Abstract Number: PB0727
Meeting: ISTH 2022 Congress
Theme: Hemostatic Systems in Cancer, Inflammation and Immunity » Coagulation Proteins Beyond Hemostasis
Background: Activated protein C (aPC) is a coagulation protease known to regulate cellular signaling in innate as well as adaptive immunity. We and others have previously shown that aPC ameliorates graft versus host disease (GvHD). aPC induced regulatory T cells (Tregs) from T-effector (Teff) cells. Interestingly, cellular metabolism is known to regulate T-cell plasticity. Accordingly, we hypothesized that the coagulation protease aPC regulates T cell metabolism and plasticity.
Aims: We aim to investigate whether aPC induces Tregs via metabolic reprogramming of Teff cells.
Methods: Activation of T cells was determined by in vitro proliferation using the MLR (mixed lymphocyte reaction model: stimulation of Teff cells with AgPC, ratio 1:3) or plate-bound anti-CD3 and CD28 stimulation. Teff and Treg cell markers were measured by FACS. 5-methyl cytosine(5mC) pattern was analyzed by dot blot and FACS. Oxygen consumption rate (OCR) was measured by Seahorse and TCA metabolites by LC/MS. Steady-state T-cell phenotyping was done by analyses of T-cells from APChigh and WT mice. GvHD was induced in BALB/c mice by transplanting 2×106 T cells with 5-6×106 bone marrow cells from C57BL/6 mice.
Results: aPC treatment suppressed the global 5mC methylation in Tregs. Furthermore, induction of Tregs from Teff cells was associated with Foxp3 promoter demethylation and increased Foxp3 expression in vitro suggesting that T-cell plasticity is associated with epigenetic changes. These epigenetic changes were associated with metabolic reprogramming in T-cells, reflected by reduced OCR and alteration in TCA cycle metabolites upon aPC treatment. In particular α-ketoglutarate (αKG) was reduced in aPC-pretreated Teff cells, which was associated with reduced expression of the glutamine transporter Snat1 and Asct2 and reduced glutamate levels. These effects of aPC were reversed by addition of α-KG and glutamine resulting in increased proliferation and decreased Foxp3 expression.
Conclusion(s): These data demonstrate that aPC metabolically re-programs T-cells and induces epigenetic changes, favoring a Treg phenotype.
To cite this abstract in AMA style:
Gupta D, Elwakiel A, Ranjan S, Pandey M, Krishnan S, Henschler R, Kohli S, Isermann B. The induction of Tregs by aPC depends on metabolic reprogramming and reduced αKG levels [abstract]. https://abstracts.isth.org/abstract/the-induction-of-tregs-by-apc-depends-on-metabolic-reprogramming-and-reduced-%ce%b1kg-levels/. Accessed September 21, 2023.« Back to ISTH 2022 Congress
ISTH Congress Abstracts - https://abstracts.isth.org/abstract/the-induction-of-tregs-by-apc-depends-on-metabolic-reprogramming-and-reduced-%ce%b1kg-levels/