Visualization and functional characterization of T-T cell interactions in vivo

To define the T-T cell interactions during differentiation of naïve
CD4+ T helper cells in vivo displaying the interactions of naïve
CD4+ and differentiated type 2 (TH2) T helper cells by measuring
specific T helper (TH) cytokines. The homing capacity of naïve T
cells bearing a T cell receptor (TCR) specific for influenza
hemagglutinin (HA) or TH2 cells bearing a TCR specific for
ovalbumin (OVA) was analyzed in draining lymph nodes of naïve
murine BALB/c recipients after immunization and peptide challenge
in vivo. T cell homing was visualized by flow cytometry (FACS) and
fluorescence microscopy. For measuring different T cell specific
cytokines we used established systems as
Enzyme-Linked-Immunosorbent Assay (ELISA) and
Enzyme-Linked-Immunospot-(ELISPOT) Assay for interferon(IFN)-γ and
CT.4S assay and ELISPOT for IL-4 analysis. We showed that naïve and
TH2 cells migrate with similar dynamics into draining lymph nodes
and were found in close vicinity in T cell areas, when driven by
antigen in complete Freunds’ adjuvant (CFA). Under these
conditions, TH2 cells prevented CFA-mediated TH1-induction and
deviated the differentiation of naïve TH cells toward a TH2
phenotype. OVA- TH2 cells induced TH2-differentiation in naïve T
cells exclusively, if naïve and TH2 memory cells were both
activated simultaneously in the same lymph node. ‘TH2’-cells from
IL-4-/- mice were unable to affect CFA-mediated TH1-induction
showing that TH2-derived IL-4 was strictly required to deviate
naïve HA TH cells toward a TH2 phenotype. Among the various signals
capable of promoting TH2 differentiation IL-4 is one of the most
important factor for differentiation of naïve T cells into TH2
phenotype. Direct, IL-4-mediated, T-T cell interactions could be
measured and demonstrated that TH2 derived IL-4 under restricted
conditions plays a major role in TH cell differentiation in vivo.