ABSTRACT
Following activation, T cells rapidly divide and acquire effector functions. This energetically demanding process depends upon the ability of T cells to undergo metabolic remodeling from oxidative phosphorylation to aerobic glycolysis, during which glucose is converted into lactate and released extracellularly. In this article, we demonstrate that extracellular lactate can be used to dynamically assess human T cell responses in vitro. Extracellular lactate levels strongly correlated with T cell proliferation, and measuring lactate compared favorably with traditional methods for determining T cell responses (i.e., [3H]thymidine incorporation and the use of cell proliferation dyes). Furthermore, we demonstrate the usefulness of measuring lactate as a read-out in conventional suppression assays and high-throughput peptide-screening assays. Extracellular lactate was stably produced over 7 d, and results were reproducibly performed over several freeze-thaw cycles. We conclude that the use of extracellular lactate measurements can be a sensitive, safe, stable, and easy-to-implement research tool for measuring T cell responses and cellular metabolic changes in vitro.
Subject(s)
CD4-Positive T-Lymphocytes/immunology , CD8-Positive T-Lymphocytes/immunology , Cell Proliferation/physiology , Lactic Acid/analysis , Cells, Cultured , Cytomegalovirus/immunology , Glycolysis/physiology , Humans , Lactic Acid/metabolism , Lymphocyte Activation/immunology , Oxidative Phosphorylation , Viral Proteins/immunologyABSTRACT
Multiple sclerosis is an autoimmune disease of the central nervous system. Genome wide association studies have identified over 100 common variants associated with multiple sclerosis, the majority of which implicate immunologically relevant genes, particularly those involved in T-cell development. SNP rs13204742 at the THEMIS/PTPRK locus is one such variant. Here, we have demonstrated mutually exclusive use of exon 1 and 2 amongst 16 novel THEMIS isoforms. We also show inverse correlation between THEMIS expression in human CD4+ T-cells and dosage of the multiple sclerosis risk allele at rs13204742, driven by reduced expression of exon 1- containing isoforms. In silico analysis suggests that this may be due to cell-specific, allele-dependent binding of the transcription factors FoxP3 and/or E47. Research exploring the functional implications of GWAS variants is important for gaining an understanding of disease pathogenesis, with the ultimate aim of identifying new therapeutic targets.
