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Overexpression of AMPKγ2 increases AMPK signaling to augment human T cell metabolism and function.

ABSTRACT: Cellular therapies are currently employed to treat a variety of disease processes. For T cell-based therapies, success often relies on the metabolic fitness of the T cell product, where cells with enhanced metabolic capacity demonstrate improved in vivo efficacy. AMP-activated protein kinase (AMPK) is a cellular energy sensor which combines environmental signals with cellular energy status to enforce efficient and flexible metabolic programming. We hypothesized that increasing AMPK activity in human T cells would augment their oxidative capacity, creating an ideal product for adoptive cellular therapies. Lentiviral transduction of the regulatory AMPKγ2 subunit stably enhanced intrinsic AMPK signaling and promoted mitochondrial respiration with increased basal oxygen consumption rates, higher maximal oxygen consumption rate, and augmented spare respiratory capacity. These changes were accompanied by increased proliferation and inflammatory cytokine production, particularly within restricted glucose environments. Introduction of AMPKγ2 into bulk CD4 T cells decreased RNA expression of canonical Th2 genes, including the cytokines interleukin (IL)-4 and IL-5, while introduction of AMPKγ2 into individual Th subsets universally favored proinflammatory cytokine production and a downregulation of IL-4 production in Th2 cells. When AMPKγ2 was overexpressed in regulatory T cells, both in vitro proliferation and suppressive capacity increased. Together, these data suggest that augmenting intrinsic AMPK signaling via overexpression of AMPKγ2 can improve the expansion and functional potential of human T cells for use in a variety of adoptive cellular therapies.

SUBMITTER: Braverman EL

PROVIDER: S-EPMC10825059 | biostudies-literature | 2024 Jan

REPOSITORIES: biostudies-literature

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Overexpression of AMPKγ2 increases AMPK signaling to augment human T cell metabolism and function.

Braverman Erica L EL McQuaid Margaret A MA Schuler Herbert H Qin Mengtao M Hani Sophia S Hippen Keli K Monlish Darlene A DA Dobbs Andrea K AK Ramsey Manda J MJ Kemp Felicia F Wittmann Christopher C Ramgopal Archana A Brown Harrison H Blazar Bruce B Byersdorfer Craig A CA

The Journal of biological chemistry 20231122 1

Cellular therapies are currently employed to treat a variety of disease processes. For T cell-based therapies, success often relies on the metabolic fitness of the T cell product, where cells with enhanced metabolic capacity demonstrate improved in vivo efficacy. AMP-activated protein kinase (AMPK) is a cellular energy sensor which combines environmental signals with cellular energy status to enforce efficient and flexible metabolic programming. We hypothesized that increasing AMPK activity in h ...[more]

PMID: 38000657

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Project description:ObjectiveDiabetic sensorimotor polyneuropathy (DSPN) affects approximately half of diabetic patients leading to significant morbidity. There is impaired neurotrophic growth factor signaling, AMP-activated protein kinase (AMPK) activity and mitochondrial function in dorsal root ganglia (DRG) of animal models of type 1 and type 2 diabetes. We hypothesized that sub-optimal insulin-like growth factor 1 (IGF-1) signaling in diabetes drives loss of AMPK activity and mitochondrial function, both contributing to development of DSPN.MethodsAge-matched control Sprague-Dawley rats and streptozotocin (STZ)-induced type 1 diabetic rats with/without IGF-1 therapy were used for in vivo studies. For in vitro studies, DRG neurons from control and STZ-diabetic rats were cultured and treated with/without IGF-1 in the presence or absence of inhibitors or siRNAs.ResultsDysregulation of mRNAs for IGF-1, AMPKα2, ATP5a1 (subunit of ATPase), and PGC-1β occurred in DRG of diabetic vs. control rats. IGF-1 up-regulated mRNA levels of these genes in cultured DRGs from control or diabetic rats. IGF-1 treatment of DRG cultures significantly (P < 0.05) increased phosphorylation of Akt, P70S6K, AMPK and acetyl-CoA carboxylase (ACC). Mitochondrial gene expression and oxygen consumption rate (spare respiratory capacity), ATP production, mtDNA/nDNA ratio and neurite outgrowth were augmented (P < 0.05). AMPK inhibitor, Compound C, or AMPKα1-specific siRNA suppressed IGF-1 elevation of mitochondrial function, mtDNA and neurite outgrowth. Diabetic rats treated with IGF-1 exhibited reversal of thermal hypoalgesia and, in a separate study, reversed the deficit in corneal nerve profiles. In diabetic rats, IGF-1 elevated the levels of AMPK and P70S6K phosphorylation, raised Complex IV-MTCO1 and Complex V-ATP5a protein expression, and restored the enzyme activities of Complex IV and I in the DRG. IGF-1 prevented TCA metabolite build-up in nerve.ConclusionsIn DRG neuron cultures IGF-1 signals via AMPK to elevate mitochondrial function and drive axonal outgrowth. We propose that this signaling axis mediates IGF-1-dependent protection from distal dying-back of fibers in diabetic neuropathy.

Project description:Polyphenols are important immunonutrients which have been investigated in the context of inflammatory and autoimmune disease due to their significant immunosuppressive properties. However, the mechanism of action of many polyphenols is unclear, particularly in human immune cells. The emerging field of immunometabolism has highlighted the significance of metabolic function in the regulation of immune cell activity, yet the effects of polyphenols on immune cell metabolic signaling and function has not been explored. We have investigated the effects of two plant-derived polyphenols, carnosol and curcumin, on the metabolism of primary human dendritic cells (DC). We report that human DC display an increase in glycolysis and spare respiratory capacity in response to LPS stimulation, which was attenuated by both carnosol and curcumin treatment. The regulation of DC metabolism by these polyphenols appeared to be mediated by their activation of the cellular energy sensor, AMP-activated Protein Kinase (AMPK), which resulted in the inhibition of mTOR signaling in LPS-stimulated DC. Previously we have reported that both carnosol and curcumin can regulate the maturation and function of human DC through upregulation of the immunomodulatory enzyme, Heme Oxygenase-1 (HO-1). Here we also demonstrate that the induction of HO-1 by polyphenols in human DC is dependent on their activation of AMPK. Moreover, pharmacological inhibition of AMPK was found to reverse the observed reduction of DC maturation by carnosol and curcumin. This study therefore describes a novel relationship between metabolic signaling via AMPK and HO-1 induction by carnosol and curcumin in human DC, and characterizes the effects of these polyphenols on DC immunometabolism for the first time. These results expand our understanding of the mechanism of action of carnosol and curcumin in human immune cells, and suggest that polyphenol supplementation may be useful to regulate the metabolism and function of immune cells in inflammatory and metabolic disease.

Project description:BackgroundChronic alcohol consumption impairs alveolar macrophage's (AM) function and increases risk for developing lung infection and pneumonia. However, the mechanism and metabolic basis of alcohol-induced AM dysfunction leading to lung infection are not well defined, but may include altered ethanol (EtOH) and reactive oxygen species metabolism and cellular energetics. Therefore, oxidative stress, endoplasmic reticulum (ER) stress, the formation of fatty acid ethyl esters [FAEEs, nonoxidative metabolites of EtOH], AMP-activated protein kinase (AMPK) signaling, and phagocytic function were examined in freshly isolated AM incubated with EtOH.MethodsAMs separated from bronchoalveolar lavage fluid samples obtained from normal volunteers were incubated with EtOH for 24 hours. AMPK signaling and ER stress were assessed using Western blotting, FAEEs by GC-MS, oxidative stress by immunofluorescence using antibodies to 4-hydroxynonenal, and phagocytosis by latex beads. Oxidative stress was also measured in EtOH-treated AMs with/without AMPK activator [5-aminoimidazole-4-carboxamide ribonucleotide (AICAR)] or inhibitor (Compound C), and in AMs incubated with FAEEs. mRNA expression for interleukins (IL-6 and IL-8), monocyte chemoattractant protein (MCP)-1, and transforming growth factor (TGF)-β was measured in AM treated with EtOH or FAEEs using RT-PCR.ResultsEtOH exposure to AM increased oxidative stress, ER stress, and synthesis of FAEEs, decreased phosphorylated AMPK, and impaired phagocytosis. Attenuation or exacerbation of EtOH-induced oxidative stress by AICAR or Compound C, respectively, suggests a link between AMPK signaling, EtOH metabolism, and related oxidative stress. The formation of FAEEs may contribute to EtOH-induced oxidative stress as FAEEs also produced concentration-dependent oxidative stress. An increased mRNA expression of IL-6, IL-8, and MCP-1 by FAEEs is key finding to suggest a metabolic basis of EtOH-induced inflammatory response.ConclusionsEtOH-induced impaired phagocytosis, oxidative stress, ER stress, and dysregulated AMPK signaling are plausibly associated with the formation of FAEEs and may participate in the pathogenesis of nonspecific pulmonary inflammation.

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Overexpression of AMPKγ2 increases AMPK signaling to augment human T cell metabolism and function.

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Overexpression of AMPKγ2 increases AMPK signaling to augment human T cell metabolism and function.

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