ABSTRACT
The purpose of this study is to further explore the effects of SI-4650, a newly discovered small molecule inhibitor of spermine oxidase (SMO) in our laboratory, on proliferation and migration of human osteosarcoma 143B cells and its underlying molecular mechanism. Chemiluminescence and high performance liquid chromatograph were used to analyze the effect of SI-4650 on SMO activity in 143B cells. DCFH-DA-staining/FCM was used to analyze the accumulation of cellular reactive oxygen species (ROS), whereas MTT and FCM were used to detect proliferation and cell cycle. Transwell culture and Western blot were used to analyze the expression levels of migration-related proteins. PI/FITC-Annexin V/FCM, fluorescence microscopy and Western blot were used to analyze apoptosis and autophagy. Our results showed that SI-4650 could significantly decrease SMO activity, inhibit cell proliferation or migration, and induce a S-phase cell cycle arrest in 143B human osteosarcoma cells. The mechanism may be related to interfering with polyamine metabolism, activating mitochondrial-mediated apoptosis and causing autophagic death. These results suggest that SI-4650 has the potential for clinical use in treatment of osteosarcoma.
ABSTRACT
Aim To investigate the effects of a novel spermine oxidase (SMO) inhibitor SI-4650 on proliferation, apoptosis and autophagy of human glioma U87MG cells as well as its possible mechanism. Methods MIT assay was used to detect cell proliferation. The chemiluminescence assay was used to determine enzyme activities of SMO and N'-acetylpolyamine oxidase (APAO). High performance liquid chromatography (HPI.C) was performed to detect cellular polyamine content. Transwell assay was used to evaluate the migration ability of U87MG cells. PI single-staining/flow cytometry( FCM ) were used to analyze cell cycle. PI/ FITC-Annexin V double-staining/ FCM and Western blot were used to analyze apoptosis. Confocal laser scanning microscopy ( LSCM) and Western blot were used to analyze autophagy. Results SI-4650 could significantly inhibit the activities of SMO and APAO, interfere with polyamine metabolism and reduce the content of total polyamine in U87MG cells. Treatment of U87MG cells with SI-4650 inhibited the proliferation and migration, caused G0/G, cell cycle arrest and induced apoptotic and autophagic cell death. Conclusions SI-4650 has the pharmacological activity of killing glioma U87MG cells, and its mechanism may be related to the interference of polyamine metabolism and induction of cell apoptosis and autophagy.
ABSTRACT
Aim To discuss the changes of polyamine metabolism and autophagy in aging rat heart and the effect of exogenous spermidine on autophay in aged rat heart. Methods Western blot assay was used to detect the expressions of ODC and SSAT-rate-limiting enzyme of polyamine anabolism and catabolism as well as the expression of autophagy-relevant protein LC3?/? and p62 in cardiac tissues from male Wistar rats aged 3, 6, 12 and 24 months, and the expressions of p16, LC3?/?, ATG5 and ATG7 protein were detected in cardiac tissues in rats aged 3 months, 24 months and 24 months with spermidine administrtion, respectively. The myocardial autophagosome formation was observed by transmission electron microscope. In addition, cell cross-sectional area, cell apoptosis rate and generation of ROS were evaluated. Results With heart aging in rats, ODC expression and LC3?/? ratio were degraded, and SSAT and p62 protein expression upgraded. In rats aged 24 months, myocardium showed increased p16 expression, cell cross-sectional area, cell apoptosis and ROS generation, while cell autophagosome formation decreased, p62 expression increased, the expression of LC3?/?, ATG5 and ATG7 all declined. Spermidine intervention obviously inhibited myocardial changes induced by aging, showing decrease in cell cross-sectional area, apoptosis, ROS generation and p62 expression, and increase in LC3?/?, ATG5 and ATG7 expression. Conclusions With heart aging in rats, polyamine anabolism is degraded, catabolism is upgraded, and cell autophagy declined. Exogenous spermidine might delay aging through inducing autophagy of cardiomyocytes.
ABSTRACT
Aim To study the effects of polyamine analogue CPENSpm on the human lung cancer line A549 in cell proliferation and apoptosis.Methods MTS was used to assay the cell proliferation,chemical analysis methods were used to determine the activities of enzymes in the polyamine metabolism,HPLC was performed to assay the intracellular concentration of polyamines,Sub-G1 and DNA fragmentation assays were used to determine the cell apoptosis.Results Treating A549 lung cancer cells by CPENSpm resulted in:①cell-growth inhibition and cell apoptosis;②inhibition of ODC(key enzyme in polyamine synthetic pathway)and activation of SSAT and SMO(key enzymes in polyamine catabolism);③great decrease of intracellular polyamine concentrations.MDL72527,the SMO inhibitor,can antagonize the effect of CPENSpm on inhibiting the proliferation of A549 cells.Conclusion CPENSpm inhibits proliferation and induces apoptosis of human A549 lung cancer cell line by interfering the polyamine metabolism,depleting intracellular polyamine contents that are need by quick-growth of cancer cells and inducing production of H2O2.