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Objectives To investigate the effects of cell aging on the disorders relating to gastric mucosa aging.Methods A treatment of 200 μmol/L H2O2 was used to induce senescence of human gastric epithelial cell line GES-1,and the cell growth curve was monitored.Senescence secretory phenotypes were observed by detecting the protein level of p53 and p16INK4a with senescence-associated β-galactosidase(SA-β gal)staining and Western blot testing.The mRNA levels of senescence-associated secretory phenotype(SASP)factors in human gastric epithelial GES-1 cell including IL-1β,IL-6,IL-8,TGF-β、IFN-γ,and VEGF-A were detected by RT PCR.The mRNA expression levels of IL-1β,IL-6,IL-8,TGF-β,IFN-γ,and VEGF in the conditioned medium were detected by ELISA analysis.Results The 200 μmol/L H2O2-induced GES-1 cells stopped proliferating after 3 days of treatment,and cells enlarged and flattened at 10 days.The increased SA-β-gal staining(P＜0.001) and the increased expression levels of p53 and p16INK4a proteins indicated the success of establishing the aging model of GES-1.The mRNA levels of IL 1β,IL6,IL8,TGF-β,and IFNγ were higher(t=2.94,3.38,3.15,3.64,2.97;P=0.015,0.000,0.000,0.000,0.000)and the mRNA level of VEGF-A was lower(t=2.31,P =0.20) in senescent GES-1 cells than in the control group.In the conditioned medium of senescent GES-1 cells,the levels of IL-1β,IL6,IL8,TGF-β1,and IFNγ were higher in the H2O2-induced group [(3.12±0.21)μg/L,(4.26±0.15)μg/L,(3.37±0.14)μg/L,(5.34±0.19)μg/L,and(2.90±0.47)μg/L]than in the negative control group[(0.24±0.04,0.04±0.07,0.52±0.02,1.05±0.10,0.52±0.02,respectively,P＜0.001)],while the level of VEGF was lower in the H2O2-induced group than in the negative control group(0.21±0.03)μg/L vs (0.59±0.07)μg/L(P＜0.05).Conclusions The changes in senescence-associated secretory phenotype factors of the aging human gastric epithelial cells induced by oxidative stress may promote chronic gastritis and gastric cancer.
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Objective To explore the potential mechanism of action of estradiol benzoate on proliferation and differentiation of rat hippocampal neural stem cells.Methods Hippocampus tissues from SD rats were dissociated and plated into culture flasks.Neural stem cells were identified by immunofluorescence against Nestin.After treatment with different concentration of estradiol benzoate (10-10,10-9,10-8,10-7,10-6 mol/L),the proliferation and differentiation of neural stem cells were assessed by MTT assay and the proportion of neurons and astrocytes in differentiated cells was assessed by flow cytometry.Western blot assay was performed to detect the expression levels of protein in neural stem cells.Results (1) The neural stem cells formed neurospheres and grew in floating,and were Nestin-positive.Estrogen receptors,including ERα and ERβ,were expressed in neural stem cells;(2)After treatment with 10-8 mol/L estradiol benzoate versus in the control group,the cell viability was significantly increased (t =5.36,P =0.003),and the ratio of neuron-specific nuclear protein(NeuN)-positive neuron cells in total cells was significantly increased (t =4.32,P =0.02),but the ratio of glial fibers acidic protein(GFAP)-positive cell in total cells was decreased(t=4.65,P=0.008).The expression levels of GDNF,GFRal and Ret proteins were enhanced in the estradiol benzoate group compared with the control group.Conclusions Appropriate concentration of estradiol benzoate promotes proliferation and differentiation ofin vitrocultured hippocampal neural stem cells and elevates the ratio between neurons and glial cells differentiated from neural stem cells.
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Objective To study adipose-derived stem cells (ADSCs)differentiation into endothelial cells and smooth muscle cells by induction with supernatant liquid from cerebral infarction tissue in rats.Methods The ADSCs were obtained from retroperitoneal adipose tissue of rats and identified by flow cytometry technology.The normal brain tissues and the infarcted cerebral tissue obtained from rats with middle cerebral artery occlusion were used to induce ADSCs differentiation,and no intervention group was as a control.The mRNA expression level of von willebrand factor (vWF),α-smooth muscle actin (α-SMA) and smooth muscle myosin heavy chain (SM-MHC) in cells after induction were detected using reverse transcriptase-polymerase chain reaction.Immunofluorescence were used to identify the expression of endothelial cells markers and smooth muscle cell markers,and positive expression cell was detected by fluorescence microscope.Results On the ADSCs surfaces,the high expressed-positive antigens included CD90 (96.7％),CD29 (84.4％),CD44 (98.9％),and the low expressed negative antigens included CD45 (6.5％),CD34 (7.4％) and CD31 (3.6％).Compared with no intervention group and normal brain tissue of supernatant liquid group,the cerebral infarction tissue of supernatant liquid-induced group showed the increased mRNA expression level of vWF,α-SMA and SM-MHC(F=5.962,6.756,6.144,P=0.001,0.004,0.003),and showed that the immunofluorescence indicated-cell expression level of vWF,α-SMA and SM-MHC was much more increased (all P ＜ 0.05).Conclusions Under the induction by supernatant liquid of cerebral infarction tissue,ADSCs highly expresses the markers of endothelial cells and smooth muscle cells.This suggests that the cerebral infarction tissue of supernatant liquid-induced ADSCs have a tendency to differentiate into endothelial cells and smooth muscle cells.
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Objective To observe expression changes of the telomeric repeat binding factor 2 (TRF2) in mouse cortical neurons during aging and its biological significance.Methods TRF2 expression in cortical neurons of young (2 months) and old (20 months) C57BL/6J mice were tested by Western blot and real time PCR.pcDNA-TRF2 was transfected in embryonal cortical neurons.Neurons viability was determined by MTT after exposure to camptothecin for 16 h.Results TRF2 expression decreased significantly in cortical neurons in old mice than that in young mice.After exposure to camptothecin for 16 h,(75.4±2.6) ％ of pcDNA TRF2 transfected neurons were viable and the transfection rate was higher in pcDNA-TRF2 transfected neurons than in control transfected neurons [(32.6 ± 9.3) ％] (t =22.85,P ＜ 0.05).Conclusions TRF2 expression decreases significantly in aging mouse,downregulation of TRF2 may participate in neurons aging,and TRF2 overexpression may be a potential therapeutic target against neurodegeneration.
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Objective To synthesize H.pylori bacterial ghosts (BG) and loaded with adriamycin.The cytotoxic effects in gastric cancer cell line were also observed.MethodsThe lysis plasmid was introduced into H.pylori by bacterial conjugation. H.pylori BG were produced by inducing H.pylori lysis at 42 ℃.After suspension and centrifuge, H.pylori BG were loaded with adriamycin.The adriamycin loading quantity was measured with spectrophotometry.The cytotoxic effects of H.pylori BG-adriamycin in gastric cancer cell line SGC7901 were evaluated with MTT assay.ResultsH.pylori BG were successfully synthesized and loaded with adriamycin.The loading quantity of adriamycin was 70.4 μg/mg.H.pylori BG were seen to be adsorbed and internalized by gastric cancer cells under confocal microscope, which distributed on the surface or cytoplasmic of SGC7901 cell line. Carried Adriamycin was delivered into gastric cancer cell line and mainly accumulated in the nucleus.IC50 of SGC7901 to H.pylori BG-adriamycin was 0.32 ± 0.15 by MTT assay, which was significantly lower than that to free adriamycin (0.44 ±0.15, P＜0.05).Conclusions The proliferation of gastric cancer cells were effectively inhibited by H.pylori BG-adriamycin.H.pylori BG are expected to be ideal carrier for anti-gastric cancer medicine.