Discussion
Activation of NLRP3 is a tightly regulated process and a key step in autoimmunity of CNS. In this study, we provide three advances that broaden our understanding of NLRP3-associated neuroinflammation. First, by using Tslpr-/- mice, we demonstrated that TSLP signaling regulates neuroinflammation and paralysis of mouse. Second, we show that TSLPR signals via JAK2 to activate NLRP3-associated neuroinflammation in EAE settings. Third, we show that patients with anti-NMDAR encephalitis have increased levels of NLRP3 and IL-1β in CSF. Taken together, the current study identifies novel functions of TSLP-dominant JAK pathways upstream of NLRP3 represent promising targets for the treatment of autoimmune disorders.
Generally, Th17 response in autoimmune disorders has been believed to cause neurons death or inflammatory response 34, 35. Besides, previous studies also have shown that type 3 innate lymphoid cell (ILC3) produce IL-17 in autoimmune disorder such as ankylosing spondylitis 36 and that ILC3 can maintains neuroinflammation by supporting T cell survival 37. Thus, both neuroinflammation and demyelination were largely believed to be mediated by Th17- and ILC3- related responses. We and other have recently shown that TSLP primes dendritic cells (DCs) maturation21, 38 and that DCs determines status of activation and survival of ILC3 39. Based on our new data demonstrating that brain tissue TSLPR signaling critically mediates neuroinflammation, we speculate that innate immune cells, such as ILC3s, play important roles in promoting autoimmune inflammation-associated demyelination, in addition to Th17 cells. Identification of the precise role of TSLP in Th17 cells- and ILC3-mediated neuroinflammation and demyelination requires further investigation.
Despite previous observation that TSLP is reported to be critical for regulatory T cells formation in autoimmune disease, we found that, in contrast to decreased IL-7R expression on conventional T cells and reduced Treg function of MS in previous study 40, deletion of Tslpr efficiently alleviated neuroinflammation displayed by decreased CD4+ cells infiltration and restored myelin expression, which coincides with one previous study showing ameliorated EAE symptoms accompanied by reduced inflammatory infiltrates in the brain of Tslp -/- mice41. In fact, the epithelial cell-derived cytokine TSLP, GM-CSF, and IL-25 have been shown to be master initiators of type 3 inflammation via their effects on a variety of cells including Th17, ILC3, and mast cells 42, 43, 44. These cytokines are believed to rapidly bind to membrane receptor in order to generate innate immune response and therefore prime adaptive immune cells. Strikingly, two recent studies have demonstrated that TSLP directly activates neurons 45, 46. Additionally, our current data highly regulated TSLP in the brain in the settings of EAE coincides with previous study 47 and go beyond by demonstrating that TSLP receptor signals by phosphorylation of JAK2 to activate NLRP3-mediated inflammation in response to MOG35-55administration. Thus, we speculate that TSLP cytokine may act as master regulator of neuroinflammation in immune cells of brain.
In immune cells, cytokine signaling by the JAK-STAT pathway causes transcriptional changes to promote cellular activation. However, although JAK inhibitor has been reported to be an alternative immunotherapy in patients with autoimmune disorders such as neuromyelitis optica 48, our data indicated that neuroinflammation to MOG35-55 injection with additional JAK inhibition by ruxolitinib failed to induce typical neuroinflammation as observed in EAE mice brain. Also, these decreased neuroinflammation was accompanied by restored expression of myelin basic protein after treatment with ruxolitinib. Thus, we predict that alterations in classic JAK-mediated NLRP3 inflammasome activation is sufficient to explain how neuroinflammation and demyelination occur. One previous study has shown that JAK1 mediates sensory neuronal responsiveness which can be enhanced by cytokines such as IL-4 18. Our data coincides with this study and further demonstrates that JAK proteins have novel functions in neurons and regulates meylination/demyelination balance at least by NLRP3-mediated pathways. However, we note that such alteration of myelin basic protein expression does not exclude the role of JAK2 or other pathways in modulating transcription or other post-transcription of myelin basic protein within CNS. Future studies will be required to better understand how changes of JAK-STAT pathway impact myelin expression and neuroinflammation in autoimmune disorders.
Clinical application for ruxolitinib, an non-selective JAK1/2 inhibitor, have been reported to induce improvement of neurologic disability in neuromyelitis optica 48. In fact, significant clinical efficacy of symptoms in autoimmune disorders has been observed in clinical trials employing other JAK inhibitor such as tofacitinib and barcitinib 49, 50. Previously, the changes in neuroinflammation observed with JAK inhibition have been attributed to the anti-inflammatory role of Th17 response 35. Recently, study from others demonstrate that transient receptor potential (TRP) plays a critical role in EAE by mediating axonal and neuronal degeneration 51 and that JAK-STAT pathway determines TRP expression 52, 53, which indicates involvement of TRP in JAK pathway-mediated neuroinflammation. Based on our and others’ studies, we speculate that the improvement of neuroinflammation in EAE mice treated with ruxolitinib may be mediated, at least in part, by disruption of these signals in the CNS and that these therapies may alleviate neuroinflammation in autoimmune disorders. Strikingly, recent studies published experimental and clinical evidence for evobrutinib, the first Bruton’s tyrosine kinase (BTK) inhibiting molecule being developed, also described reduced disease severity in clinical and mouse model of multiple sclerosis (MS) by impairment of encephalitogenic T cells 54, 55. Given our current study demonstrating a direct role of JAK in neuroinflammation, whether combinations of TSLP-, JAK-, and BTK-blockade can lead to synergistic therapeutic improvements of neuroinflammation in autoimmune disorders such as MS demands further investigation.
In conclusion, our data establish and highlight the capability of TSLPR-JAK signaling inhibition to control disease-driving neuroinflamamtion of NLRP3 in inflammatory CNS demyelination. This is demonstrated here for ruxolitinib, a non-selective JAK inhibitor clinically tested in several autoimmune diseases, and the elucidated immunological effects may similar apply to other JAK inhibitors in clinical development. Based on this, the mechanistic data provided here will be instrumental in facilitate how this molecule is integrated into the current autoimmune disorder treatment.