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Peroxisome proliferator-activated receptor γ ligands induce cell cycle arrest and apoptosis in human renal carcinoma cell lines

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Acta Pharmacologica Sinica 2005年第6期26卷 753-761页

作者： Feng-guang YANG~2, Zhi-wen ZHANG~3, Dian-qi XIN~2, Chang-jin SHI~2, Jie-ping WU~2, Ying-lu GUO~(2,4), You-fei GUAN~(3,4) ~2 Department of Molecular biology, Institute of Urology Peking University First Hospital ~3 Department of Physiology and Pathophysiology, Peking University Health Science center, Beijing 100034, China Department of Molecular biology Institute of Urology Peking University First Hospital Beijing 100034 China Department of Physiology and Pathophysiology Peking University Health Science Center Beijing 100034 China

Aim: To study the effect of peroxisome proliferator-actived receptor y （PPARγ） ligands on cell proliferation and apoptosis in human renal carcinoma cell lines. Methods: The expression of PPARγ was investigated by reverse transcriptase polymerase chain reaction （RT-PCR）, Western blot and immunohistochemistry. The effect of thiazolidinedione （TZD） PPARγ ligands on growth of renal cell carcinoma （RCC） cells was measured by MTT assay and flow cytometric analysis. Cell death ELISA, Hoechst 33342 fluorescent staining and DNA ladder assay were used to observe the effects of PPARγ ligands on apoptosis. Regulatory proteins of cell cycle and apoptosis were detected by Western blot analysis. Results: PPARγ was expressed at much higher levels in renal tumors than in the normal kidney （2.16±0.85 vs 0.90±0.73; P32 μmol/L for pioglitazone, and 5.71 μmol/L and 8.38 μmol/L for troglitazone in 786-0 and A498 cells, respectively. Cell cycle analysis showed a G0/G1arrest in human RCC cells following 24-h exposure to TZD. Analysis of cell cycle regulatory proteins revealed that TZD decreased the protein levels of proliferating cell nuclear antigen, pRb, cyclin D1, and Cdk4 but increased the levels of p21 and p27 in a time-dependent manner. Furthermore, high doses of TZD induced massive apoptosis in renal cancer cells, with increased Bax expression and decreased Bcl-2 expression. Conclusion: TZD PPARγligands showed potent inhibitory effect on proliferation, and could induce apoptosis in RCC cells. These results suggest that ligands for PPARγ have potential antitumor effects on renal carcinoma cells.

关键词： renal cell carcinoma peroxisome proliferator activated receptor γ cell cycle apoptosis cyclin cdk

Impaired reproduction in transgenic mice overexpressing γ-aminobutyric acid transporter Ⅰ (GAT1)

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Cell Research 2004年第1期14卷 54-59页

作者： Jia Hua HU~(1,*), Jin Fu ZHANG~(2,*), Ying Hua MA~1, Jie JIANG~1, Na YANG~1, Xin Bo LI~3, Zhi Guang YU CHI~4, Jian FEI~(1,5,**), Li He GUO~(1,**) ~1Laboratory of Molecular Cell biology, Institute of Biochemistry and Cell biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031, China. ~2Department of Urology, Tongji Hospital, Tongji University, 389 Xincun Road, Shanghai 200065, China. ~3New Drug R & D center, North China Pharmaceutical Group Corporation, 388 Heping East Road, Shijia Zhuang 050015, Hebei, China. ~4School of life sciences, Fudan University, 220 Handan Road, Shanghai 200433, China. ~5Shanghai Research center for Biomodel Organism, Shanghai 201203, China. Laboratory of Molecular Cell biology Institute of Biochemistry and Cell Biology Shanghai Institutes for Biological Sciences Chinese Academy of Sciences Shanghai 200031 China.

It is well documented that γ-aminobutyric acid (GABA) system existed in reproductive organs. Recent researches showed that GABA_A and GABA_B receptors were present in testis and sperm, and might mediate the acrosome reaction induced by GABA and progesterone. GABA transporter Ⅰ (GAT1) also existed in testis and sperm, but its physiological function was unknown. In the present study, we used GAT1 overexpressing mice to explore GAT1 function in male reproductive system. We found that the expression level of GAT1 continuously increased in wild-type mouse testis from 1 month to 2 months after birth. GAT1 overexpression in mouse affected testis development, which embodied reduced testis mass and slowed spermatogenesis in transgenic mice. Moreover, transgenic mice showed increase of the percentage of broken sperm. The further study revealed that the reproductive capacity was impaired in GAT1 overexpressing mice. In addition, testosterone level was significantly low in transgenic mice compared with that in wi

关键词： GABA GAT1 testes sperm reproduction transgenic.

The intracellular mechanism of alpha-fetoprotein promoting the proliferation of NIH 3T3 cells

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Cell Research 2002年第2期12卷 151-156页

作者： MENG SEN LI, PING FENG LI, FBI YI YANG, SHI PENG HE, Guo GUANG DU, GANG LI1 Department of Biochemistry and Molecular biology, 2 Department of Biophysics, Health Science center, Peking University, Beijing 100083, China 1Department of Biochemistry and Molecular biology Health Science Center Peking University Beijing 100083 China 1Department of Biochemistry and Molecular biology Health Science Center Peking University Beijing 100083 China 1Department of Biochemistry and Molecular biology Health Science Center Peking University Beijing 100083 China 2Department of Biophysics Health Science Center Peking University Beijing 100083 China 1Department of Biochemistry and Molecular biology Health Science Center Peking University Beijing 100083 China 1Department of Biochemistry and Molecular biology Health Science Center Peking University Beijing 100083 China

AIM The existence and properties of alpha-fetoprotein (AFP) receptor on the surface of NIH 3T3 cells and the effects of AFP on cellular signal transduction pathway were investigated. METHODS The effect of AFP on the proliferation of NIH 3T3 cells was measured by incorporation of 3H-TdR. Receptor-binding assay of 125I-AFP was performed to detect the properties of AFP receptor in NIH 3T3 cells. The influences of AFP on the [cAMP]i and the activities of protein kinase A (PKA) were determined. Western blot was used to detect the change of K-ras P21 protein expression. RESULTS The proliferation of NIH 3T3 cells treated with 0-80 mg/L of AFP was significantly enhanced. The Scatchard analysis indicated that there were two classes of binding sites with KD of 2.722×10-9M (Bmax=12810 sites per cell) and 8.931× 10-8M (Bmax=119700 sites per cell) respectively. In the presence of AFP (20 mg/L), the content of cAMP and activities of PKA were significantly elevated . The level of K-ras P21 protein was upregulated by

关键词： Alpha-fetoprotein receptor signal transduction monoclonal antibody gene expression.

MicroRNA miR-24 inhibits erythropoiesis by targeting activin type Ⅰ receptor ALK4

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MicroRNA miR-24 inhibits erythropoiesis by targeting activin...

Cloning of rat sp56, the homologue of mouse sperm ZP3 receptor-sp56

中国遗传学会“发育、遗传和疾病”研讨会

作者： Qiang Wang~1 Zheng Huang~2 Huiling Xue~2 Chengcheng Jin~1 Xiu-Li Ju~3 Jing-Dong J.Han~2 Ye-Guang Chen~(1#) 1 State Key Laboratory of Biomembrane and Membrane Biotechnology,Department of Biological Sciences and Biotechnology,Tsinghua University,Beijing 100084,China 2 CAS Key Laboratory of Molecular Developmental biology and center for Molecular systems biology,Institute of Genetics and Developmental biology,the Chinese Academy of Sciences,Beifing 100101, China 3 Department of Pediatrics,Qilu Hospital of Shandong University,Jinan 250012,China

MicroRNAs have been suggested to modulate a variety of cellular *** we report that miR-24 regulates erythroid differentiation by influencing the expression of human activin type I receptor ALK4(bALK4).Ectopic expression of miR-24 reduces the mRNA and protein levels of bALK4 by targeting its 3'-untranslated region and interferes with activin-induced Smad2 phosphorylation and reporter ***,miR-24 represses the activin-mediated accumulation of hemoglobin,an erythroid differentiation marker,in erythroleukemic K562 cells and decreases erythroid colony-forming and burst-forming units of CD34+ hematopoietic progenitor ***4 expression is reversely correlated with miR-24 expression during erythroid ***,our findings define a regulation mode of miR-24 on erythropoiesis by impeding ALK4 expression.

关键词： ALK receptor ALK4 MicroRNA miR-24 inhibits erythropoiesis by targeting activin type miR

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Cell Research 2003年第2期13卷 121-129页

作者： SS KOIDE The Population Council center for Biomedical Research 1230 York Avenue New York New York 100021 USA

Mouse sp56 is considered as one of the candidates for mouse zona pellucida 3 (mZP3) receptor. Up to date, its homologue has only been cloned from guinea pig. namely AM67. Based on the cDNA sequence of mouse sp56, we designed a pair of primer to amplify its homologue from rat testis cDNA. Using RT-PCR, two fragments of 743 bp and 938 bp were amplified. The PCR products show very high homology to mouse sp56. However, the 743 bp product completely lacks one of the seven Sushi domains of mouse sp56. Using the 743 bp product as the probe to detect the expression profile of sp56 in rat tissues, Northern blot shows that a -2.0 kb mRNA expresses specifically in testis. Employed the RACE method, two full cDNA sequences of rat sp56 were obtained. A Mr -42 KD band was detected in denatured and non-reducing protein sample of rat testis and sperm with anti-mouse sp56 monoclonal antibody by Western blot method. Rat sp56 was localized on rat sperm head by the indirect immunofluorescence method. Rat sp56 immunoreactivi

关键词： ZP3 receptor sp56 rat testis sperm cloning.

A20 overexpression under control of mouse osteocalcin promoter in MC3T3-E1 cells inhibited tumor necrosis factor-alpha-induced apoptosis

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Acta Pharmacologica Sinica 2006年第9期27卷 1231-1237页

作者： Yue-juan QIN~2 Zhen-lin ZHANG~(2,4)Lu-yang YU~3 Jin-wei HE~2 Ya-nan HOU~3 Tian-jin LIU~3 Jia-cai WU~3 Song-hua WU~2 Li-he GUO~3 2 center for Preventing and Treating Osteoporosis,Osteoporosis Research Unit,Shanghai Sixth People's Hospital,Shanghai Jiao Tong University,Shanghai 200233,China 3 Institute of Biochemistry and Cell biology,Shanghai Institutes for Biological Science,Chinese Academy of Sciences,Shanghai 200031,China centerforPreventingandTreatingOsteoporosis OsteoporosisResearchUnitShanghaiSixthPeople'sHospitalShanghaiJiaoTongUniversityShanghai200233China InstituteofBiochemistryandCellbiology ShanghaiInstitutesforBiological

Aim:To construct an A20 expression vector under the control of mouse osteocalcinpromoter(OC-A20),and investigate osteoblastic MC3T3-EI cell line,whichstably overexpresses A20 protein prevented tumor necrosis factor(TNF)-alpha-induced ***:OC-A20 vector was constructed by fusing a frag-ment of the mouse osteocalcin gene-2 promoter with human A20 *** the mouse MC3T3-E1 cell line,stably transfected by A20,*** expression of A20 mRNA and A20 protein in the cells weredetected by reverse transcription-polymerase chain reaction(RT-PCR)and West-ern blot analysis,*** determine the specificity of A20 expression inosteoblast,the mouse osteoblastic MC3T3-E1 cell line and mouse embryo fibro-blast NIH3T3 cell line were transiently transfected with *** anti-apoptoticrole of A20 in MC3T3-E1 cells was determined by Flow cytometric analysis(FACS),terminal dUTP nick endo-labeling(TUNEL)and DNA gel electrophoresis analysis(DNA Ladder),***:Weak A20 expression was found in MC3T3-E1 cells with the primers of mouse A20.A20 mRNA and A20 protein expressionwere identified in MC3T3-E1 cells transfected with OC-A20 using RT-PCR andWestern blot *** A20 mRNA expression was found in MC3T3-E1 cellafter MC3T3-E1 cells and NII-/3T3 cells were transient transfected with *** obviously occurred in the rate of apoptosis in the OC-A20 group com-pared with the empty vector(pcDNA3)group by FACS(P3T3-E1 cells andconfirmed that A20 protects osteoblast against TNF-alpha-induced apoptosis.

关键词： A20 osteoblast apoptosis tumor necrosis factor-alpha

Effects of ohmefentanyl stereoisomers on phosphorylation of cAMP-response element binding protein in cultured rat hippocampal neurons

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Acta Pharmacologica Sinica 2003年第12期24卷 79-84页

作者： GAO Can~3, CHEN Li-Wei, TAO Yi-Min, CHEN Jie, XU Xue-Jun, CHI Zhi-Qiang~2 Shanghai Institute of Materia Medica, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031 ~3Research center for Biochemistry and Molecular biology, Xuzhou Medical College, Xuzhou, Jiangsu 221002, China Shanghai Institute of Materia Medica Shanghai Institutes for Biological Sciences Chinese Academy of Sciences Shanghai 200031 China

AIM: To define the effects and signal pathways of ohmefentanyl stereoisomers [(-)-cis-(3R,4S,2’R) OMF (F9202), (+)-cis-(3R,4S,2’S) OMF (F9204), and (-)-cis-(3S,4S,2’R) OMF (F9203)] on the phosphorylation of cAMP-response element binding protein (CREB) in cultured rat hippocampal neurons. METHODS: The effects of the three OMF stereoisomers and morphine (Mor) on cAMP accumulation and CREB phosphorylation were monitored by radioimmunoassay and Western blot analysis, respectively. RESULTS: The three OMF stereoisomers and Mor could all partially inhibit forskolin-stimulated (25μmol/L, 15min) cAMP accumulation in a dose-dependent manner and this effect could be reversed by naloxone. F9202, F9204, and Mor could significantly increase CREB phosphorylation from 2.88 to 3.59 folds over control levels after 30-min exposure. This effect was reversed by naloxone, but F9203 failed to increase CREB phosphorylation. KN-62 and staurosporine significantly blocked the opioidsinduced CREB phosphorylation, while H-89 and P

关键词： ohmefentanyl stereoisomers cAMP-response element binding protein mu opioid receptor cyclic AMP hippocampal neurons phosphorylation

Upregulation of TNF-αand IL-6 mRNA in mouse liver induced by bacille Calmette-Guerin plus lipopolysaccharide

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Acta Pharmacologica Sinica 2006年第4期27卷 460-468页

作者： Dao-fang LIU~(1,2,3) Wei WEI~(1,4) Li-hua SONG~(1,2,3,4) ~1Institute of Clinical Pharmacology,Key Laboratory of Anti-inflammatory and Immunopharmacology in Anhui Province.Key Laboratory of Research and Development of Chinese Medicine in Anhui Province.Anhui Medical University,Hefei 230032,China:~2Molecular biology Laboratory,Anhui biology Institute,Hefei 230088,China ~3center of Research and Development,Anhui Anke Biotechnology Group,Hefei 230088.China Institute of Clinical Pharmacology Key Laboratory of Anti-inflammatory and Immunopharmacology in Anhui Province Key Laboratory of Research and Development of Chinese Medicine in Anhui Province Anhui Medical University Hefei 230032 China Molecular biology Laboratory Anhui Biology Institute Hefei 230088 China center of Research and Development Anhui Anke Biotechnology Group Hefei 230088 China

Aim:To investigate the mechanism of immunological liver injury induced by bacilleCalmette-Guerin(BCG)plus lipopolysaccharide(LPS).Methods:Mice were in-jected via the tail vein with 125 mg/kg BCG,and 12 d later,the mice were injectedintravenously with different doses of LPS(125,250,or 375 μg/kg).Serum alanineaminotransferase(ALT)activity and liver pathological changes were *** expression of tumor necrosis factor(TNF)-α interleukin(IL)-6,lipopolysac-charide binding protein(LBP)and CD14 mRNA,and NF-κB and IκB-α protein inmouse liver at different time points after BCG and LPS injection were measuredusing RT-PCR,immunohistochemistry and Western blotting analysis,***:The activity of serum ALT in mice treated with BCG and LPS was signifi-cantly *** degrees of liver injury,such as inflammatory cellinfiltration,spotty necrosis,piecemeal necrosis,even bridging necrosis,could beseen in liver sections from mice after BCG and LPS ***,thelevels of TNF-α and IL-6 mRNA in mouse liver were significantly elevated afteradministration of BCG plus LPS(P<0.05).The levels of LBP and CD14 mRNA inmouse liver were markedly upregulated after treatment with BCG and LPS,andtreatment with BCG alone led to an increase in CD14 InRNA in mouse ***,immunoreactivity for NF-κB p65 was predominantly detected in hepatocyte nucleifrom mice treated with BCG plus LPS,compared with the normal *** of IκB-α were strikingly decreased by LPS or BCG plus LPS treatment,compared with the normal group or BCG ***:TNF-α and IL-6mRNA were partially involved in early immunological liver injury induced by chal-lenge with small doses of LPS after BCG *** of TNF-α and IL-6 mRNA might be related to increases in LBP and CD 14 mRNA expression andactivation of NF-κ***,BCG priming in immunological liver injury mayoccur via upregulation of CD 14 mRNA expression in monon uclear cell infiltrationinto t

关键词： liver injury lipopolysaccharide binding protein CD14 tumor necrosis factor-alpha interleukin-6 NF-kappa B

Signaling mechanisms for regulation of chemotaxis

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Cell Research 2005年第1期15卷 52-56页

作者： Dianqing WU* Department of Genetics and Developmental biology, University of Connecticut Health center, Farmington, CT 06030, USA Department of Genetics and Developmental biology University of Connecticut Health Center Farmington CT 06030 USA. dwu@neuron.uchc.edu

Chemotaxis is a fascinating biological process, through which a cell migrates along a shallow chemoattractant gradient that is less than 5% difference between the anterior and posterior of the cell. Chemotaxis is composed of two independent, but interrelated processes-motility and directionality, both of which are regulated by extracellular stimuli, chemoattractants. In this mini-review, recent progresses in the understanding of the regulation of leukocyte chemotaxis by chemoattractant signaling are reviewed.

关键词： chemotaxis leukocyte signaling chemoattractant.