Leukocytes have been shown to utilize different modes of motility in vitro, depending on the nature of their surrounding environment. Using a system of intravital 2-photon microscopy to image live T cells within the inflamed dermis of a mouse, we have observed two distinct interstitial crawling phenotypes that were displayed by functionally discrete CD4+ T cell populations. Th1 cells moved rapidly with amoeboid-like motility while Th2 cells crawled with mesenchymal-like motility. These two modes suggest the mechanisms underlying movement of these functionally distinct T cells are distinct. Given the feedback between the extracellular matrix and intracellular signaling through surface receptors (such as integrins), these observations have relevance in a model of Leishmania major, in which our previous data has suggests that there are functional alterations to T cells in the locally infected tissue that may restrict Th1 accumulation. Thus, we will test the hypotheses that 1) Th1 and Th2 effectors T cells utilize different molecular machinery for motility during in vivo interstitial crawling and 2) motility is modified during parasitic L. major infection.
Specific Aim 1 : Do effector Th1 and Th2 cells utilize distinct mechanisms to crawl in sites of inflammation? Currently, two main modes of interstitial crawling have been described by cell biologists;integrin-dependent (mesenchymal) and integrin-independent, myosin II-dependent (amoeboid) motility. The objective of this aim is to determine the molecular means by which distinct helper T cell populations crawl within the interstitial space of the inflamed dermis. Candidate molecular pathways will be disrupted by antibody-blockade, genetic deletion and siRNA and the effects on motility determined by intravital 2-photon microscopy.
Specific Aim 2 : Can the migratory and crawling patterns of helper T cells be modulated by local changes induced by a pathogen? In addition to the immunological responses that accompany chronic infection, pathogens such as L. major can induce significant changes in the local milieu. L. major could modulate T cell motility in trans through pathogen-driven changes in architecture and composition of the ECM of the local tissue and in cis through modulating the chemokines and adhesion molecules expressed by the infected host cells. These changes can result local microenvironments that are dramatically different from steady state and even non-parasitized inflamed tissue. The goal of this aim is to evaluate the effects of the L. major-infected tissue microenvironment on Th1 and Th2 cell crawling, as well as the ability of parasitized antigen presenting cells to interact with the different effector T cells subsets.
Using a system of intravital microscopy to image live T cells within the inflamed dermis of a mouse, we have observed two unique crawling phenotypes that were displayed by functionally distinct CD4+ T cell populations. In the current research training plan, we will test whether the different crawling phenotypes observed utilize unique molecular mechanisms in vivo. An understanding of mechanisms employed by functionally distinct effector T cells could present means of preferentially manipulating subsets of tissue-resident effector cells, potentially offering great therapeutic potential for many disease states.
|Mock, David J; Hollenbaugh, Joseph A; Daddacha, Waaqo et al. (2012) Leishmania induces survival, proliferation and elevated cellular dNTP levels in human monocytes promoting acceleration of HIV co-infection. PLoS Pathog 8:e1002635|