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The ion channel TRPV1 regulates the activation and proinflammatory properties of CD4+ T cells

Abstract

TRPV1 is a Ca2+-permeable channel studied mostly as a pain receptor in sensory neurons. However, its role in other cell types is poorly understood. Here we found that TRPV1 was functionally expressed in CD4+ T cells, where it acted as a non–store-operated Ca2+ channel and contributed to T cell antigen receptor (TCR)-induced Ca2+ influx, TCR signaling and T cell activation. In models of T cell–mediated colitis, TRPV1 promoted colitogenic T cell responses and intestinal inflammation. Furthermore, genetic and pharmacological inhibition of TRPV1 in human CD4+ T cells recapitulated the phenotype of mouse Trpv1−/− CD4+ T cells. Our findings suggest that inhibition of TRPV1 could represent a new therapeutic strategy for restraining proinflammatory T cell responses.

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Figure 1: TRPV1 is expressed in CD4+ T cells.
Figure 2: TRPV1 is a functional Ca2+ channel in CD4+ T cells.
Figure 3: TRPV1CD4 acts as a non–store-operated Ca2+ channel and contributes to TCR-induced Ca2+ influx.
Figure 4: TRPV1CD4 participates in TCR signaling.
Figure 5: Genetic deletion or pharmacological inhibition of TRPV1CD4 decreases TCR-induced production of cytokines.
Figure 6: Genetic and pharmacological inhibition of TRPV1CD4 decrease the activation of human primary CD4+ T cells.
Figure 7: TRPV1CD4 regulates T cell–mediated colitis.
Figure 8: TRPV1 expression in non-CD4+ T cells does not affect colitis severity in an adoptive-transfer model.

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References

  1. Oh-hora, M. & Rao, A. Calcium signaling in lymphocytes. Curr. Opin. Immunol. 20, 250–258 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  2. Gallo, E.M., Cante-Barrett, K. & Crabtree, G.R. Lymphocyte calcium signaling from membrane to nucleus. Nat. Immunol. 7, 25–32 (2006).

    Article  CAS  PubMed  Google Scholar 

  3. Hogan, P.G., Lewis, R.S. & Rao, A. Molecular basis of calcium signaling in lymphocytes: STIM and ORAI. Annu. Rev. Immunol. 28, 491–533 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Omilusik, K. et al. The Ca(v)1.4 calcium channel is a critical regulator of T cell receptor signaling and naive T cell homeostasis. Immunity 35, 349–360 (2011).

    Article  CAS  PubMed  Google Scholar 

  5. Omilusik, K.D., Nohara, L.L., Stanwood, S. & Jefferies, W.A. Weft, warp, and weave: the intricate tapestry of calcium channels regulating T lymphocyte function. Front. Immunol. 4, 164 (2013).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  6. Schwarz, E.C. et al. TRP channels in lymphocytes. Handb. Exp. Pharmacol. 179, 445–456 (2007).

    Article  CAS  Google Scholar 

  7. Wenning, A.S. et al. TRP expression pattern and the functional importance of TRPC3 in primary human T-cells. Biochim. Biophys. Acta 1813, 412–423 (2011).

    Article  CAS  PubMed  Google Scholar 

  8. Venkatachalam, K. & Montell, C. TRP channels. Annu. Rev. Biochem. 76, 387–417 (2007).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Owsianik, G., Talavera, K., Voets, T. & Nilius, B. Permeation and selectivity of TRP channels. Annu. Rev. Physiol. 68, 685–717 (2006).

    Article  CAS  PubMed  Google Scholar 

  10. Caterina, M.J. et al. The capsaicin receptor: a heat-activated ion channel in the pain pathway. Nature 389, 816–824 (1997).

    Article  CAS  PubMed  Google Scholar 

  11. Touska, F., Marsakova, L., Teisinger, J. & Vlachova, V. A “cute” desensitization of TRPV1. Curr. Pharm. Biotechnol. 12, 122–129 (2011).

    Article  CAS  PubMed  Google Scholar 

  12. Gunthorpe, M.J. et al. Identification and characterisation of SB-366791, a potent and selective vanilloid receptor (VR1/TRPV1) antagonist. Neuropharmacology 46, 133–149 (2004).

    Article  CAS  PubMed  Google Scholar 

  13. Arenkiel, B.R., Klein, M.E., Davison, I.G., Katz, L.C. & Ehlers, M.D. Genetic control of neuronal activity in mice conditionally expressing TRPV1. Nat. Methods 5, 299–302 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Parekh, A.B. & Penner, R. Store depletion and calcium influx. Physiol. Rev. 77, 901–930 (1997).

    Article  CAS  PubMed  Google Scholar 

  15. Smith, G.D. et al. TRPV3 is a temperature-sensitive vanilloid receptor-like protein. Nature 418, 186–190 (2002).

    Article  CAS  PubMed  Google Scholar 

  16. Valenzano, K.J. et al. N-(4-tertiarybutylphenyl)-4-(3-chloropyridin-2-yl)tetrahydropyrazine-1(2H)-carbox-amide (BCTC), a novel, orally effective vanilloid receptor 1 antagonist with analgesic properties: I. in vitro characterization and pharmacokinetic properties. J. Pharmacol. Exp. Ther. 306, 377–386 (2003).

    Article  CAS  PubMed  Google Scholar 

  17. Wahl, P., Foged, C., Tullin, S. & Thomsen, C. Iodo-resiniferatoxin, a new potent vanilloid receptor antagonist. Mol. Pharmacol. 59, 9–15 (2001).

    Article  CAS  PubMed  Google Scholar 

  18. Barr, V.A., Bernot, K.M., Shaffer, M.H., Burkhardt, J.K. & Samelson, L.E. Formation of STIM and Orai complexes: puncta and distal caps. Immunol. Rev. 231, 148–159 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Oh-hora, M. Calcium signaling in the development and function of T-lineage cells. Immunol. Rev. 231, 210–224 (2009).

    Article  CAS  PubMed  Google Scholar 

  20. Hanke, J.H. et al. Discovery of a novel, potent, and Src family-selective tyrosine kinase inhibitor. Study of Lck- and FynT-dependent T cell activation. J. Biol. Chem. 271, 695–701 (1996).

    Article  CAS  PubMed  Google Scholar 

  21. Jin, X. et al. Modulation of TRPV1 by nonreceptor tyrosine kinase, c-Src kinase. Am. J. Physiol. Cell Physiol. 287, C558–C563 (2004).

    Article  CAS  PubMed  Google Scholar 

  22. Yao, X., Kwan, H.Y. & Huang, Y. Regulation of TRP channels by phosphorylation. Neurosignals 14, 273–280 (2005).

    Article  CAS  PubMed  Google Scholar 

  23. Zhang, X., Huang, J. & McNaughton, P.A. NGF rapidly increases membrane expression of TRPV1 heat-gated ion channels. EMBO J. 24, 4211–4223 (2005).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Straus, D.B. & Weiss, A. Genetic evidence for the involvement of the lck tyrosine kinase in signal transduction through the T cell antigen receptor. Cell 70, 585–593 (1992).

    Article  CAS  PubMed  Google Scholar 

  25. Wirtz, S. & Neurath, M.F. Mouse models of inflammatory bowel disease. Adv. Drug Deliv. Rev. 59, 1073–1083 (2007).

    Article  CAS  PubMed  Google Scholar 

  26. Berg, D.J. et al. Rapid development of colitis in NSAID-treated IL-10-deficient mice. Gastroenterology 123, 1527–1542 (2002).

    Article  CAS  PubMed  Google Scholar 

  27. Launay, P. et al. TRPM4 regulates calcium oscillations after T cell activation. Science 306, 1374–1377 (2004).

    Article  CAS  PubMed  Google Scholar 

  28. Philipp, S. et al. TRPC3 mediates T-cell receptor-dependent calcium entry in human T-lymphocytes. J. Biol. Chem. 278, 26629–26638 (2003).

    Article  CAS  PubMed  Google Scholar 

  29. Caterina, M.J. et al. Impaired nociception and pain sensation in mice lacking the capsaicin receptor. Science 288, 306–313 (2000).

    Article  CAS  PubMed  Google Scholar 

  30. Schumacher, M.A., Moff, I., Sudanagunta, S.P. & Levine, J.D. Molecular cloning of an N-terminal splice variant of the capsaicin receptor. Loss of N-terminal domain suggests functional divergence among capsaicin receptor subtypes. J. Biol. Chem. 275, 2756–2762 (2000).

    Article  CAS  PubMed  Google Scholar 

  31. Saunders, C.I., Kunde, D.A., Crawford, A. & Geraghty, D.P. Expression of transient receptor potential vanilloid 1 (TRPV1) and 2 (TRPV2) in human peripheral blood. Mol. Immunol. 44, 1429–1435 (2007).

    Article  CAS  PubMed  Google Scholar 

  32. Engler, A. et al. Expression of transient receptor potential vanilloid 1 (TRPV1) in synovial fibroblasts from patients with osteoarthritis and rheumatoid arthritis. Biochem. Biophys. Res. Commun. 359, 884–888 (2007).

    Article  CAS  PubMed  Google Scholar 

  33. Spinsanti, G. et al. Quantitative real-time PCR detection of TRPV1–4 gene expression in human leukocytes from healthy and hyposensitive subjects. Mol. Pain 4, 51 (2008).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  34. Bachiocco, V. et al. Lymphocyte TRPV 1–4 gene expression and MIF blood levels in a young girl clinically diagnosed with HSAN IV. Clin. J. Pain 27, 631–634 (2011).

    Article  PubMed  Google Scholar 

  35. Shin, J.S. et al. Differences in sensitivity of vanilloid receptor 1 transfected to human embryonic kidney cells and capsaicin-activated channels in cultured rat dorsal root ganglion neurons to capsaicin receptor agonists. Neurosci. Lett. 299, 135–139 (2001).

    Article  CAS  PubMed  Google Scholar 

  36. Voolstra, O. & Huber, A. Post-translational modifications of TRP channels. Cells 3, 258–287 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  37. Armstrong, D.L., Erxleben, C. & White, J.A. Patch clamp methods for studying calcium channels. Methods Cell Biol. 99, 183–197 (2010).

    Article  CAS  PubMed  Google Scholar 

  38. Gad, M., Pedersen, A.E., Kristensen, N.N., Fernandez Cde, F. & Claesson, M.H. Blockage of the neurokinin 1 receptor and capsaicin-induced ablation of the enteric afferent nerves protect SCID mice against T-cell-induced chronic colitis. Inflamm. Bowel Dis. 15, 1174–1182 (2009).

    Article  PubMed  Google Scholar 

  39. Moran, M.M., McAlexander, M.A., Biro, T. & Szallasi, A. Transient receptor potential channels as therapeutic targets. Nat. Rev. Drug Discov. 10, 601–620 (2011).

    Article  CAS  PubMed  Google Scholar 

  40. Kedei, N. et al. Analysis of the native quaternary structure of vanilloid receptor 1. J. Biol. Chem. 276, 28613–28619 (2001).

    Article  CAS  PubMed  Google Scholar 

  41. Moqrich, A. et al. Impaired thermosensation in mice lacking TRPV3, a heat and camphor sensor in the skin. Science 307, 1468–1472 (2005).

    Article  CAS  PubMed  Google Scholar 

  42. González-Navajas, J.M. et al. Interleukin 1 receptor signaling regulates DUBA expression and facilitates Toll-like receptor 9-driven antiinflammatory cytokine production. J. Exp. Med. 207, 2799–2807 (2010).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  43. Franco, A., Shimizu, C., Tremoulet, A.H. & Burns, J.C. Memory T cells and characterization of peripheral T cell clones in acute Kawasaki disease. Autoimmunity 43, 317–324 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  44. Srikanth, S., Jung, H.J., Ribalet, B. & Gwack, Y. The intracellular loop of Orai1 plays a central role in fast inactivation of Ca2+ release-activated Ca2+ channels. J. Biol. Chem. 285, 5066–5075 (2010).

    Article  CAS  PubMed  Google Scholar 

  45. Fu, G. & Gascoigne, N.R. Multiplexed labeling of samples with cell tracking dyes facilitates rapid and accurate internally controlled calcium flux measurement by flow cytometry. J. Immunol. Methods 350, 194–199 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  46. Fu, G. et al. Themis controls thymocyte selection through regulation of T cell antigen receptor-mediated signaling. Nat. Immunol. 10, 848–856 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  47. González-Navajas, J.M. et al. TLR4 signaling in effector CD4+ T cells regulates TCR activation and experimental colitis in mice. J. Clin. Invest. 120, 570–581 (2010).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  48. Li, X. et al. Divergent requirement for Gαs and cAMP in the differentiation and inflammatory profile of distinct mouse Th subsets. J. Clin. Invest. 122, 963–973 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

We thank C. Conche and K. Sauer for help with measurements of Ca2+; A. Patapoutian (The Scripps Research Institute) for Chinese hamster ovary cells transfected to express TRPV1; E. Garcia and T. Snutch for access to electrophysiological equipment; J. Lee and C. Quinley for discussions; M. Scholl for animal breeding; T. Rambaldo for cell sorting; S. Shenouda for tissue processing; N. Varki and L. Eckmann for help with histological evaluations; and J. Santini for technical assistance with confocal imaging at the Neuroscience microscopy shared facility of the University of California San Diego (supported by the National Institute of Neurological Disorders and Stroke of the US National Institutes of Health (P30 NS047101)). Supported by the US National Institutes of Health (U01 AI095623 and P01 DK35108 to E.R., and AI083432 to Y.G.), the Canadian Institutes of Health Research (MOP-102698 to W.A.J.), the Broad Foundation (IBD-0342R to E.R.), the Crohn's & Colitis Foundation of America (SRA 3038 to E.R., RFA 3574 to S.B., and RFA 2927 to P.R.d.J.), the European Molecular Biology Organization (ALTF 288-2009 to S.B.), the Fulbright Association (S.B.), the Philippe Foundation (S.B.) and the Japan Society for the Promotion of Science (Y.A.-N.).

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Contributions

S.B., P.R.d.J., A.F. and E.R. designed the study; S.B., Y.A.-N., P.R.d.J., K.T., L.A., T.Y., T.H., X.L. and J.M.G.-N. performed most of the in vitro and in vivo experiments; S.B. measured the Ca2+ flux by flow cytometry, with the technical assistance of X.L. and G.F.; S.S. and Y.G. performed single-cell Ca2+ imaging in mouse CD4+ T cells; S.B. and J.L., with the help of H.D., performed single-cell Ca2+ imaging in Jurkat cells; L.L.N., H.X., S.R.S. and W.A.J. planned and designed the electrophysiological assays, L.L.N., H.X. and S.R.S. performed these assays, and L.L.N., H.X., S.R.S. and W.A.J. wrote the corresponding sections of the manuscript; R.T. and A.F. performed the human T cell experiments with TRPV1 antagonists; S.H. and M.C. took care of the mouse colony and genotyped the mice; L.L.N., H.X., S.R.S., W.A.J., S.B., Y.A.-N., P.R.d.J., T.Y., K.T., L.A., A.F. and E.R. analyzed and interpreted the data; S.B., E.R. and W.A.J. revised the manuscript for publication; and S.B. and E.R. wrote the manuscript.

Corresponding authors

Correspondence to Wilfred A Jefferies or Eyal Raz.

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Competing interests

H. Dong is cofounder of AddexBio.

Integrated supplementary information

Supplementary Figure 1 TRPV1 is expressed and functional in CD4+ T cells.

(a) TRPV1 protein expression in mouse splenic (SP) CD4+ T cells, in human primary CD4+ T cells enriched from peripheral blood of healthy donors and in Jurkat human T cell line (clone E6.1) was determined by immunoblot on a 4-12% SDS-PAGE gel. Chinese hamster ovary (CHO) cells either control (Ctrl) or overexpressing rat TRPV1 (TRPV1+) were used as negative and positive controls, respectively. Doublets at ~95 kilodaltons (kDa) and 115 kDa correspond to the nonglycosylated and glycosylated forms of the TRPV1 channel and upper bands likely correspond to TRPV1 multimers40. (b) Confocal microscopy images of wild-type spleen CD4+ T cells stained with the DNA-binding dye DAPI (far left), goat anti-TRPV1 primary antibody and Alexa Fluor 546–conjugated anti-goat secondary antibody (middle left) and Alexa Fluor 488–conjugated anti-CD4 (middle right) antibodies, after pre-incubation of the TRPV1 antibody with its specific blocking peptide (TRPV1 IgG + peptide) or when using the isotype-matched control antibody immunoglobulin G (Ctrl IgG). Right, merged image; Scale bar = 5 μm. (c) Confocal microscopy images of CHO-Ctrl or CHO-TRPV1+ cells stained with the DNA-binding dye DAPI (left), goat anti-TRPV1 primary antibody and Alexa Fluor 546–conjugated anti-goat secondary antibody (middle); right, merged image. Scale bar = 10 μm. (d) Immunoblot analysis of TRPV1, β-actin (a cytosolic protein) and CD3ɛ (a plasma membrane protein) in total lysates or in plasma membrane protein fractions (methods, Biotinylation of Cell Surface Proteins) of Jurkat T cells either untransduced (wild-type, WT) or transduced with nontargeting control shRNA lentiviral particles (copGFP, GFP) or with TRPV1 shRNA lentiviral particles (TRPV1 knockdown, Trpv1-KD). (e) [Ca2+]i in wild-type and Trpv1-KD Jurkat T cells loaded with the calcium indicator Fura-2 AM and treated with 10 μM capsaicin (CAP, gray bar) in the presence of 2 mM CaCl2 (2 Ca; black bar), monitored by confocal imaging and presented as the ratio of Fura-2 emission at an excitation of 340 nm to that at 380 nm (340/380); right, quantification of the Ca2+-influx peak at left (mean ± s.e.m.); ****P <0.0001 (two-tailed Student t-test). Data are representative of two (b,c) or three (a,d-e) independent experiments.

Supplementary Figure 2 TRPV1 channel contributes to TCR-induced Ca2+ influx in CD4+ T cells.

(a) [Ca2+]i in wild-type (WT) and Trpv1–/– spleen CD4+ T cells loaded with the fluorescent Ca2+ indicator dye Indo-1 AM and stimulated with soluble anti-CD3 (10 μg/ml) and anti-CD28 (1 μg/ml) antibodies (α-CD3 + α-CD28) in Ca2+-free medium, with 5 mM CaCl2 (5 Ca) added to the extracellular medium during the acquisition, monitored by flow cytometry; results are presented as the ratio of Indo-1 AM emission at 405 nm to that at 510 nm (405/510). (b) Quantification of the Ca2+-influx profile shown in a. *P <0.05; ***P <0.001 (two-tailed Student t-test). (c) [Ca2+]i in WT and Trpv1–/– spleen CD4+ T cells as in a with 0.5 mM CaCl2 (0.5 Ca) added to the extracellular medium prior to α-CD3 + α-CD28 stimulation. (d) [Ca2+]i in WT and Trpv1–/– spleen CD4+ T cells as in a with α-CD3 + α-CD28 stimulation in RPMI medium (0.42 Ca). (e) [Ca2+]i in WT and Trpv1–/– spleen CD4+ T cells stimulated with soluble anti-CD3 (20 μg/mL) and anti-CD28 (2 μg/mL) Abs in Ca2+-free medium. (f) [Ca2+]i in WT and Trpv1-TG spleen CD4+ T cells stimulated as in a. (g) [Ca2+]i in WT and Trpv1-TG spleen CD4+ T cells stimulated with ionomycin (500 nM) in Ca2+-free medium, with 5 mM CaCl2 (5 Ca) added to the extracellular medium during the acquisition. (h) [Ca2+]i in WT and Trpv1–/– spleen CD4+ T cells stimulated with ionomycin (500 nM) or (i) thapsigargin (TPSG, 1 μM) in Ca2+-free medium, with 5 mM CaCl2 (5 Ca) added to the extracellular medium during the acquisition. (j) [Ca2+]i in WT and Trpv3–/– spleen CD4+ T cells stimulated as in a. (k) [Ca2+]i in WT CD4+ T cells loaded with Indo-1 AM and pretreated for 5 min with the TRPV1 antagonist I-RTX (0.1 or 1 μM) or the vehicle (veh; 0.1% dimethyl sulfoxide (DMSO)), then stimulated as in a.

Supplementary Figure 3 Cytokine profiles of Trpv1-TG CD4+ T cells and Trpv1–/– OT-II CD4+ T cells.

(a) Enzyme-linked immunosorbent assay (ELISA) of cytokines (horizontal axes) in wild-type (WT) and Trpv1-TG splenic CD4+ T cells stimulated for 24 h (IFN-γ, IL-17A, IL-2 and TNF) or 48 h (IL-10 and IL-4) with plate-bound anti-CD3 (10 μg/ml) and soluble anti-CD28 (1 μg/ml) (mean ± s.e.m. (n = 4 mice/group)); *P <0.05; **P <0.01; ***P <0.001 (two-tailed Student t-test). (b) Flow cytometry analysis of IFN-γ and IL-2 intracellular production in CD4+ T cells recovered from cocultures of OVA-loaded wild-type splenic CD11c+ DCs incubated for 5 d with OVA-specific Trpv1+/+ or Trpv1–/– OT-II splenic CD4+ T cells (n = 4 mice per group). Equal numbers of the CD4+ T cells recovered were restimulated or not for 5 h with anti-CD3 plus anti-CD28 before analysis. Representative panels of intracellular cytokine production in the different conditions are shown.

Supplementary Figure 4 Trpv1–/– and WT CD4+ T cells display similar apoptosis and proliferative response.

(a) Apoptosis of wild-type (WT) and Trpv1–/– spleen CD4+ T cells stimulated with anti-CD3 (10 μg/mL plate-bound) and anti-CD28 (1 μg/mL soluble) for 24 h as in Figure 5a, stained for CD4, annexin V and the DNA-intercalating dye 7-AAD and analyzed by flow cytometry. NS, not significant (two-tailed Student t-test) (mean ± s.e.m. (n = 3-4 mice/group)). (b) Proliferation of WT and Trpv1–/– spleen CD4+ T cells left unstimulated (US) or stimulated with anti-CD3 (10 μg/mL plate-bound) plus anti-CD28 (2 μg/mL soluble) (top panel) or PMA (25 ng/mL) and ionomycin (Iono, 500 nM) (bottom panel) for 72 h, stained with CD4 and CFSE and analyzed by flow cytometry. Representative panels of 3 independent experiments and percentages of cells that underwent 0-5 divisions are shown (mean ± s.e.m. (n = 3-4 mice/group)).

Supplementary Figure 5 Treatment with a TRPV1 antagonist reduces colitis severity in Il10–/– mice.

(a) Wasting disease in Il10–/– treated daily with the TRPV1 antagonist SB366791 (3 mg/kg, i.p) or with its vehicle (VEH) starting 3 days prior and daily during the 14 days of colitis induction by piroxicam (PXC). (b) Microcopy of colon sections from mice as in a, stained with hematoxylin and eosin. Scale bar, 1 mm (main images; 20× objective) or 100 μm (insets; 1× objective). (c) Colitis scores of the mice in a. (d) Enzyme-linked immunosorbent assay (ELISA) of IFN-γ and IL-17A production by spleen (SP) CD4+ T cells recovered from mice as in a, and restimulated for 24 h with anti-CD3 (10 μg/mL plate-bound) plus anti-CD28 (1 μg/mL soluble). (mean ± s.e.m. (n = 7-8 mice/group)). NS, not significant; **P <0.01 (two-tailed Student t-test). Data are representative of two independent experiments.

Supplementary Figure 6 Trpv1–/– naive CD4+ T cells have impaired colitogenic properties in an adoptive transfer model.

(a) Representative pictures of colons from Rag1–/– recipient mice 4 weeks after adoptive transfer of 3 × 105 wild-type (WT) or Trpv1–/– naive (CD4+CD45RBhiCD25) T cells or a mixture of 3 × 105 WT naive CD4+ T cells plus 1.5 × 105 WT Treg (CD4+CD45RBloCD25+) cells (WT naive T cells + Treg) as in Figure 7d. (b) Colon length in each experimental group. (c) Microcopy of colon sections from mice as in a, stained with the DNA-binding dye DAPI, Alexa Fluor 546–conjugated anti-CD45 and Alexa Fluor 488–conjugated anti-Claudin-3 antibodies. Scale bar, 50 μm (40× objective). (d) Real-time PCR analysis of several proinflammatory cytokines (top panel) and chemokines (bottom panel) in colon samples. The Treg control group was used as reference and assigned to 1. (mean ± s.e.m. (n = 6-8 mice/group)). NS, not significant; *P < 0.05, **P < 0.01 and ***P < 0.001 (one-way analysis of variance (ANOVA) with post-hoc Bonferroni’s test).

Supplementary Figure 7 Trpv1–/– naive CD4+ T cells have impaired inflammatory responses in an adoptive transfer model.

Flow cytometry analysis of IFN-γ, IL-17A, FOXP3 and IL-10 intracellular expression in CD4+ T cells recovered from Rag1–/– recipient mice 4 weeks after adoptive transfer of 3 × 105 wild-type (WT) or Trpv1–/– naive (CD4+CD45RBhiCD25) T cells as in Figure 7d, and restimulated for 5 h with anti-CD3 (10 μg/mL plate-bound) plus anti-CD28 (1 μg/mL soluble). Percentages and numbers of IFN-γ+, IL-17A+, FOXP3+ or IL-10+ CD4+ T cells in (a) the spleen (SP), (b,c) the mesenteric lymph nodes (MLN) and (d) the lamina propria (LP) are shown (pooled organs (n = 2 mice/group)).

Supplementary Figure 8 Deletion of Trpv1 decreases the capacity of naive CD4+ T cells to differentiate into TH1, TH2 and TH17 effector cells in vitro.

(a) Flow cytometry analysis of IFN-γ, IL-4, IL-17A and IL-10 intracellular production in CD4+ T cells recovered from cocultures of OVA-loaded wild-type bone marrow CD11c+ DCs incubated for 5 d with OVA-specific Trpv1+/+ or Trpv1–/– OT-II splenic CD4+ T cells (n = 4 mice per group) under Th1, Th2, Th17 and Treg-polarizing conditions (methods, in vitro T cell differentiation). Equal numbers of the CD4+ T cells recovered were restimulated or not for 5 h with anti-CD3 (10 μg/mL plate-bound) plus anti-CD28 (1 μg/mL soluble) before analysis. Representative panels of intracellular cytokine production in the different conditions are shown. (b) Percentages of CD4+IFN-γ+ (Th1), CD4+IL-4+ (Th2), CD4+IL-17A+ (Th17), and CD4+IL-10+ (Treg) cells as in a (mean ± s.e.m.). NS, not significant; *P <0.05 (two-tailed Student t-test).

Supplementary Figure 9 Trpv1-TG naive CD4+ T cells induce exacerbated colitis in an adoptive transfer model.

(a) Body weight of Rag1–/– recipient mice 4 weeks after adoptive transfer of 3 × 105 wild-type (WT) or Trpv1-TG naive (CD4+CD45RBhiCD25) T cells or a mixture of 3 × 105 WT naive CD4+ T cells plus 1.5 × 105 WT Treg (CD4+CD45RBloCD25+) cells (WT naive T cells + Treg). (b) Disease activity index (DAI) of the mice in a. (c) Microcopy of colon sections from the mice in a, stained with hematoxylin and eosin. Scale bar, 1 mm (main images; 20× objective) or 100 μm (insets; 1× objective). (d) Colitis scores of the mice in a. (e) Cytokine concentrations in colonic explants from the mice in a after 24 h of culture. (f) Cytokine production by splenic or (g) MLN CD4+ T cells isolated from the mice in a and restimulated for 24 h with anti-CD3 (10 μg/ml plate-bound) plus anti-CD28 (1 μg/ml soluble). *P < 0.05, **P < 0.01 and ***P < 0.001 (one-way (b,dg) or two-way (a) ANOVA with post-hoc Bonferroni's test). Data are from one experiment representative of two experiments (mean ± s.e.m. (n = 5–6 mice per group)).

Supplementary Figure 10 Proposed model for the regulation of T cell activation by TRPV1.

TCR stimulation induces the activation of protein tyrosine kinases, such as Lck, which initiate a cascade of phosphorylation events. Our data indicate that TRPV1 is part of this signaling cascade and that tyrosine phosphorylation by Lck is likely the gating mechanism of TRPV1 after TCR stimulation. In addition to CRAC, Voltage-gated Ca2+ channels (CaV) and other TRPs channels, we identified that TRPV1 is required for the proper transduction of TCR induced-Ca2+ influx, TCR-mediated signaling events (for example, MAPKs, NFAT-1 and NF-κB), and downstream activation and acquisition of proinflammatory properties by CD4+ T cells.

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Bertin, S., Aoki-Nonaka, Y., de Jong, P. et al. The ion channel TRPV1 regulates the activation and proinflammatory properties of CD4+ T cells. Nat Immunol 15, 1055–1063 (2014). https://doi.org/10.1038/ni.3009

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