Faculty Bibliography

Immune modulation is a hallmark of patent filarial infection, including suppression of antigen-presenting cell function and downmodulation of filarial antigen-specific T cell responses. The mammalian target of rapamycin (mTOR) signaling pathway has been implicated in immune regulation, not only by suppressing T cell responses but also by regulating autophagy (through mTOR sensing amino acid availability). Global proteomic analysis (liquid chromatography-tandem mass spectrometry) of microfilaria (mf)-exposed monocyte-derived dendritic cells (DC) indicated that multiple components of the mTOR signaling pathway, including mTOR, eIF4A, and eIF4E, are downregulated by mf, suggesting that mf target this pathway for immune modulation in DC. Utilizing Western blot analysis, we demonstrate that similar to rapamycin (a known mTOR inhibitor), mf downregulate the phosphorylation of mTOR and its regulatory proteins, p70S6K1 and 4E-BP1, a process essential for DC protein synthesis. As active mTOR signaling regulates autophagy, we examined whether mf exposure alters autophagy-associated processes. mf-induced autophagy was reflected in marked upregulation of phosphorylated Beclin 1, known to play an important role in both autophagosome formation and autolysosome fusion, in induction of LC3II, a marker of autophagosome formation, and in induced degradation of p62, a ubiquitin-binding protein that aggregates protein in autophagosomes and is degraded upon autophagy that was reduced significantly by mf exposure and by rapamycin. Together, these results suggest that Brugia malayi mf employ mechanisms of metabolic modulation in DC to influence the regulation of the host immune response by downregulating mTOR signaling, resulting in increased autophagy. Whether this is a result of the parasite-secreted rapamycin homolog is currently under study.

Filarial infection in humans is initiated when a mosquito deposits third-stage parasite larvae (L3) in the skin. Langerhans cells (LCs) and dermal dendritic cells (DDCs) are the first cells that the parasite encounters, and L3s must evade these highly effective antigen-presenting cells to establish infection. To assess LC and DDC responses to L3 in human skin, we employed three models of increasing physiologic relevance: in vitro-generated LCs, epidermal blister explants and full-thickness human skin sections. In vitro-generated LCs expressed TLR1-10 and robustly produced IL-6 and TNF-α in response to PolyI:C, but pre-exposure to L3s did not alter inflammatory cytokine production or TLR expression. L3s did not modulate expression of LC markers CDH1, CD207, or CD1a, or the regulatory products TSLP or IDO in epidermal explants or in vitro-generated LC. LC, CD14+ DDC, CD1c+ DC and CD141+ DC from human skin sections were analysed by flow cytometry. While PolyI:C potently induced CCL22 production in LC, CD1c+ DC, and CD141+ DC, and IL-10 production in LC, L3s did not modulate the numbers of or cytokine production by any skin DC subset. L3s broadly failed to activate or modulate LCs or DDCs, suggesting filarial larvae expertly evade APC detection in human skin.

To characterize the function and plasticity of the major human circulating monocyte populations and to explore their role in systemic helminth infection, highly purified (by flow-based sorting) human monocyte subsets (CD14(hi)/CD16(neg) [classical], CD14(+ or hi)/CD16(med) [intermediate], and CD14(neg)/CD16(hi) [nonclassical]) were examined at homeostasis and after activation. Among these three subsets the classical and intermediate subsets were found to be the major sources of inflammatory and regulatory cytokines, as well as cytokines/chemokines associated with alternative activation, whereas the nonclassical and classical populations demonstrated an ability to transmigrate through endothelial monolayers. Moreover, it was primarily the classical subset that was the most efficient in promoting autologous T cell proliferation. The distribution of these subsets changed in the context of a systemic helminth (Wuchereria bancrofti) infection such that patent infection altered the frequency and distribution of these monocyte subsets with the nonclassical monocytes being expanded (almost 2-fold) in filarial infection. To understand further the filarial/monocyte interface, in vitro modeling demonstrated that the classical subset internalized filarial antigens more efficiently than the other two subsets but that the parasite-driven regulatory cytokine interleukin-10 was exclusively coming from the intermediate subset. Our data suggest that monocyte subsets have a differential function at homeostasis and in response to helminth parasites.