IFN-γ induced by IL-12 administration prevents diabetes by inhibiting pathogenic IL-17 production in NOD mice

Abstract

Interleukin 12 (IL-12) is a pivotal Th1-associated cytokine and a potent immunoregulatory molecule. However, the role of IL-12 in inducing immune tolerance that prevents insulitis and inhibits type 1 diabetes (T1D) remains unknown. The aim of this study was to investigate whether intermittent administration of IL-12 could prevent the development of T1D in nonobese diabetic (NOD) mice. We examined whether IL-12 treatment prevented diabetes by injecting different doses of IL-12 into NOD mice and compared the incidence of diabetes and insulitis in NOD mice with the incidence in control mice. Furthermore, we investigated the potential mechanisms of IL-12-mediated prevention of diabetes and insulitis. The expression of pro-inflammatory and immunoregulatory cytokines was measured before and following therapeutic administration of IL-12 in NOD mice. Our data demonstrated that both the absolute number and the function of DCs were impaired in NOD mice and that the levels of the Th17-associated pro-inflammatory cytokines, IL-1β, IL-6 and IL-23, were elevated in NOD mice compared with age-matched BALB/c and C57BL/6 mice. However, treatment of NOD mice with IL-12 suppressed insulitis and increased the number of healthy islets, and the levels of IL-17, IL-1β, IL-6 and IL-23 were significantly decreased. Moreover, IL-12 treatment of NOD mice induced the secretion of IFN-γ, a potent inhibitor of Th17 cells. These data indicated that intermittent administration of IL-12 prevented diabetes by inducing IFN-γ, suppressing the pathogenic IL-17-producing cells and reducing the expression of Th17-associated pro-inflammatory cytokines. Our results suggest a promising strategy for the treatment of human T1D and other Th17 cell-mediated autoimmune diseases.

Introduction

Type 1 diabetes (T1D) is an autoimmune disease thought to be caused by autoantigen-reactive T lymphocytes that mediate the destruction of insulin-producing β-cells located in pancreatic islets, eventually resulting in β cell loss, insulin deficiency, and hyperglycemia [1]. The nonobese diabetic (NOD) mouse spontaneously develops insulin-dependent diabetes that strongly resembles human T1D [2], [3]. Long-term administration of insulin in appropriate doses is necessary to manage the blood glucose levels in T1D patients. However, use of exogenous insulin cannot precisely match endogenous insulin secretion, and this often leads to the risk of hypoglycemia and other severe complications [4]. The events that initiate T1D and the precise mechanisms of pancreatic β cell destruction are incompletely understood. Therefore, safe and effective therapies for T1D are urgently needed.

DCs are professional antigen-presenting cells that initiate both innate and adaptive immunity [5]. DCs have the ability to produce large amounts of IL-12 and induce T cell maturation as well as Th1 responses, and these functions have been demonstrated to be abnormal in both humans with T1D [6], [7] and NOD mice [8]. Hence, modulation of DC biology with the purpose of reshaping the repertoire of T cells may be an attractive therapeutic option for the treatment of T1D.

Increasing evidence from NOD mouse and human T1D studies suggests that Th17 cells play a crucial role in the pathogenesis of autoimmune diabetes. Several studies have shown an increase in the number of IL-17-producing cells and the secretion of IL-17 in NOD mice [9], [10] as well as in the peripheral blood of patients with T1D [11], [12]. However, the mechanism behind this increase and its relationship to the pathogenesis of T1D remain obscure. Substantial evidence has indicated that IFN-γ plays a protective role in the experimental autoimmune encephalitis mouse model [13], [14]. Here, mice lacking IFN-γ develop severe autoimmune disease compared with wild-type mice, and this is attributed to the inhibitory activity of IFN-γ against Th17 cells [15], [16], [17]. A similar effect of IFN-γ on the inhibition of IL-17 production has been reported in autoimmune diabetes [9]. However, the potent inducer of IFN-γ, IL-12, has been shown to be impaired in diabetes patients.

IL-12 is an immunoregulatory cytokine that promotes cell-mediated immunity [18] and is produced mainly by activated antigen-presenting cells [19]. It has been demonstrated that IL-12 plays a particularly important role in antitumor immunity [20], [21], [22]. Results from mouse models of intracellular protozoan, fungal and bacterial infections have indicated that IL-12 has a key role in protection against pathogens [23], [24], [25]. The role of IL-12 in autoimmunity is attracting increased attention. Previous studies have shown that IL-12 administration induces Th1 cells and accelerates autoimmune diabetes [26]. Consistent with these studies, it has been shown that daily administration of IL-12 to NOD mice induces a rapid onset of T1D in 100% of treated mice [27]. In addition, recent study revealed an IL-12 specific antibody protected transplanted islets from inflammatory damage [28]. However, another study showed that intermittent administration of IL-12 markedly reduced the incidence of diabetes [29]. Moreover, IL-12 treatment can directly induce high levels of IFN-γ in the circulation. Taken together, the role of IL-12 is controversial, as it has been shown to have both disease-promoting and disease-protective roles in autoimmune diabetes. The reason for these opposing roles of IL-12 is unclear, but administration of IL-12 likely affects systemic immune regulation.

In the current study, we found that both the absolute number and the function of DCs were impaired in NOD mice and that the levels of the Th17-associated pro-inflammatory cytokines, IL-1β, IL-6 and IL-23, were elevated in NOD mice. We showed that the intermittent administration of IL-12 to NOD mice suppressed insulitis and increased the number of healthy islets. Finally, we demonstrated that the IFN-γ induced by IL-12 administration prevented diabetes through a mechanism of inhibition of pathogenic IL-17 production in NOD mice.