ABSTRACT
Previously, we have reported that high mobility group box 1 (HMGB1), a proinflammatory mediator in sepsis, is released via the IFN-beta-mediated JAK/STAT pathway. However, detailed mechanisms are still unclear. In this study, we dissected upstream signaling pathways of HMGB1 release using various molecular biology methods. Here, we found that calcium/calmodulin-dependent protein kinase (CaM kinase, CaMK) is involved in HMGB1 release by regulating IFN-beta production. CaMK inhibitor, STO609, treatment inhibits LPS-induced IFN-beta production, which is correlated with the phosphorylation of interferon regulatory factor 3 (IRF3). Additionally, we show that CaMK-I plays a major role in IFN-beta production although other CaMK members also seem to contribute to this event. Furthermore, the CaMK inhibitor treatment reduced IFN-beta production in a murine endotoxemia. Our results suggest CaMKs contribute to HMGB1 release by enhancing IFN-beta production in sepsis.
Subject(s)
Benzimidazoles , Cytokines , Endotoxemia , HMGB1 Protein , Inflammation , Interferon Regulatory Factor-3 , Interferon-beta , Molecular Biology , Naphthalimides , Phosphorylation , Phosphotransferases , Protein Kinases , Sepsis , Signal TransductionABSTRACT
The objective of this study is to examine the effects of ANISpm, a novel polyamine naphthalimide conjugate, with acetylsalicylic acid against hepatocellular carcinoma in vivo and in vitro and elucidate its potential molecular mechanism. The proliferation inhibition was detected by MTT assay. Cell apoptosis, intracellular fluorescence intensity and mitochondrial membrane potential (MMP) were detected by high content screening (HCS) analysis. Polyamines content was analyzed by reverse-phase high performance liquid chromatography Protein expression levels were quantified by Western blotting assay. The combination treatment strongly inhibited cell proliferation, induced cell apoptosis in HepG2 cells and H22 hepatoma cells, which was mediated by enhanced ANISpm uptake via up-regulation of spermidine/spermine N1-acetyltransferase (SSAT) and depression of intracellular polyamine. Furthermore, this synergistic apoptosis was involved in mitochondria and death-receptor signal pathway. All these findings demonstrated that the combination treatment with acetylsalicylic acid and ANISpm resulted in synergistic antitumor effects on hepatoma cells. Thus, combination therapy with these agents may be useful as a potential template for the development of better chemotherapeutic strategy against hepatoma.
Subject(s)
Animals , Female , Humans , Mice , Acetyltransferases , Metabolism , Antineoplastic Agents , Pharmacology , Apoptosis , Aspirin , Pharmacology , Caspase 8 , Metabolism , Caspase 9 , Metabolism , Cell Line, Tumor , Cell Proliferation , Drug Synergism , Hep G2 Cells , Liver Neoplasms, Experimental , Pathology , Membrane Potential, Mitochondrial , Naphthalimides , Metabolism , Pharmacology , Neoplasm Transplantation , Polyamines , Metabolism , Pharmacology , Random Allocation , Spermine , Metabolism , Pharmacology , Tumor Burden , Up-RegulationABSTRACT
Six naphthalimide polyamine conjugates were synthesized and their structures were confirmed by elemental analysis, 1H NMR, 13C NMR and MS. Antitumor activities were evaluated in vitro using MTT assay on Leukemia cells (K562), human breast cancer cells (MB-231) and prostate cancer cells (Ln cap cell). The results showed that most of the six compounds were superior to the control (amonafide), 6d, 6e, and 6f exhibited nice selectivity in a screen of hepatoma cells (BEL-7402) and human normal hepatocytes (QSG-7701).
Subject(s)
Humans , Male , Antineoplastic Agents , Pharmacology , Cell Line, Tumor , Molecular Structure , Naphthalimides , Pharmacology , Polyamines , PharmacologyABSTRACT
<p><b>OBJECTIVE</b>To evaluate the effects of Bisnaphthalimide (C8) on the proliferation and apoptosis of SMMC-7721 cells.</p><p><b>METHODS</b>The effects of C8 on the proliferation of SMMC-7721 cells were evaluated by MTT. Cell cycle and apoptotic cell percentage were studied by flow cytometry. The protein of Bcl-2 was detected by Western blot. The intra-cellular protein of Bcl-2 was detected by flow cytometry. The proteins of caspase9 and caspase3 were detected by ELISA.</p><p><b>RESULTS</b>C8 inhibited the growth of SMMC-7721 cells. The IC50 of C8 on SMMC-7721 cells was 15 micromol/L. C8 initiated apoptosis of SMMC-7721 cells. After SMMC-7721 cells were exposed to C8 in concentrations of 10, 15, 20 micromol/L, the apoptosis rates were 16.8%, 29.4% and 35.8%, respectively, significantly higher than those of the controls (P less than 0.01). Flow cytometry and Western blot analysis showed that Bcl-2 protein level was inhibited after treatment with C8. The ELISA analysis showed that caspase9 and caspase3 were activated in the SMMC-7721 cells after the C8 treatment.</p><p><b>CONCLUSION</b>C8 could induce apoptosis of human liver cancer SMMC-7721 cells. C8 might be a potential efficient anticancer drug.</p>