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Logo of nihpaAbout Author manuscriptsSubmit a manuscriptNIH Public Access; Author Manuscript; Accepted for publication in peer reviewed journal;
J Immunol. Author manuscript; available in PMC Oct 15, 2011.
Published in final edited form as:
PMCID: PMC3132880

IL-17 Promotes Immune Privilege of Corneal Allografts


Corneal allograft rejection has been described as a Th1-mediated process involving IFN-γ production. However, it has been reported that corneal allograft rejection soars in IFN-γ−/− mice or mice treated with anti–IFN-γ mAb. Th17 is a recently described IL-17A–producing Th cell population that has been linked to renal and cardiac graft rejection, which was originally thought to be Th1-mediated. We tested the hypothesis that Th17 cells mediate corneal allograft rejection in an IL-17A–dependent fashion and unexpectedly found that depletion of IL-17A increased the incidence of rejection to 90%. We demonstrate that the exacerbated rejection following depletion of IL-17A did not result from a loss of cross-regulation of Th1 cells or exaggerated delayed-type hypersensitivity responses. Instead, inhibition of the Th1 or Th17 cell lineages promoted the emergence of a Th2 cell subset that independently mediated allograft rejection. These findings demonstrate that IL-17A is not required for corneal allograft rejection and may instead contribute to the immune privilege of corneal allografts.

Corneal allografts enjoy an immune privilege that is unrivaled in the field of transplantation. Every year in the United States alone, >30,000 corneal transplants are performed (http://www.restoresight.org/donation/statistics). The surgical procedure is routinely performed without histocompatibility matching and without use of systemic immunosuppressive drugs, yet corneal transplants experience a 90% success rate under the cover of topical steroids (1, 2). Nonetheless, 10% of corneal transplants undergo immune rejection, and use of corticosteroids carries the added risk of developing cataracts and glaucoma.

The precise immune mechanism underlying graft failure is incompletely understood. It is thought to involve an initial sensitization phase during which host APCs migrate into the graft and internalize soluble Ag derived from the donor’s corneal endothelium. Subsequently, the APCs can induce an oligoclonal T cell response via either the direct or indirect pathway of allorecognition. In the direct pathway, recipient T cells recognize MHC class II molecules expressed on donor APCs. In contrast, indirect allorecognition requires presentation of processed MHC or minor Ags by host APCs (3). Allograft rejection is believed to be mediated by allospecific CD4+ Th1 lymphocytes and is closely associated with the development of delayed-type hypersensitivity (DTH) to donor alloantigens and production of IFN-γ (4). However, recent studies have implicated additional CD4+ Th cell subsets in the process. For example, the incidence and tempo of corneal allograft rejection rises steeply in hosts with either allergic conjunctivitis or allergic asthma (5). In the allergic host, rejection is associated with Th2 alloantigen-specific immune responses involving the production of IL-4, IL-5, and IL-13 (5, 6).

The Th17 cell subset is a recently described IL-17A–producing CD4+ T cell population that has been implicated in the resistance to certain bacterial pathogens (7). IL-17A has also been shown to have several regulatory and protective effects. In mouse models of colitis and graft-versus-host disease, IL-17A has been shown to mitigate the severity of the disease by limiting Th1-associated inflammation (8, 9). Additionally, IL-17A has been shown to negatively regulate asthma via inhibition of dendritic cells and chemokine synthesis in sensitized hosts (10). Nonetheless, dysfunctional Th17 responses have been linked to the pathogenesis of several autoimmune diseases conventionally thought to be Th1 mediated (1113). In the transplantation setting, Th17 cells have been associated with lung and renal allograft rejection and have been shown to accelerate cardiac allograft rejection in T-bet–deficient hosts (14, 15). Based on the observation that Th1 cell responses are not necessary for corneal allograft rejection, we hypothesized that Th17 cells can also mediate corneal allograft rejection.

We report that the Th17 cell subset contributes to the ocular immune privilege and that the interplay between Th1, Th2, and Th17 cells determines the course of corneal allograft rejection.

Materials and Methods


Eight-to-ten-week-old female mice were used in all experiments. BALB/c (H-2d) and C57BL/6 (H-2b) mice were purchased from Taconic Farms (Germantown, NY), and BALB/c nude mice were obtained from the National Cancer Institute (Frederick, MD). STAT6 knockout (KO) mice were purchased from The Jackson Laboratories (Bar Harbor, ME). All experimental animals were treated in accordance with the Association for Research in Vision and Ophthalmology ARVO Statement for the Use of Animals in Ophthalmic and Visual Research (ARVO Animal Policy).

Orthotopic corneal allograft and clinical evaluation of grafted corneas

Naive C57BL/6 corneas were grafted onto the right eye of BALB/c mice as described previously (16). Corneal grafts were scored for their graft opacity, neovascularization, and edema twice per week (16). Briefly, degree of opacification ranged between 0 and 4+; with 0, clear; 1+, minimal superficial opacity; 2+, mild deep stromal opacity with pupil margin and iris visible; 3+, moderate stromal opacity with pupil margin visible, but iris structure obscured; and 4+, complete opacity, with pupil and iris totally obscured. Corneal grafts were considered rejected upon two successive scores of 3+.

Cytokine depletion protocol

Anti–IFN-γ hybridoma (R4-6A2; catalog number HB-170) was purchased from American Type Culture Collection (Rockville, MD). Anti–IL-17A monoclonal and polyclonal Abs were produced by the University of Texas Southwestern Hybridoma Facility as described previously (17). mAbs were isolated from hybridoma cultures and affinity purified. Rat IgG was purchased from Sigma-Aldrich (St. Louis, MO). BALB/c mice were injected intraperitoneally with 500 μg Ab daily from day −4 to day −2, and corneal transplants were grafted on day 0. Biweekly injections of the Abs were continued up to day 60.

DTH assays

An ear swelling assay was used to measure DTH responses to C57BL/6 alloantigens as described previously (5). A cell suspension of 4 × 106 mitomycin C-treated C57BL/6 splenocytes in 20 μl HBSS was injected into the right-ear pinna of BALB/c mice. The left-ear pinna received 20 μl HBSS without cells and served as a negative control. Results were expressed according to the following: specific ear swelling = (24 h measurement − 0 h measurement) for experimental ear − (24 h measurement − 0 h measurement) for negative control ear.

Preparation of APCs

APCs were isolated from spleen cells of naive C57BL/6 mice. Briefly, cells were incubated with NH4Cl erythrocyte lysis solution, washed, and resuspended at 2 × 106 cells/ml HBSS with 400 μg/ml mitomycin C. The cell suspension was incubated at 37°C for 1 h and washed three times with HBSS and was used as a source of APCs in direct MLRs. For the indirect MLR, C57BL/6 cell lysate was initially generated by resuspending C57BL/6 splenocytes at 3 × 107 cells/ml HBSS and sonicating the suspension with ten 1-s pulsations. Lysates were frozen at −80°C for 10 min and thawed at 37°C for 5 min for two cycles. BALB/c APCs were isolated by incubating the cell suspension of splenocytes onto two 100-mm Primaria plates (Franklin Lakes, NJ) (5 ml each plate) at 37°C for 1 h. Nonadherent cells were removed by vigorous washing. Adherent APCs were cultured in a 100-mm Primaria plate containing 4 ml complete RPMI 1640 medium supplemented with 10% FBS and pulsed with the C57BL/6 cell lysate (1 ml). Cell cultures were incubated at 37°C overnight.

MLRs and cytokine ELISA

Spleen cells were harvested from BALB/c mice 4–7 d after rejection of the C57BL/6 corneal allografts or at 3 wk posttransplantation in acceptors. CD4+ T cells were enriched by positive selection using rat anti-mouse CD4-conjugated magnetic microbeads (Miltenyi Biotec, Auburn, CA). Purified CD4+ T cells were incubated at 1 × 106 per well with respective APCs at a 1:1 ratio for 5 d at 37°C in 2 ml complete RPMI 1640. ELISAs for IL-4, IL-5, IL-13, IL-17A, IFN-γ, and TNF-α were performed on culture supernatants according to the manufacturer’s instructions (R&D Systems, Minneapolis, MN).


Eyes from corneal allograft rejector mice were enucleated after two successive scores of 3+ and fixed in 10% formalin and were processed for histology. Sections (4 μm) of paraffin-embedded tissue were labeled with mAb against T1/ST2 (DJ8; MD Biosciences) to detect Th2 cells using the Vectastain Elite ABC system (Vector Laboratories, Burlingame, CA). Ab specificity was validated using a rat IgG1 isotype control (BD Pharmingen, St. Paul, MN/eBioscience, San Diego, CA).

Adoptive transfer of Th2 CD4+ T cells to nude mice

Spleen cell suspensions were obtained from anti–IL-17A–treated BALB/c recipients 4–7 d after rejection of C57BL/6 corneal allografts. CD4+ T cell enrichment was carried out using the magnetic microbead system as described earlier. Each nude mouse received an adoptive transfer of one-donor equivalent of the CD4+ T spleen-cell population intravenously (10 × 106 to 15 × 106 cells/recipient). Nude mice were grafted with C57BL/6 corneal allografts within 24 h of the adoptive transfer of CD4+ T cells.

Statistical analysis

The log-rank test was used for statistical analysis of the differences in the tempo of corneal graft rejection using Kaplan-Meier survival curves (18). Comparisons yielding p < 0.05 were considered significantly different.


Inhibition of the Th1 and/or Th17 cytokines abrogates immune privilege of corneal allografts

To test the hypothesis that elimination of the Th1 pathway promotes graft survival, we treated BALB/c mice systemically with either anti–IFN-γ mAb or an IgG isotype control Ab given intraperitoneally twice per week before and after the application of orthotopic C57BL/6 corneal allografts. The isotype control-treated BALB/c hosts rejected 50% of their C57BL/6 corneal allografts with a mean rejection time (MRT) of 35.2 ± 8.0 d (Fig. 1). By contrast, depletion of IFN-γ in BALB/c hosts resulted in a 90% incidence of rejection and MRT of 22.2 ± 7.3 d (Fig. 1). The median survival time (MST) for the anti–IFN-γ–treated group was significantly reduced (p = 0.014) compared with that of the IgG isotype control (MST = 23.5 d and 52 d, respectively). This suggests that conventional Th1 cells are not necessary for corneal allograft rejection and implies that either Th2 or Th17 cells are capable of mediating corneal allograft rejection. Accordingly, the role of Th17 cells was evaluated. BALB/c mice were treated with either monoclonal or polyclonal Abs specific for IL-17A. In multiple experiments, we observed an increased tempo and incidence of corneal allograft rejection in BALB/c mice treated with anti–IL-17A (Fig. 2A). Mice treated with either monoclonal or polyclonal anti–IL-17A Abs rejected 90% of their corneal allografts, with MRTs of 26 ± 7.9 d and 24.7 ± 12.8 d and MSTs of 24 and 22.5 d, respectively. The rates of corneal allograft rejection between the rat IgG isotype control and anti–IL-17A–treated groups were significantly different (p < 0.05).

C57BL/6 corneal allograft survival in BALB/c mice treated with either anti–IFN-γ or a rat IgG isotype control Ab. BALB/c mice treated with anti–IFN-γ rejected 90% of the C57BL/6 corneal allografts, which had an MST of 23.5 ...
C57BL/6 corneal allograft survival in BALB/c mice treated with anti–IL-17A, anti–IFN-γ, or a rat IgG isotype control Ab. A, C57BL/6 corneal allografts underwent rejection in 50% of hosts treated with the isotype control IgG (n ...

As a cross-regulation between Th17 and Th1 cell subsets has been suggested (9, 19), we tested the hypothesis that elimination of both Th1 and Th17 cytokines would enhance graft survival. BALB/c mice were simultaneously treated with anti–IL-17A and anti–IFN-γ mAbs prior to and after the application of corneal allografts. Treatment with both Abs did not prevent allograft rejection but instead resulted in a 90% incidence of rejection with an MRT of 30 ± 11 d and an MST of 26 d (Fig. 2B). The tempo of rejection was significantly different compared with that of the rat IgG isotype Ab-treated group (p = 0.04). Importantly, neither anti–IFN-γ nor anti–IL-17A Ab treatment affected the survival of syngeneic BALB/c corneal grafts (data not shown). To our knowledge, these results demonstrate that elimination of the signature cytokines for Th1 and Th17 T cell subsets abolishes ocular immune privilege and exacerbates corneal allograft rejection.

Depletion of Th17 cytokine does not exaggerate DTH responses

Corneal allograft rejection is closely correlated with the development of DTH responses to donor alloantigens (2024). Based on the earlier observation that depletion of IL-17A exacerbated the incidence and tempo of corneal allograft rejection, we performed additional experiments to address the possibility that mice treated with anti–IL-17A might develop exaggerated DTH responses to donor alloantigens. In these experiments, mice were treated with either a rat IgG isotype control or anti–IL-17A on days −4, −2, and twice per week during the course of the experiment. BALB/c mice were immunized subcutaneously (s.c.) with C57BL/6 splenocytes on day 0 and challenged with mitomycin C-treated C57BL/6 splenocytes in the ear pinnae 14 d later. Mice treated with anti–IL-17A did not display an exaggerated DTH response (Fig. 3A). To confirm that anti–IL-17A treatment did not promote an exaggerated DTH response in corneal allograft recipients, rat IgG isotype control and anti–IL-17A–treated BALB/c corneal allograft rejector mice were challenged with mitomycin C-treated C57BL/6 splenocytes in the ear pinnae 14 d postallograft rejection. As in s.c. immunized mice, corneal allograft rejector mice treated with anti–IL-17A did not develop DTH responses that were any greater than those of similar mice treated with the isotype control IgG (Fig. 3B).

Anti–IL-17A treatment does not exacerbate DTH. A, BALB/c animals were treated with either anti–IL-17A or rat IgG isotype control on days −4, −2, and twice per week over 2 wk after s.c. immunization with C57BL/6 splenocytes ...

Depletion of IL-17A and IFN-γ promotes emergence of Th2 alloimmune responses

In nonmanipulated hosts, corneal allograft rejection was characterized by the production of the Th1 cytokine IFN-γ (Fig. 4A). CD4+ T cell production of IL-4 and IL-5 was barely detectable, whereas moderate levels of IL-13 and small quantities of IL-17A and TNF-α were found. To confirm that disabling the Th1 subset with anti–IFN-γ did not exacerbate rejection via the Th17 lineage, we evaluated the cytokine profile of CD4+ T cells that were collected from anti–IFN-γ–treated graft rejector mice and stimulated in vitro with C57BL/6 alloantigens in an MLR. The cytokine profile of the allospecific CD4+ T cells indicated a preferential production of Th2 cytokines IL-4, IL-5, and IL-13 and negligible expression of IL-17A, IFN-γ, and TNF-α (Fig. 4B). Mice treated with anti–IL-17A showed a similar cytokine profile, implicating Th2 cells in corneal allograft rejection in hosts lacking IL-17 (Fig. 4C). Although IL-17A is known to cross-regulate IFN-γ, in vivo treatment with anti–IL-17A Abs did not result in an increased production of IFN-γ (Fig. 4C). Thus, disabling the Th1 and Th17 alloimmune responses favors the expression of Th2-mediated corneal allograft rejection. Collectively, these results support the notion that IL-17A and IFN-γ can independently cross-regulate the activity and cytokine secretion of Th2 cells.

Th1, Th2, and Th17 cytokine production by CD4+ spleen cells from corneal allograft rejector mice. CD4+ spleen cells were isolated from BALB/c mice 4–7 d after their rejection of C57BL/6 corneal allografts. CD4+ spleen cells were stimulated with ...

Based on the Th2 cytokine profiles observed from the MLRs, we hypothesized that the rejected corneas of anti–IFN-γ–treated and/or anti–IL-17A–treated BALB/c corneal allograft rejector mice would display a predominant infiltration of Th2 cells and eosinophils into the graft rejection site. To assess infiltration of Th2 cells within the rejection site, we used Abs specific for T1/ST2, which is expressed on Th2 cells but not Th1 cells (25). Mononuclear cells infiltrating the corneas of anti–IL-17A–treated animals stained positively for the T1/ST2 Ag (Fig. 4DG), and no significant eosinophilic infiltrate was detected at the rejection site (data not shown).

The Th2 pathway is sufficient to mediate corneal graft rejection

The results up to this point suggested that the exacerbation of corneal allograft rejection associated with depletion of either IL-17A or IFN-γ was the result of a Th2 alloimmune response. To explore whether Th2 cells could independently cause graft rejection, we collected CD4+ T cells from anti–IL-17A–treated mice that had rejected their corneal allografts (MRT = 16.7 ± 4.7 d; MST = 13 d) and transferred them into naive BALB/c nude mice. The transferred CD4+ T cells were predominantly of the Th2 phenotype as confirmed by their preferential production of Th2 cytokines when confronted with C57BL/6 alloantigens in both indirect and direct MLRs (Fig. 5A). Nude mice that received adoptively transferred CD4+ Th2 cells rejected 100% of their C57BL/6 corneal allografts with an MRT of 23.6 ± 9.2 d and an MST of 17 d (Fig. 5B). Immunohistochemical analysis of the rejected corneas revealed a significant infiltration of T1/ST2+ mononuclear cells (Fig. 5C, 5D). These results suggest that anti–IL-17A treatment elicits an immune deviation that favors the emergence of allospecific Th2 cells that are sufficient to mediate corneal allograft rejection. With this in mind, we examined the fate of corneal allografts transplanted into STAT6 KO mice, which do not generate IL-4–mediated functions including Th2 cell differentiation (26, 27). Accordingly, STAT6 KO mice on a BALB/c background were treated with a combination of anti–IFN-γ and anti–IL-17A mAbs or were untreated and then challenged with C57BL/6 corneal allografts. For comparison, wild-type (WT) BALB/c mice were treated with either anti–IFN-γ or anti–IL-17A mAbs and then challenged with C57BL/6 corneal allografts. As expected, WT mice treated with either anti–IFN-γ or anti–IL-17A mAbs rejected 90% of their corneal allografts. By contrast, only one of the eight STAT6 KO mice treated with both anti–IFN-γ and anti–IL-17A rejected their C57BL/6 corneal allograft (Fig. 6). STAT6 KO mice grafted with C57BL/6 corneal allografts rejected 50% of their corneal allografts. These results support the hypothesis that blockade of Th17 and Th1 pathways favors the emergence of a Th2-mediated form of immune rejection of corneal allografts. Moreover, simultaneous blockade of Th1, Th2, and Th17 pathways inhibits the CD4+ T cell-mediated rejection of corneal allografts.

Corneal allograft survival in anti–IL-17A–treated nude BALB/c mice after adoptive transfer of CD4+ T splenocytes from corneal allograft rejector BALB/c mice treated with anti–IL-17A. A, Cell supernatant cytokine profiles of CD4 ...
Effect of anti–IL-17A and/or anti–IFN-γ Ab treatment on corneal allograft survival in Th2-impaired hosts. STAT6 KO BALB/c mice were treated with anti–IL-17A and anti–IFN-γ mAbs and transplanted with C57BL/6 ...

Corneal allograft survival correlates with CD4+ T cell expression of IL-17A

Our initial hypothesis proposed that Th17 T cells mediated corneal allograft rejection. However, the weight of evidence shown here indicates the opposite and suggests that IL-17 may contribute to the immune privilege of corneal allografts.

To test the hypothesis that IL-17A and IFN-γ production correlates with corneal allograft survival, CD4+ T cells were isolated from corneal allograft acceptors and were stimulated in vitro with C57BL/6 alloantigens, and cytokine production was measured by ELISA. Unlike CD4+ T cells from anti–IFN-γ–treated or anti–IL-17A–treated mice, which preferentially produce IL-4, IL-5, and IL-13 and reject 90% of their corneal allografts, CD4+ T cells from untreated mice that had accepted their corneal allografts produced large amounts of IL-17A and IFN-γ and moderate levels of IL-13 and TNF-α (Fig. 7).

Cytokine production by CD4+ spleen cells from corneal allograft acceptor BALB/c mice. CD4+ spleen cells were isolated from BALB/c mice bearing clear C57BL/6 corneal allografts on day 21 post-transplantation and were stimulated in vitro with C57BL/6 alloantigens ...


Corneal transplantation is arguably one of most successful forms of solid organ transplantation performed in humans. The high success rate of keratoplasty is attributable to the immune privilege of the cornea and rests on factors intrinsic to the tissue and the ocular environment (2, 28). When the delicate balance required for the maintenance of immune privilege is disrupted, graft rejection ensues by an immune response classically thought to be mediated by Th1 cells. However, in contrast with this prevailing paradigm, our attempt to inhibit the Th1 lineage by depletion of its signature cytokine, IFN-γ, abolished the immune privilege of corneal allografts and exacerbated rejection. The observation that the Th1 subset was not required for graft rejection led us to examine the role of Th17 T cells via its effector cytokine, IL-17A, in corneal immune privilege.

Our findings demonstrate that the Th17 T cell subset is not necessary for allograft rejection and are in keeping with a recent report by Yamada and coworkers (29) who reported that MHC-matched corneal allografts from the 129 mouse strain underwent rejection in 100% of C57BL/6 IL-17 KO or IFN-γ KO mice. However, because 100% of the 129 mouse strain corneal allografts underwent rejection in WT C57BL/6 mice, it was not possible to discern an effect of either IL-17 or IFN-γ on immune privilege of corneal allografts. A recent report by Chen and coworkers (30) also demonstrated that 100% of the C57BL/6 corneal allografts underwent rejection in IL-17 KO BALB/c mice. However, unlike our findings, Chen et al. noted that although 100% of the corneal allografts underwent rejection in IL-17–deficient hosts, the MST was significantly prolonged. On first blush, one might wonder if the in vivo Ab treatment used in the current study did not completely disable the IL-17– or IFN-γ–dependent immune processes. However, it should be noted that we and others have shown that either IFN-γ KO mice or WT mice treated with anti–IFN-γ Ab reject 100% of their fully allogeneic corneal allografts, suggesting that the anti–IFN-γ Ab treatment in WT mice recapitulated the effect of IFN-γ gene deletion (29). The recent study by Chen et al. (30) suggested that IL-17 was produced during the early stage of corneal allograft rejection and that deletion of the IL-17 gene delayed but did not prevent the rejection of corneal allografts. Thus, deletion of the IL-17 gene (29) or in vivo treatment with anti–IL-17 (current findings) fails to reduce the incidence of corneal allograft rejection. If treatment with monoclonal anti–IL-17 Ab produces only a partial diminution of IL-17A, one would expect that corneal allograft survival would be unaffected or perhaps delayed in such hosts. However, neither occurred, which further supports the notion that IL-17A is necessary for the immune privilege of corneal allografts that occurs in WT BALB/c mice. Unlike other mouse strains, such as BALB/c, corneal allografts do not enjoy immune privilege in C57BL/6 hosts, which have an exceptionally high incidence of corneal graft rejection that is 100% in most cases. By using BALB/c hosts, we were able to reveal that immune privilege is associated with IL-17A. Neutralization of IL-17 robbed the corneal allografts of their immune privilege. Moreover, IL-17A appeared to be preferentially expressed by CD4+ T cells from mice with surviving allografts. Our observations add support to the growing body of evidence suggesting a protective role of IL-17A in some immune-mediated diseases. In T cell-dependent colitis and graft-versus-host disease, IL-17A appears to limit Th1-associated inflammation and differentiation (8, 9). However, our observations suggest that a completely distinct regulatory mechanism is involved in corneal transplantation. It appears that the increased incidence of rejection is not due to a loss of Th1 cross-regulation or an exaggeration of DTH. The cytokine and the immunohistological analyses suggest that anti–IL-17A treatment tilts the alloimmune response toward a Th2 pathway. Adoptive transfer of CD4+ T cells from anti–IL-17A–treated hosts to nude mice culminates in accelerated corneal allograft rejection and the appearance of a putative Th2 cell infiltrate in the rejected corneal allografts, which is further evidence that anti–IL-17A treatment promotes preferential emergence of Th2 alloimmunity and Th2-mediated corneal allograft rejection.

Interestingly, the Th2-mediated graft rejection observed in IL-17A–depleted hosts appears to be independent of an eosinophilic infiltrate into the rejecting corneal allografts. Although eosinophils are often associated with Th2-based inflammation, recent studies indicate that hosts with Th2-mediated airway hyperreactivity have a similar exacerbation of corneal allograft rejection, which is associated with a Th2 cytokine profile, yet rejection occurs in the absence of infiltrating eosinophils (5).

It appears that the cytokine signature of the CD4+ T cells from grafted mice reflects the mechanism of allograft rejection. In untreated WT mice, the Th1 arm of the alloimmune response predominates and mediates corneal allograft rejection in ~50% of the hosts. Corneal allografts in the remaining 50% of WT hosts enjoy immune privilege, which correlates with concomitant expression of IFN-γ and IL-17A. It should also be noted that although the CD4+ T cells from acceptor mice also produced IFN-γ, the amount of cytokine produced in acceptor mice was higher than that produced by isotype IgG-treated mice that rejected their corneal allografts. Additional experiments are needed to elucidate the role of IL-17A in graft survival. Our report is not the first to suggest that IL-17 might mitigate Th2-based inflammation. Using a murine model of allergic asthma, Schnyder-Candrian et al. (10) found that in vivo neutralization of IL-17 exacerbated allergic asthma, whereas administration of exogenous IL-17 reduced pulmonary eosinophil recruitment and bronchial hyperreactivity. Studies on human peripheral blood have revealed the presence of IL-17–producing regulatory T cells (31, 32). We have determined that IL-17A is required for the generation of CD4+CD25+ Foxp3+ regulatory T cells (Tregs), which suppress CD4+ T cells in a contact-dependent manner and are required for corneal allograft survival (K. Cunnusamy, P.W. Chen, and J.Y. Niederkorn, manuscript in preparation). It has been reported that activation of the Tregs appears to be dependent on Th1 cytokines such as IFN-γ, and their function appears to be stifled in the presence of Th2 cytokines (33, 34). Additional studies are needed to determine if IL-17A can potentiate Tregs. It has also been reported that IL-17A acts synergistically with TGF-β to upregulate antiapoptotic molecules such as Bcl-XL and Bcl-2 in mouse mammary carcinomas (35). Based on the fact that the donor corneal endothelium is generously bathed with TGF-β from the aqueous humor, it would be interesting to investigate whether IL-17A might be acting in situ to enhance Treg activity and to protect the corneal endothelial cells from immune-mediated apoptosis.

In summary, our study demonstrates that IL-17A is required for corneal allograft survival and is preferentially expressed by CD4+ T cells from mice with surviving allografts. Our data also indicate that neither the Th1 nor the Th17 subset is required for allograft rejection and that inhibition of these lineages promotes emergence of a Th2 subset that can independently mediate allograft rejection.


This work was supported by National Institutes of Health Grants EY 007641 and EY020799 and by an unrestricted grant from Research to Prevent Blindness.

Abbreviations used in this paper

delayed-type hypersensitivity
mean rejection time
median survival time
regulatory T cell



The authors have no financial conflicts of interest.


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