The immunosuppressive drug Rapamycin (RAPA) has been shown to promote expansion of CD4+IL-10+ natural human regulatory T cells (nTreg) in vitro and in vivo. RAPA effects on inducible Treg (Tri) are unknown, and this study explores in vitro responses of Tr1 to this drug. CD4+CD25neg T cells isolated from PBMC of normal donors were used to generate Tr1 cells. Expanded Tr1 cells were tested for surface markers, expression of survival proteins, resistance to apoptosis and the ability to suppress proliferation of autologous CD4+CD25neg responder T cells (RC) in functional assays. RAPA was found to promote the generation of human Tr1 cells from autologous CD4+CD2Sneg precursors in peripheral blood. Tr1 cells + RAPA mediated higher suppression (p<0.01) of RC proliferation than Tr1 cells cultured without RAPA. Tr1 cells + RAPA also expressed higher levels of FasL and granzyme B (p<0.002), produced more IL-10 and TGF-βl and were more resistant to activation-induced cell death (p<0.02). RAPA up-regulated expression of the Bcl-2 family anti-apoptotic proteins in Tri. In addition, stimulation of Tr1 by LPS + RAPA resulted in increased proliferation and resistance to apoptosis. RAPA favors in vitro generation of inducible human Treg (Tr1) from CD4+CD25neg precursor cells and significantly enhances their survival and suppressor functions.
RoncaroloMGGregoriSBattagliaMBacchettaRFleischhauerKLevingsMK. Interleukin-10-secreting type 1 regulatory T cells in rodents and humans. Immunol Rev2006; 212:28–50.
2.
BattagliaMStabiliniARoncaroloMG. Rapamycin selectively expands CD4+CD25+FoxP3+ regulatory T cells. Blood2005; 105:4743–48.
3.
BattagliaMStabiliniAMigliavaccaBHorejs-HoeckJKaupperTRoncaroloMG. Rapamycin promotes expansion of functional CD4+CD25+FOXP3+ regulatory T cells of both healthy subjects and type 1 diabetic patients. J Immunol2006; 177:8338–47.
4.
BergmannCStraussLWangYSzczepanskiMJLangSJohnsonJTWhitesideTL. T regulatory type 1 cells in squamous cell carcinoma of the head and neck: Mechanisms of suppression and expansion in advanced disease. Clin Cancer Res2008; 14:3706–15.
5.
SempereJMSorianoVBenitoJM. T regulatory cells and HIV infection. AIDS Rev2007; 9:54–60.
6.
BergmannCStraussLZeidlerRLangSWhitesideTL. Expansion and characteristics of human T regulatory type 1 cells in co-cultures simulating tumor microenvironment. Cancer Immunol Immunother2007; 56:1429–42.
7.
CaoXCaiSFFehnigerTASongJCollinsLIPiwnica-WormsDRLeyTJ. Granzyme B and perforin are important for regulatory T cell-mediated suppression of tumor clearance. Immunity2007; 27:635–46.
8.
MandapathilMHilldorferBSzczepanskiMJ. Generation and accumulation of immunosuppressive adenosine by human CD4+CD25highFOXP3+ regulatory T cells. J Biol Chem2010; 285:7176–86.
9.
RoncaroloMGBattagliaM. Regulatory T-cell immunotherapy for tolerance to self antigens and alloantigens in humans. Nat Rev Immunol2007; 7:585–98.
10.
TrzonkowskiPSzarynskaMMysliwskaJMysliwskiA. Ex vivo expansion of CD4(+)CD25(+)T regulatory cells for immunosuppressive therapy. Cytometry A2009; 75:175–88.
11.
BrunsteinCGMillerJSCaoQ. Infusion of ex vivo expanded T regulatory cells in adults transplanted with umbilical cord blood: Safety profile and detection kinetics. Blood2011; 117:1061–70.
12.
ZieglerSF. FOXP3: Not just for regulatory T cells anymore. Eur J Immunol2007; 37:21–23
13.
MandapathilMLangSGorelikEWhitesideTL. Isolation of functional human regulatory T cells (Treg) from the peripheral blood based on the CD39 expression. J Immunol Methods2009; 346:55–63.
14.
SeddikiNSantner-NananBMartinsonJ. Expression of interleukin (IL)-2 and IL-7 receptors discriminates between human regulatory and activated T cells. J Exp Med2006; 203:1693–700.
15.
SchulerPJHarasymczukMSchillingBLangSWhitesideTL. Separation of human CD4+CD39+ T cells by magnetic beads reveals two phenotypically and functionally different subsets. J Immunol Meth2011; 369:59–68.
16.
TheofllopoulosANDummerWKonoDH. T cell homeostasis and systemic autoimmunity. J Clin Invest2001; 108:335–40.
17.
PetersJHKoenenHJHilbrandsLBJoostenI. Immunotherapy with regulatory T cells in transplantation. Immunotherapy2009; 1:855–71.
18.
BattagliaMStabiliniADraghiciEGregoriSMocchettiCBonifacioERoncaroloMG. Rapamycin and interleukin-10 treatment induces T regulatory type 1 cells that mediate antigen-specific transplantation tolerance. Diabetes2006; 55:40–49.
19.
StraussLCzystowskaMSzajnikMMandapathilMWhitesideTL. Differential responses of human regulatory T cells (Treg) and effector T cells to rapamycin. PLoS One2009; 4:e5994.
20.
ZeiserRLeveson-GowerDBZambrickiEAKambhamNBeilhackALohJHouJZNegrinRS. Differential impact of mammalian target of rapamycin inhibition on CD4+CD25+Foxp3+ regulatory T cells compared with conventional CD4+ T cells. Blood2008; 111:453–62.
21.
ThomsonAWTurnquistHRRaimondiG. Immunoregulatory functions of mTOR inhibition. Nat Rev Immunol2009; 9:324–37.
22.
MandapathilMSzczepanskiMJSzajnikMRenJJacksonEKJohnsonJTGorelikELangSWhitesideTL. Adenosine and prostaglandin E2 cooperate in the suppression of immune responses mediated by adaptive regulatory T cells. J Biol Chem2010; 285(36):27571–80.
23.
CzystowskaMStraussLBergmannCSzajnikMRabinowichHWhitesideTL. Reciprocal granzyme/perforin-mediated death of human regulatory and responder T cells is regulated by interleukin-2 (IL-2). J Mol Med2010; 88:577–88.
24.
TaylorALWatsonCJBradleyJA. Immunosuppressive agents in solid organ transplantation: Mechanisms of action and therapeutic efficacy. Crit Rev Oncol Hematol2005; 56:23–46.
25.
Meier-KriescheHUSteffenBJChuAHLovelandJJGordonRDMorrisJAKaplanB. Sirolimus with neoral versus mycophenolate mofetil with neoral is associated with decreased renal allograft survival. Am J Transplant2004; 4:2058–66.
26.
ChapmanJRO'ConnellPJNankivellBJ. Chronic renal allograft dysfunction. J Am Soc Nephrol2005; 16:3015–26.
27.
GeisslerEKSchlittHJ. The potential benefits of rapamycin on renal function, tolerance, fibrosis, and malignancy following transplantation. Kidney Int2010; 78:1075–79.
28.
ZhengXXSanchez-FueyoAShoMDomenigCSayeghMHStromTB. Favorably tipping the balance between cytopathic and regulatory T cells to create transplantation tolerance. Immunity2003; 19:503–14.
29.
SakaguchiSMiyaraMCostantinoCMHaflerDA. FOXP3+ regulatory T cells in the human immune system. Nat Rev Immunol2010; 10:490–500.
30.
YoungJA. Epidemiology and management of infectious complications of contemporary management of chronic leukemias. Infect Disord Drug Targets2011; 11:3–10.
31.
AllanSEBroadyRGregoriSHimmelMELockeNRoncaroloMGBacchettaRLevingsMK. CD4+ T-regulatory cells: Toward therapy for human diseases. Immunol Rev2008; 223:391–421.
32.
GrossmanWJVerbskyJWBarchetWColonnaMAtkinsonJPLeyTJ. Human T regulatory cells can use the perforin pathway to cause autologous target cell death. Immunity2004; 21:589–601.
33.
StraussLBergmannCWhitesideTL. Human circulating CD4+CD25highFoxp3+ regulatory T cells kill autologous CD8+ but not CD4+ responder cells by Fas-mediated apoptosis. J Immunol2009; 182:1469–80.
34.
PasseriniLDi NunzioSGregoriS. Functional type 1 regulatory T cells develop regardless of FOXP3 mutations in patients with IPEX syndrome. Eur J Immunol2011; 41:1120–31.
35.
Leveson-GowerDBOlsonJASegaEILuongRHBakerJZeiserRNegrinRS. Low doses of natural killer T cells provide protection from acute graft-versus-host disease via an IL-4 dependent mechanism. Blood2011; 117:3220–29.
