Voie d'immunisation et séquence d'administration de l ... - TEL

tel-00827710, version 1 - 29 May 2013
II. CHARACTERIZATION OF THE T CELL RESPONSE
Once the tetramer-enrichment technique was developed and validated in our hands, it was
used to characterize the CD8 + T cell response in our model of cross-presentation to examine
the impact of route of immunization, i.d. or i.v., on the establishment of the T cell response.
A. Kinetic of CD8 + T cell response
To determine the optimal conditions for achieving cross-priming, we compared the effects of
immunizing with a local versus systemic dissemination of cell-associated antigen. C57BL/6
mice were injected i.d. or i.v. with live splenocytes from H-2K bm1 mice engineered to express
a membrane-bound form of chicken ovalbumin in all tissues (referred to as K bm1 mOva). Use
of membrane-associated Ovalbumin (mOva) ensured that our model was not confounded by
secreted protein captured by endocytosis; and an altered K b molecule (known as K bm1 )
ensured a role for host presenting cells in the cross-priming of CD8 + T cells. In order to
precisely monitor the priming of the endogenous T cell repertoire, we utilized K b -SIINFEKL
tetramer-based enrichment, thus allowing precise enumeration and phenotypic analysis of
Ovalbumin peptide-specific T cells at early time points after immunization. Accumulation of
tetramer-positive cells could be observed as early as day 5 for i.v. immunization (Figure
21A), with cells showing downregulation of CD62L (Figure 21B) and expression of CD25
(Figure 21C). In contrast, the kinetics of T cell priming was delayed when cell-associated
antigen was delivered via the i.d. route. In the latter condition, accumulation of Ova-specific
CD8 + T cells was not observed until day 7 post-immunization. For both routes of
immunization, antigen-specific T cells accumulated over time, with day 9-12 being the peak
of the response (Figure 21A). These data demonstrated that the local delivery of cell-
associated antigen results in delayed T cell cross-priming.
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