Phenotypical and Functional Analysis of Intraepithelial Lymphocytes from Small Intestine of Mice in Oral Tolerance
Figure 6
Quantitative mRNA analysis of cytokine expression in intestinal IELs. Total RNA was extracted from freshly isolated IELs of BALB/c and DO11.10 mice (), and cDNA of IL-2, IL-4, IL-6, IL-10, IL-17, IFN-γ, TNF-α, and TGB-β was made using PureLink Micro-to-Midi Total RNA Purification System (Invitrogen, SP, Brazil,). Quantitative PCR analysis was performed with the ABI 7500 Fast Real-Time PCR system (Applied Biosystems), in four replicates, with the TaqMan Mastermix (Applied Biosystems). Samples were normalized to 18S rRNA, and an arbitrary value of 1 was given to control group (naïve mice) for the normalization, and the remaining samples were plotted relative to that value. IELs from BALB/c mice immunized by ip route show an increase in the expression of mRNA of cytokines IL-10, IL-2, IFN-γ, TGF-β, and TNF-α. IELs from DO11.10 mice fed with OVA and challenged by ip route showed increased expression of mRNA for cytokines IFN-γ, TGF-β, and TNF-α. The results are representative of three independent experiments.
