Deletion of AMPK minimizes graft-versus-host disease through an early impact on effector donor T cells
Darlene A. Monlish, Kevin J. Beezhold, Pailin Chiaranunt, Katelyn Paz, Nathan J. Moore, Andrea K. Dobbs, Rebecca A. Brown, John A. Ozolek, Bruce R. Blazar, Craig A. Byersdorfer
Darlene A. Monlish, Kevin J. Beezhold, Pailin Chiaranunt, Katelyn Paz, Nathan J. Moore, Andrea K. Dobbs, Rebecca A. Brown, John A. Ozolek, Bruce R. Blazar, Craig A. Byersdorfer
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Research Article Metabolism Transplantation

Deletion of AMPK minimizes graft-versus-host disease through an early impact on effector donor T cells

Abstract

Allogeneic hematopoietic stem cell transplantation is a viable treatment for multiple hematologic diseases, but its application is often limited by graft-versus-host disease (GVHD), where donor T cells attack host tissues in the skin, liver, and gastrointestinal tract. Here, we examined the role of the cellular energy sensor AMP kinase (AMPK) in alloreactive T cells during GVHD development. Early posttransplant, AMPK activity increased more than 15-fold in allogeneic T cells, and transplantation of T cells deficient in both AMPKα1 and AMPKα2 decreased GVHD severity in multiple disease models. Importantly, a lack of AMPK lessened GVHD without compromising antileukemia responses or impairing lymphopenia-driven immune reconstitution. Mechanistically, absence of AMPK decreased both CD4+ and CD8+ effector T cell numbers as early as day 3 posttransplant, while simultaneously increasing regulatory T cell (Treg) percentages. Improvements in GVHD resulted from cell-intrinsic perturbations in conventional effector T cells as depletion of donor Tregs had minimal impact on AMPK-related improvements. Together, these results highlight a specific role for AMPK in allogeneic effector T cells early posttransplant and suggest that AMPK inhibition may be an innovative approach to mitigate GVHD while preserving graft-versus-leukemia responses and maintaining robust immune reconstitution.

Authors

Darlene A. Monlish, Kevin J. Beezhold, Pailin Chiaranunt, Katelyn Paz, Nathan J. Moore, Andrea K. Dobbs, Rebecca A. Brown, John A. Ozolek, Bruce R. Blazar, Craig A. Byersdorfer

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Figure 7

AMPK is activated in human allogeneic T cells.

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AMPK is activated in human allogeneic T cells. (A) T cells from 2 health...
(A) T cells from 2 healthy donors were labeled with CellTrace and placed in an MLR with Mitomycin-C–treated allogeneic stimulator cells. At 96 hours, donor T cells undergoing either 0–1 or more than 4 cell divisions were flow-sorted and cell lysates immunoblotted for total and p-AMPK (n = 2/group). (B) Peripheral blood T cells were plated with Mitomycin-C–treated allogeneic stimulator cells for 96 hours with varying concentrations of tacrolimus, followed by comparing divided donor T cell percentages in the treated versus untreated groups (n = 2). Data are representative of results from 4 independent donor/stimulator pairs in 2 separate experiments. (C) T cells were harvested, labeled with CellTrace, and placed into an allogeneic MLR ± 0.3 ng/mL tacrolimus. After 6 days, cells undergoing more than 2 divisions were flow-sorted, and AMPK activation was assessed by immunoblot analysis for p-AMPK (n = 2–3 responder/stimulator pairs). Bar graphs represent densitometry measurements from a second technical replicate where n = 3 responder/stimulator pairs. (D) Ten million human PBMCs were transplanted into lightly irradiated NSG mice, and human T cells were recovered on day 7 and 11 posttransplant, followed by immunoblotting for AMPK activity in CellTracelo cells (n = 3 mice for naive or 6–8 recipients/time point pooled into 3 individual sets, days 7 and 11). *P < 0.05, **P < 0.01 by Student’s t test.