Genes activated by nitric oxide during Th17 cell polarisation
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ABSTRACT: CD4+ T cells were purified from the pool of spleens and lymph nodes of naive mice by negative selection (routine purity >98%) using an AutoMacs (Miltenyi Bioscience). Culture medium was RPMI-1640 supplemented with 10% (vol/vol) FCS (LONZA), 2 mM L-glutamine, 100 U/ml penicillin, 100 ug/ml streptomycin, and 0.05 M 2-mercaptoethanol. For Th17 cell differentiation, 5 x 105cells/ml were cultured in round-bottom 96-well plates with mitomycin-C-treated spleen cells (antigen-presenting cells, 1:1 ratio APC:T cells), 1 ug/ml soluble anti-CD3, I mg/ml soluble anti CD28, 1 ng/ml TGF-beta, 10 ng/ml IL-6, 10 ng/ml IL-1beta, 10 ug/ml anti-IFNg and 10 ug/ml anti IL-4. NO-donor (NOC-18) was added at the beginning of the culture (immediately before cells) at the indicated doses. At the end of 3 days culture, the cells were harvested and used for RNA extraction.
ORGANISM(S): Mus musculus
SUBMITTER: PAULA Donate
PROVIDER: E-MEXP-3959 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
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