DMS/NIGMS 1: An Experimental Mathematical Framework for understanding and Controlling Regulatory Delay in Self- Nonself Determination in the Immune System - How does the immune system differentiate self from nonself? Our hypothesis, based on circuit analogues, and supported by mathematical models, is that the delay between helper (Th) and regulatory T cell (Treg) response allows the immune system to respond to the rate of increase in Ag, and that it is this signal which triggers a broader immune response. The goal of this project, then, is to provide definitive experimental evidence verifying this hypothesis. Specifically, we propose an innovative in vivo experimental framework for isolating and controlling delay in immune response and use this framework to show that by changing this delay, we can alter the self-nonself determination. This project proposes a novel experimental framework for isolating and controlling the effect of delay in immune response. The obvious challenge is that, practically, it is not possible to eliminate delay in vivo. Our approach, then, is not to eliminate Treg delay, but rather to induce delay in the Th response. Unfortunately, this too is impractical, due to the myriad of pathways involved in Th response and inability to slow down these pathways without altering other parts of the Th response. To address this, we isolate the Th and Treg populations, and prove that introducing delay in the Vaccine Schedule (VS) presented to the Th circuit, but not to the Treg circuit, can be used to negate or even reverse the PD response - implying that immune decisions can be reversed. This presents the problem of how to delay the VS for the Th circuit, but not the Treg circuit. Our solution is to decouple these circuits in vivo (E2). Specifically, we use a donor mouse (M0) with a normal immune system. We then remove the immune system from M0 (similar to a bone-marrow transplant), use flow cytometry to separate Tregs from Ths, and then transplant the Th cells to Mouse H (MH) and the Treg cells to mouse R (MR), where neither MH nor MR have a pre-existing immune response to Ag. We then use microparticles developed by co-PI Acharya to apply the VSs to MR and MH, but delaying the VS for MH. Our circuit model then predicts that the effect of VS on response will be negated or reversed if Th delay is sufficiently large - verifying our hypothesis. A critical part of E2 is selecting markers by which to sort the M0 immune system. To identify these markers, we use E1, which replicates E2, but with a single mouse. We then use flow cytometry and feature selection algorithms developed by PI Peet to identify targets for sorting.
