Suppression assays with human T regulatory cells: A technical guide

European Journal of Immunology

Volumn 42, Issue 1, 2012, Pages 27-34

  Mcmurchy, Alicia N ^a   Levings, Megan K ^a  

^a UNIVERSITY OF BRITISH COLUMBIA   (Canada)

Author keywords

Division index; Human; Suppression; Tolerance; Treg cells

Indexed keywords

CYTOKINE; THYMIDINE; TRANSCRIPTION FACTOR FOXP3;

CD4+ T LYMPHOCYTE; CELL DEATH; CELL PROLIFERATION; CYTOKINE PRODUCTION; FLOW CYTOMETRY; HUMAN; IMMUNOSUPPRESSIVE TREATMENT; NONHUMAN; PERIPHERAL BLOOD MONONUCLEAR CELL; PRIORITY JOURNAL; REGULATORY T LYMPHOCYTE; REVIEW;

FLOW CYTOMETRY; FORKHEAD TRANSCRIPTION FACTORS; HUMANS; IMMUNE TOLERANCE; T-LYMPHOCYTES, REGULATORY;

EID: 84855233207     PISSN: 00142980     EISSN: 15214141     Source Type: Journal    
DOI: 10.1002/eji.201141651     Document Type: Review

References (62)

1. Sakaguchi, S., Yamaguchi, T., Nomura, T. and Ono, M., Regulatory T cells and immune tolerance. Cell 2008. 133: 775-787.
2. Shevach, E. M., Mechanisms of foxp3+ T regulatory cell-mediated suppression. Immunity 2009. 30: 636-645.
3. Sakaguchi, S., Miyara, M., Costantino, C. M. and Hafler, D. A., FOXP3+ regulatory T cells in the human immune system. Nat. Rev. Immunol. 2010. 10: 490-500.
4. Allan, S. E., Broady, R., Gregori, S., Himmel, M. E., Locke, N., Roncarolo, M. G., Bacchetta, R. et al., CD4+ T-regulatory cells: toward therapy for human diseases. Immunol. Rev. 2008. 223: 391-421.
5. Wang, J., Ioan-Facsinay, A., van der Voort, E. I., Huizinga, T. W. and Toes, R. E., Transient expression of FOXP3 in human activated nonregulatory CD4+ T cells. Eur. J. Immunol. 2007. 37: 129-138.
6. Tran, D. Q., Ramsey, H. and Shevach, E. M., Induction of FOXP3 expression in naive human CD4+FOXP3 T cells by T-cell receptor stimulation is transforming growth factor-beta dependent but does not confer a regulatory phenotype. Blood 2007. 110: 2983-2990.
7. Allan, S. E., Crome, S. Q., Crellin, N. K., Passerini, L., Steiner, T. S., Bacchetta, R., Roncarolo, M. G. et al., Activation-induced FOXP3 in human T effector cells does not suppress proliferation or cytokine production. Int. Immunol. 2007. 19: 345-354.
8. Gavin, M. A., Torgerson, T. R., Houston, E., DeRoos, P., Ho, W. Y., Stray-Pedersen, A., Ocheltree, E. L. et al., Single-cell analysis of normal and FOXP3-mutant human T cells: FOXP3 expression without regulatory T cell development. Proc. Natl. Acad. Sci. USA 2006. 103: 6659-6664.
9. Morgan, M. E., van Bilsen, J. H., Bakker, A. M., Heemskerk, B., Schilham, M. W., Hartgers, F. C., Elferink, B. G. et al., Expression of FOXP3 mRNA is not confined to CD4+CD25+ T regulatory cells in humans. Hum. Immunol. 2005. 66: 13-20.
10. Sakaguchi, S., Sakaguchi, N., Asano, M., Itoh, M. and Toda, M., Immunologic self-tolerance maintained by activated T cells expressing IL-2 receptor alpha-chains (CD25). Breakdown of a single mechanism of self-tolerance causes various autoimmune diseases. J. Immunol. 1995. 155: 1151-1164.
11. Thornton, A. M. and Shevach, E. M., CD4+CD25+ immunoregulatory T cells suppress polyclonal T cell activation in vitro by inhibiting interleukin 2 production. J. Exp. Med. 1998. 188: 287-296.
12. Takahashi, T., Kuniyasu, Y., Toda, M., Sakaguchi, N., Itoh, M., Iwata, M., Shimizu, J. et al., Immunologic self-tolerance maintained by CD25+CD4+ naturally anergic and suppressive T cells: induction of autoimmune disease by breaking their anergic/suppressive state. Int. Immunol. 1998. 10: 1969-1980.
13. Crellin, N. K., Garcia, R. V., Hadisfar, O., Allan, S. E., Steiner, T. S. and Levings, M. K., Human CD4+ T cells express TLR5 and its ligand flagellin enhances the suppressive capacity and expression of FOXP3 in CD4+CD25+ T regulatory cells. J. Immunol. 2005. 175: 8051-8059.
14. Schneider, A. and Buckner, J. H., Assessment of suppressive capacity by human regulatory T cells using a reproducible, bi-directional CFSE-based in vitro assay. Methods Mol. Biol. 2011. 707: 233-241.
15. Baecher-Allan, C., Wolf, E. and Hafler, D. A., Functional analysis of highly defined, FACS-isolated populations of human regulatory CD4+ CD25+ T cells. Clin. Immunol. 2005. 115: 10-18.
16. Levings, M. K., Sangregorio, R. and Roncarolo, M. G., Human cd25(+)cd4(+) t regulatory cells suppress naive and memory T cell proliferation and can be expanded in vitro without loss of function. J. Exp. Med. 2001. 193: 1295-1302.
17. Walker, L. S., Chodos, A., Eggena, M., Dooms, H. and Abbas, A. K., Antigen-dependent proliferation of CD4+ CD25+ regulatory T cells in vivo. J. Exp. Med. 2003. 198: 249-258.
18. Allan, S. E., Alstad, A. N., Merindol, N., Crellin, N. K., Amendola, M., Bacchetta, R., Naldini, L. et al., Generation of potent and stable human CD4+ T regulatory cells by activation-independent expression of FOXP3. Mol. Ther. 2008. 16: 194-202.
19. Oberle, N., Eberhardt, N., Falk, C. S., Krammer, P. H. and Suri-Payer, E., Rapid suppression of cytokine transcription in human CD4+CD25 T cells by CD4+Foxp3+ regulatory T cells: independence of IL-2 consumption, TGF-beta, and various inhibitors of TCR signaling. J. Immunol. 2007. 179: 3578-3587.
20. Ruitenberg, J. J., Boyce, C., Hingorani, R., Putnam, A. and Ghanekar, S. A., Rapid assessment of in vitro expanded human regulatory T cell function. J. Immunol. Methods 2011. 372: 95-106.
21. Venken, K., Thewissen, M., Hellings, N., Somers, V., Hensen, K., Rummens, J. L. and Stinissen, P., A CFSE based assay for measuring CD4+CD25+ regulatory T cell mediated suppression of auto-antigen specific and polyclonal T cell responses. J. Immunol. Methods 2007. 322: 1-11.
22. Parish, C. R., Fluorescent dyes for lymphocyte migration and proliferation studies. Immunol. Cell Biol. 1999. 77: 499-508.
23. Crome, S. Q., Clive, B., Wang, A. Y., Kang, C. Y., Chow, V., Yu, J., Lai, A. et al., Inflammatory effects of ex vivo human Th17 cells are suppressed by regulatory T cells. J. Immunol. 2010. 185: 3199-3208.
24. Sakaguchi, S., Wing, K., Onishi, Y., Prieto-Martin, P. and Yamaguchi, T., Regulatory T cells: how do they suppress immune responses? Int. Immunol. 2009. 21: 1105-1111.
25. Taams, L. S., Smith, J., Rustin, M. H., Salmon, M., Poulter, L. W. and Akbar, A. N., Human anergic/suppressive CD4(+)CD25(+) T cells: a highly differentiated and apoptosis-prone population. Eur. J. Immunol. 2001. 31: 1122-1131.
26. Baecher-Allan, C., Brown, J. A., Freeman, G. J. and Hafler, D. A., CD4+CD25high regulatory cells in human peripheral blood. J. Immunol. 2001. 167: 1245-1253.
27. Janson, P. C., Winerdal, M. E., Marits, P., Thorn, M., Ohlsson, R. and Winqvist, O., FOXP3 promoter demethylation reveals the committed Treg population in humans. PLoS One 2008. 3: e1612.
28. Floess, S., Freyer, J., Siewert, C., Baron, U., Olek, S., Polansky, J., Schlawe, K. et al., Epigenetic control of the foxp3 locus in regulatory T cells. PLoS Biol. 2007. 5: e38.
29. Miyara, M., Yoshioka, Y., Kitoh, A., Shima, T., Wing, K., Niwa, A., Parizot, C. et al., Functional delineation and differentiation dynamics of human CD4+ T cells expressing the FoxP3 transcription factor. Immunity 2009. 30: 899-911.
30. Hoffmann, P., Eder, R., Boeld, T. J., Doser, K., Piseshka, B., Andreesen, R. and Edinger, M., Only the CD45RA+ subpopulation of CD4+CD25high T cells gives rise to homogeneous regulatory T-cell lines upon in vitro expansion. Blood 2006. 108: 4260-4267.
31. Dwyer, K. M., Hanidziar, D., Putheti, P., Hill, P. A., Pommey, S., McRae, J. L., Winterhalter, A. et al., Expression of CD39 by human peripheral blood CD4+ CD25+ T cells denotes a regulatory memory phenotype. Am. J. Transplant. 2010. 10: 2410-2420.
32. Liu, W., Putnam, A. L., Xu-Yu, Z., Szot, G. L., Lee, M. R., Zhu, S., Gottlieb, P. A. et al., CD127 expression inversely correlates with FoxP3 and suppressive function of human CD4+ T reg cells. J. Exp. Med. 2006. 203: 1701-1711.
33. Seddiki, N., Santner-Nanan, B., Martinson, J., Zaunders, J., Sasson, S., Landay, A., Solomon, M. et al., Expression of interleukin (IL)-2 and IL-7 receptors discriminates between human regulatory and activated T cells. J. Exp. Med. 2006. 203: 1693-1700.
34. Baecher-Allan, C., Wolf, E. and Hafler, D. A., MHC class II expression identifies functionally distinct human regulatory T cells. J. Immunol. 2006. 176: 4622-4631.
35. Marwaha, A. K., Crome, S. Q., Panagiotopoulos, C., Berg, K. B., Qin, H., Ouyang, Q., Xu, L. et al., Cutting edge: increased IL-17-secreting T cells in children with new-onset type 1 diabetes. J. Immunol. 2010. 185: 3814-3818.
36. Kimura, A. and Kishimoto, T., IL-6: regulator of Treg/Th17 balance. Eur. J. Immunol. 2010. 40: 1830-1835.
37. Broady, R., Yu, J. and Levings, M. K., ATG-induced expression of FOXP3 in human CD4(+) T cells in vitro is associated with T-cell activation and not the induction of FOXP3(+) T regulatory cells. Blood 2009. 114: 5003-5006.
38. Levings, M. K., Gregori, S., Tresoldi, E., Cazzaniga, S., Bonini, C. and Roncarolo, M. G., Differentiation of Tr1 cells by immature dendritic cells requires IL-10 but not CD25+CD4+ Tr cells. Blood 2005. 105: 1162-1169.
39. Pillai, V., Ortega, S. B., Wang, C. K. and Karandikar, N. J., Transient regulatory T-cells: a state attained by all activated human T-cells. Clin. Immunol. 2007. 123: 18-29.
40. Walker, M. R., Kasprowicz, D. J., Gersuk, V. H., Benard, A., Van Landeghen, M., Buckner, J. H. and Ziegler, S. F., Induction of FoxP3 and acquisition of T regulatory activity by stimulated human CD4+CD25- T cells. J. Clin. Invest. 2003. 112: 1437-1443.
41. Golovina, T. N., Mikheeva, T., Suhoski, M. M., Aqui, N. A., Tai, V. C., Shan, X., Liu, R. et al., CD28 costimulation is essential for human T regulatory expansion and function. J. Immunol. 2008. 181: 2855-2868.
42. Dige, A., Hvas, C. L., Kelsen, J., Deleuran, B., Dahlerup, J. F. and Agnholt, J., Ethylene-Diamine-Tetra-Acetate (EDTA) mimics the effect of regulatory T cells in suppression assays: a potential pitfall when using AutoMACS-separated cells. J. Immunol. Methods 2009. 353: 141-144.
43. Baecher-Allan, C., Viglietta, V. and Hafler, D. A., Inhibition of human CD4(+)CD25(+high) regulatory T cell function. J. Immunol. 2002. 169: 6210-6217.
44. Baecher-Allan, C. M., Costantino, C. M., Cvetanovich, G. L., Ashley, C. W., Beriou, G., Dominguez-Villar, M. and Hafler, D. A., CD2 costimulation reveals defective activity by human CD4+CD25hi regulatory cells in patients with multiple sclerosis. J. Immunol. 2011. 186: 3317-3326.
45. Shalev, I., Schmelzle, M., Robson, S. C. and Levy, G., Making sense of regulatory T cell suppressive function. Semin. Immunol. 2011. 23: 282-292.
46. Onishi, Y., Fehervari, Z., Yamaguchi, T. and Sakaguchi, S., Foxp3+ natural regulatory T cells preferentially form aggregates on dendritic cells in vitro and actively inhibit their maturation. Proc. Natl. Acad. Sci. USA 2008. 105: 10113-10118.
47. Wang, A. Y., Crome, S. Q., Jenkins, K. M., Medin, J. A., Bramson, J. L. and Levings, M. K., Adenoviral-transduced dendritic cells are susceptible to suppression by T regulatory cells and promote interleukin 17 production. Cancer Immunol. Immunother. 2011. 60: 381-388.
48. Putnam, A. L., Brusko, T. M., Lee, M. R., Liu, W., Szot, G. L., Ghosh, T., Atkinson, M. A. et al., Expansion of human regulatory T-cells from patients with type 1 diabetes. Diabetes 2009. 58: 652-662.
49. Levings, M. K., Sangregorio, R., Sartirana, C., Moschin, A. L., Battaglia, M., Orban, P. C. and Roncarolo, M. G., Human CD25+CD4+ T suppressor cell clones produce transforming growth factor beta, but not interleukin 10, and are distinct from type 1 T regulatory cells. J. Exp. Med. 2002. 196: 1335-1346.
50. Jonuleit, H., Schmitt, E., Stassen, M., Tuettenberg, A., Knop, J. and Enk, A. H., Identification and functional characterization of human CD4(+)CD25(+) T cells with regulatory properties isolated from peripheral blood. J. Exp. Med. 2001. 193: 1285-1294.
51. Chaturvedi, V., Collison, L. W., Guy, C. S., Workman, C. J. and Vignali, D. A., Cutting edge: human regulatory T cells require IL-35 to mediate suppression and infectious tolerance. J. Immunol. 2011. 186: 6661-6666.
52. Kubach, J., Lutter, P., Bopp, T., Stoll, S., Becker, C., Huter, E., Richter, C. et al., Human CD4+CD25+ regulatory T cells: proteome analysis identifies galectin-10 as a novel marker essential for their anergy and suppressive function. Blood 2007. 110: 1550-1558.
53. Andersson, J., Tran, D. Q., Pesu, M., Davidson, T. S., Ramsey, H., O'Shea, J. J. and Shevach, E. M., CD4+ FoxP3+ regulatory T cells confer infectious tolerance in a TGF-beta-dependent manner. J. Exp. Med. 2008. 205: 1975-1981.
54. Jonuleit, H., Schmitt, E., Kakirman, H., Stassen, M., Knop, J. and Enk, A. H., Infectious tolerance: human CD25(+) regulatory T cells convey suppressor activity to conventional CD4(+) T helper cells. J. Exp. Med. 2002. 196: 255-260.
55. Francis, R. S., Feng, G., Tha-In, T., Lyons, I. S., Wood, K. J. and Bushell, A., Induction of transplantation tolerance converts potential effector T cells into graft-protective regulatory T cells. Eur. J. Immunol. 2011. 41: 726-738.
56. McClymont, S. A., Putnam, A. L., Lee, M. R., Esensten, J. H., Liu, W., Hulme, M. A., Hoffmuller, U. et al., Plasticity of human regulatory T cells in healthy subjects and patients with type 1 diabetes. J. Immunol. 2011. 186: 3918-3926.
57. Beriou, G., Costantino, C. M., Ashley, C. W., Yang, L., Kuchroo, V. K., Baecher-Allan, C. and Hafler, D. A., IL-17-producing human peripheral regulatory T cells retain suppressive function. Blood 2009. 113: 4240-4249.
58. Crome, S. Q., Wang, A. Y. and Levings, M. K., Translational mini-review series on Th17 cells: function and regulation of human T helper 17 cells in health and disease. Clin. Exp. Immunol. 2010. 159: 109-119.
59. Sakaguchi, S., Immunology: Conditional stability of T cells. Nature 2010. 468: 41-42.
60. Brunstein, C. G., Miller, J. S., Cao, Q., McKenna, D. H., Hippen, K. L., Curtsinger, J., Defor, T. et al., Infusion of ex vivo expanded T regulatory cells in adults transplanted with umbilical cord blood: safety profile and detection kinetics. Blood 2010. 117: 1061-1070.
61. Di Ianni, M., Falzetti, F., Carotti, A., Terenzi, A., Castellino, F., Bonifacio, E., Del Papa, B. et al., Tregs prevent GvHD and promote immune reconstitution in HLA-haploidentical transplantation. Blood 2011. 117: 3921-3928.
62. Palomares, O., Yaman, G., Azkur, A. K., Akkoc, T., Akdis, M. and Akdis, C. A., Role of Treg in immune regulation of allergic diseases. Eur. J. Immunol. 2010. 40: 1232-1240.