In a recent study published in Nature, researchers developed a library of blood nutrient molecules and performed screens to find dietary components that influence anti-tumor immunity.
To work: Trans-vaccenic acid reprograms CD8+ T cells and anti-tumor immunity. Image Credit: Anusorn Nakdee/Shutterstock.com
Background
Dietary nutrients are intricately related to human physiological processes because they provide energy, biosynthesize building blocks, and act as intermediaries.
However, the ways in which circulating human nutrients influence specific physiological pathways is unclear and requires further investigation.
About the study
In this study, researchers investigated the effect of dietary-derived trans-vaccenic acid (TVA) on effector cytotoxic T lymphocyte functions and anti-tumor immunity in the brain. in vivo settings.
Researchers examined the effects of various nutrients on human T lymphocyte cells using a screening technique based on a nutrient molecule library in the blood.
All cell lines (Jurkat T, human Plat-E, B16-OVA mouse melanoma cancer cells, B16F10 mouse melanoma cancer cells, E0771 breast cancer cells and LLC1 Lewis lung carcinoma cells, and MC38 mouse colorectal adenocarcinoma cells) were validated using genomic short tandem repeats ( STR) profiling.
Initial screening was used to identify foods that promote Jurkat T lymphocyte activation triggered by cluster of differentiation 23 (CD23) and CD28 antibodies. Screen 1b revealed programmed death ligand 1 (PD-L1)-dependent Jurkat T cells stably expressing programmed cell death protein 1 (PD-1) produced by co-cultured PD-L1-expressing human H596 lung cancer cells. identified nutrients that reversed depletion.
The team performed magnetic bead purification to identify CRISPR-associated protein 9 (Cas9)-expressing OT-I cells from the spleen and peripheral lymph nodes of Cas9-OT-I animals.
Recipients of commercial chimeric antigen receptor (CAR) T lymphocyte therapy provided serum samples. Researchers infected Jurkat T lymphocytes with pre-prepared PD-1-expressing lentivirus and then selected 2.0 g/mL puromycin to obtain PD-1-expressing Jurkat T lymphocytes.
Western blotting revealed the presence of PD-1. The study involved feeding Jurkat T lymphocytes with a nutritional library for two days, then stimulating them with anti-CD3 and anti-CD28 antibodies for 12 hours. Interleukin-2 (IL-2) levels were measured using enzyme-linked immunosorbent assays (ELISA).
In the second screen, Jurkat T lymphocytes were cocultured with PD-1-expressing cells for 60 h and then stimulated with anti-CD23 and anti-CD28 antibodies for 12 h. Gpr43fl/fl, Gpr43−/− and Cd8acre mouse animals were bred to generate Gpr43−/flCd8acre conditional knockout mice (Gpr43/flCd8acre) and develop the Gpr43/ animal tumor model.
The team obtained buccal bleeds on days 3.0, 12, and 18 and performed flow cytometry to assess CD8+ T cell knockdown efficiency using antibodies targeting noncompetitive CD8 epitopes (BV711 anti-mouse CD8).
In a laboratory environment [13C] fatty acid monitoring, seahorse fatty acid oxidation assay, measured calcium (Ca2+) level of cytotoxic (CD8+) T lymphocytes, clustered regularly interspaced short palindromic repeats (CRISPR) editing of mouse OT-I cells, pull-down assay to identify cross-linked protein TVA complexes, Co-culture assay and CAR-T cell expansion assay were performed with blinatumomab. TVA levels were measured by nuclear magnetic resonance (NMR) spectroscopy.
Results
TVA, a trans fatty acid component of human milk, is derived predominantly from ruminant foods such as lamb, beef, and dairy products. Humans and mice convert only 12% to 19% of dietary TVA into rumenic acid. TVA inhibited immunoregulatory G protein-coupled receptor 43 (GPR43), a molecule stimulated by SCFA ligands.
TVA increases cytotoxic T lymphocyte function by triggering the cyclic AMP (cAMP)-protein kinase A (PKA)-cAMP-response element binding protein (CREB) axis; this indicates that dietary TVA rather than within-host gut microbiota-derived SCFAs. Host extrinsic reprogramming pathway for cytotoxic T lymphocytes.
TVA attenuated the activity of Gαi-associated GPR43 and increased cAMP levels; antagonized the effect of short-chain fatty acid molecules on cyclic AMP to enhance the function of effector-type cytotoxic T lymphocytes. TVA promoted anti-tumor immunity through CD8+ T lymphocyte regulation and selectively potentiated stimulated cytotoxic T lymphocyte function.
The increase in cytotoxic T lymphocyte function induced by TVA was regulated by the GPCR-CREB pathway, with positive feedback increasing PKA and CREB expression at the gene level. TVA activity required CREB and may enhance cytotoxic T lymphocyte function and anti-tumor immunity in vivo. CREB inhibition antagonized the effect of dietary TVA on anti-tumor immunity.
TVA developed T cell-based therapies. For cytotoxic T lymphocytes, the GPR43-CREB pathway may be cell type specific. For example, TVA treatment increased interleukin-2 synthesis by helper T (CD4+) lymphocytes but did not affect the formation of effector molecules such as tumor necrosis factor-alpha (TNF-α) and interferon-gamma (IFN-γ). Proliferation or apoptosis of helper T lymphocytes.
Therefore, the effects of dietary TVA on helper T lymphocytes were less compared to the effects on cytotoxic T lymphocytes; this may be due to low GPR43 expression in helper T lymphocytes.
Solution
Overall, study findings showed that dietary trans-vaccenic acid increased effector cytotoxic T lymphocyte activity and anti-tumor immunity. in vivo settings.
Extraorganismal TVA reprogrammed CD8+ T lymphocytes via extrinsic regulation to inactivate GPR43, in contrast to gut microbe-derived intraorganismic SCFAs that work as GPR43 agonists.
Study findings contribute to a better understanding of the molecular links between nutrition and human pathophysiology, with implications for future research on the function of circulating nutrients in health and disease.
Further research is needed to improve understanding of the downstream effector pathways of GPR43 and elucidate the underlying processes.