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Keywords:

  • Macrophages;
  • Self-/non-self-discrimination;
  • Tolerance;
  • Vaccination

Abstract

  1. Top of page
  2. Abstract
  3. Conflict of interest
  4. References

A promising therapeutic approach for inducing tolerance in autoreactive T cells is the use of APCs such as DCs and macrophages. In this issue of the European Journal of Immunology, Zheng et al. [Eur. J. Immunol. 2013. 43: 219–227] study the concept of “tolerogenic adjuvants” to induce tolerance via vaccination. These authors have previously identified dexamethasone (Dex) as an effective “tolerogenic adjuvant” and, in this study, they have identified a population of peripheral macrophages that is enriched by Dex treatment and that mediates Dex's tolerogenic effect. In addition to performing a phenotypic characterization of this population, the authors noted an increase in serum levels of IL-10 and Treg cells after Dex treatment of mice. As discussed in this Commentary, by employing Dex as a tolerogenic adjuvant in the presence of relevant peptides, we may have a means of restoring specific immune tolerance in cases of autoimmune disease and allergy.

The pathogenesis of some autoimmune diseases such as multiple sclerosis and diabetes is thought to be mediated by autoreactive T cells that overcome the mechanisms of immune tolerance and inappropriately target self proteins resulting in injury [1, 2]. Current treatments for autoimmune diseases are not completely effective and can result in general immune suppression. There is thus a need to develop therapeutic methods that more selectively target the aberrant response of the disease-mediating autoreactive lymphocytes. Ideally, such methods would involve the restoration of immune tolerance to self proteins. A promising therapeutic target for inducing tolerance in autoreactive T cells is that of APCs, such as DCs and macrophages, since these cells not only activate T cells, but can also induce T-cell tolerance [3-5].

In this issue of the European Journal of Immunology, Zheng et al. [6] study the elegant concept of “tolerogenic adjuvants” to induce tolerance via vaccination with a specific peptide immunogen in the presence of an immunosuppressant. In contrast to traditional immunogenic adjuvants that enhance immunity (Table 1), tolerogenic adjuvants are used in a process of suppressed immunization to dampen antigen-specific immune responses [7]. This current study builds on this group's previous cutting edge work in 2008 in which they identified dexamethasone (Dex) as a true “tolerogenic adjuvant” [7]. Dex is a synthetic glucocorticoid steroid with well-known immunosuppressive activities. It is also well known for its ability to induce Treg cell-mediated immune suppression [8]. In their 2008 study, Zheng and colleagues utilized Dex as an adjuvant to induce tolerogenic APCs in vivo [7]. Other groups had previously demonstrated Dex's ability to induce a tolerogenic phenotype in APCs in vitro [9-11]; however, in vivo studies had been lacking and thus the results of Zheng and colleagues in 2008 [7] moved the concept of a tolerogenic adjuvant from a hypothetical idea to practical application. Mice were first sensitized to OVA using incomplete Freund's adjuvant, thus establishing a model of delayed-type hypersensitivity (DTH) [7]. These mice were subsequently immunized with OVA-derived, MHC class II-restricted peptide in the presence of Dex as a tolerogenic adjuvant [7]. A similar treatment regimen was used with a second autoimmune disease model in which NOD mice were vaccinated with a combination of Dex and insulin-derived, MHC class II-restricted peptide [7]. The use of Dex in combination with disease-specific peptide antigens during active immunization decreased the symptoms of both DTH and autoimmune diabetes [7]. These treatments induced a predictable expansion of antigen-specific CD4+CD25+Foxp3+ regulatory T (Treg) cell numbers and these cells were able to persist long term and to respond to antigenic rechallenge [7]. The clinically relevant outcome of this therapy was the long-term desensitization of mice to antigen, in this case either OVA or insulin-peptide [7]. Additionally, the authors demonstrated that Dex treatment altered the composition and function of APC populations in vivo [7]. Despite the provocative demonstration of Dex as a valid tolerogenic adjuvant, there remained a gap in understanding the precise mechanism by which Dex mediated its effects. In their current article, Zheng et al. [6] present new findings that specifically characterize the type of tolerogenic APCs induced by Dex in vivo and further dissect the mechanism of Dex-mediated tolerogenicity of APCs.

Table 1. Comparison of adjuvants for immunomodulation
 Immunogenic adjuvantsTolerogenic adjuvants
ExampleFreund's adjuvantDexamethasone
Impact on T cellsExpansion of cytotoxic T cellsExpansion of Treg-cell numbers
Impact on APCsPromotion of immunogenic APCsSelection of tolerogenic APCs
 (e.g. Maturation and/or activation of DCs)(e.g. CD11clo CD40lo macrophages)

Zheng et al. [6] identify a population of APCs in the periphery that are enriched by Dex treatment and mediate Dex's tolerogenic effect. In their 2008 paper, Zheng's group had characterized Dex-induced tolerogenic APCs as DCs based on the expression of CD11c. In this current paper, they delved deeper in their exploration of lineage markers on the tolerogenic APCs and were able to more carefully define the Dex-enriched cell population as CD11clo monocyte-derived macrophages. They found that Dex treatment depleted peripheral DCs [6], as has been reported by other groups [12, 13]. The novelty of the observations of Zheng et al. [6] lies in the fact that Dex specifically depletes nontolerogenic DCs, while selectively enriching tolerogenic APCs, macrophages in this case. The tolerogenic macrophages were characterized by low expression of both CD11c and CD40 as well as high expression of F4/80 and CD68 [6, 14]. Further investigation by Zheng et al. [6] revealed that the Dex-enriched population of CD11clo CD40lo macrophages displayed tolerogenic markers, namely, a decrease in MHC class II expression, lower levels of CD83 (a marker of monocyte maturation), and upregulation of CD68 (a marker for phagocytosis) in comparison with CD11clo cells in animals that did not receive Dex treatment [6]. As the authors state, these cells appear to be immature macrophages with increased phagocytic potential.

The functional capability of these Dex-selected CD11clo CD40lo macrophages was assessed using a DTH model with OVA peptide and Dex as a tolerogenic adjuvant [6]. When mice were rechallenged with antigen, the CD11clo CD40lo cells maintained low levels of MHC class II and, in addition, IL-10 levels were increased in the serum, while IL-12p70 levels fell [6] which is noteworthy since IL-10 expression is a hallmark of tolerogenic APCs, while IL-12 is characteristic of immunogenic APCs [8]. When macrophages were depleted in vivo, serum IL-10 levels decreased suggesting that the CD11clo CD40lo macrophages were the source of the cytokine [6]. Furthermore, the authors demonstrated that the Dex-selected CD11clo CD40lo cells were functionally tolerogenic in vivo [6]. In the presence of these macrophages, DTH symptoms were lessened, and antigen-specific Treg cell numbers expanded [6]. Depletion of Dex-selected CD11clo CD40lo cells reversed the ability of Dex to attenuate DTH, and Treg cell numbers no longer expanded [6]. It will be of interest to learn more about the properties of these tolerogenic macrophages and future studies should pay special attention to the dosage of Dex and the duration of treatment in order to optimize the most efficacious response in terms of enriching tolerogenic APCs. The ability to induce tolerance in an antigen-specific manner is a clinically desirable tool especially in fields such as autoimmune disease and allergy. Dex is already used clinically as a general immunosuppressant for the treatment of a variety of autoimmune diseases including chronic idiopathic thrombo-cytopenic purpura, systemic lupus erythematosus, and Graves’ disease [15-17]. As expected Dex treatment in these patients results in an enriched Treg-cell population. The tolerization approach indicated by the studies of Zheng et al. [6] will be most promising clinically if lower doses of Dex given over a shorter period prove effective.

A recent clinical report described the functional improvement of CD4+CD25+Treg cells after patients received dexamethasone for the treatment of autoimmune hyperthyroidism [15]. Treg cells are known to play a role in Graves’ disease based on studies using mouse models of this disease—Treg-cell depletion in such models increases susceptibility to disease induction [18, 19]. Mao et al. [20] introduced a new treatment approach for Graves’ disease using Dex in combination with methimazole and, using this approach, intrathyroid injection of Dex reduced the relapse rate in patients over a 2-year period. Such Dex treatment of Graves’ disease also improved Treg-cell function as well as decreasing the proportion of Th2 cells [15]. These clinical findings are quite interesting when examined in light of the new report by Zheng et al. [6], which demonstrates the capacity of Dex to support a tolerogenic phenotype in APCs. We would predict to see an enrichment of tolerogenic APCs, as well as a corresponding decrease in nontolerogenic APCs, following Dex treatment of Graves’ disease. While Mao et al.'s [20] studies did not focus on antigen specificity, their administration of Dex directly into the thyroid suggests that the immune response may have involved disease-specific antigen in the context of a tolerogenic adjuvant which may, in part, account for the reduction in relapse rate. This, and other similar clinical studies, could readily be extended to determine whether tolerogenicity of APCs is influenced by Dex when it is used as a tolergenic adjuvant in immunotherapy in the presence of disease-specific antigen.

Antigen-specific immunotherapy is also of special interest in the field of allergy and asthma. Van Overtvelt et al. [21] employed Dex in a similar fashion as Zheng et al. did in their current [6] and previous studies [7]. In an attempt to augment the efficacy of immunotherapy in an animal model of asthma, Van Overtvelt et al. [21] combined 1,25-dihydroxyvitamin D3 and dexamethasone as adjuvants in the presence of allergen and found that these adjuvants triggered IL-10 production as well as an increase in CD4+CD25+Foxp3+ Treg-cell numbers. Their results mirror the results of Zheng and colleagues’ 2008 study [7]; however, Van Overtvelt et al. [21] did not investigate the impact of Dex on APCs. As sublingual administration of an antigen is commonly used in allergy treatment and is somewhat unique in the sense that the target organ, the oral mucosa, favors tolerance induction [22], it would be interesting to revisit the use of Dex as an adjuvant in sublingual immunotherapy with the aim of examining the role of APCs in facilitating protection against asthma and allergy.

The study of factors capable of inducing tolerance in APCs is not only relevant to situations associated with autoimmune disease and allergy, but also to cancer studies. Tolerogenic APCs have been identified in tumors and may be involved in supporting tumor growth. Recently, Watkins et al. [23] identified a population of tumor-associated dendritic cells (TADCs) in both murine and human tumors that mediate the tolerization and suppressive function of tumor-specific CD8+ T cells. The authors found that these tolerogenic TADCs express the transcription factor FOXO3 and knocking down FOXO3 expression by siRNA abrogated the tolerogenic effect of the TADCs [23]. In such cases, understanding the detailed mechanisms by which APCs become tolerogenic is of use to understand how the cells are mediating an undesirable effect such as promoting tumor growth. Countering the tolerance-inducing factors, such as those identified by Zheng et al. [6], may provide further clues that could also be used to inhibit the tolerogenic pathways used by APCs to promote the growth and spread of cancer. Consideration should also be given to therapeutics other than Dex that might serve as tolerogenic adjuvants, particularly therapeutic agents with fewer undesirable side effects than Dex. Adverse effects resulting from Dex treatment can affect almost every system in the body including metabolic, gastrointestinal, musculoskeletal, and the CNS [24, 25]. As suggested by Zheng et al. [6] in this issue, as well as by others [9], rapamycin and vitamin D3 show promise in this arena.

To sum up, the clinically applicable message of the findings of Zheng et al. [6] is that administration of Dex in conjunction with antigen, such as insulin peptide in the case of diabetes, may result in more prolonged disease remission for patients. This approach, which is presently restricted to animal studies, warrants further exploration and exploitation but nonetheless Zheng et al. [6] have succeeded in showing that an “old drug” can be exploited therapeutically to perform useful “new tricks.”

Conflict of interest

  1. Top of page
  2. Abstract
  3. Conflict of interest
  4. References

The authors declare no financial or commercial conflict of interest.

References

  1. Top of page
  2. Abstract
  3. Conflict of interest
  4. References
Abbreviations
TADC

tumor-associated dendritic cell

Dex

dexamethasone

DTH

delayed-type hypersensitivity