IL-36 was predominantly expressed in epithelial cells in experimental colitis, allergic lung inflammation, chronic rhinosinusitis and influenza A virus infection (22, 36, 41, 44, 45) and was upregulated by proinflammatory cytokines, such as IL-17 (41, 46). Reproductive tract epithelial cells also increased IL-36γ and IL-36R expression following treatment with microbial products (47). It is notable that epithelial IL-36 could be expressed as a full-length form and required a cleavage to the biologically active form (41). Epithelial cells can secret inflammatory cytokines in response to IL-36. Subcutaneous injection of IL-36α induced various inflammatory factors including IL-17, IL-20, IL-22, IL-23, interferon (IFN)-γ, TNF-α and KCs (48). IL-17 production by Th17 cells may upregulate all three IL-36 expression from human keratinocytes, creating a feedback loop that drives inflammation and disease development (49). Human keratinocytes were potent sources of chemokines following the exposure of IL-36 cytokines, leading to the recruitment of macrophages, T cells, and neutrophils (43). Notably, human keratinocytes upregulated type I and II IFN-responsive genes in response to IL-36, leading to potent cytokine production (35, 43). These findings indicate that IL-36 is critical for the early regulation of IFN and immune cell recruitment in the skin (50–52). Expression of IL-36R by skin-resident cells (e.g., keratinocytes and fibroblasts), but not the hematopoietic cells (e.g., T cells and DCs) is pivotal for the cutaneous pathology (51). Consistently, using a conditional knockout murine model, Goldstein et al. demonstrated that IL-36R signaling in keratinocytes played a major role in the induction of psoriasis-like dermatitis (52).

Activated neutrophils were considered a source of IL-36 in various diseases such as experimental autoimmune encephalomyelitis (EAE), chronic rhinosinusitis and influenza infection (21, 41, 53), while neutrophils from naïve mice express low levels of IL-36γ (53). Importantly, IL-36R is abundant on murine neutrophils derived from bone marrow, spinal cord and spleen (38, 53). However, IL-36R was not detectable in blood neutrophils in both mice and patients with inflammatory diseases (41, 53). Consistently, healthy human blood neutrophils failed to express IL-36R and did not respond to IL-36 cytokines (43, 50). Interestingly, IL-36R expression on human peripheral neutrophils could be induced by IL-1β, IL-6, and Der p1 (41), suggesting that both IL-36 cytokines and receptor might be inducible on neutrophils by the local inflammatory milieu. However, a study also reported that IL-36-triggered human bronchial epithelial cell can release neutrophil-associated chemokines such as CXCL8, and promote infiltration, activation, and inflammatory activity of neutrophils (54). In addition, IL-36 may active neutrophils and amplify lung inflammation in mice (55)

Dermal macrophages expressed high amounts of IL-36R transcript (37), indicating that the expression of IL-36R might be associated with its anatomical localization and immune microenvironment. IL-36β was as potent as IL-1β in stimulating human M2 macrophages, but not M1 and dermal macrophages (37). In addition, both human M1 macrophages and mouse lung macrophages were reported to produce IL-36 ligands following bacterial infection and LPS exposure (5, 24), indicating that macrophages might be the source of IL-36 similar to IL-1β and IL-33. Bone marrow-derived macrophages have undetectable levels or express much lower IL-36R compared to DCs (38, 41). Both human and mouse DCs were found to express IL-36R and become activated by IL-36 agonists stimulation (41, 43, 50, 56). Human monocyte-derived DCs expressed 6-fold more IL-36R mRNA than their monocyte precursors and accelerated maturation by IL-36α, β and γ (43, 56). IL-36R was also detectable in human Langerhans cells, which responded strongly to IL-36β stimulation (37). Additionally, plasmacytoid DCs (pDCs) can highly express IL-36R (50, 56). These pDCs bound by IL-36 potentiated Toll-like Receptor (TLR)-9 activation and IFN-α production (50).

T cells are most likely not the main source of IL-36 cytokines, but can respond to IL-36 due to their expression of IL-36R. Unlike the receptors of other IL-1 family members, such as IL-33R and IL-1R, whose expression are upregulated during T cell activation, IL-36R expression is detectable on naïve T cells, but is negligible in differentiated Th cells (14, 57). It is reported that IL-36γ synergized with IL-12 to facilitate Th1 differentiation, but suppressed Th17 differentiation in vitro in murine experiments (36, 38). Interestingly, IL-36γ inhibited Foxp3-expression in murine regulatory T cell development through the IL-36R/MyD88/NF-κBp50 axis, while concomitantly promoted the differentiation of Th9 and Th22 cells (58, 59). Therefore, IL-36 plays a critical role in mouse T cell differentiation.

Whether IL-36 have effect on human T cells is still unclear. It is reported that IL-36 may induced IFN-γ production in human CD3⁺ lymphocytes in vitro (14, 56). Penha et al. found the IL-36R expression on CD4⁺ T cells in the human blood and intestines, and IL-36β stimulation promoted CD4⁺ T cell proliferation in vitro (42). On the contrary, other researchers reported that IL-36R transcripts were undetectable in blood CD4⁺ T cells from healthy donors, and IL-36 failed to effect on resting or activated human T cells (43). Similarly, no obvious colocalization of IL-36R with human T cells in nasal polyps (41). Further study is needed to elucidate the regulation of IL-36R as well as the role of IL-36 in human T cell activation and differentiation. Similar to mouse CD4⁺ T cells, mouse effector CD8⁺ T cells increased IFN-γ production by IL-36γ stimuli (14, 60), and this process required IL-12 or IL-2 synergy (60). In addition, IL-36γ promoted IFN-γ production in vitro by murine NK cells and γδ T cells, which were able to express IL-36R (14). IL-36R mRNA was undetectable in mouse B cells in a previous study (38); Resident B cells and plasma cells in inflamed human tissues were found to express IL-36α (61). CD138⁺ and CD79α⁺ plasma cells were identified as the cellular sources of IL-36α in the synovial tissues and psoriatic skin in patients, respectively (62). How IL-36 regulates B cell functions is still not understood.

IL-36R mRNA has been detected in mouse astrocytes and microglia in the brain, but not in primary neurons (39, 40). However, IL-36 cytokines were dispensable for microglia activation and disease development of EAE (39, 40, 53). Increased IL-36α γ expression was also observed in murine hepatocytes following IL-1β/TNF-α/IFN-γ stimulation (12, 63), indicating that IL-36 may play a role in liver diseases. In addition, IL-36β has a pro-inflammatory effect on human synovial fibroblasts and articular chondrocytes in RA, suggesting the potential role of IL-36 in inflammatory responses of autoimmune diseases (64).

IL-36 was first identified as an inducible inflammatory cytokine in mouse keratinocytes following herpes simplex virus type 1 (HSV-1) infection (75). IL-36β-deficient mice developed more severe secondary zosteriform lesions and succumbed more frequently to HSV-1 infection (65). IL-36γ treatment protected mice from lethal intravaginal challenge, as evidenced by limited vaginal viral replication, delayed disease onset, decreased disease severity, and significantly increased survival (66). Further analysis demonstrated that IL-36β promoted type I IFN production through upregulation of IFN-α receptor expression and activation of the STAT signaling pathway in animal model (76). Indeed, IL-36 also promoted type I IFN in IL-36R⁺ pDC (50). Therefore, these studies indicate that IL-36 plays a critical role in innate immunity by boosting type I IFN signaling, inducing pro-inflammatory cytokines, and attracting innate immune cells, such as neutrophils.

Using a murine epicutaneous infection model, Nakagawa and Liu et al. found that S. aureus induced IL-1 and IL-36α from keratinocytes via secretion of S. aureus-expressed phenol-soluble modulin α, leading to the induction of IL-17 and recruitment of neutrophils in the skin (34, 69). Interestingly, IL-36α may not only regulate Th17 cell activity, but also modulate IL-17-production by γδ T cells and type 3 innate lymphoid cells (ILC3) (34, 69). Skin inflammation was dependent on IL-1R and IL-36R signals as well as their signaling adaptor MyD88. Satoh et al. also demonstrated that Cutibacterium acnes can induce IL-36γ through NF-κB in keratinocytes and subsequently IL-8, leading to cutaneous neutrophilia (77).

Fungal infection can induce IL-36 expression in epithelial cells and human PBMC (74, 78, 79). In oral candidiasis, IL-36α, β and γ transcript levels were all increased in the tongue of the sublingually challenged mice at 2 days post-injection (74). Candida albicans (C. albicans) infection resulted in increased IL-36 cytokines in human oral epithelial cells via NF-κB, MAPK and PI3K-dependent pathways (74). IL-36R-deficient mice were susceptible to acute oral candidiasis as evidenced by higher fungal loads and greater body weight loss, indicating the protective role of IL-36 in C. albicans infection (74).

Influenza virus infection can trigger epithelial cell-derived IL-36 cytokines (22, 46, 80), which activated NF-κB signaling and increased inflammatory cytokines (e.g. IL-6 and IL-8) in the lung (46). However, the role of IL-36 in influenza virus infection is incompletely understood. Aoyagi et al. reported that IL-36R-deficient mice were protected from influenza virus-induced lung injury and mortality accompanied by reduced lymphocyte activation, accumulation of myeloid cells, pro-inflammatory cytokine and chemokine production (e.g., IL-6, IL-17, CXCL1, and CXCL10) and permeability of the alveolar-epithelial barrier (22). However, IL-36γ was upregulated in the lungs and played a protective role in severe H1N1 and H3N2 influenza infection via modulating macrophage polarization and activity (21). Lack of IL-36γ resulted in increased viral titers, higher levels of IL-6, and more severe pathology in the lungs (21). Interestingly, macrophages in IL-36γ-deficient mice exhibited an M2-like phenotype and were likely to undergo apoptosis by infection, whereas adoptive transfer of WT alveolar macrophages protected IL-36γ-deficient mice against influenza infection (21). The reason for the discrepancies from the studies using IL-36R- and IL-36γ-deficient mice are not known at present. Different animal models and interfering strategies, such as neutralizing antibodies, should be used to further confirm these results.

The role of IL-36 in Mycobacterium tuberculosis (M. tuberculosis) has been documented in several studies. M. tuberculosis infection induced IL-36γ expression in human macrophages in vitro, and in the lungs of infected mice in vivo (72, 81). Its expression was induced through microbial ligands, which triggered host TLR and MyD88-dependent pathways, and was further amplified by endogenous IL-1β and IL-18 (81). Increased IL-36γ transcriptional expression was also observed in the plasma and bronchoalveolar lavage (BAL) samples of patients with Pseudomonas aeruginosa (P. aeruginosa)- or Streptococcus pneumoniae (S. pneumoniae)-induced acute respiratory distress syndrome (ARDS) (23, 24). Animal studies revealed that IL-36 signaling pathway may play a protective role in the lung with bacterial infection. The induction of IL-36 contributed to antimicrobial peptide production and M. tuberculosis growth restriction through promoting the accumulation of Liver X Receptor and modulating cholesterol biosynthesis and efflux (82). Activation of autophagy in macrophages was considered another hallmark by IL-36γ in restricting M. tuberculosis growth (71). However, IL-36R deficiency showed negligible impact on M. tuberculosis infection in mice, as demonstrated by similar survival rates and bacterial loads (72). Additionally, IL-36γ-deficient mice were more susceptible to S. pneumoniae infection, as evidenced by increased mortality, ameliorated lung bacterial clearance and increased bacterial dissemination, which might be due to the reduced type-1 cytokine expression and impaired lung macrophage M1 polarization (23). Similarly, the protective effect of IL-36γ was also demonstrated in a Klebsiella pneumoniae (K. pneumoniae) mouse model (23). Interestingly, Sequeira et al. revealed that microbiota Bacteroidetes protected against K. pneumoniae colonization (83) via IL-36 signals and macrophages (83). In addition, administration of Legionella pneumophila to IL-36R-deficient mice resulted in more severe disease as evidenced by higher mortality, delayed lung bacterial clearance, increased bacterial dissemination to the spleen, and impaired innate immune responses compared to that in infected wild-type mice (73). In contrast, IL-36R^-/- and IL-36γ^-/-, but not IL-36α^-/-, mice were resistant to during P. aeruginosa infection, as demonstrated by the reduction of bacterial burden, pro-inflammatory cytokine production and lung injury. Further investigation is needed to determine the role of IL-36 in intracellular bacterial infection using various interfering methods, such as IL-36 cytokine knockout mice and neutralizing antibodies.

Clinical evidence showed that ulcerative colitis patients had higher IL-36α in the colonic mucosa (36). Lack of IL-36R resulted in defective recovery following DSS-induced damage and impaired closure of colonic mucosal biopsy wounds due to the profound reduction of IL-22 (84). Interestingly, IL-36 can also regulate Treg/Th9 balance and the IL-23/IL-22 network in model of colitis induced by oxazolone, indicating that IL-36γ has multiple functions in modulating antigen-presenting cell function and in regulating T cell differentiation in a mouse model (58, 59). Russel et al. reported that infection with Citrobacter rodentium resulted in reduced CD11b⁺F4/80⁺Gr-1⁺ inflammatory cell recruitment, imbalanced Th1/Th17 responses and increased bacterial colonization of the colon in IL-36R^-/- mice (36). Accordingly, suppressed Th17, but enhanced Th1 differentiation was observed in vitro by IL-36α supplement (36, 57). However, since IL-36 is necessary for IL-22 production in DSS-induced colitis, it is not clear whether IL-36 differently regulates Th17 and Th22 differentiation in vivo among various animal models of gastrointestinal dysregulation.

Although IL-36 has been detected in hepatocytes (12, 63), the function of IL-36 in the liver remains unclear. Higher levels of IL-36α were observed in chronic hepatitis B virus (HBV) patients compared with that in healthy individuals (85). The positive correlation between IL-36α and HBV-DNA titers may indicate the potential involvement of IL-36 in antiviral immunity during chronic infection (85). Additionally, hepatitis C virus infection significantly increased the production of IL-36Ra but not IL-36 agonist ligands in human monocytes, leading to reduced NK cell activation (86). Further research is needed to dissect the role of IL-36 in liver resident cells (e.g., kupffer cells, hepatic stellate cells and sinusoidal endothelial cells) as well as in different liver disease models.

In addition to lung infections, IL-36α and IL-36γ were also upregulated in the mouse cornea in early responses to P. aeruginosa challenge (70). Exogenous IL-36γ treatment enhanced corneal innate immunity and alleviated P. aeruginosa keratitis. The protective role of IL-36γ required S100A9 and was partially dependent on the CXCL10/CXCR3 axis (70). On the contrary, IL-36Ra treatment exacerbated the outcome of P. aeruginosa keratitis (70).

Louis et al. reported that a truncated IL-36γ-encoded plasmid can act as a potent adjuvant for a DNA-encoded Zika virus (ZIKV) vaccine. Immunization with truncated IL-36γ promoted antiviral T cell responses and protected mice from ZIKV challenge (68). Moreover, co-delivery of truncated IL-36γ can also enhance antiviral immunity against HIV and influenza DNA vaccines (68). Besides, both in vivo and in vitro studies have proved that IL-36 treatment reduced HSV-2 replication in a lethal genital infection model and in human vaginal epithelial cells (66, 67). The absence of IL-36γ led to reduced mature neutrophil recruitment to the vaginal microenvironment at early times in HSV-2 infection (66). These findings set the stage for IL-36 in infectious diseases and shed light on IL-36 in the next generation of vaccines.