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In vitro cultivation of schistosoma japonicum germinal cells
S60
Abstracts / Cell Biology International 32 (2008) S1eS67
Apoptosis was detected in paraffin sections by the TUNEL technique. Expression of PCNA protein was examined by immunohistochemistry. As compared with group C, the indexes of apoptosis increased significantly in other groups (p<0.01). The indexes of apoptosis occurred in groups Hilo more than groups HE and T (p<0.01). Compared with group C, PCNA protein expressed slightly ascend in spite of groups HE, group T and groups Hilo. And compared with groups HE, it showed that there was prominent significance in groups Hilo (p<0.01). In conclusion, there were liver apoptosis in purely training, hypoxic exposure and Hilo. This research demonstrated that apoptosis occurred significantly in different hypoxic time. There were liver proliferation in hypoxic exposure, purely training and Hilo. It showed there was different appearance with different hypoxia time in hypoxic exposure and Hilo. Proliferation increased significantly after training intervention. Training avail liver cells to renewing. It is important to keep a dynamic equilibrium between liver apoptosis and proliferation.
IN VITRO CULTIVATION OF SCHISTOSOMA JAPONICUM GERMINAL CELLS Yan Liu 1,2, Yan Hui Gong 1, Qing Ren Zeng 1, Zu Ping Zhang 1, Sheng Hui Yang 1, Tie Bing Zeng 1,2, Li Ting Cai 1, Ben Wen Li 3, Shun Ke Zhang 1 1 Cell & Molecular Biological Experimental Centre, Xiangya School of Medicine, Central South University, Changsha, Hunan 410013, China 2 Dept. of Parasitology, School of Medicine, Nanhua University, Hengyang, Hunan 421001, China 3 Infectious Diseases Division, Washington University School of Medicine, Washington 63110, USA The possibility of cultivating schistosoma japoncum germinal cells in vitro by using modified human primordial germ cells (MHPGC) medium was evaluated in this study. Cells from 12-day-old juvenile worms and 36-day-old adult worms of S. j were selectively cultured by using MHPGC medium. Growth characteristics, general morphology, ultrastructures, alkaline phosphatase (AKP) staining, chromosomes karyotype and proliferations of cultured cells were observed, respectively. Biological identification of cultivation of cells from larva worms displayed that the cells grew in a semi-floating or accumulative way, and were round with slick-surface and clear-circumsciption after cultured for 8 hours. Ultrastructure showed that most of the cells revealed normal shape and structure with integrity cell envelope, clear caryotheca and chromatospherite during the period of 30 days. Chromosome karyotype analysis of the cells cultured for 60-day showed diploid, a typical character of blood-fluke. The growth features of the cells from adult worms were semifloating and accumulative, too. Cells grew faster and had great morphological differences in initial 4 weeks. BrdU incorporation assay was carried out in 4th week. The result revealed that the DNA synthesis in cells was athletic. Ultrastructure showed that most cells cultured for 5 weeks possessed the normal morphology with numerous vacuoles in different sizes in cytoplasm, caryon became bigger and caryotina became puffer, the characteristics of germinal cells. Chromosome karyotype of the cells cultured for 5 weeks showed the characters of the diploid and haploid. The cells cultured for 6 weeks were strong positive for AKP staining. It could be concluded from our research that some types of cells from S. j possessed some characteristics of germinal cells, and could be successfully propagated in vitro by using MHPGC medium.
PROTECTIVE ROLE OF A NEW VANADIUM COMPLEXES IN DIABETIC NEPHROPATHY Ying Liu 1, Yong Heng Xing 2, Wei Zou 1,2 1 College of Life Science , Liaoning Normal University, Dalian, China 2 The Key laboratory of Biotechnology and Molecular Pharmacy in Liaoning Province, Liaoning Normal University, Dalian, China Vanadium is an early first-row transition metal and forms colorful compounds in its many different oxidation states. The catalytic and material properties of vanadium compounds and their effects in biological systems have long provided the impetus and fuel to studies of vanadium science. We previously synthesized three kinds of vanadium complexes and tested the cytotoxicity by
MTT in vitro, which showed that VOpz’ (TP’)(SCN)3 was more feasible to cure the diabetes. Here we focused on fundamental biological studies in vivo to investigate the role of vanadium in the pathogenesis of diabetic nephropathy. Diabetic mice were induced by alloxan and in vanadium-treatment group diabetic mice were treated by intraperitoneal injection for three weeks with 5mg/kg of VOpz’(TP’)(SCN)3. The results indicated that the glucose level was decreased significantly in the vanadium-treatment mice, compared with diabetic mice, which protected from progressive glomerular damage by diabetes. Furthermore, the expression of caveolin-1, the marker protein of caveolae that related to insulin signaling, was decreased significantly in the kidney after the treatment. Also, the protein expression of in PI3k-Akt pathway was decreased in Vanadium treated group. These results suggest that VOpz ’ (TP’)(SCN)3 had a better effect of decreasing of glucose level and curative effect to pathology changes of glomerulus, indicating potential use of VOpz’ (TP’)(SCN) 3 in the treatment of diabetes. Cav-1 may play an important role in the process of insulin signal transduction.
THE RESEARCH OF SERUM KYNURENINE AND TRYPTOPHAN IN CHRONIC RENAL FAILURE PATIENTS Xi Bo Luo, Rui Wang, Ai Guo Tang Department of Clinical Laboratory, The Second Xiangya Hospital, Central South University, Changsha, 410011 China The aim of this study was to explore the clinical significance of serum kynurenine (KYN) and tryptophan (TRP) determined in chronic renal failure (CRF) patients. The concentrations of serum KYN, TRP, creatinine (Cr), urua and basical blood parameters in chronic glomerulonephritis (CG) patients and CRF patients were determined, and the clinical significance of serum KYN and TRP determined in different progresses of CRF was explored. Compared with healthy subjects, there were significantly increased concentrations of serum KYN and significantly decreased concentrations of TRP in the patients with CRF and CG. Compared with the patients with CG, the altered concentration of serum KYN and TRP in the patients with CRF were significant. It showed the levels of serum KYN and TRP may be correlated with the development of the renal function or blood parameters. It showed there was severe unbalance of kynurenine pathway in peripheral tissues in CRF patients, and it suggested that accumulation of some KYN metabolites in CRF patients blood may somewhat be responsible for symptoms.
EXPRESSION OF UTEROGLOBIN-BINDING PROTEIN IS UPREGULATED IN MURINE LUNGS AND IN NIH 3T3 FIBROBLAST FOLLOWING FIBROTIC STIMULI Chen Li, Jian Zhong Han, Lian Li, Hua She, Xiao Juan Shi, Wan Xiang Hu, Wei Liu, Dan Dan Feng, Zi Qiang Luo* Department of Physiology, Xiangya School of Medicine, Central South University, Changsha 410078, P.R. China As a small, secreted, multifunctional protein, uteroglobin (UG) plays important roles in preventing pulmonary fibrogenesis. Recent studies have suggested that UG-binding protein, a protein with nine transmembrane domains, is a putative receptor of UG, and mediates some cellular functions of UG on several cell types. In this study, we demonstrated the expression of mouse UG-binding protein (mUGBP) in bleomycin-induced pulmonary fibrosis in mice as well as its induction by profibrotic TGF-b1 in murine fibroblast NIH 3T3 cells. Realtime PCR showed that mUGBP mRNA in whole lung homogenates began to increase gradually from day 14 to day 28 following bleomycin challenge. In situ hybridization revealed that increased mUGBP positive labeling was preferentially observed in interstitial sites of bleomycin-injured lungs. In vitro studies further showed that treatment of mouse NIH 3T3 cells with TGF-b1 for 48 hours led to increased expressions of mUGBP mRNA and protein. TGF-b1 stimulated procollagen 1 (I) (pro-colla1(I)) expression in NIH 3T3 cells was inhibited by 10mM of antiflammin-1 (AF-1, MQMKKVLDS), a known UG-derived bioactive fragment. However, this UG-bioactive fragment did not change the expression levels of pro-colla1 (I) under basal condition without TGF-b1. Antisense oligonucleotides (AS-OND) of mUGBP partly blocked the inhibition of TGF-b1-stimulated pro-colla1(I) protein expression by AF-1. These results pointed to the likelihood that the induction of mUGBP was necessary for AF-1 to inhibit of procollagen I expression,
Abstracts / Cell Biology International 32 (2008) S1eS67
Apoptosis was detected in paraffin sections by the TUNEL technique. Expression of PCNA protein was examined by immunohistochemistry. As compared with group C, the indexes of apoptosis increased significantly in other groups (p<0.01). The indexes of apoptosis occurred in groups Hilo more than groups HE and T (p<0.01). Compared with group C, PCNA protein expressed slightly ascend in spite of groups HE, group T and groups Hilo. And compared with groups HE, it showed that there was prominent significance in groups Hilo (p<0.01). In conclusion, there were liver apoptosis in purely training, hypoxic exposure and Hilo. This research demonstrated that apoptosis occurred significantly in different hypoxic time. There were liver proliferation in hypoxic exposure, purely training and Hilo. It showed there was different appearance with different hypoxia time in hypoxic exposure and Hilo. Proliferation increased significantly after training intervention. Training avail liver cells to renewing. It is important to keep a dynamic equilibrium between liver apoptosis and proliferation.
IN VITRO CULTIVATION OF SCHISTOSOMA JAPONICUM GERMINAL CELLS Yan Liu 1,2, Yan Hui Gong 1, Qing Ren Zeng 1, Zu Ping Zhang 1, Sheng Hui Yang 1, Tie Bing Zeng 1,2, Li Ting Cai 1, Ben Wen Li 3, Shun Ke Zhang 1 1 Cell & Molecular Biological Experimental Centre, Xiangya School of Medicine, Central South University, Changsha, Hunan 410013, China 2 Dept. of Parasitology, School of Medicine, Nanhua University, Hengyang, Hunan 421001, China 3 Infectious Diseases Division, Washington University School of Medicine, Washington 63110, USA The possibility of cultivating schistosoma japoncum germinal cells in vitro by using modified human primordial germ cells (MHPGC) medium was evaluated in this study. Cells from 12-day-old juvenile worms and 36-day-old adult worms of S. j were selectively cultured by using MHPGC medium. Growth characteristics, general morphology, ultrastructures, alkaline phosphatase (AKP) staining, chromosomes karyotype and proliferations of cultured cells were observed, respectively. Biological identification of cultivation of cells from larva worms displayed that the cells grew in a semi-floating or accumulative way, and were round with slick-surface and clear-circumsciption after cultured for 8 hours. Ultrastructure showed that most of the cells revealed normal shape and structure with integrity cell envelope, clear caryotheca and chromatospherite during the period of 30 days. Chromosome karyotype analysis of the cells cultured for 60-day showed diploid, a typical character of blood-fluke. The growth features of the cells from adult worms were semifloating and accumulative, too. Cells grew faster and had great morphological differences in initial 4 weeks. BrdU incorporation assay was carried out in 4th week. The result revealed that the DNA synthesis in cells was athletic. Ultrastructure showed that most cells cultured for 5 weeks possessed the normal morphology with numerous vacuoles in different sizes in cytoplasm, caryon became bigger and caryotina became puffer, the characteristics of germinal cells. Chromosome karyotype of the cells cultured for 5 weeks showed the characters of the diploid and haploid. The cells cultured for 6 weeks were strong positive for AKP staining. It could be concluded from our research that some types of cells from S. j possessed some characteristics of germinal cells, and could be successfully propagated in vitro by using MHPGC medium.
PROTECTIVE ROLE OF A NEW VANADIUM COMPLEXES IN DIABETIC NEPHROPATHY Ying Liu 1, Yong Heng Xing 2, Wei Zou 1,2 1 College of Life Science , Liaoning Normal University, Dalian, China 2 The Key laboratory of Biotechnology and Molecular Pharmacy in Liaoning Province, Liaoning Normal University, Dalian, China Vanadium is an early first-row transition metal and forms colorful compounds in its many different oxidation states. The catalytic and material properties of vanadium compounds and their effects in biological systems have long provided the impetus and fuel to studies of vanadium science. We previously synthesized three kinds of vanadium complexes and tested the cytotoxicity by
MTT in vitro, which showed that VOpz’ (TP’)(SCN)3 was more feasible to cure the diabetes. Here we focused on fundamental biological studies in vivo to investigate the role of vanadium in the pathogenesis of diabetic nephropathy. Diabetic mice were induced by alloxan and in vanadium-treatment group diabetic mice were treated by intraperitoneal injection for three weeks with 5mg/kg of VOpz’(TP’)(SCN)3. The results indicated that the glucose level was decreased significantly in the vanadium-treatment mice, compared with diabetic mice, which protected from progressive glomerular damage by diabetes. Furthermore, the expression of caveolin-1, the marker protein of caveolae that related to insulin signaling, was decreased significantly in the kidney after the treatment. Also, the protein expression of in PI3k-Akt pathway was decreased in Vanadium treated group. These results suggest that VOpz ’ (TP’)(SCN)3 had a better effect of decreasing of glucose level and curative effect to pathology changes of glomerulus, indicating potential use of VOpz’ (TP’)(SCN) 3 in the treatment of diabetes. Cav-1 may play an important role in the process of insulin signal transduction.
THE RESEARCH OF SERUM KYNURENINE AND TRYPTOPHAN IN CHRONIC RENAL FAILURE PATIENTS Xi Bo Luo, Rui Wang, Ai Guo Tang Department of Clinical Laboratory, The Second Xiangya Hospital, Central South University, Changsha, 410011 China The aim of this study was to explore the clinical significance of serum kynurenine (KYN) and tryptophan (TRP) determined in chronic renal failure (CRF) patients. The concentrations of serum KYN, TRP, creatinine (Cr), urua and basical blood parameters in chronic glomerulonephritis (CG) patients and CRF patients were determined, and the clinical significance of serum KYN and TRP determined in different progresses of CRF was explored. Compared with healthy subjects, there were significantly increased concentrations of serum KYN and significantly decreased concentrations of TRP in the patients with CRF and CG. Compared with the patients with CG, the altered concentration of serum KYN and TRP in the patients with CRF were significant. It showed the levels of serum KYN and TRP may be correlated with the development of the renal function or blood parameters. It showed there was severe unbalance of kynurenine pathway in peripheral tissues in CRF patients, and it suggested that accumulation of some KYN metabolites in CRF patients blood may somewhat be responsible for symptoms.
EXPRESSION OF UTEROGLOBIN-BINDING PROTEIN IS UPREGULATED IN MURINE LUNGS AND IN NIH 3T3 FIBROBLAST FOLLOWING FIBROTIC STIMULI Chen Li, Jian Zhong Han, Lian Li, Hua She, Xiao Juan Shi, Wan Xiang Hu, Wei Liu, Dan Dan Feng, Zi Qiang Luo* Department of Physiology, Xiangya School of Medicine, Central South University, Changsha 410078, P.R. China As a small, secreted, multifunctional protein, uteroglobin (UG) plays important roles in preventing pulmonary fibrogenesis. Recent studies have suggested that UG-binding protein, a protein with nine transmembrane domains, is a putative receptor of UG, and mediates some cellular functions of UG on several cell types. In this study, we demonstrated the expression of mouse UG-binding protein (mUGBP) in bleomycin-induced pulmonary fibrosis in mice as well as its induction by profibrotic TGF-b1 in murine fibroblast NIH 3T3 cells. Realtime PCR showed that mUGBP mRNA in whole lung homogenates began to increase gradually from day 14 to day 28 following bleomycin challenge. In situ hybridization revealed that increased mUGBP positive labeling was preferentially observed in interstitial sites of bleomycin-injured lungs. In vitro studies further showed that treatment of mouse NIH 3T3 cells with TGF-b1 for 48 hours led to increased expressions of mUGBP mRNA and protein. TGF-b1 stimulated procollagen 1 (I) (pro-colla1(I)) expression in NIH 3T3 cells was inhibited by 10mM of antiflammin-1 (AF-1, MQMKKVLDS), a known UG-derived bioactive fragment. However, this UG-bioactive fragment did not change the expression levels of pro-colla1 (I) under basal condition without TGF-b1. Antisense oligonucleotides (AS-OND) of mUGBP partly blocked the inhibition of TGF-b1-stimulated pro-colla1(I) protein expression by AF-1. These results pointed to the likelihood that the induction of mUGBP was necessary for AF-1 to inhibit of procollagen I expression,