Transforming growth factor‑β2 increases the capacity of retinal pigment epithelial cells to induce the generation of regulatory T cells

Yan,Feng; He,Jin; Tang,Li; Kong,Yi; Shi,Yuhua; Chen,Suihua; Huang,Zhenping

doi:10.3892/mmr.2015.4666

Transforming growth factor‑β2 increases the capacity of retinal pigment epithelial cells to induce the generation of regulatory T cells

Authors:
- Feng Yan
- Jin He
- Li Tang
- Yi Kong
- Yuhua Shi
- Suihua Chen
- Zhenping Huang
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Affiliations: Department of Ophthalmology, Jinling Hospital, School of Medicine, Nanjing University, Nanjing, Jiangsu 210002, P.R. China, Department of Neurosurgical Laboratory, Jinling Hospital, School of Medicine, Nanjing University, Nanjing, Jiangsu 210002, P.R. China, Department of Ophthalmology, No. 97 Hospital, Xuzhou, Jiangsu 221004, P.R. China
Published online on: December 9, 2015 https://doi.org/10.3892/mmr.2015.4666
Pages: 1367-1372

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Abstract

The present study investigated the underlying mechanism of the induction of regulatory T cells (Tregs) by retinal pigment epithelial (RPE) cells and the characteristics of these Tregs. Human RPE cells were cultured in the presence or absence of transforming growth factor‑β 2 (TGF‑β2), and reverse-transcription quantitative PCR was performed to determine the mRNA expression of indoleamine 2,3‑dioxygenase (IDO) and nuclear factor erythroid 2‑related factor (Nrf2). Supernatants of RPE cell cultures were added to CD4+ T cells to induce Tregs. The RPE‑induced Tregs were purified by two‑step magnetic cell sorting. The natural Tregs were isolated from the peripheral blood mononuclear cells of healthy volunteers. Purified CD4+ CD25- T cells (2x105/well) were cultured alone or with Tregs (various densities, natural or RPE‑induced). The proliferation of CD4+ CD25- T cells was determined by 3H‑thymidine incorporation. After 24 h of stimulation with TGF‑β2, the mRNA expression of IDO in RPE cells was upregulated. The highest level of IDO mRNA expression was reached after 72 h of stimulation with TGF‑β2. However, the Nrf2 mRNA expression was slightly decreased after 24 h of stimulation with TGF‑β2 and significantly increased after 48‑72 h of TGF‑β2 stimulation. Increased levels of CD25 expression were observed on CD4+ T cells exposed to supernatants of RPE cell cultures treated with TGF‑β2 and recombinant interleukin‑2. The RPE‑induced Tregs were more effective at suppressing the proliferation of CD4+ CD25- T cells compared with native Tregs. These findings suggested that IDO may be a signaling protein in RPE cells which is implicated in the induction of Tregs. RPE‑induced Tregs have the potential to be applied for immunotherapy for ocular inflammatory diseases.

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1	Sugita S: Role of ocular pigment epithelial cells in immune privilege. Arch Immunol Ther Exp (Warsz). 57:263–268. 2009. View Article : Google Scholar
2	Imai A, Sugita S, Kawazoe Y, Horie S, Yamada Y, Keino H, Maruyama K and Mochizuki M: Immunosuppressive properties of regulatory T cells generated by incubation of peripheral blood mononuclear cells with supernatants of human RPE cells. Invest Ophthalmol Vis Sci. 53:7299–7309. 2012. View Article : Google Scholar : PubMed/NCBI
3	Detrick B and Hooks JJ: Immune regulation in the retina. Immunol Res. 47:153–161. 2010. View Article : Google Scholar : PubMed/NCBI
4	Gregerson DS, Heuss ND, Lew KL, McPherson SW and Ferrington DA: Interaction of retinal pigmented epithelial cells and CD4 T cells leads to T-cell anergy. Invest Ophthalmol Vis Sci. 48:4654–4663. 2007. View Article : Google Scholar : PubMed/NCBI
5	Sugita S, Horie S, Nakamura O, Futagami Y, Takase H, Keino H, Aburatani H, Katunuma N, Ishidoh K, Yamamoto Y and Mochizuki M: Retinal pigment epithelium-derived CTLA-2alpha induces TGFbeta-producing T regulatory cells. J Immunol. 181:7525–7536. 2008. View Article : Google Scholar : PubMed/NCBI
6	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. 155:1151–1164. 1995.PubMed/NCBI
7	Sakaguchi S: Naturally arising Foxp3-expressing CD25+CD4+ regulatory T cells in immunological tolerance to self and non-self. Nat Immunol. 6:345–352. 2005. View Article : Google Scholar : PubMed/NCBI
8	Yamazaki S and Morita A: Dendritic cells in the periphery control antigen-specific natural and induced regulatory T cells. Front Immunol. 4:1512013. View Article : Google Scholar : PubMed/NCBI
9	Golshayan D, Jiang S, Tsang J, Garin MI, Mottet C and Lechler RI: In vitro-expanded donor alloantigen-specific CD4+CD25+ regulatory T cells promote experimental transplantation tolerance. Blood. 109:827–835. 2007. View Article : Google Scholar
10	Yamazaki S, Inaba K, Tarbell KV and Steinman RM: Dendritic cells expand antigen-specific Foxp3+ CD25+ CD4+ regulatory T cells including suppressors of alloreactivity. Immunol Rev. 212:314–329. 2006. View Article : Google Scholar : PubMed/NCBI
11	Singer-Sam J, Robinson MO, Bellvé AR, Simon MI and Riggs AD: Measurement by quantitative PCR of changes in HPRT, PGK-1, PGK-2, APRT, MTase and Zfy gene transcripts during mouse spermatogenesis. Nucleic Acids Res. 18:1255–1259. 1990. View Article : Google Scholar : PubMed/NCBI
12	Horie S, Sugita S, Futagami Y, Yamada Y and Mochizuki M: Human retinal pigment epithelium-induced CD4+CD25+ regulatory T cells suppress activation of intraocular effector T cells. Clin Immunol. 136:83–95. 2010. View Article : Google Scholar : PubMed/NCBI
13	Bodaghi B, Goureau O, Zipeto D, Laurent L, Virelizier JL and Michelson S: Role of IFN-gamma-induced indoleamine 2,3 dioxygenase and inducible nitric oxide synthase in the replication of human cytomegalovirus in retinal pigment epithelial cells. J Immunol. 162:957–964. 1999.PubMed/NCBI
14	Livak KJ and Schmittgen TD: Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method. Methods. 25:402–408. 2001. View Article : Google Scholar
15	Grohmann U, Fallarino F and Puccetti P: Tolerance: DCs and tryptophan: Much ado about IDO. Trends Immunol. 24:242–248. 2003. View Article : Google Scholar : PubMed/NCBI
16	Chen W: IDO: More than an enzyme. Nat Immunol. 12:809–811. 2011. View Article : Google Scholar : PubMed/NCBI
17	Harden JL and Egilmez NK: Indoleamine 2,3-dioxygenase and dendritic cell tolerogenicity. Immunol Invest. 41:738–764. 2012. View Article : Google Scholar : PubMed/NCBI
18	Pallotta MT, Orabona C, Volpi C, Vacca C, Belladonna ML, Bianchi R, Servillo G, Brunacci C, Calvitti M, Bicciato S, et al: Indoleamine 2,3-dioxygenase is a signaling protein in long-term tolerance by dendritic cells. Nat Immunol. 12:870–878. 2011. View Article : Google Scholar : PubMed/NCBI
19	Cousins SW, McCabe MM, Danielpour D and Streilein JW: Identification of transforming growth factor-beta as an immunosuppressive factor in aqueous humor. Invest Ophthalmol Vis Sci. 32:2201–2211. 1991.PubMed/NCBI
20	Park CY, Yang SH, Chuck RS, Gehlbach PL and Park CG: The role of indoleamine 2,3-dioxygenase in retinal pigment epithelial cell-mediated immune modulation. Ocul Immunol Inflamm. 18:24–31. 2010. View Article : Google Scholar : PubMed/NCBI
21	Chan K, Han XD and Kan YW: An important function of Nrf2 in combating oxidative stress: Detoxification of acetaminophen. Proc Natl Acad Sci USA. 98:4611–4616. 2001. View Article : Google Scholar : PubMed/NCBI
22	Wang X, Lu L and Jiang S: Regulatory T cells: Customizing for the clinic. Sci Transl Med. 3:83ps192011. View Article : Google Scholar : PubMed/NCBI
23	Okubo Y, Mera T, Wang L and Faustman DL: Homogeneous expansion of human T-regulatory cells via tumor necrosis factor receptor 2. Sci Rep. 3:31532013. View Article : Google Scholar : PubMed/NCBI
24	Hippen KL, Merkel SC, Schirm DK, Sieben CM, Sumstad D, Kadidlo DM, McKenna DH, Bromberg JS, Levine BL, Riley JL, et al: Massive ex vivo expansion of human natural regulatory T cells [T(regs)] with minimal loss of in vivo functional activity. Sci Transl Med. 3:83ra412011. View Article : Google Scholar
25	Karlsson F, Martinez NE, Gray L, Zhang S, Tsunoda I and Grisham MB: Therapeutic evaluation of ex vivo-generated versus natural regulatory T-cells in a mouse model of chronic gut inflammation. Inflamm Bowel Dis. 19:2282–2294. 2013. View Article : Google Scholar : PubMed/NCBI