OBJECTIVETo investigate the differential expression of isocitrate lyase in Penicillium marneffei phagocytized by nonstimulated and stimulated murine macrophages, and explore the role of glyoxylate pathway in pathogenesis of Penicilliosis marneffei.

METHODSPenicillium marneffei conidia and Raw264.7 cells were incubated in 16 cultures, which were divided to 4 groups for treatment with N-monomethyl-L-arginine (LNMMA, CI group), murine interferon-gamma (IFN-gamma) plus lipopolysaccharide (LPS) (T group), IFN-gamma plus LPS and LNMMA (TI group), or the same volume of culture medium (C group). The transcriptional levels of isocitrate lyase were detected using real-time RT-PCR, and its expression levels detected biochemically.

RESULTSThe transcriptional levels of isocitrate lyase in C, CI, T, TI groups were 1.00, 1.42, 33.09, and 74.88 (P<0.05), while the expression levels were 0.06, 0.07, 0.18, and 0.93, respectively (P<0.05). The content of nitric oxide in T group was significantly higher than that in the other groups (P<0.01), but the CFU of T group was the lowest (P<0.01).

CONCLUSIONReactive nitrogen intermediates induced by stimulated murine macrophages restrain the expression of isocitrate lyase of Penicillium marneffei and development of Penicillium marneffei, in which process the glyoxylate pathway may play an important role.

Animals ; Cell Line ; Fungal Proteins ; genetics ; Gene Expression Profiling ; Gene Expression Regulation, Enzymologic ; drug effects ; Gene Expression Regulation, Fungal ; drug effects ; Host-Pathogen Interactions ; Interferon-gamma ; pharmacology ; Isocitrate Lyase ; genetics ; Lipopolysaccharides ; pharmacology ; Macrophages ; drug effects ; immunology ; microbiology ; Mice ; Nitric Oxide ; immunology ; Penicillium ; genetics ; immunology ; physiology ; Phagocytosis ; immunology ; Reverse Transcriptase Polymerase Chain Reaction ; omega-N-Methylarginine ; pharmacology

Excessive production of nitric oxide (NO) and proinflammatory cytokines from activated microglia play an important role in human neurodegenerative disorders. Here, we investigated whether celastrol, which has been used as a potent anti-inflammatory and anti-oxidative agent in Chinese medicine, attenuates excessive production of NO and proinflammatory cytokines such as TNF-alpha and IL-1beta in LPS-stimulated BV-2 cells, a mouse microglial cell line. We report here that the LPS-elicited excessive production of NO, TNF-alpha, and IL-1beta in BV-2 cells was largely inhibited in the presence of celastrol, and the attenuation of inducible iNOS and these cytokines resulted from the reduced expression of mRNAs of iNOS and these cytokines, respectively. The molecular mechanisms that underlie celastrol-mediated attenuation were the inhibition of LPS-induced phosphorylation of MAPK/ERK1/2 and the DNA binding activity of NF-kappaB in BV-2 cells. The results indicate that celastrol effectively attenuated NO and proinflammatory cytokine production via the inhibition of ERK1/2 phosphorylation and NF-kappaB activation in LPS-activated microglia. Thus, celastrol may be an effective therapeutic candidate for use in the treatment of neurodegenerative human brain disorders.
Animals ; Cell Line ; Cytokines/*biosynthesis/drug effects ; Gene Expression Regulation, Enzymologic/drug effects/immunology ; Inflammation/immunology ; Inflammation Mediators/immunology ; Mice ; Microglia/*drug effects/immunology ; Mitogen-Activated Protein Kinases/*physiology ; NF-kappa B/metabolism/*physiology ; Nitric Oxide/*metabolism ; Nitric Oxide Synthase Type II/biosynthesis/drug effects ; RNA, Messenger/analysis ; Signal Transduction/*drug effects/physiology ; Transcription, Genetic/drug effects/immunology ; Triterpenes/*pharmacology

Triptolide (TPT), an active compound extracted from Chinese herb Tripterygium wilfordii , has been used in therapy of rheumatoid arthritis (RA). In this study, after synovial fibroblasts from rheumatoid arthritis (RASFs) were treated with TPT, we investigated its effect on the differentiation of Th17 cells. Firstly, the mRNA level of cyclooxygenase (COX) wad detected by qRT-PCR and the protein level of prostaglandin E2 (PGE2) was tested by ELISA in RASFs treated with different concentrations (0, 10, 50, 100 nmol L-1 ) of TPT. Then after TPT pre-treated RASFs and RA CD4 + T cells wer e co-cultured for 3 days in the presence or absence of PGE2, IL-17 and IFN-gamma production in CD4 T cell subsets were detected by flow cytometry. The results showed TPT decreased the mRNA experssion of COX2 and the secretion of PGE2 in RASFs in a dose-dependent manner(P <0. 05). We further found that differentiation of Thl7 cells was downregulated in a dose-dependent manner, and exogenous PGE2 could reverse the inhibition of Th17 cell differentiation(P <0. 05). Taken together, our results demonstrated that TPT inhibited the mRNA level of COX2 and the secretion of PGE2 in RASFs, which partly led to impaired Th17 cell differentiation in vitro.
Arthritis, Rheumatoid ; drug therapy ; enzymology ; immunology ; Cell Differentiation ; drug effects ; Cell Line ; Cyclooxygenase 2 ; genetics ; metabolism ; Dinoprostone ; metabolism ; Diterpenes ; pharmacology ; Epoxy Compounds ; pharmacology ; Fibroblasts ; drug effects ; immunology ; Gene Expression Regulation, Enzymologic ; drug effects ; Humans ; Middle Aged ; Phenanthrenes ; pharmacology ; Synovial Fluid ; drug effects ; Th17 Cells ; drug effects ; pathology

OBJECTIVETo explore the action mechanism of the immune active components of sappon wood (SWE) for antagonizing reject reaction by observing the influence of its ethyl acetate extract on mRNA expression of myocardial GrB in rat model of allogenic ectopic cardiac transplantation.

METHODSAnimal model of abdominal cardiac ectopic transplantation was established taking Wistar rat as the donor and SD rat as the receptor. The successfully modeled rats were randomly divided into the model group, the SWE group and the CsA group. GrB mRNA expression was detected by RT-PCR method and myocardial pathomorphologic picture was observed in routine.

RESULTSThe pathologic changes in the SWE group (23 scores) and the CsA group (14 scores) were milder than in the model group (31 scores), the former two could markedly alleviate the myocardial pathologic injury (P<0.05, P<0.01). The GrB mRNA expression in the model group was 1.3000 +/- 0.1207, the SWE group 0. 7070 +/- 0.1215, and the CsA group 0.6700 +/- 0.0997, respectively; compared with the model group, the latter two could obviously down-regulate the expression of GrB mRNA (P<0.01) and no significant difference was found between the latter two groups (P>0.05).

CONCLUSIONSWE could alleviate the pathologic change, down-regulate the mRNA expression of myocardial GrB in allogenic ectopic transplanted myocardium of rats, it is possibly one of the factors for its antagonizing effect against reject reaction.

Acetates ; chemistry ; Animals ; Caesalpinia ; chemistry ; Gene Expression Regulation, Enzymologic ; drug effects ; Graft Rejection ; drug therapy ; Granzymes ; genetics ; Heart ; drug effects ; Heart Transplantation ; immunology ; Male ; Models, Animal ; Myocardium ; metabolism ; pathology ; Plant Extracts ; pharmacology ; therapeutic use ; RNA, Messenger ; genetics ; metabolism ; Rats ; Transplantation, Homologous