1.Construction of fusion expression vector AR-GFP and its expression in Hek293 cells
Jianwei LIU ; Ling YE ; Jing LIU ; Dongbin NIU ; Mingmei DU ; Yuhong GAO
Chinese Journal of Pathophysiology 2000;0(11):-
AIM:To construct a hAR and GFP fusion gene vector and to observe the AR-GFP gene expression in Hek293 cells. METHODS: A recombined vector pcDNA3.1/myc-HisA-AR-GFP (pH-AG) was constructed by gene engineering technique. The recombined vector was transfected into Hek293 cells using calcium phosphate. RESULTS: AR-GFP fusion protein was successfully expressed in Hek293 cells without biologic activity, which was confirmed by fluorescence microscopy and Western blotting. CONCLUSION: The Hek293 cells transfected by AR-GFP fusion gene can express its protein successfully. However, it is not a cellular model for ARI screening.
2.The role of nephroblastoma overexpressed gene on renal cell carcinoma
Zhihong NIU ; Shuai LIU ; Dongbin BI ; Zheng LIU ; Xiaowen LIU ; Xiaodong YUAN ; Jiaju Lü
Chinese Journal of Urology 2012;33(4):250-253
Objective To investigate the effects of nephroblastoma overexpressed (NOV) on proliferation,adhesion,migration and invasion of remal cell curcinomai (RCC) cells. Methods We constructed a NOV expression plasmid and transfected the plasmid into RCC cell line 786-O and analyzed the effects of NOV expression on proliferation,adhesion,migration and invasion of RCC cells by growth curve assay,WST-1 assay,cell adhesion assay,matrigel invasion assay and transwell migration assay. Results The stable NOV transfected 786-O cells (786-O-NOV) showed decreased growth rate,at 48 h and 72 h,the proliferation activities of 786-O-NOV cells were inhibited by 29.14% and 32.46% the proliferation activities of empty vector cells were inhibited by 9.25% and - 8.16%,respectively,compared to 786-O cells (P <0.05); while the 786-O cells transfected with empty vector (786-O-mock) had no difference with 786-Ocells.Adhesion assay indicated significantly increased adhesion of 786-O-NOV cells to fibronectin (0.26 ±0.03) and laminin (0.28 ±0.04),compared to 786-O cells (0.15 ±0.01,0.12±0.10) and 786-O-mock cells (0.14 ±0.02,0.13 ± 0.08).Invasion assay displayed that the numbers of cells penetrated through matrigel membrane were significantly higher in 786-O-NOV cells (240.25 ± 23.12) compared to 786-O cells ( 56.16 ± 6.25 ) and 786-O-mock cells ( 50.28 ± 7.13 ).Migration assay displayed that the numbers of cells passed through polycarbonate filters were significantly higher in 786-O-NOV cells (267.25 ± 20.94) compared to 786-O cells ( 66.10 ± 5.68 ) and 786-O-mock cells ( 56.28 ± 4.11 ).Conclusion NOV exhibits anti-proliferative effects on RCC cells; however,it promotes adhesion,migration and invasion of RCC cells.
3.Electroacupuncture up-regulated arcuate nucleus ?-MSH expression in the rat of diet-induced obesity
Derun TIAN ; Xiaodong LI ; Dongbin NIU ; Yushun SHI ; Zhaokang ZHANG ; Jisheng HAN ;
Journal of Peking University(Health Sciences) 2003;0(05):-
Objective: To investigate the effect of electroacupuncture (EA) on rat with diet induced obesity (DIO) and to explore the possible neurochemical mechanisms using the technique of immumohistochemisty. Methods:To establish DIO rat model by feeding the animals with high fat diet for 14 weeks. DIO rats were randomly divided into 4 groups: (1) 2Hz EA group, (2) 100Hz EA group, (3) restrain control group,(4) diet resistance (DR) group,(5) DIO group and (6) normal control group. EA treatment: (1) The acupoints used were Zusanli and Sanyinjiao on both legs. (2) The intensities of stimulation were 0.5, 1.0 and 1.5mA for 10 mins each. EA treatment was administered 3 times per week. Food intake and body weight were measured daily for 4 weeks. (3) The changes of the expression of ? melanocyte stimulating hormone (? MSH) in the hypothalamic arcuate nucleus (ARC) were measured with immunohistochemical semiquantitative analysis. Results: (1) The food intake and body weight of 2 Hz EA group and 100 Hz EA group were decreased significantly compared with the restrain control group and DIO group. (2) The number of ? MSH positive cells in hypothalamic ARC in 2 Hz EA and 100 Hz EA group was significantly higher than that in restrain control group and DIO group. The number of ? MSH positive cells in hypothalamic ARC in DIO group is significantly lower than those in DR group or normal control group. Conclusion: A decrease of ? MSH level in hypothalamus may be associated with diet induced obesity. The therapeutic effect on obesity produced by EA may be accounted for by the stimulation of pro opio melanocortin neurons in hypothalamic ARC to release ? MSH, which inhibits food intake , resulting in a decrease of body weight.