IL-17A has been implicated in the pathogenesis of many autoimmune diseases, especially in MS (Mills, 2022). The main sources of IL-17A, TH 17 cells and γδ T cells contribute pivotally to MS and EAE (Cua et al., 2003; Cua & Tato, 2010; Langrish et al., 2005; McGinley et al., 2018). Nevertheless, IL-17A alone does not vitally contribute to neuroinflammation as we demonstrated earlier, that chronic IL-17A expression in the CNS neither induces demyelination nor leukocyte infiltration (Zimmermann et al., 2013). Ex vivo stimulation of microglia with Th17 cytokines does not influence their activation state (Prajeeth et al., 2014). On the other hand, however, we could show that chronic IL-17A stimulation in the context of other neuroinflammatory diseases, such as a LPS-sepsis model or cuprizone-induced demyelination, enhances neuroinflammatory responses (Zimmermann et al., 2018). In GF/IL17 mice, toxic demyelination was accelerated and synthesis of myelin proteins was reduced during cuprizone exposure.
The pathogenic role of IL-17A during EAE has been discussed intensively in the literature. Whereas a study showed that overexpression of IL-17A in CD4 + and CD8 + T cells does not enhance disease (Haak et al., 2009) others clearly demonstrated the contribution of IL-17A to EAE pathogenesis. Blockade of IL-17A attenuated EAE disease course as demonstrated in several studies and blood brain barrier (BBB) integrity is disrupted by IL-17A via reduced expression of tight junction related genes (Mardiguian et al., 2013; McGinley et al., 2020; Setiadi et al., 2019). Interestingly, T cells do not need to synthesize IL-17 in the CNS to achieve their encephalithogenic capacity. Gut targeted production of IL-17 or reconstitution of a wild-type like microbiota in mice with IL-17A/F deficient TH cells is sufficient to reestablish their susceptibility to EAE again (Regen et al., 2021). Another recent paper links encephalithogenicity of TH 17 cells with the microbial composition of the intestine. Homeostatic SLAMF6 + TH 17 cell populations are converted to pathogenic CXCR6 + TH 17 cells in intestinal tissues, which migrate to the CNS during EAE (Schnell et al., 2021). This is consistent to our findings, as we clearly demonstrated effects of this cytokine especially at early time points of EAE. The first events during EAE are T-cell priming and thereafter disruption of BBB and trafficking of autoreactive T-cells into the CNS (Sospedra & Martin, 2016). Unfortunately, we were not able to differentiate at which of these early points IL-17 synthesis exactly mediates its effect. Nevertheless, we were able to show that chronic IL-17A expression resulted in accelerated EAE onset and higher peak disease scores in classically induced EAE. Remission after peak disease was unchanged in GF/IL17 and WT mice, as indicated by the parallel clinical course of the two groups. We speculate that in our model, local IL-17 synthesis mediates a direct CNS-specific effect and does not systemically affect T cell priming. Although priming is IL-17-mediated in the gut, it is also dependent on the gut microbial flora, which is influenced by IL-17 (Regen et al., 2021). A brain microbiome has yet not been demonstrated and we were able to rule out IL-17 secretion from the brain into the plasma (Bedarf et al., 2021). General, well described effector functions of IL-17A in autoimmune diseases are induction of inflammatory mediators from keratinocytes and fibroblasts (Mills, 2011). Furthermore, IL-17A is believed to disrupt BBB tight junctions and promote central nervous system inflammation through CD4 + lymphocyte recruitment (Kebir et al., 2007). Our results add further evidence to the hypothesis that IL-17A plays a crucial role, especially in the early phase of EAE. Genetically IL17-receptor deficient mice are resistant to the induction of MOG EAE. However, neutralization of IL-17A during the effector phase no longer affects disease progression (McGinley et al., 2020). The same work found high expression levels of the IL-17A receptor IL-17RA on neutrophils and Ly6C+-inflammatory monocytes early in EAE. Genetic deficiency of IL17 in turn lead to ameliorated EAE course and reduced numbers of these cell types in the spleen after EAE induction. Interestingly, we showed in our work that neutrophils and Ly6C+-inflammatory monocytes are recruited to the CNS at a particularly early stage by local IL-17A synthesis, and are thus clearly target cell types. Also in Cuprizone induced demyelination we were able to demonstrate the early recruitment of neutrophils into the CNS by local IL-17A production (Zimmermann et al., 2018).
Local IL-17A synthesis in the CNS was not only able to accelerate the disease development of EAE, but also to facilitate the induction of EAE with a suboptimal immunization protocol without addition of the co-adjuvant PTX. Though immunostimulatory effects of PTX are widely used in EAE and other models of autoimmune inflammatory diseases for more than half a century, the exact mechanism how this toxin confers its potent adjuvant activity remains elusive (Levine et al., 1966). An early hypothesis was that PTX increased the permeability of blood vessels, particularly the BBB (Kügler et al., 2007; Linthicum et al., 1982; Schellenberg et al., 2012; Yong et al., 1993). Meanwhile PTX was identified to activate pyrin, thereby inducing neutrophil adhesion to cerebral capillaries (Dumas et al., 2014). The same group demonstrated that PTX increases the plasma level of IL-6, which in turn increases the expression of endothelial adhesion molecules and chemokines, leading to the recruitment of leukocytes that patrol the cerebral vasculature by crawling on the luminal endothelial surface (Richard et al., 2011; Roy et al., 2012). Interestingly, a crucial PTX mechanism is the activation of the inflammasome, whereby PTX causes the cleavage of pro-IL-1β into its active form IL-1β. As discussed earlier, IL-17A by itself induces IL-1β synthesis from granulocytes, thereby activating encephalithogenic T-cells (McGinley et al., 2020). Similarly, we demonstrated that in the context of other inflammatory stimuli, IL-1β synthesis is induced in the CNS of GF/IL17 mice. We speculate that local IL-17A synthesis renders PTX-induced IL-1β synthesis by neutrophils unnecessary and thus facilitates EAE induction.
The same mechanism might be the explanation of the spontaneous recruitment of encephalithogenic T-cells with a MOG-specific T-cell receptor in GF17-2D2 mice. 2D2 mice do not develop spontaneous T-cell infiltration without PTX administration (Bettelli et al., 2003; Waldner et al., 2004). PTX driven priming of encephalithogenic 2D2 cells for the induction EAE is driven via IL-1β (Ronchi et al., 2016). In the context of chronic CNS-specific IL-17A synthesis, Vβ11-TCR positive MOG-specific T-cells were spontaneously recruited into the brains of double transgenic mice, accompanied by microglial activation and infiltration of neutrophils and inflammatory monocytes.
In an experimental stoke model IL-17A drives CXCL-1 mediated recruitment of neutrophils into the CNS (Gelderblom et al., 2012). After cerebral ischemia conventional dendritic cells populate the CNS becoming the major source of IL-23, thereby promoting IL-17 induction in γδ T cells and neutrophil infiltration (Gelderblom et al., 2018). Though, IL-23 is well known to contribute to the development and differentiation of Th17 cells, CNS targeted overexpression of this cytokine using the same promoter does not lead to spontaneous granulocyte infiltration into the CNS but to a different phenotype with spontaneous accumulation of T and B cells into the brain and the clinical development of cerebellar ataxia (Nitsch et al., 2019). These slightly divergent results suggest a strong context dependence of these cytokines on the disease involved. In the context of EAE, astrocyte-specific expression of IL-23 results in a particular accumulation of B cells and γδ T cells, demonstrating that the CNS-specific effects of IL-23 are much more extensive than local IL-17A synthesis (Nitsch et al., 2021). Similar to IL-17A, local IL-23 induces spontaneous infiltration of MOG-specific T-cells, interestingly also displaying high numbers of infiltrating B cells again (Nitsch et al., 2022). These results again show that IL-23 mediates its effects further upstream compared to IL-17A and that blocking IL-17A could lead to a more targeted MS therapy than blocking IL-23, which failed in a phase II clinical trial to treat patients with relapsing remitting MS (Havrdová et al., 2016; Segal et al., 2008).
Taken together, CNS targeted expression of IL-17A facilitated induction of T-cell-mediated autoimmunity in the CNS. In the GF/IL17-mouse induction of MOG-EAE was accelerated and peak disease aggravated. CNS production of IL-17A was sufficient to induce MOG-EAE without the co-adjuvant PTX and to mediate spontaneous CNS infiltration of MOG-specific transgenic T-cells in a double transgenic mouse model. Our data support the evidence that IL-17A plays an important role in the early phase of EAE and MS, in particular contributing to the infiltration of immune cells into the CNS. Our findings should reinforce efforts in the development of IL-17A directed therapies to provide more targeted therapeutic options for individualized treatment of MS patients.