We have observed that systemic infection with the protozoan parasite Toxoplasma gondii in addition to leading to an increase in activated CD4+ Th1 cells also triggers a decrease in the size of the naive CD4+ T lymphocyte population as a consequence of a rapid and persistent thymic atrophy triggered by parasite-induced destruction of the thymic epithelium. Importantly, the resulting deficiency in naive CD4 T cells leads to an immunocompromised state in which animals chronically infected with T. gondii are susceptible to challenge with an unrelated pathogen. In 2015, we further characterized the population of naive CD4 T cells in T. gondii chronically infected mice and showed that they express decreased levels of CD5 surface expression. This work revealed that chronically infected hosts have not only fewer naive CD4+ T lymphocyte but that those remaining display TCR repertoires with lower avidities for antigen. Taken together, our results raised the possibility that thymic atrophy contributes to parasite persistence itself. To test this hypothesis we implanted neonatal thymii from RAG-/- mice into nave or chronically WT infected animals. Despite the expected significant difference in the size of the endogenous orthotopic thymii, both the cellularity and histology of the thymic implants were indistinguishable in the two sets of recipients. Moreover, we found that chronically T. gondii exposed animals implanted with thymii from naive RAG-/- mice display a significant 3-fold reduction in brain cyst loads compared with non-implanted controls. The latter finding supports the concept that parasite induced thymic involution promotes chronic infection. The innate recognition of T. gondii has been extensively characterized in the mouse but little is known about how human innate cells, which do not express the major murine sensors TLR11 and TLR12, respond to the parasite. While mice are a natural reservoir of T. gondii, humans are thought to be an accidental host and an end stage for the pathogen and as such may employ a distinct parasite detection strategy. We have been working on both characterizing the cytokine/chemokine response elicited in human peripheral blood myeloid cells upon exposure to T. gondii tachyzoites and identifying the signaling pathways involved. We found that purified elutriated CD14+ monocytes produce proinflammatory cytokines (e.g. TNF and p40 IL-12) and chemokines when exposed to live tachyzoites and established that this response occurs selectively in the CD16high (patrolling) and CD16 intermediate but not CD16low (classical) monocyte subsets. In addition, we determined that induction of cytokine/chemokine production requires live but not replicating parasites and direct contact with host cells. During 2015, we demonstrated that tachyzoite phagocytosis, but not host cell invasion, is required for cytokine responses. This finding clearly distinguishes recognition of T. gondii by human cells from the pathway occurring in mouse cells, which can be mediated by a soluble parasite extract. Moreover, we tested human primary dendritic cells (DC) and found that the CD1c+, but not CD141+ subset secrete IL-12 and TNF alpha in response to tachyzoites revealing yet another difference in toxoplasma sensing between human and mouse myeloid cells since the murine homolog of the non-responding human DC population are CD8 alpha+ DC, the major source of IL-12 in mice. The distinct molecular mechanisms and responding subsets utilized by human and murine myeloid cells for T. gondii detection may reflect differences in the nature of the host-parasite adaptation between the two species. The NOD receptors are cytoplasmic pattern recognition receptors previously characterized as recognizing bacterial cell wall peptidoglycans. Unexpectedly, when infected i.p. with T. gondii NOD1, but not NOD2, deficient mice display rapid mortality. The uncontrolled parasitemia in NOD1KO mice was a consequence of defective adaptive rather innate immunity and was accompanied by a systemic decrease in the number of lymphocytes. Subsequent analyses revealed that uninfected NOD1-/- animals display smaller lymphoid organs and reduced numbers of T and B lymphocytes. Experiments with mixed bone marrow chimeras demonstrated that NOD1-/- animals display a selective decrease in expansion of common lymphoid progenitors (CLP), a defect due to an intrinsic role of NOD1 in these cells. In 2015, we showed that CLP from NOD1-/- animals display selective hypo-responsiveness when stimulated with Flt3 and IL-7. Phenotypic analysis demonstrated that differences in expression of Flt3 or IL-7R cannot account for the poor ligand responses observed with NOD1 deficient CLP. Instead, it appears that NOD1 is required for optimal signaling downstream of Flt3/IL-7R. mRNA and protein comparative analyses revealed that CLP from NOD1-/- deficient mice express a truncated form of NOD1 that lacks the normal CARD signaling domain, but expresses the N-terminal ligand binding portion of the NOD domain and thus may serve as a dominant negative form. These findings reveal a novel role for NOD1 in cytokine-mediated homeostatic proliferation of lymphoid progenitors in BM at steady state, a function that becomes even more important under conditions of immune stress induced by infection.
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