Metabolic Reprogramming of

Immune Cells

Ricardo Silvestre
ricardosilvestre@med.uminho.pt
Cellular metabolism

Pearce EL, Science 2013

Lötscher and M. L. Balmer, Clin Exp Immunology 2019
Molecular Players

Mesquita I, PhD Thesis, 2020

Lötscher and M. L. Balmer, Clin Exp Immunology 2019
Cell Type

Kramer et al, Redox Biology 2014

Macrophages

Wculek et al, Cell Mol Immunol 2022

O’Neill et al, Nat Rev Immunol 2016
Mills et al, Cell 2016
Yin and O’Neill et al, FASEB J 2021
Macrophages

Wculek et al, Cell Mol Immunol 2022

Macrophages

Thapa and Lee, BMB Reports 2019

Macrophages

Thapa and Lee, BMB Reports 2019

Innate Cells

Ferreira C, PhD Thesis, 2021

T cells

Buck et. al., J Exp Med 2015

T cell metabolism

Pearce EL, Science 2013

T cell metabolism

Prlic and Bevan, Nature 2019

T cell metabolism

Palmer et al, EBioMedicine 2016

T cell metabolism

Zhang and Romero, Trends in Molecular Medicine 2016

T cell metabolism

Kouidhi, Front Immunol 2016

T cell metabolism
 Metabolism controls cell function
Upon activation

Control at post-transcriptional level

Cheng, Cell 2013
Metabolism controls cell function

O’Neill, Nat Rev Immunol 2016

Pearce EL, Science 2013
 Fatty acid synthesis and oxidation in immunity

• Inflammatory signals drive fatty acid

synthesis, which is important for
immune cell proliferation and
inflammatory cytokine production.
• Tolerogenic stimuli from the immune
system drive fatty acid oxidation,
which is required for the production of
suppressive cytokines leading to immune
tolerance and the inhibition of
inflammation.
• Effector T cells show enhanced fatty acid
synthesis and this is needed for their
growth. Memory T cells show fatty acid
oxidation, which limits their growth and
allows them to persist.
O’Neill, Nat Rev Immunol 2016
 To sum up…

O’Neill, Nat Rev Immunol 2016

 Epigenetic-metabolic interface
The epigenetic-metabolic interface in immune cells

McCall CE. et al, J. Leukocyte Biol 201

 Epigenetic-metabolic interface
Diet and energy metabolism affect gene expression, which influences human health and
disease.

Bioenergetics provides the interface between the environment and the

epigenome
A number of key energy metabolites including SAM, acetyl-CoA, NAD+ and ATP serve as
essential co-factors for many, perhaps most, epigenetic enzymes that regulate DNA
methylation, posttranslational histone modifications, and nucleosome position.

The relative abundance of these energy metabolites allows a cell to sense its energetic state.
And as co-factors, energy metabolites act as rheostats to modulate the activity of epigenetic
enzymes and upregulate/downregulate transcription as appropriate to maintain homeostasis.
Targeting Immunometabolism
as therapy
O’Neill, Nat Rev Immunol 2016
O’Neill, Nat Rev Immunol 2016
Pålsson-McDermott and O’Neill, Cell Research 2020
Pålsson-McDermott and O’Neill, Cell Research 2020
Pålsson-McDermott and O’Neill, Cell Research 2020
 Systemic to Cellular Immunometabolic Crosstalk

Norata et al, Immunity 2015

 The Gut Microbiota at the Service of
Immunometabolism

Tan et al, Nutrients 2021

 The Gut Microbiota at the Service of
Immunometabolism

Alzharani et al, EBIoMedicine 2019

 Impact of dietary-derived metabolites on host
immunometabolism and gut immune cell
function

Tan et al, Nutrients 2021

 Impact of bacterial-derived metabolites on host
immunometabolism and gut immune cell function.

Tan et al, Nutrients 2021

Norata et al, Immunity 2015
 Adipose tissue

Kammoun et al, Reviews in Endocrine and Metabolic Disorders 2014

 Adipose tissue

Davanzo et al, Current Opinion in Pharmacology 2021

 Adipose tissue

Kohlgruber et al, Seminars in Immunology 2016

 How to address Immunometabolism at the lab

Voss et al, Nat Rev Immunol 2021

Extracellular Flux Analysis
Extracellular Flux Analysis
 How to address Immunometabolism at the lab

Voss et al, Nat Rev Immunol 2021

 Cell metabolism by flow cytometry

O’Connor et al, IUBMB Life. 2001

Sun et al, Cell Death Dis. 2014
Lipid metabolism by flow cytometry
 Measurement of mitochondrial membrane potential
The accumulation of dye is dependent and proportional to
mitochondrial membrane potential

ΔΨm: hyperpolarization Signal

ΔΨm: hypopolarization Signal

MitoTracker Deep Red staining of HeLa cells

TMRE (tetramethylrhodamine, ethyl ester);

JC-1 (fluorescence emission shift from green (~529 nm) to red (~590 nm).
Mitochondrial depolarization is indicated by a decrease in the red/green fluorescence
intensity ratio)

Mitotracker (Deep Red, Orange)

Stained

JC-1 staining of NIH 3T3 cells

Unstained
 Mitochondria mass analysis
What if mitochondria alterations are not related to changes in membrane potential? What if it
is related to the number of mitochondria per cell?

Borodkina et al, Aging, 2014

How can we assess differences in mitochondrial mass by flow cytometry?