RESUMEN
N(2)-[alpha-O-benzyl-N-(acetylmuramyl)-L-alanyl-D-isoglutaminyl]-N(6)-trans-(m-nitrocinnamoyl)-L-lysine (muramyl dipeptide C, or MDP-C) has been synthesized as a novel, nonspecific immunomodulator. The present study shows that MDP-C induces strong cytolytic activity by macrophages on P388 leukemia cells and cytotoxic activity by cytotoxic T lymphocytes (CTLs) on P815 mastocytoma cells. Our results also indicate that MDP-C is an effective stimulator for production of interleukin-2 and interleukin-12 by murine bone marrow derived dendritic cells (BMDCs) and production of interferon-gamma by CTLs. Additionally, MDP-C increases the expression levels of several surface molecules, including CD11c, MHC class I, and intercellular adhesion molecule-1 in BMDCs. Moreover, MDP-C remarkably enhances the immune system's responsiveness to hepatitis B surface antigen (HBsAg) in hepatitis B virus transgenic mice for both antibody production and specific HBsAg T-cell responses ex vivo. Our results indicate that MDP-C is an apyrogenic, nonallergenic, and low-toxicity immunostimulator with great potential for diagnostic, immunotherapeutic, and prophylactic applications in diseases such as hepatitis B and cancers.
Asunto(s)
Acetilmuramil-Alanil-Isoglutamina/análogos & derivados , Adyuvantes Inmunológicos/síntesis química , Antígenos de Superficie de la Hepatitis B/inmunología , Acetilmuramil-Alanil-Isoglutamina/efectos adversos , Acetilmuramil-Alanil-Isoglutamina/síntesis química , Acetilmuramil-Alanil-Isoglutamina/farmacología , Adyuvantes Inmunológicos/efectos adversos , Adyuvantes Inmunológicos/farmacología , Animales , Formación de Anticuerpos , Células de la Médula Ósea/efectos de los fármacos , Células de la Médula Ósea/metabolismo , Antígeno CD11c/biosíntesis , Línea Celular Tumoral , Citotoxicidad Inmunológica , Células Dendríticas/efectos de los fármacos , Células Dendríticas/metabolismo , Hepatitis B/inmunología , Vacunas contra Hepatitis B/inmunología , Antígenos de Histocompatibilidad Clase I/biosíntesis , Técnicas In Vitro , Molécula 1 de Adhesión Intercelular/biosíntesis , Interleucina-12/biosíntesis , Interleucina-2/biosíntesis , Macrófagos/efectos de los fármacos , Macrófagos/inmunología , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Conejos , Ratas , Ratas Wistar , Linfocitos T Citotóxicos/efectos de los fármacos , Linfocitos T Citotóxicos/inmunología , Pruebas de Toxicidad AgudaRESUMEN
AIM: To examine the inhibitory effect of resveratrol on matrix metalloroteinase-9 (MMP-9) and explore its mechanism. METHODS: MMP-9 activity was analyzed by gelatin zymography; MMP-9 protein was detected by Western blot; MMP-9 mRNA expression was investigated by RT-PCR. Activation of activator protein -1 (AP-1) was measured by electrophoretic mobility shift assay (EMSA). RESULTS: MMP-9 activity in U937 cells increased significantly after exposed to PMA at 10 nmol/L for 24 h without FCS (P<0.01). Resveratrol at 1 and 10 micromol/L showed significant inhibition on MMP-9 activity (P<0.05 and P<0.01, respectively). Western blot and RT-PCR experiments displayed that MMP-9 protein (P<0.01) and mRNA expression (P<0.01) increased significantly in PMA-treated U937 cells. Resveratrol at 1 and 10 micromol/L showed inhibitory effects on MMP-9 protein production and MMP-9 mRNA expression (P<0.05). The activation of AP-1 induced by PMA was also extensively inhibited by resveratrol at 0.1, 1, and 10 micromol/L. CONCLUSION: The inhibitory effect of resveratrol on MMP-9 activity may be partly through suppression of activation of nuclear transcription factor AP-1, and inhibition of MMP-9 mRNA expression and MMP-9 protein production.
Asunto(s)
Antiinflamatorios no Esteroideos/farmacología , Metaloproteinasa 9 de la Matriz/metabolismo , Estilbenos/farmacología , Factor de Transcripción AP-1/metabolismo , Dexametasona/farmacología , Humanos , Metaloproteinasa 9 de la Matriz/genética , ARN Mensajero/genética , Resveratrol , Transcripción Genética , Células U937RESUMEN
AIM: To study the effects of lipopolysaccharide (LPS), the supernatant of U937 cells stimulated with LPS and dexamethasone on interleukin-6 (IL-6) expression in the synoviocyte from patients with rheumatoid arthritis (RA). METHODS: Fibroblast-like synoviocytes (FLS) from the joint tissue of patients with rheumatoid arthritis were cultured and incubated for 24 h with LPS (1 mg.L-1) or the supernatant of U937 cells stimulated with LPS (1 mg.L-1) for 24 h. Dexamethasone was added to the supernatant of U937 cells and FLS was incubated for 24 h. The expression of IL-6 protein was detected by radioimmunoassay. The mRNA expression of IL-6 was accessed by RT-PCR. RESULTS: The growth of FLS was not markedly affected by LPS, and the protein secretion and mRNA expression of IL-6 were not markedly changed in FLS treated with LPS. The IL-6 secretion and IL-6 mRNA expression were significantly increased in FLS cultured with the supernatant from U937 cell treated with LPS. Dexamethasone markedly inhibited the protein secretion and mRNA expression of IL-6 in FLS cultured with the supernatant from U937 cell stimulated with LPS. The inhibitory effects were increased as the concentration of dexamethasone increased. CONCLUSION: LPS was not shown to directly affect the expression of IL-6 in FLS, but it indirectly causes the increase of the IL-6 expression in FLS by stimulating U937 cell. Dexamethasone can inhibit this increase of the IL-6 expression.
Asunto(s)
Artritis Reumatoide/patología , Interleucina-6/biosíntesis , Lipopolisacáridos/farmacología , Membrana Sinovial/efectos de los fármacos , División Celular/efectos de los fármacos , Células Cultivadas , Dexametasona/farmacología , Fibroblastos/efectos de los fármacos , Fibroblastos/metabolismo , Expresión Génica/efectos de los fármacos , Humanos , Interleucina-6/genética , ARN Mensajero/genética , Membrana Sinovial/metabolismo , Células U937RESUMEN
AIM: To study the effects of lipopolysaccharide (LPS), the supernatant of U937 cells stimulated with LPS and dexamethasone on matrix metalloproteinase-9 (MMP-9) expression in the synoviocyte from patients with rheumatoid arthritis(RA). METHODS: Fibroblast-like cells (FLS) from the joint tissue of patients with rheumatoid arthritis were cultured and incubated for 24 h with LPS (1 mg.L-1) or the supernatant of U937 cells stimulated with LPS (1 mg.L-1) for 24 h. Dexamethasone was added to the supernatant of U937 cells and FLS was incubated for 24 h. The activity of MMP-9 was analyzed by gelatin zymography. Protein expression of MMP-9 was detected by Western blot using special polyclonal antibodies. The mRNA expression of MMP-9 was detected by RT-PCR. RESULTS: The expression of MMP-9 was not markedly changed in FLS treated with LPS. The MMP-9 activity, MMP-9 secretion and MMP-9 mRNA expression were significantly increased in FLS cultured with the supernatant from U937 cell treated with LPS. Dexamethasone markedly inhibited the activity, protein secretion and mRNA expression of MMP-9 in FLS cultured with the supernatant from U937 cell stimulated with LPS, and the inhibitory effects were increased as the concentration of dexamethasone increased. CONCLUSION: LPS did not directly affect the expression of MMP-9 in FLS, but it was found to indirectly cause the increase of MMP-9 expression in FLS by stimulating U937 cell. Dexamethasone was found to inhibit this increase of MMP-9 expression.
Asunto(s)
Artritis Reumatoide/patología , Expresión Génica/efectos de los fármacos , Lipopolisacáridos/farmacología , Metaloproteinasa 9 de la Matriz/biosíntesis , Membrana Sinovial/efectos de los fármacos , Antiinflamatorios/farmacología , División Celular/efectos de los fármacos , Células Cultivadas , Dexametasona/farmacología , Fibroblastos/patología , Humanos , Metaloproteinasa 9 de la Matriz/genética , Metaloproteinasa 9 de la Matriz/metabolismo , ARN Mensajero/efectos de los fármacos , ARN Mensajero/genética , Membrana Sinovial/enzimología , Membrana Sinovial/patología , Células U937RESUMEN
AIM: To study the effects of indomethacin on interleukin-6 (IL-6) expression stimulated with lipopolysaccharide (LPS) in rheumatoid arthritic patients' synoviocyte. METHODS: Fibroblast-like cells (FLS) from rheumatoid arthritic patients' joint tissue were cultured for 24 h and incubated 24 h with LPS (1 mg.L-1) or the supernatant of U937 cells stimulated by LPS (1 mg.L-1). After indomethacin or dexamethasone added into the supernatant of U937 cells, FLS was incubated with the super natant for 24 h. The expression of IL-6 protein was detected by radioimmunoassay. The mRNA expression of IL-6 was accessed by RT-PCR. RESULTS: LPS did not obviously affect the growth of FLS, and the protein secretion and mRNA expression of IL-6 were not changed in FLS treated with LPS. The IL-6 secretion and IL-6 mRNA expression were significantly increased in FLS cultured with the supernatant from U937 cell treated with LPS. Indomethacin at concentrations of 1 x 10(-7)-1 x 10(-5) mol.L-1 obviously inhibited the protein secretion and mRNA expression of IL-6 in FLS cultured with the supernatant from U937 cell stimulated with LPS, and the inhibitory effects increased as the concentrations of indomethacin increased. CONCLUSION: Indomethacin can inhibit the increase of IL-6 expression caused by supernatant of U937 cells stimulated with LPS in FLS.
Asunto(s)
Antiinflamatorios no Esteroideos/farmacología , Artritis Reumatoide/patología , Indometacina/farmacología , Interleucina-6/biosíntesis , Membrana Sinovial/metabolismo , Artritis Reumatoide/metabolismo , Células Cultivadas , Fibroblastos/metabolismo , Fibroblastos/patología , Humanos , Interleucina-6/genética , Lipopolisacáridos/farmacología , ARN Mensajero/genética , Membrana Sinovial/patología , Células U937RESUMEN
AIM: To investigate the effects of growth inhibition of human gastric cancer SGC-7901 cell with RRR-alpha-tocopheryl succinate (VES), a derivative of natural Vitamin E, via inducing apoptosis and DNA synthesis arrest. METHODS: Human gastric cancer SGC-7901 cells were regularly incubated in the presence of VES at 5, 10 and 20mg x L(-1) (VES was dissolved in absolute ethanol and diluted in RPMI 1640 complete condition media correspondingly to a final concentration of VES and 1 mL x L(-1) ethanol), succinic acid and ethanol equivalents as vehicle (VEH) control and condition media only as untreated (UT) control. Trypan blue dye exclusion analysis and MTT assay were applied to detect the cell proliferation. Cells were pulsed with 37kBq of tritiated thymidine and (3H) TdR uptake was measured to observe DNA synthesis. Apoptotic morphology was observed by electron microscopy and DAPI staining. Flow cytometry and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) assay were performed to detect VES-triggered apoptosis. RESULTS: VES inhibited SGC-7901 cell growth in a dose-dependent manner. The growth curve showed suppression by 24.7%, 49.2% and 68.7% following 24h of VES treatment at 5, 10 and 20 mg x L(-1), respectively, similar to the findings from MTT assay. DNA synthesis was evidently reduced by 35%, 45% and 98% after 24h VES treatment at 20mg x L(-1) and 48 h at 10 and 20mg x L(-1), respectively. VES induced SGC-7901 cells to undergo apoptosis with typically apoptotic characteristics, including morphological changes of chromatin condensation, chromatin crescent formation/margination, nucleus fragmentation and apoptotic body formation, typical apoptotic sub-G1 peak by flow cytometry and increase of apoptotic cells by TUNEL assay in which 90% of cells underwent apoptosis after 48 h of VES treatment at 20 mg x L(-1). CONCLUSION: VES can inhibit human gastric cancer SGC-7901 cell growth by inducing apoptosis and DNA synthesis arrest. Inhibition of SGC-7901 cell growth by VES is dose- and time-dependent. Therefore VES can function as a potent chemotherapeutic agent against human gastric carcinogenesis.