Evaluating memory CD8 T cell quantity, distribution and migration

Abstract

Memory CD8 T cells protect against intracellular pathogens by scanning host cell surfaces, thus infection detection rates depend on memory cell number and distribution. Many cell population analyses rely on isolation from whole organs and interpretation is predicated on presumptions of near-complete cell recovery. Paradigmatically, T cell memory is parsed into central, effector, and resident subsets, ostensibly defined by immunosurveillance patterns, but in practice identified by phenotypic markers. Because isolation methods and subsequent phenotypic marker-based analyses ultimately inform models of memory T cell differentiation, protection, and vaccine translation, we tested their validity via quantitative immunofluorescence microscopy of a murine memory CD8 T cell population. We found that lymphocyte isolation fails to recover most cells and recovery is biased against certain subsets. Applying this approach to parabiotic mice we found that the overwhelming majority of memory CD8 T cells in non-lymphoid tissues are resident, rather than recirculating. Residence was not absolutely predicted by common phenotypic markers (CD103 & CD69), a finding that demonstrates heterogeneity in the resident memory population and insists that migration rather than solely phenotype be used for identification. Despite tissue-specific immune regulation, establishment of resident memory CD8 T cells was extended to male genital tract tissues, where they maintain local cytokine production in the presence of rechallenge. Our studies of male genital tract organs revealed non-canonical migration of effector CD8 T cells directly into visceral non-lymphoid tissues of recently infected mice. Together, these results provide a systematic quantification of the distribution and compartmentalization of virus-specific memory CD8 T cell subsets and highlight the relative numerical abundance of resident memory CD8 T cells, indicating that host immunosurveillance by memory CD8 T cells is conducted in a highly localized manner.