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• W2066500966 abstract "Recently, the anti-CD3 antibody has been shown to be a promising candidate for the efficient treatment of overt autoimmunity. However, the mechanisms underlying this effect remain unclear. Our previous studies demonstrated that natural killer (NK)T cells and transforming growth factor (TGF)-β were key elements in anti-CD3 F(ab′)2-mediated re-establishment of glucose homeostasis and restoration of self tolerance to islets in type 1 diabetes. In this report, we further investigate the regulatory pathways involved, especially the cellular source of TGF-β production. The treatment of new-onset nonobese diabetic mice with anti-CD3 F(ab′)2 resulted in a significant increase in the numbers of NK cells in spleen and pancreatic lymph nodes that secrete TGF-β. Depletion of this cell population with a specific anti-AsGM1 antibody abrogated anti-CD3 F(ab′)2 therapeutic effects and splenic TGF-β production. When fractionated from recovered mice after CD3 antibody therapy, these NK cells actively suppressed diabetogenic cell proliferation and prevented the cotransfer of diabetes into nonobese diabetic-severe combined immunodeficient mice in a TGF-β-dependent manner. In addition, the regulatory NKT cells from remitting mice were capable of causing NK cells to exhibit a TGF-β-producing phenotype by the secretion of the T helper 2 cytokines interleukins 4 and 10. Overall, these data indicate that NK cells are the main source of TGF-β production after anti-CD3 F(ab′)2 treatment, which are controlled by a population of T helper 2-like NKT cells. Recently, the anti-CD3 antibody has been shown to be a promising candidate for the efficient treatment of overt autoimmunity. However, the mechanisms underlying this effect remain unclear. Our previous studies demonstrated that natural killer (NK)T cells and transforming growth factor (TGF)-β were key elements in anti-CD3 F(ab′)2-mediated re-establishment of glucose homeostasis and restoration of self tolerance to islets in type 1 diabetes. In this report, we further investigate the regulatory pathways involved, especially the cellular source of TGF-β production. The treatment of new-onset nonobese diabetic mice with anti-CD3 F(ab′)2 resulted in a significant increase in the numbers of NK cells in spleen and pancreatic lymph nodes that secrete TGF-β. Depletion of this cell population with a specific anti-AsGM1 antibody abrogated anti-CD3 F(ab′)2 therapeutic effects and splenic TGF-β production. When fractionated from recovered mice after CD3 antibody therapy, these NK cells actively suppressed diabetogenic cell proliferation and prevented the cotransfer of diabetes into nonobese diabetic-severe combined immunodeficient mice in a TGF-β-dependent manner. In addition, the regulatory NKT cells from remitting mice were capable of causing NK cells to exhibit a TGF-β-producing phenotype by the secretion of the T helper 2 cytokines interleukins 4 and 10. Overall, these data indicate that NK cells are the main source of TGF-β production after anti-CD3 F(ab′)2 treatment, which are controlled by a population of T helper 2-like NKT cells. Type 1 diabetes in human and nonobese diabetic (NOD) mice is an autoimmune disease in which pancreatic islet β cells are destroyed by the cellular immune system.1Cameron MJ Meagher C Delovitch TL Failure in immune regulation begets IDDM in NOD mice.Diabetes Metab Rev. 1998; 14: 177-185Crossref PubMed Scopus (13) Google Scholar Based on our understanding of the pathogenesis of β cell destruction in type 1 diabetes, many strategies targeted to immune cells have been developed, including antibodies recognizing antigens expressed on the surface of T cells. CD3-specific antibodies have been believed to be promising candidates to treat overt diabetes.2Chatenoud L Thervet E Primo J Bach JF Anti-CD3 antibody induces long-term remission of overt autoimmunity in nonobese diabetic mice.Proc Natl Acad Sci USA. 1994; 91: 123-127Crossref PubMed Scopus (558) Google Scholar, 3Keymeulen B Vandemeulebroucke E Ziegler AG Mathieu C Kaufman L Hale G Gorus F Goldman M Walter M Candon S Schandene L Crenier L De Block C Seigneurin JM De Pauw P Pierard D Weets I Rebello P Bird P Berrie E Frewin M Waldmann H Bach JF Pipeleers D Chatenoud L Insulin needs after CD3-antibody therapy in new-onset type 1 diabetes.N Engl J Med. 2005; 352: 2598-2608Crossref PubMed Scopus (947) Google Scholar, 4Herold KC Gitelman SE Masharani U Hagopian W Bisikirska B Donaldson D Rother K Diamond B Harlan DM Bluestone JA A single course of anti-CD3 monoclonal antibody hOKT3γ1(Ala-Ala) results in improvement in C-peptide responses and clinical parameters for at least 2 years after onset of type 1 diabetes.Diabetes. 2005; 54: 1763-1769Crossref PubMed Scopus (529) Google Scholar Short-term administration of an anti-CD3 antibody resulted in acquisition of immune tolerance to islets and long-lasting normoglycemia. In surveying the underlying mechanisms, our previous study has identified that natural killer (NK)T cells are key players in the immunoregulation of autoimmunity after anti-CD3 F(ab′)2 therapy.5Chen G Han G Wang J Wang R Xu R Shen B Qian J Li Y Active tolerance induction and involvement of CD1d-restricted NKT cells in anti-CD3 F(ab′)2 treatment reversed new onset diabetes in NOD mice.Am J Pathol. 2008; 172: 972-979Abstract Full Text Full Text PDF PubMed Scopus (8) Google Scholar Furthermore, anti-CD3 F(ab′)2 treatment heightened the level of production of transforming growth factor (TGF)-β, which is widely accepted as a critical immunoregulatory cytokine in controlling pathogenic cells and maintaining immune homeostasis.6Chen G Han G Wang J Wang R Xu R Shen B Qian J Li Y Essential roles of TGF-β in anti-CD3 antibody therapy: reversal of diabetes in nonobese diabetic mice independent of Foxp3+CD4+ regulatory T cells.J Leukoc Biol. 2008; 83: 280-287Crossref PubMed Scopus (25) Google Scholar Interestingly, up-regulated TGF-β appears not to derive from NKT cells or CD4+CD25+ regulatory T cells, as depletion of this regulatory subset does not affect TGF-β secretion in tolerized NOD mice.6Chen G Han G Wang J Wang R Xu R Shen B Qian J Li Y Essential roles of TGF-β in anti-CD3 antibody therapy: reversal of diabetes in nonobese diabetic mice independent of Foxp3+CD4+ regulatory T cells.J Leukoc Biol. 2008; 83: 280-287Crossref PubMed Scopus (25) Google Scholar Thus, it is necessary to clarify the identity of lymphocyte population responsible for producing TGF-β after CD3 antibody treatment. NK cells have been shown to be important components in bridging innate and adaptive immunity. Although this kind of cell plays an effector role in cleaning virally infected cells and rejection of allogenic grafts through cytotoxic capacity and producing pro-inflammatory cytokines,7Trinchieri G Biology of natural killer cells.Adv Immuol. 1989; 47: 187-376Crossref PubMed Scopus (2717) Google Scholar, 8Kos FJ Regulation of adaptive immmunity by natural killer cells.Immunol Res. 1998; 17: 303-312Crossref PubMed Scopus (68) Google Scholar in some settings, their role is regulatory, as they can also produce multiple immunomodulatory cytokines, eg, interferon-γ, TGF-β, and interleukin (IL)-10.9Kelly JM Takeda K Darcy PK Yagita H Smith MJ A role of IFN-γ in primary and secondary immunity generated by NK cell-sensitive tumor-expressing CD80 in vivo.J Immunol. 2002; 168: 4472-4479Crossref PubMed Scopus (57) Google Scholar Recently, the deficiency of NK cell function in NOD mice has been reported, which contributes to diabetes development.10Poulton LD Smyth MJ Hawke CG Silveira P Shepherd D Naidenko OV Godfrey DI Baxter AG Cytometric and functional analyses of NK and NKT cell deficiencies in NOD mice.Int Immunol. 2001; 13: 887-896Crossref PubMed Scopus (132) Google Scholar, 11Johansson SE Hall H Björklund J Höglund P Broadly impaired NK cell function in nonobese diabetic mice is partially restored by NK cell activation in vivo and by IL-12/IL-18 in vitro.Int Immunol. 2004; 16: 1-11Crossref PubMed Scopus (49) Google Scholar Accordingly, it is conceivable to prevent the onset of diabetes by modulating NK cells. In fact, a recent study demonstrated that administration of the NK cell activator poly (I:C) in young NOD mice potentially reduced diabetes incidence and insulitis by secreting TGF-β.12Zhou R Wei H Tian Z NK3-like NK cells are involved in protective effect of polyinosinic-polycytidylic acid on type 1 diabetes in nonobese diabetic mice.J Immunol. 2007; 178: 2141-2147Crossref PubMed Scopus (35) Google Scholar Based on the regulatory function of NK cells in autoimmune disorders, this study examined the role of NK cells in anti-CD3 F(ab′)2-mediated therapeutic effects. We found that anti-CD3 F(ab′)2 antibody treatment increased the frequency and number of NK cells with a hallmark of producing TGF-β and depletion of NK cells abolished anti-CD3 F(ab′)2 effects. Furthermore, NK cells from treated mice inhibited diabetogenic cell response to autoantigen stimulation in vitro and prevented the transfer of diabetes in vivo in a TGF-β-dependent manner. NOD and NOD-severe combined immunodeficient (NOD. scid) mice were obtained originally from the Jackson Laboratory and bred in our facilities under specific pathogen-free conditions. Care, use and treatment of mice in this study were in strict agreement with the guidelines in the care and use of laboratory animals set forth by Institute of Basic Medical Sciences. The incidence of diabetes in these mice is 80% to 90% by 30 weeks of age. At 10 week of age, NOD mice were monitored for fasting blood glucose weekly. Diabetes was defined ≥11.3mmol/L on two consecutive measurements. Polyclonal rabbit anti-AsGM1 antibody (Ab) and rabbit IgG were purchased from Wako Pure Chemical. Glutamic acid decarboxylase was prepared in our lab. Ovalbumin (OVA) and porcine insulin were purchased from Sigma-Aldrich. α-Galactosylceramide was purchased from Toronto Research Chemicals Inc. α-Galactosylceramide-loaded dimer was prepared as previous description.5Chen G Han G Wang J Wang R Xu R Shen B Qian J Li Y Active tolerance induction and involvement of CD1d-restricted NKT cells in anti-CD3 F(ab′)2 treatment reversed new onset diabetes in NOD mice.Am J Pathol. 2008; 172: 972-979Abstract Full Text Full Text PDF PubMed Scopus (8) Google Scholar Fluorescein isothiocyanate-labeled anti-murine CD3 (145-2C11) and T cell receptor-β (H57-597), phycoerythrin-labeled anti-murine DX5 (DX5), and NKG2D (CX5) were purchased from eBioscience. Biotinylated polyclonal Anti-TGF-β1 antibody (1D11) and neutralizing anti-TGF-β Ab were purchased from R&D Systems. The anti-mouse IL-4 (11B11) and IL-10 (JES2A5) neutralizing Ab was purchased from Peprotech. As described previously,5Chen G Han G Wang J Wang R Xu R Shen B Qian J Li Y Active tolerance induction and involvement of CD1d-restricted NKT cells in anti-CD3 F(ab′)2 treatment reversed new onset diabetes in NOD mice.Am J Pathol. 2008; 172: 972-979Abstract Full Text Full Text PDF PubMed Scopus (8) Google Scholar anti-CD3 F(ab′)2 fragments were obtained by pepsin digestion. The diabetic NOD mice within 7 days of the onset of overt diabetes were treated intravenously with anti-CD3 F(ab′)2 (40 μg/mouse) for 5 consecutive days. The untreated diabetic littermates were regarded as controls. Lymphocytes were stained with fluorescence-labeled antibody in PBS with 2% heat-inactivated fetal calf serum and 0.2% sodium azide on ice for 25 minutes, and fixed using PBS with 1% paraformaldehyde. Data collection and analysis were performed on a FACS Calibur flow cytometer using CellQuest software (Becton Dickinson). NOD mice were injected i.v. with 20 μl of anti-AsGM1 Abs or control rabbit IgG once every 5 days. The elimination of NK cells in mice was confirmed by flow cytometry, and the majority (>90%) was depleted. Single cell suspension was prepared from spleen by passing them through nylon mesh. NK cells (CD3−DX5+) were enriched by magnetic-labeled microbeads (Miltenyi Biotec, Germany) according to manufacturer’s instructions. For purifying NKT cells, hepatic lymphocytes were pooled by 40%/80% percoll isolation and further obtained by gated on TCR-β+α-galactosylceramide-loading dimer+ in flow cytometry. The purity (>90%) of NK and NKT cells was confirmed by flow cytometry. In some experiments, CD3−DX5+ cells were further divided into CD3−CD11c−DX5+ and CD3−CD11c+DX5+ population by CD11c-positive beads (Dynal). For measurement of islet Antigen-specific T cell responses, spleen cells (5 × 105/ml) from acutely diabetic NOD mice were co-cultured with glutamic acid decarboxylase (10 μg/ml) or insulin protein (40 μg/ml) for 72 hours at 37°C. As a control, the same number of responder cells was stimulated with OVA (20 μg/ml) for 72 hours. For evaluation of the inhibitory effects of NK or DX5+ dendritic cells (DC) on the islet Ag-specific T cell responses of spleen cells, NK or DX5+ DCs (105/ml) were purified from NOD mice 5 weeks after anti-CD3 F(ab′)2 treatment or control mice and then added in the cultures of spleen cells. In some experiments, anti-TGF-β Ab (20 μg/ml) was added to the culture. As positive controls, OVA-specific T cells were stimulated with anti-CD3 mAb (0.5 μg/ml). On day 3 cultures were pulsed with 0.5μCi/well of [3H] thymidine for the last 16 hours, and the cells were harvested and counted by standard liquid scintillation. NK (CD3−CD11c−DX5+) cells were isolated from acutely diabetic NOD mice and cultured alone, with syngenic hepatic NKT cells from anti-CD3 F(ab′)2-treated remitting mice or autologous NKT cells (at 2:1 ratio) for 3 days. In some setting, neutralizing antibodies to IL-4 and/or IL-10 (5 μg/ml) or isotype IgG were added into the culture. At the end of co-culture, the supernatants were collected for determination of TGF-β production. Splenocytes (5 × 105), pancreas lymph node cells (2 × 105), or purified NK cells (1 × 105) were incubated in 96-well flat-bottom microtiter plates in the presence of 40 μg/ml insulin protein or not. Supernatants were harvested after 48 hours. The levels of TGF-β and IL-12 were determined in triplicate in 0.1 ml of supernatant by sandwich enzyme-linked immunosorbent assay. The enzyme-linked immunosorbent assay kits used in this study were purchased from R&D Systems. TGF-β enzyme-linked immunospot assay was conducted according to the instructions of the manufacturer. In brief, plates were coated with purified anti-TGF-β overnight and blocked. NK cells (2 × 105) derived from pancreatic lymph nodes of treated or untreated mice were suspended in medium and co-cultured with stimulus. After 72 hours in 5% CO2 at 37C, cells were removed and the reaction was visualized by addition of the individual biotinylated anticytokine antibody and subsequent alkaline phosphatase–conjugated streptavidin. Spots were counted using an immunospot image analyzer (Bioreader 4000 PRO-X). NOD mice were sacrificed by cervical dislocation, and the pancreas was immediately removed. Each pancreas was fixed with 10% buffered formalin, embedded in paraffin, sectioned at 4.5 μm, and stained with H&E. Insulitis grade was determined as follows: 0, normal islet; 1, mononuclear infiltration, largely in the periphery, in <25% of the islet; 2, 25% to 50% of the islets showing mononuclear infiltration; 3, >50% of the islet showing mononuclear infiltration; 4, small, retracted islet with few mononuclear cells. Purified NK or DX5+ DCs from the spleen of anti-CD3 F(ab′)2-treated or control mice were mixed with splenocytes (1 × 107 cells) from untreated, acutely diabetic NOD mice and given i.v. into the tail veins of 4- to 8-week-old NOD.scid mice. Age-matched NOD.scid mice receiving only 1 × 107 diabetic splenocytes were used as a positive control. In combination with adoptive transfer, some mice were injected intraperitoneally with anti-TGF-β Abs at a dose of 1 mg on day 0, 3, 5, 8, and thereafter every 5 days. Recipients were tested every week for diabetes and diagnosed as described above. Kaplan-Meier method was used to compare diabetes remission. Student’s t-test was used to compare mean values. Values of P < 0.05 were considered significant. First, as described previously,5Chen G Han G Wang J Wang R Xu R Shen B Qian J Li Y Active tolerance induction and involvement of CD1d-restricted NKT cells in anti-CD3 F(ab′)2 treatment reversed new onset diabetes in NOD mice.Am J Pathol. 2008; 172: 972-979Abstract Full Text Full Text PDF PubMed Scopus (8) Google Scholar, 6Chen G Han G Wang J Wang R Xu R Shen B Qian J Li Y Essential roles of TGF-β in anti-CD3 antibody therapy: reversal of diabetes in nonobese diabetic mice independent of Foxp3+CD4+ regulatory T cells.J Leukoc Biol. 2008; 83: 280-287Crossref PubMed Scopus (25) Google Scholar the efficacy of anti-CD3 F(ab′)2 antibody on treating new-onset diabetes was tested. Freshly diabetic NOD mice were randomized into two groups and injected intravenously with a low dose of anti-CD3 F(ab′)2 antibodies on five consecutive days or left untreated. The results showed that over 80% of anti-CD3 F(ab′)2-treated mice returned to normal glucose level and exhibited diabetes-free survival by 9 weeks post-treatment (Figure 1A and B). In contrast, all untreated control mice showed a progressively heightened glycemia accompanied by typical symptoms of type 1 diabetes. To determine whether the restoration of normoglycemia by CD3-specific antibody is caused by interference with cell infiltration, the mice were subjected to histopathological analysis at various time points after completion of the treatment regimen. Most of the islets in control hyperglycemic and diabetic mice exhibited intra-insulitis (Figure 2A), the majority of islets in treated mice were not inflamed (Figure 2, B–D). Also, the insulitis scores for the islets 10 weeks post-treatment indicated that the treated group had a higher number of islets with peri-insulitis (35% vs. 14%) or no insulitis (31% vs. 6%) relative to the untreated diabetic mice (Figure 2E). On the other hand, the number of islets with severe and mild intra-insulitis were reduced in the treated versus control mice (20% and 14% vs. 53% and 27%, respectively). Overall, the treatment with anti-CD3 F(ab′)2 antibody led to the establishment of benign milieus in pancreatic islets facilitating recovery of endogenous insulin-producing cell function. Previous studies indicated that the improved histology of pancreatic islets and restoration of normoglycemia was not attributed to immunosuppression, but to re-establishing immune tolerance to self islets.2Chatenoud L Thervet E Primo J Bach JF Anti-CD3 antibody induces long-term remission of overt autoimmunity in nonobese diabetic mice.Proc Natl Acad Sci USA. 1994; 91: 123-127Crossref PubMed Scopus (558) Google Scholar, 5Chen G Han G Wang J Wang R Xu R Shen B Qian J Li Y Active tolerance induction and involvement of CD1d-restricted NKT cells in anti-CD3 F(ab′)2 treatment reversed new onset diabetes in NOD mice.Am J Pathol. 2008; 172: 972-979Abstract Full Text Full Text PDF PubMed Scopus (8) Google Scholar TGF-β, whose production is significantly elevated after treatment, has been proved to be pivotal for maintaining this process by controlling pathogenic cells. Neutralization of TGF-β activity dramatically abrogated anti-CD3 therapeutic effects.13Belghith M Bluestone JA Barriot S Megret J Bach JF Chatenoud L TGF-β-dependent mechanisms mediate restoration of self-tolerance induced by antibodies to CD3 in overt autoimmune diabetes.Nat Med. 2003; 9: 1202-1208Crossref PubMed Scopus (540) Google Scholar However, the cellular source of TGF-β is still controversial, although several studies reported that CD4+CD25+ regulatory T cells were able to produce TGF-β.14Green EA Gorelik L McGregor CM Tran EH Flavell RA CD4+CD25+ T regulatory cells control anti-islet CD8+ T cells through TGF-β–TGF-β receptor interactions in type 1 diabetes.Proc Natl Acad Sci USA. 2003; 100: 10878-10883Crossref PubMed Scopus (361) Google Scholar, 15Nakamura K Kitani A Fuss I Pedersen A Harada N Nawata H Strober W TGF-β1 play an important role in the mechanism of CD4+CD25+ regulatory T cell activity in both humans and mice.J Immunol. 2004; 172: 834-842Crossref PubMed Scopus (553) Google Scholar Our observation demonstrated that depletion of this regulatory subset had minimal influence in TGF-β production and glucose homeostasis after anti-CD3 F(ab′)2 treatment.6Chen G Han G Wang J Wang R Xu R Shen B Qian J Li Y Essential roles of TGF-β in anti-CD3 antibody therapy: reversal of diabetes in nonobese diabetic mice independent of Foxp3+CD4+ regulatory T cells.J Leukoc Biol. 2008; 83: 280-287Crossref PubMed Scopus (25) Google Scholar Thus, in this system TGF-β appears not to be mainly produced by CD4+CD25+ regulatory T cells. It is known that all leukocyte types can secrete TGF-β. Of these lymphocyte populations, NK cells are emerging as an important origin of this kind of cytokine.16Horwitz DA Gray JD Ohtsuka K Role of NK cells and TGF-β in the regulation of T-cell-dependent antibody production in health and autoimmune disease.Microbes Infect. 1999; 1: 1305-1311Crossref PubMed Scopus (42) Google Scholar, 17Gray JD Hirokawa M Ohtsuka K Horwitz DA Generation of an inhibitory circuit involving CD8+ T cells. IL-2, and NK cell-derived TGF-β: contrasting effects of anti-CD2 and anti-CD3.J Immunol. 1998; 160: 2248-2254Crossref PubMed Google Scholar Hence, to determine whether NK cells are involved in anti-CD3 F(ab′)2-mediated diabetes reversion and TGF-β production, we observed the change of NK cells in quantity and quality after anti-CD3 F(ab′)2 treatment. The data showed that not only the proportion but also the number of NK cells in spleen and pancreatic lymph nodes did increase remarkably from treated mice compared with the control diabetic mice (Figure 3A and B). Interestingly, this effect was more significant on 12th week post-treatment, indicating these NK cells might play a role in maintaining long-lasting tolerance and glucose homeostasis in remitting mice. So, next we addressed the capacity of these NK cells to produce TGF-β. NK cells from mice 3 or 12 weeks after treatment remarkably secreted much more TGF-β than those from control age-matched mice (Figure 3, C and D). This phenomenon could also be seen in mice 20 weeks or much longer after treatment. As a result, anti-CD3 F(ab′)2 administration induces a population of TGF-β-producing NK cells, which may regulate peripheral immune balance and retard the attack to self islets by pathogenic cells. In addition, as IL-12 is a powerful stimulator for NK cell expansion,18Brunda MJ Interleukin-12.J Leukoc Biol. 1994; 55: 280-288Crossref PubMed Scopus (401) Google Scholar we investigated whether increased number of NK cells after anti-CD3 F(ab′)2 treatment was due to up-regulated expression of IL-12. Intriguingly, IL-12 production by pancreas lymph node lymphocytes of anti-CD3 F(ab′)2-treated mice was much lower than the untreated diabetic controls at different time points post-treatment (see supplemental Figure S1 at http://ajp.amjpathol.org), which indicates the increase of NK cell number is not attributed to enhanced expression of IL-12. On the other hand, recent reports showed a crucial role of membrane-bound TGF-β for the function of CD4+CD25+ regulatory T cells.19Xia Z Xu L Zhong W Wei J Li N Shao J Li Y Yu S Zhang Z Heme oxygenase-1 attenuates ovalbumin-induced airway inflammation by up-regulation of Foxp3 T-regulatory cells, interleukin-10, and membrane-bound transforming growth factor-β1.Am J Pathol. 2007; 171: 1904-1914Abstract Full Text Full Text PDF PubMed Scopus (77) Google Scholar, 20Ostroukhova M Qi Z Oriss TB Dixon-McCarthy B Ray P Ray A Treg-mediated immunosuppression involves activation of the Notch-HES1 axis by membrane-bound TGF-β.J Clin Invest. 2006; 116: 996-1004Crossref PubMed Scopus (150) Google Scholar, 21Ostroukhova M Segiun-Devaux C Oriss TB Dixon-McCarthy B Yang L Ameredes BT Corcoran TE Ray A Tolerance induced by inhaled antigen involves CD4+ T cells expressing membrane-bound TGF-β and FOXP3.J Clin Invest. 2004; 114: 28-38Crossref PubMed Scopus (250) Google Scholar To determine whether NK cells also express cell-bound TGF-β, we detected TGF-β expression on the surface of NK cells by fluorescence-activated cell sorting. Consequently, we did not find TGF-β expression on the NK cell surface (Figure 3E), even those producing large amounts of soluble TGF-β after anti-CD3 F(ab′)2 treatment. These data indicated that NK cells primarily produced soluble TGF-β, which mediated NK cell regulatory function. Another question to be clarified is whether anti-CD3 F(ab′)2 treatment led to NK cell activation. To address this, active status of NK cells was examined. Because NK cells express a diverse array of activating receptors, here we focused on NKG2D expression in NK cells by fluorescence-activated cell sorting, which is critical for function of mouse NK cells22Lanier LL Up on the tightrope: natural killer cell activation and inhibition.Nat Immunol. 2008; 9: 495-502Crossref PubMed Scopus (1206) Google Scholar and whose expression is impaired in NOD NK cells.23Ogasawara K Hamerman JA Hsin H Chikuma S Bour-Jordan H Chen T Pertel T Carnaud C Bluestone JA Lanier LL Impairment of NK cell function by NKG2D modulation in NOD mice.Immunity. 2003; 18: 41-51Abstract Full Text Full Text PDF PubMed Scopus (240) Google Scholar The result showed that NKG2D expression in NK cells from anti-CD3 F(ab′)2-treated mice was low and comparable with control counterparts (see supplemental Figure S2 at http://ajp.amjpathol.org). These data indicate that these NK cells with increased production of TGF-β are likely not activated. It is worthy to note that a previous report showed the presence of a population of cells co-expressing NK and DC markers (DX5 and CD11c) in naive mice. These cells exhibit regulatory function in some settings such as CD40L blockade to prevent autoimmune disease in the RIP-LCMV mouse model for virally induced type 1 diabetes.24Homann D Jahreis A Wolfe T Hughes A Coon B van Stipdonk MJ Prilliman KR Schoenberger SP von Herrath MG CD40L blockade prevents autoimmune diabetes by induction of bitype NK/DC regulatory cells.Immunity. 2002; 16: 403-415Abstract Full Text Full Text PDF PubMed Scopus (147) Google Scholar To uncover the virtual players, CD3−DX5+ population was further divided into CD11c+ and CD11c− subsets by magnetic sorting then the abilities of these two subsets (CD3−CD11c−DX5+ and CD3−CD11c+DX5+, respectively) to secrete TGF-β were tested. The results showed that soluble TGF-β after anti-CD3 F(ab′)2 treatment was mainly produced by CD3−CD11c−DX5+ population, that was NK cells, instead of CD3−CD11c+DX5+ population (see supplemental Figure S3A at http://ajp.amjpathol.org), indicating that key players in anti-CD3 F(ab′)2-mediated diabetes reversion and tolerance restoration were NK cells rather than DX5+ DC population. To further address the role of NK cells in anti-CD3 F(ab′)2 therapeutic effects, we depleted AsGM1-positive cells with specific antibodies and investigated the influence on anti-CD3 F(ab′)2 effects as well as TGF-β production. The AsGM1-positive cell elimination significantly abrogated diabetes remission following anti-CD3 F(ab′)2 therapy while treatment with isotype antibodies had no such effect (Figure 4A). Moreover, AsGM1-positive cell depletion also decreased TGF-β production of splenocytes from treated mice (Figure 4B), indicating in anti-CD3 F(ab′)2-treated remitting NOD mice a subset of NK cells produces immunoregulatory TGF-β thereby modulates overt autoimmunity to homeostasis. In addition, as described above, we also detected cell-bound TGF-β on the surface of both AsGM1-positive cell-depleted and non-depleted splenocytes. In agreement with the above data, we did not find TGF-β expression on the NK cell surface or the alteration of cell-bound TGF-β expression in splenocytes before and after anti-AsGM1 depletion (data not shown). In summary, these data support an immunoregulatory function of NK cells in anti-CD3 F(ab′)2-mediated diabetes reversion and tolerance restoration through producing soluble TGF-β. For antibody therapy, it is logical to contemplate that the resolution of the inflammatory infiltration is caused by modulation of β-cell autoantigen-reactive diabetogenic T cells. Thus, a plausible hypothesis postulates that the regulatory NK cells induced by antibody treatment maybe inhibit the proliferation of diabetogenic T cells and block diabetes progression. To test this premise, the splenic cells from diabetic mice were pooled and the proliferative response of these diabetogenic cells to antigen stimulation co-cultured with NK cells was assessed. Two critical autoantigens to islet β cells, glutamic acid decarboxylase, and insulin,25Kim J Richter W Aanstoot H-J Shi Y Fu Q Rajotte R Warnock G Baekkeskov S Differential expression of GAD65 and GAD67 in human, rat, and mouse pancreatic islets.Diabetes. 1993; 42: 1799-1808Crossref PubMed Google Scholar, 26Lernmark A Agardh CD Immunomodulation with human recombinant autoantigens.Trends Immunol. 2005; 26: 608-612Abstract Full Text Full Text PDF PubMed Scopus (22) Google Scholar as well as control antigens OVA were used. Splenocytes from diabetic mice actively cloned in response to glutamic acid decarboxylase or insulin incubation, showing their effector function to destroy β cells. When added recovered mice-derived NK cells to the culture, this proliferative responses were suppressed strongly, whereas addition of NK cells from control mice did not show such effects (Figure 5). Interestingly, treated NK cells just slightly inhibited responses of effector cells to OVA stimulation, indicating the inhibitory effects of NK cells are β cell antigen-specific, which is consistent with previous observations that anti-CD3 administration blocked β cell-reactive T cell activity specifically instead of renderin" @default.
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• W2066500966 date "2009-09-01" @default.
• W2066500966 modified "2023-10-18" @default.
• W2066500966 title "Natural Killer Cells Modulate Overt Autoimmunity to Homeostasis in Nonobese Diabetic Mice after Anti-CD3 F(ab′)2 Antibody Treatment through Secreting Transforming Growth Factor-β" @default.
• W2066500966 cites W1519232981 @default.
• W2066500966 cites W1556867369 @default.
• W2066500966 cites W1658220892 @default.
• W2066500966 cites W1898102248 @default.
• W2066500966 cites W1931140415 @default.
• W2066500966 cites W1933331438 @default.
• W2066500966 cites W1957860739 @default.
• W2066500966 cites W1969950463 @default.
• W2066500966 cites W1970959631 @default.
• W2066500966 cites W1980278108 @default.
• W2066500966 cites W1990242210 @default.
• W2066500966 cites W2002801575 @default.
• W2066500966 cites W2009389800 @default.
• W2066500966 cites W2017917253 @default.
• W2066500966 cites W2024275584 @default.
• W2066500966 cites W2030715890 @default.
• W2066500966 cites W2040355823 @default.
• W2066500966 cites W2041266914 @default.
• W2066500966 cites W2050793959 @default.
• W2066500966 cites W2057043713 @default.
• W2066500966 cites W2067067228 @default.
• W2066500966 cites W2087171945 @default.
• W2066500966 cites W2110949372 @default.
• W2066500966 cites W2113313714 @default.
• W2066500966 cites W2116383269 @default.
• W2066500966 cites W2117002902 @default.
• W2066500966 cites W2117113925 @default.
• W2066500966 cites W2118803877 @default.
• W2066500966 cites W2122412498 @default.
• W2066500966 cites W2126892029 @default.
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