1.Impact of 4HPR on the expression of E-Cad in human bladder transitional epithelial cancer cells T24.
Eyou, WANG ; Jun, LI ; Guohua, YANG ; Shan, ZHONG ; Tongzu, LIU
Journal of Huazhong University of Science and Technology (Medical Sciences) 2012;32(2):237-41
Previous researches showed that the expression level of E-Cad in most infiltrating cancer cells was reduced or negative. This study explored whether 4HPR restrained the infiltration of bladder cancer cells through regulating the expression of E-Cad. The infiltrating bladder cancer cells T24 were cultured, and then treated by a proper dosage of drug. Their viability was a determined by MTT method. Western blotting and RT-PCR were adopted to detect the changes of E-Cad gene expression at both protein and mRNA levels. Moreover, immunofluorescent staining and confocal fluorescence microscopy were employed for the observation of the expression of E-Cad. The result showed that, at both mRNA and protein levels, the expression level of E-Cad in T24 cells treated by 4HPR was significantly higher than that of control group, while the β-Cat expression was also relocated from the cell nucleus to cytoplasm. Our findings suggested that the regulatory function of 4HPR on infiltration of bladder cancer cells T24 is at least partly achieved by regulating the expression of E-Cad.
2.Impact of 4HPR on the Expression of E-Cad in Human Bladder Transitional Epithelial Cancer Cells T24
WANG EYOU ; LI JUN ; YANG GUOHUA ; ZHONG SHAN ; LIU TONGZU
Journal of Huazhong University of Science and Technology (Medical Sciences) 2012;32(2):237-241
Previous researches showed that the expression level of E-Cad in most infiltrating cancer cells was reduced or negative.This study explored whether 4HPR restrained the infiltration of bladder cancer cells through regulating the expression of E-Cad.The infiltrating bladder cancer cells T24 were cultured,and then treated by a proper dosage of drug.Their viability was a determined by MTT method.Western blotting and RT-PCR were adopted to detect the changes of E-Cad gene expression at both protein and mRNA levels.Moreover,immunofluorescent staining and confocal fluorescence microscopy were employed for the observation of the expression of E-Cad.The result showed that,at both mRNA and protein levels,the expression level of E-Cad in T24 cells treated by 4HPR was significantly higher than that of control group,while the β-Cat expression was also relocated from the cell nucleus to cytoplasm.Our findings suggested that the regulatory function of 4HPR on infiltration of bladder cancer cells T24 is at least partly achieved by regulating the expression of E-Cad.
3.Effect of mAb2G4-ODN-lip on myocardial ischemia-reperfusion injury in rats
Guoqing JING ; Yun WU ; Qingtao MENG ; Eyou WANG ; Zhongyuan XIA
Chinese Journal of Anesthesiology 2018;38(5):614-617
Objective To evaluate the effect of anti-myosin monoclonal antibodies-nuclear factor kappa B (NF-κB) decoy oligodeoxynucleotides-lipofectamine compound (mAb2G4-ODN-lip) on myocardial ischemia-reperfusion (I/R) injury in rats.Methods Forty clean-grade healthy adult male Sprague-Dawley rats,aged 8-10 weeks,weighing 240-260 g,were divided into 4 groups (n=10 each) using a random number table:sham operation group (group S),myocardial I/R group (group I/R),ODN-lip group (group ODN) and mAb2G4-ODN-lip group (group mAb2G4).Myocardial I/R was induced by occlusion of the left anterior descending branch of coronary artery for 30 min followed by 120 min reperfusion.In ODN and mAb2G4 groups,ODN-lip (100 μg ODN) and mAb2G4-ODN-lip (100 μg ODN) compounds were injected via the femoral vein,respectively,immediately after onset of ischemia.The left anterior descending branch of coronary artery was only occluded but not ligated in group S.The animals were sacrificed at 120 min of reperfusion and myocardial specimens of the left ventricle on the ischemic side were obtained for examination of the pathological changes (using haematoxylin and eosin staining) and for determination of the expression of NF-κB (by Western blot) and contents of tumor necrosis factor-alpha (TNF-α) and interleukin-6 (IL-6) (by enzyme-linked immunosorbent assay).Results Compared with group S,the expression of NF-κB was significantly up-regulated,and the contents of TNF-α and IL-6 were increased in I/R,ODN and mAb2G4 groups (P< 0.05).Compared with group I/R,the expression of NF-κB was significantly down-regulated,and the contents of TNF-α and IL-6 were decreased in ODN and mAb2G4 groups (P<0.05).Compared with group ODN,the expression of NF-κB was significantly down-regulated,and the contents of TNF-α and IL-6 were decreased in group mAb2G4 (P<0.05).The pathological changes of myocardial tissues were significantly attenuated in group I/R,group ODN and group mAb2G4 in turn.Conclusion mAb2G4-ODN-lip can mitigate myocardial I/R injury in rats.
4.Role of caveolin-1 in penehyclidine hydrochioride-induced inhibition of lipopolysaccharide-induced activation of TLR4/p38 MAPK signaling pathway in macrophages of mice
Qian KONG ; Xiaojing WU ; Lei ZHANG ; Bo ZHAO ; Ying JIANG ; Qingtao MENG ; Liying ZHAN ; Eyou WANG ; Zhongyua XIA
Chinese Journal of Anesthesiology 2017;37(9):1132-1135
Objective To evaluate the role of caveolin-1 (Cav-1) in penehyclidine hydrochioride (PHC)-induced inhibition of lipopolysaccharide(LPS)-induced activation of Toll-like receptor 4 (TLR4) /p38 mitogen-activated protein kinase (p38 MAPK) signaling pathway in macrophages of mice.Methods Macrophages of mice were seeded in 6 em diameter dishes (5 ml per dish) and divided into 5 groups (n=20 each) using a random number table:Scr-siRNA group (S group),Scr-siRNA + LPS group (LPS group),Ser-siRNA+LPS +PHC group (LPS+P group),Cav-1-siRNA+LPS group (C+LPS group) and Cav-1-siRNA+LPS+PHC group (C+LPS+P group).Macrophages were transfected with Scr-siRNA for 24 h in S,LPS and LPS+P groups and with Smart pool Cav-1 siRNAs for 24 h in C+LPS and C+LPS+P groups.LPS at the final concentration of 1 μg/ml was added after the end of transfection,and macrophages were then incubated for 2 h in LPS,LPS+P,C+LPS and C+LPS+P groups.In LPS+P and C+LPS+P groups,PHC at the final concentration of 2 μg/ml was added at 2 h of incubation with LPS,and macrophages were then incubated for 2 h.The expression of Cav-1 and TLR4 was detected by Western blot.The expression of p38 MAPK was determined by immunofluorescence.The level of tumor necrosis factor-alpha (TNF-α) in the culture medium was determined by enzyme-linked immunosorbent assay.The activity of myeloperoxidase (MPO) in macrophages was measured by colorimetry.Results Compared with group S,the expression of TLR4 and p38 MAPK was significantly up-regulated,and the concentration of TNF-α in the culture medium and activity of MPO were increased in the other four groups,the expression of Cav-1 was significantly downregulated in LPS and C+LPS groups (P <0.05),and no significant change was found in the expression of Cav-1 in group LPS+P (P>0.05).Compared with group LPS,the expression of Cav-1 was significantly up-regulated,the expression of TLR4 and p38 MAPK was down-regulated,and the concentration of TNF-α in the culture medium and activity of MPO were decreased in group LPS+P,and the expression of Cay-1 was significantly down-regulated,the expression of TLR4 and p38 MAPK was up-regulated,and the concentration of TNF-α in the culture medium and activity of MPO were increased in group C+LPS (P<0.05).Compared with group LPS+P,the expression of Cav-1 was significantly down-regulated,the expression of TLR4 and p38 MAPK was up-regulated,and the concentration of TNF-α in the culture medium and activity of MPO were increased in group C+LPS+P (P<0.05).Compared with group C+LPS,the expression of Cav-1 was significantly up-regulated,the expression of TLR4 and p38 MAPK was down-regulated,and the concentration of TNF-α in the culture medium and activity of MPO were decreased in group C+LPS+P (P<0.05).Conclusion The mechanism by which PHC inhibits LPS-induced activation of TLR4/p38 MAPK signaling pathway in macrophages is related to up-regulating Cav-1 expression in mice.