drug group,the inhibitory effect in combined treatment group was more significantly.After treatment for 12 h,the typical morphological changes of apoptosis were found in juglone group and cisplatin group by Hoechst 33258 staining,especially in combined treatment group. The Western blotting results showed that the expression levels of Bcl-2 and pAKt in HeLa cells in juglone group and cisplatin group 12 h after treatment were decreased obviously,whereas the expression levels of Bax,Caspase-3,and AKt were increased significantly, especially in combined treatment group compared with control group. Conclusion:Juglone combined with cisplatin could inhibit the PI3K/AKt pathway,thereby promoting the apoptpsis of HeLa cells.
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AIM:To explore the effect of peptidyl-prolyl cis/trans isomerase (Pin1) inhibitor juglone on apop-tosis of human cervical cancer SiHa cells.METHODS:Cultured SiHa cells were incubated with juglone at concentrations of 10, 20, 50, 80 and 100 μmol/L for 24 h.The SiHa cell activity was detected by methyl thiazolyl tetrazolium ( MTT) assay.The cell apoptosis was analyzed by flow cytometry with Hoechst 33258 staining.The protein levels of cleaved caspase-3,8,9 and PTEN was determined by Western blotting.RESULTS:In different doses of juglone groups, the SiHa cell growth was greatly inhibited ( P<0.05) in a dose-dependent manner as compared with control group.The IC50 of ju-glone was 20.4 μmol/L.After treatment with juglone at concentration of 20 μmol/L for 12 h, the apoptosis of SiHa cells was induced, and the typical morphological changes of cell apoptosis such as karyopyknotic pyknic hyperfluorescence bolus, nuclear fragmentation and apoptotic body were observed by Hoechst 33258 staining.The early apoptotic rate was increased significantly as compared with the control.The protein levels of cleaved caspase-3, 8, 9 and PTEN were also increased sig-nificantly as compared with control group.CONCLUSION:Juglone significantly inhibits the cell activity and induces the apoptosis of SiHa cells in vitro by inhibiting the caspase pathway and increasing the expression of anti-oncogene.
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[ ABSTRACT] AIM:To investigate the effect of microwave radiation at different intensities on the rat myocardium and its possible mechanism.METHODS:The rats were radiated by the intensity of 500, 1 000, 1 500 and 2 000 W/m2 with 2 450 MHz microwave for 6 min.The heart tissue was collected 6 h after microwave radiation.ATP and mitochondria complexⅣandⅤwere measured.The changes of the tissue structures were observed under transmission electron micro-scope.The apoptosis of the myocardial cells was detected by a cell analyzer.The protein level of cleaved caspase-3 was de-termined by Western blotting.RESULTS:The concentration of ATP and activity of mitochondria complexⅣandⅤsigni-ficantly decreased compared with control group in the cardiac tissues.The decreased number, morphological abnormalities such as dissolved cavitation, matrix and obvious tumefaction of mitochondria were observed under transmission electron mi-croscope.The microwave radiation induced the apoptosis of myocardial cells in the rats.The cell apoptotic rate and the pro-tein level of cleaved caspase-3 increased with increasing intensity of microwave radiation ( P<0.05 ) .CONCLUSION:Microwave radiation has obvious injury effect on the rat heart, which can cause cardiac energy metabolism dysregulation and cardiac myocyte apoptosis.
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OBJECTIVE To explore the pro-apoptotic mechanism of juglone in SiHa cells and to in?vestigate its associated signal transduction pathways. METHODS SiHa cells were treated with juglone 20μmol·L-1 for 24,48 and 72 h. Cellular morphology was detected by inverted microscopy.The cell viability was detected by methyl thiazolyl tetrazolium (MTT) assay. After 24 h treatment with juglone 20μmol·L-1,the cell apoptosis was detected by flow cytometry while the expressions of apopto?sis-related protein BCL-2,BAX and cleave-caspase-3,PI3K/AKt pathway-related protein PTEN,AKT and pAKT were detected by Western blotting. RESULTS After treatment with juglone for 24, 48 and 72 h,the growth of SiHa cells was significantly inhibited. Compared with cell control group,cells in juglone treated gruop were sparse,slipped off the wall,became round and the cell proliferation inhibitory rate was 43.3%,63.0%and 73.1%(P<0.05,P<0.01),respectively. Twenty-four hours post treatment, the early apoptosis rate of juglone treated gruop cells was increased by(6.47±1.79)%(P<0.01)compared with cell control group. Western blotting results showed that the expression of BCL-2 decreased by 53.0%while the expression of BAX and caspase-3 increased by 85.5%and 183.3%,respectively. The expression of PTEN was increased by 75.0% but the pAKt was decreased by 45.8%(P<0.01). CONCLUSION Juglone can upregulate the expression of PTEN, thus inhibiting PI3K/AKt pathway and promoting apoptosis of SiHa cells.
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Objective To investigate different media-use habits and different media information comprehension of the audiences.Methods 127 inpatients and 33 outpatients were sampled for questionnaire survey,learning such information as the basics of the audience,medical experience and current media use,participation intension analysis.Results Internet,television,newspapers and other mass media constitute main channels of the audiences to learn health knowledge; audiences depend on the media to know the departments and specialists of the hospital Conclusion in the meantime of professional disciplines development and innovation,the hospital should not ignore its branding and innovation; an important management tool in its branding strategy is to integrate hospital and media resources,build positive image of the hospital,in order to improve hospital brand awareness,credibility and reputation
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OBJECTIVE To investigate the protective effect of velvet antler polypeptides(VAP)on hydrogen peroxide( H2 O2 )-induced injury in vascular endothelial cells and the possible mechanism.METHODS The EVC-304 cells cultured invitrowere incubated with H2 O2 for another 12 h after pretreat-ment with VAP 20,40 and 80 mg·L-1 for 24 h. Cell viability was determined by MTT assay, Hoechst333258 staining was used to observe cell morphology,the activity of superoxide dismutase (SOD)and the level of malondialdehyde( MDA)were detected with kits and the expression of heat shock protein(HSP70)and caspase 3 was detected by Western blotting. RESULTS Compared with the normal control group,the cell survival rate was decreased significantly in H2 O2 injury group( P ﹤0.01),cell shrinkage,chromatin condensation,and nuclear fragmentation were seen,the intracellular SOD activity decreased while MDA content increased(P﹤0.01),and caspase 3 and HSP70 expression increased(P﹤0.01). Compared with H2 O2 group,the cell survival rate in VAP 20,40 and 80 mg·L-1 pre-treatment groups increased significantly(P﹤0.01),the apoptosis ratio declined from(25.3±1.0)% to (15.2±1.2)%,(10.3±0.9)% and(7.9±1.4)%(P﹤0.01),the SOD activity increased to 19.2±0.5,22.3± 1.7 and(24.9±0.6)kU·g-1 protein(P﹤0.01),MDA concentration decreased to 1.51±0.2,1.48±0.3 and (1.02±0.1)μmol·g-1 protein(P﹤0.01),and the expression of caspase 3 and HSP70 declined significant-ly(P﹤0.01). CONCLUSION VAP has exert protective effect on H2 O2-induced injury in vascular endothe-lial cells. The possible mechanism might be related to improvement of intracellular oxidative stress level.
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Purpose Melittin was expressed in prokaryotic vector pGEX-2T for production. Methods Melittin gene synthesized with enterokinase digested sequence,the gene was cloned into vector pGEX-2T,and constructed a recombinant plasmid of pGEX-MEL. Then the recombinant vector was introduced into E. coli BL21 (DE3)for expression. Fusion protein was purified by affinity chromatography. Hemolytic activity of Melittin was detected. Results Analysis result showed that the expression products accumulate in the cells to about 29.5 % of total cell protein. Detection of western blot using ant-GST as the first antibody showed that a special blot was revealed among the expression products. It certified that we have succeeded in expressing the fusion protein. SDS-PAGE showed that most part of the products is resoluble. The purity of obtained protein is 95% , by through GST affinity chromatography system. Melittin is harvested with a recovery of 80% by EK digestion. Test results showed melittin has good hemolytic activity. Conclusion We have expressed Melittin successfully by prokaryotic expression system.