Expression Of Th17 Cells Research Articles

Dietary glutamine supplementation improves both Th1 and Th17 responses via CARD11-mTORC1 pathway in murine model of atopic dermatitis

Glutamine (GLN) is considered an immunomodulatory nutrient, while caspase recruitment domain 11 (CARD11) is a susceptibility locus for atopic dermatitis (AD). T-cell antigen receptor (TCR)-stimulated GLN uptake requires CARD11. However, the specific pathogenesis of AD via GLN uptake remains unclear. This study aimed to elucidate the association between dietary GLN supplementation and the CARD11 pathway in the pathogenesis of AD, focusing on T helper type 1 (Th1) and Th17 cell expression in AD. Herein, wild-type (WT) mice with house dust mite epidermal-sensitized skin exhibited increased expression of interferon-gamma (IFN-gamma) and interleukin (IL)-17, whereas CARD11 deficiency impaired Th1 and Th17 responses at the same site. CARD11 is a key mediator of Th1 and Th17 expression in AD. Additionally, we suppressed mammalian target of rapamycin complex 1 (mTORC1) signaling, downstream of CARD11, to underscore the critical role of CARD11 in mediating Th1 and Th17 expression in AD. Further, dietary supplementation of GLN to CARD11−/− mice restored Th1 and Th17 responses, whereas inflammatory expression was reduced in WT mice, and p-CARD11 expression and mTORC1 signaling activity were increased in JPM50.6 cells and CARD11−/− mice. Upon inhibiting the GLN transporter, alanine-serine-cysteine transporter carrier 2 (ASCT2), we observed that the Th1 and Th17 response in AD was reduced. Conclusively, ASCT2-mediated GLN uptake improves the expression of Th1 and Th17 cells via CARD11-mTORC1 signaling pathway in AD, suggesting the potential of glutamine supplementation for AD treatment.

Immune mechanism of Daphnes Cortex and its processed products on CIA rats based on B7/CD28/CTLA-4 pathway

This study investigates the effects of Daphnes Cortex and its processed products on the differentiation of Th17/Treg cells in SD rats with type Ⅱ collagen-induced arthritis(CIA).Sixty-four SD rats were randomly divided into the normal group(normal),model group(model),fried Daphne giraldii Nitsche low-dose and high-dose groups(FDGN-L group, FDGN-H group),raw D. giraldii Nitsche low-dose and high-dose groups(RDGN-L group, RDGN-H group),daphnetin group(DAPH group),and tripterygium glycosides group(GTW group).Except for the normal group, the CIA model was immunized on the seventh day after the first immunization, and was gavaged for 28 days after the second immunization.After sampling, the inflammation of articular synovial membrane in CIA rats was observed by hematoxylin-eosin(HE)staining; the levels of transforming growth factor-β(TGF-β),interferon-γ(IFN-γ),interleukin(IL)-2,IL-4,and IL-10 in serum were detected by enzyme-linked immunosorbent assay(ELISA); real-time reverse transcription-PCR(qRT-PCR)and Western blot were used to detect the mRNA and protein expressions of cluster of differentiation(CD) 80(B7-1),CD 86(B7-2),CD28,and cytotoxic T lymphocyte-associated antigen 4(CTLA-4)in the synovial membrane of rats; flow cytometry was used to detect the proportion of Th17 and Treg cells in the synovial membrane of rats.The results showed that compared with the normal group, the joint synovial inflammation of rats in the model group was significantly aggravated, the arthritis index was significantly increased, and the immune organ index was increased(P<0.01).Compared with the model group, each drug administration group could improve the joint inflammation of rats to varying degrees, reduce the arthritis index, inhibit synovial hyperplasia, and reduce the immune organ index; compared with the model group, the serum levels of IL-2 and IFN-γ in each drug administration group were significantly decreased(P<0.01),TGF-β,IL-4,and IL-10 were significantly increased(P<0.01),the mRNA and protein expressions of B7-1 and CTLA-4 in the synovial membrane were significantly increased(P<0.01),and the proportion of Th17 cells and Treg cells in the joint tissue was significantly decreased(P<0.01).In conclusion, Daphnes Cortex inhibits the expression of Th17 cells in CIA rats and promotes the expression of Treg cells by regulating the B7/CD28/CTLA-4 pathway and the balance of Th17/Treg, thereby treating rheumatoid arthritis.

Deciphering Immune Landscape Remodeling Unravels the Underlying Mechanism for Synchronized Muscle and Bone Aging.

Evidence from numerous studies has revealed the synchronous progression of aging in bone and muscle; however, little is known about the underlying mechanisms. To this end, human muscles and bones are harvested and the aging-associated transcriptional dynamics of two tissues in parallel using single-cell RNA sequencing are surveyed. A subset of lipid-associated macrophages (triggering receptor expressed on myeloid cells 2, TREM2+ Macs) is identified in both aged muscle and bone. Genes responsible for muscle dystrophy and bone loss, such as secreted phosphoprotein 1 (SPP1), are also highly expressed in TREM2+ Macs, suggesting its conserved role in aging-related features. A common transition toward pro-inflammatory phenotypes in aged CD4+ T cells across tissues is also observed, activated by the nuclear factor kappa B subunit 1 (NFKB1). CD4+ T cells in aged muscle experience Th1-like differentiation, whereas, in bone, a skewing toward Th17 cells is observed. Furthermore, these results highlight that degenerated myocytes produce BAG6-containing exosomes that can communicate with Th17 cells in the bone through its receptor natural cytotoxicity triggering receptor 3 (NCR3). This communication upregulates CD6 expression in Th17 cells, which then interact with TREM2+ Macs through CD6-ALCAM signaling, ultimately stimulating the transcription of SPP1in TREM2+ Macs. The negative correlation between serum exosomal BCL2-associated athanogene 6 (BAG6) levels and bone mineral density further supports its role in mediating muscle and bone synchronization with aging.

Hydroxychloroquine exacerbates imiquimod-induced psoriasis-like dermatitis through stimulating overexpression of IL-6 in keratinocytes

Objective Hydroxychloroquine (HCQ) is a US Food and Drug Administration (FDA)-approved treatment for systemic lupus erythematosus (SLE) through inhibition of antigen presentation and subsequent reduction in T cell activation. Psoriasis relapse after antimalarial therapy have been reported in up to 18% of patients with psoriasis. Here, we explored the role of HCQ on exacerbating dermatitis utilizing an imiquimod (IMQ)-induced psoriasis-like dermatitis mouse model. Methods Thirty-six C57BL/6 female mice were divided into six groups: wild-type control, IMQ-Only, pre-treat HCQ (30 mg/kg and 60 mg/kg HCQ), and co-treat HCQ with IMQ (30 mg/kg and 60 mg/kg HCQ). Besides control, all were topically treated with IMQ for 5 days. Pharmacological effects and mechanisms of HCQ were assessed by clinical severity of dermatitis, histopathology, and flow cytometry. HaCaT cells were co-treated with both HCQ and recombinant IL-17A, followed by the detection of proinflammatory cytokine expression and gene profiles through enzyme-linked immunosorbent assay and next-generation sequencing. Results In the pre-treated and co-treated HCQ groups, skin redness and scaling were significantly increased compared to the IMQ-Only group, and Th17 cell expression was also upregulated. Acanthosis and CD11b+IL23+ dendritic cell (DC) infiltration were observed in the HCQ treatment group. IL-6 overexpression was detected in both the HaCaT cells and skin from the experimental mice. Psoriasis-related genes were regulated after being co-treated with HCQ and recombinant IL-17A in HaCaT cells. Conclusions HCQ exacerbates psoriasis-like skin inflammation by increasing the expression of IL-6, stimulating DC infiltration, and promoting Th17 expression in the microenvironment of the skin. Key messages This study provided possible mechanisms for inducing psoriasis during HCQ treatment through an animal model.

CCL5 promotes LFA-1 expression in Th17 cells and induces LCK and ZAP70 activation in a mouse model of Parkinson's disease.

Parkinson's disease (PD), which is associated to autoimmune disorders, is characterized by the pathological deposition of alpha-synuclein (α-Syn) and loss of dopaminergic (DA) neurons. Th17 cells are thought to be responsible for the direct loss of DA neurons. C-C chemokine ligand 5 (CCL5) specifically induces Th17 cell infiltration into the SN. However, the specific effect of CCL5 on Th17 cells in PD and the relationship between CCL5 and lymphocyte function-associated antigen-1 (LFA-1) expression in Th17 cells are unknown. We evaluated the effects of CCL5 on LFA-1 expression in Th17 cells in mice treated with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and examined Th17 cell differentiation upon CCL5 stimulation in vitro. Furthermore, we assessed the effects of CCL5 on tyrosine kinase zeta-chain-associated protein kinase 70 (ZAP70) and lymphocyte-specific protein tyrosine kinase (LCK) activity in CCL5-stimulated Th17 cells in vivo and in vitro. CCL5 increased the proportion of peripheral Th17 cells in MPTP-treated mice, LFA-1 expression on Th17 cells, and Th17 cell levels in the SN of MPTP-treated mice. CCL5 promoted Th17 cell differentiation and LFA-1 expression in naive T cells in vitro. Moreover, CCL5 increased Th17 cell differentiation and LFA-1 expression by stimulating LCK and ZAP70 activation in naive CD4+ T cells. Inhibiting LCK and ZAP70 activation reduced the proportion of peripheral Th17 cells and LFA-1 surface expression in MPTP-treated mice, and Th17 cell levels in the SN also significantly decreased. CCL5, which increased Th17 cell differentiation and LFA-1 protein expression by activating LCK and ZAP70, could increase the Th17 cell number in the SN, induce DA neuron death and aggravate PD.

LKB1 isoform expression modulates T cell plasticity downstream of PKCθ and IL-6

Following activation, CD4 T cells undergo metabolic and transcriptional changes as they respond to external cues and differentiate into T helper (Th) cells. T cells exhibit plasticity between Th phenotypes in highly inflammatory environments, such as colitis, in which high levels of IL-6 promote plasticity between regulatory T (Treg) cells and Th17 cells. Protein Kinase C theta (PKCθ) is a T cell-specific serine/threonine kinase that promotes Th17 differentiation while negatively regulating Treg differentiation. Liver kinase B1 (LKB1), also a serine/threonine kinase and encoded by Stk11, is necessary for Treg survival and function. Stk11 can be alternatively spliced to produce a short variant (Stk11S) by transcribing a cryptic exon. However, the contribution of Stk11 splice variants to Th cell differentiation has not been previously explored. Here we show that in Th17 cells, the heterogeneous ribonucleoprotein, hnRNPLL, mediates Stk11 splicing into its short splice variant, and that Stk11S expression is diminished when Hnrnpll is depleted using siRNA knock-down approaches. We further show that PKCθ regulates hnRNPLL and, thus, Stk11S expression in Th17 cells. We provide additional evidence that exposing induced (i)Tregs to IL-6 culminates in Stk11 splicing downstream of PKCθAltogether our data reveal a yet undescribed outside-in signaling pathway initiated by IL-6, that acts through PKCθ and hnRNPLL to regulate Stk11 splice variants and facilitate Th17 cell differentiation. Furthermore, we show for the first time, that this pathway can also be initiated in developing iTregs exposed to IL-6, providing mechanistic insight into iTreg phenotypic stability and iTreg to Th17 cell plasticity.

Downregulation of HMGB1 in thymoma cells affects T cell differentiation.

Thymoma is the most common anterior mediastinal tumor and is closely associated with myasthenia gravis (MG). Our previous study showed that the expression of Th17 cells increased and the expression of Treg decreased in MG-associated thymoma tissues and peripheral blood. High mobility group box 1 (HMGB1) is an inflammatory mediator and participates in the pathogenesis of various autoimmune diseases. However, its function in thymoma is still unclear. Material and methods: We first analyzed immune indices in peripheral blood of patients with MG-associated thymoma and patients with thymoma alone. Next, we explored the expression of HMGB1 in MG-associated thymoma and thymoma alone tissues. Furthermore, we transfected si-HMGB1 in thymoma cell line Thy0517 and co-cultured Thy0517 with peripheral blood mononuclear cells (PBMC). In this study, the levels of IgG, C3, C4, CRP and globulins in peripheral blood of patients with MG-associated thymoma were different from those of patients with thymoma alone (p < 0.05). The expression of HMGB1 in MG-associated thymoma tissues was higher than thymoma alone. Co-culture of Thy0517 and PBMC showed that the percentage of Th17 cells in PBMC was lower than that in the control group, and the percentage of Treg cells was higher than that in the control group. These findings demonstrate that HMGB1 is involved in the mechanism of abnormal Th17/Treg cell differentiation in thymoma and affects the occurrence of immune abnormalities in MG-associated thymoma.

Cholinergic control of Th17 cell pathogenicity in experimental autoimmune encephalomyelitis.

Experimental autoimmune encephalomyelitis (EAE) is a mouse model of multiple sclerosis (MS) in which Th17 cells have a crucial but unclear function. Here we show that choline acetyltransferase (ChAT), which synthesizes acetylcholine (ACh), is a critical driver of pathogenicity in EAE. Mice with ChAT-deficient Th17 cells resist disease progression and show reduced brain-infiltrating immune cells. ChAT expression in Th17 cells is linked to strong TCR signaling, expression of the transcription factor Bhlhe40, and increased Il2, Il17, Il22, and Il23r mRNA levels. ChAT expression in Th17 cells is independent of IL21r signaling but dampened by TGFβ, implicating ChAT in controlling the dichotomous nature of Th17 cells. Our study establishes a cholinergic program in which ACh signaling primes chronic activation of Th17 cells, and thereby constitutes a pathogenic determinant of EAE. Our work may point to novel targets for therapeutic immunomodulation in MS.

003 Interleukin (IL)-26 drives pathogenic IL-17A responses through a TH17-keratinocyte crosstalk

Interleukin (IL)-26 is a TH17 cytokine with known antimicrobial and pro-inflammatory functions and is highly expressed in tissues of TH17-mediated inflammatory diseases including psoriasis. However, the expression of IL-26 with respect to IL-17A and its precise function in the context of pathogenic TH17 responses are still unclear. In this study, we used flow cytometry-based fluorescent in situ hybridization of blood TH17 cells along with single-cell and spatial transcriptomics of psoriatic skin lesions to analyze the dynamic of the regulation of IL-26 expression in TH17 cells. We show that blood TH17 cells contain a large population of IL-26-producing cells that do not produce IL-17A. Unlike IL-17A-producing cells, these IL-26-producing cells develop at early stages of TH17 differentiation and do not require TGF-β. However, when exposed to TGF-β, IL-26-producing cells transition into IL-17 producers, a process also observed in psoriatic skin lesions, where TGF-β is highly expressed by basal keratinocytes in close vicinity to IL-26-producing T cells. T cell-derived IL-26 was able to induce TGF-β expression in keratinocytes themselves, suggesting a mechanism by which TH17 cells control IL-17A responses in tissues via the expression of IL-26. Thus, our study identifies IL-26-producing cells as an early differentiation stage of TH17 cells that can further differentiate into IL-17A producing cells upon tissues infiltration via a paracrine loop involving TGF-β.

Th17 Cell-Derived Amphiregulin Promotes Colitis-Associated Intestinal Fibrosis Through Activation of mTOR and MEK in Intestinal Myofibroblasts

Intestinal fibrosis is a significant complication of Crohn's disease (CD). Gut microbiota reactive Th17 cells are crucial in the pathogenesis of CD; however, how Th17 cells induce intestinal fibrosis is still not completely understood. In this study, T-cell transfer model with wild-type (WT) and Areg-/- Th17 cells and dextran sulfate sodium (DSS)-induced chronic colitis model in WT and Areg-/- mice were used. CD4+ T-cell expression of AREG was determined by quantitative reverse-transcriptase polymerase chain reaction and enzyme-linked immunosorbent assay. The effect of AREG on proliferation/migration/collagen expression in human intestinal myofibroblasts was determined. AREG expression was assessed in healthy controls and patients with CD with or without intestinal fibrosis. Although Th1 and Th17 cells induced intestinal inflammation at similar levels when transferred into Tcrβxδ-/- mice, Th17 cells induced more severe intestinal fibrosis. Th17 cells expressed higher levels of AREG than Th1 cells. Areg-/- mice developed less severe intestinal fibrosis compared with WT mice on DSS insults. Transfer of Areg-/- Th17 cells induced less severe fibrosis in Tcrβxδ-/- mice compared with WT Th17 cells. Interleukin (IL)6 and IL21 promoted AREG expression in Th17 cells by activating Stat3. Stat3 inhibitor suppressed Th17-induced intestinal fibrosis. AREG promoted human intestinal myofibroblast proliferation, motility, and collagen I expression, which was mediated by activating mammalian target of rapamycin and MEK. AREG expression was increased in intestinal CD4+ T cells in fibrotic sites compared with nonfibrotic sites from patients with CD. These findings reveal that Th17-derived AREG promotes intestinal fibrotic responses in experimental colitis and human patients with CD. Thereby, AREG might serve as a potential therapeutic target for fibrosis in CD.

Androgen Plays a Potential Novel Hormonal Therapeutic Role in Th17 Cells Predominant Neutrophilic Severe Asthma by Attenuating BECs Regulated Th17 Cells Differentiation via MBD2 Expression.

T helper 17 (Th17) cells subtype of non-T2 asthma is less responsive (resistant) to inhaled corticosteroids (ICS), so also called severe asthma. Methyl-CpG-binding domain protein 2 (MBD2) regulates the differentiation of the Th17 cells, showing the possibility of a therapeutic target in severe asthma. Androgen tends to show beneficial therapeutic effects and is a "hot research topic," but its role in the differentiation and expression of Th17 cells via MBD2 is still unknown. The aim of this study was to evaluate how sex hormone interacts with MBD2 and affects the differentiation and expression of Th17 cells in severe asthma. Here, first, we measured the concentration of androgen, estrogen, and androgen estrogen ratio from subjects and correlated it with severe asthma status. Then, we established an animal model and bronchial epithelial cells (BECs) model of severe asthma to evaluate the role of MBD2 in the differentiation and expression of Th17 cells (IL-17), the therapeutic potential of sex hormones in severe asthma, and the effect of sex hormones in BECs regulated Th17 cells differentiation via MBD2 at the cellular level. Increased MBD2 expression and Th17 cells differentiation were noted in the animal and the BECs severe asthma models. Th17 cell differentiation and expression were MBD2 dependent. Androgen attenuated the differentiation of BECs regulated Th17 cells via MBD2 showing BECs as a therapeutic target of androgen, and these findings postulate the novel role of androgen in Th17 cells predominant neutrophilic severe asthma therapy through targeting MBD2.

Therapeutic Potential of Curcumin in a Rat Model of Dextran Sulfate Sodium–Induced Ulcerative Colitis by Regulating the Balance of Treg/Th17 Cells

The pathogenesis of ulcerative colitis (UC) remains unclear, and it is believed that an imbalance of regulatory T (Treg) cells and T helper 17 (Th17) cells is related to the occurrence of UC. Curcumin has been confirmed to exert anti-inflammatory effects in bronchial asthma and osteoarthritis by regulating the balance of Treg/Th17 cells. This study aimed to explore the therapeutic potential of curcumin in dextran sulfate sodium (DSS)-induced UC rats by regulating the balance of Treg/Th17 cells. Disease activity index (DAI) scores were calculated. Changes in colon inflammation were observed using hematoxylin and eosin staining. Treg and Th17 cells in the spleen were detected by flow cytometry, and the levels of interleukin (IL)-10 and IL-17A were determined using enzyme-linked immunosorbent assay. In DSS-induced colitis, curcumin significantly ameliorated colitis symptoms by reducing the DAI and increasing colon length. Additionally, curcumin significantly increased the expression of Treg cells and decreased the expression of Th17 cells and the extent of histopathological damage. Furthermore, curcumin increased the expression of IL-10 and decreased the expression of IL-17A. Curcumin attenuates DSS-induced UC injury by regulating Treg/Th17 balance and related cytokine secretion. Thus, curcumin may be a promising therapeutic drug for treating UC.

Th17 cell-derived miR-155-5p modulates interleukin-17 and suppressor of cytokines signaling 1 expression during the progression of systemic sclerosis.

Background miR‐155‐5p is associated with autoimmune diseases. T helper 17 (Th17) cells, interleukin (IL)‐17, and suppressor of cytokines signaling 1 (SOCS1) have important roles in the pathogenesis of systemic sclerosis (SSc). The purpose of this study was to explore the role of miR‐155‐5p in the regulation of IL‐17 and SOCS1 expression in Th17 cells and the subsequent effect on SSc disease progression.MethodsTh17 cells were isolated from peripheral blood mononuclear cells of SSc patients and healthy controls (HCs). RT‐qPCR and western blotting were used to examine the expression patterns of miR‐155‐5p, IL‐17, and SOCS1. Luciferase reporter assays were performed to confirm SOCS1 as a target of miR‐155‐5p. RNA pull‐down assays were performed to detect the interaction of IL‐17 and SOCS1 with miR‐155‐5p. In situ hybridization was performed to analyze the co‐expression pattern of miR‐155‐5p and IL17A in Th17 cells.ResultsThe levels of Th17 cell‐derived miR‐155‐5p were significantly up‐regulated in SSc patients compared with HCs, and its levels were negatively correlated with SOCS1 levels. Meanwhile, miR‐155‐5p positively regulated IL‐17 expression levels in Th17 cells isolated from SSc patients as the disease progressed. Using pmirGLO vectors, SOCS1 was confirmed as a target of miR‐155‐5p. The binding status of IL‐17 and SOCS1 to miR‐155‐5p was related to SSc progression. An increase in the co‐localization of miR‐155‐5p and IL‐17 was associated with greater SSc progression.ConclusionsIL‐17 and SOCS1 expression modulated by Th17 cell‐derived miR‐155‐5p are critical for SSc progression, which may provide novel insights into the pathogenesis of SSc.

Adipose-derived stem cells ameliorate atopic dermatitis by suppressing the IL-17 expression of Th17 cells in an ovalbumin-induced mouse model

BackgroundMesenchymal stem cells (MSCs) have therapeutic potential for atopic dermatitis (AD) owing to their immunoregulatory effects. However, the underlying mechanisms associated with the therapeutic efficacy of MSCs on AD are diverse and related to both cell type and delivery method.ObjectivesThis study investigated the therapeutic effect and mechanisms of adipose-derived stem cells (ADSCs) on AD using an ovalbumin (OVA)-induced AD mouse model.MethodsAD mice were subcutaneously injected with mouse ADSCs, cortisone, or PBS, and the therapeutic effects were determined by gross and histological examinations and serum IgE levels. Additionally, qPCR, RNA-sequencing analyses of skin samples and co-culture of ADSCs and Th17 cells were conducted to explore the underlying therapeutic mechanisms.ResultsADSCs treatment attenuated the AD pathology, decreased the serum IgE levels, and decreased mast cells infiltration in the skin of the model mice. Moreover, tissue levels of IL-4R and Th17-relevant products (IL-17A, CCL20, and MMP12) were suppressed in the ADSC- and cortisone-treated groups. Genomics and bioinformatics analyses demonstrated significant enrichment of inflammation-related pathways in the downregulated genes of the ADSC- and cortisone-treated groups, specifically the IL-17 signaling pathway. Co-culture experiments revealed that ADSCs significantly suppressed the proliferation of Th17 cells and the expression of proinflammatory cytokines (IL-17A and RORγT). Furthermore, expression levels of PD-L1, TGF-β, and PGE2 were significantly upregulated in co-cultured ADSCs relative to those in monocultured ADSCs.ConclusionADSCs ameliorate OVA-induced AD in mice mainly by downregulating IL-17 secretion of Th17 cells.

Interleukin-17 Receptor E and C-C Motif Chemokine Receptor 10 Identify Heterogeneous T Helper 17 Subsets in a Mouse Dry Eye Disease Model

Dry eye disease (DED) features the inflammatory response of the ocular surface. Pro-inflammatory T helper 17 (Th17) cells are important for the pathogenesis of DED. In the present study a mouse DED model was used to discover two Th17 subsets in draining lymph nodes and conjunctivae based on the expression of IL-17 receptor E (IL-17RE) and CCR10: IL-17RElowCCR10- Th17 and IL-17REhighCCR10+ Th17. IL-17REhighCCR10+ Th17 expressed more retinoic acid-related orphan receptor gamma t but fewer T-box-expressed-in-T-cells than IL-17RElowCCR10- Th17. In addition, the former expressed higher IL-17A, IL-21, and IL-22 but fewer IFN-γ than the latter. Further analysis showed that IL-17REhighCCR10+ Th17 did not express IFN-γ invivo, whereas IL-17RElowCCR10- Th17 contained IFN-γ-expressing Th17/Th1 cells. Moreover, IL-17REhighCCR10+ Th17 possessed more phosphorylated p38 mitogen-activated proteinkinase (MAPK) and Jnk than IL-17RElowCCR10- Th17, suggesting higher activation of MAPK signaling in IL-17REhighCCR10+ Th17. Invitro treatment with IL-17C effectively maintained IL-17A expression in Th17 cells through p38 MAPK rather than Jnk MAPK. Furthermore, the adoptive transfer of the two Th17subpopulations indicated their equivalent pathogenicity in DED. Interestingly, IL-17REhighCCR10+ Th17 cells were able to phenotypically polarize to IL-17RElowCCR10- Th17 cells invivo. In conclusion, the current study revealed novel Th17 subsets with differential phenotypes, functions, and signaling status in DED, thus deepening the understanding of Th17 pathogenicity, and exhibited Th17 heterogeneity in DED.

TH17 CELLS PROMOTE INTESTINAL FIBROSIS THROUGH THE PRODUCTION OF AMPHIREGULIN

Abstract BACKGROUND &amp; AIMS Intestinal fibrosis is a major serious complication of Crohn’s disease (CD). Gut microbiota-reactive Th17 cells have been established as crucial in the pathogenesis of CD, however, how Th17 cells induce fibrosis is still not completely understood. METHODS Wild-type Th1 cells and Th17 cells, or amphiregulin (Areg)-/- Th17 cells, were transferred into Tcrbxd-/- mice to induce colitis and intestinal fibrosis. CD4+ T cell expression of Areg was determined by qRT-PCR and ELISA. Intestinal fibroblasts were isolated from normal and CD patients and used to determine the effects of Areg on cell proliferation and migration in vitro. Video microscopy and single-cell tracking were used to assess cell migration kinetically. Areg expression was compared in peripheral blood CD4+ T cells, intestinal biopsy tissues, and lamina propria lymphocyte (LPL) from healthy controls and CD patients with or without intestinal fibrosis. RESULTS When transferred into TCRβ/δ-/- mice, although Th1 and Th17 cells induced intestinal inflammation at similar levels, Th17 cells induced more severe intestinal fibrosis than Th1 cells. RNA-seq analysis showed higher levels of AREG expression in Th17 cells compared to Th1 cells, which was further confirmed by qRT-PCR. Transfer of Areg-/- Th17 cells induced less severe fibrosis in TCRβ/δ-/- mice compared with WT Th17 cells. IL-6 but not IL-23 promoted Areg expression in Th17 cells in a time and dose-dependent manner. Treatment with Th17 signature cytokine IL-21, but not IL-17, upregulated T cell expression of Areg. Consistently, IL-21R-/- T cells produced lower levels of Areg. Mechanically, STAT3 mediates Areg expression in Th17 cells, in that IL-6 promoted STAT3 activation, and IL-6 did not induce AREG expression in STAT3-/- T cells. Areg expression was lower in STAT3-/- T cells compared with WT T cells under Th17 conditions. Treatment with RORgt inhibitor, which inhibits Th17 differentiation, suppressed Th17 cell Areg expression, indicating that higher Areg expression requires fully Th17 differentiation. Areg promoted human intestinal fibroblast proliferation and motility, which was mediated by activation of mTOR, and mTOR inhibitors inhibited Areg-induced human intestinal fibroblast proliferation and motility. Finally, Areg expression is increased in peripheral blood CD4+ T cells, intestinal biopsy tissues, and lamina propria lymphocytes of CD patients with fibrosis. CONCLUSION Collectively, these findings reveal that Th17-derived Areg promotes fibrotic responses in both experimental colitis and human CD patients. Thereby, Areg could serve as a potential therapeutic target for fibrosis in CD.

TH17 CELLS PROMOTE INTESTINAL FIBROSIS THROUGH THE PRODUCTION OF AMPHIREGULIN

Intestinal fibrosis is a major serious complication of Crohn’s disease (CD). Gut microbiota-reactive Th17 cells have been established as crucial in the pathogenesis of CD, however, how Th17 cells induce fibrosis is still not completely understood. Wild-type Th1 cells and Th17 cells, or amphiregulin (Areg)-/- Th17 cells, were transferred into Tcrbxd-/- mice to induce colitis and intestinal fibrosis. CD4+ T cell expression of Areg was determined by qRT-PCR and ELISA. Intestinal fibroblasts were isolated from normal and CD patients and used to determine the effects of Areg on cell proliferation and migration in vitro. Video microscopy and single-cell tracking were used to assess cell migration kinetically. Areg expression was compared in peripheral blood CD4+ T cells, intestinal biopsy tissues, and lamina propria lymphocyte (LPL) from healthy controls and CD patients with or without intestinal fibrosis. When transferred into TCRβ/δ-/- mice, although Th1 and Th17 cells induced intestinal inflammation at similar levels, Th17 cells induced more severe intestinal fibrosis than Th1 cells. RNA-seq analysis showed higher levels of AREG expression in Th17 cells compared to Th1 cells, which was further confirmed by qRT-PCR. Transfer of Areg-/- Th17 cells induced less severe fibrosis in TCRβ/δ-/- mice compared with WT Th17 cells. IL-6 but not IL-23 promoted Areg expression in Th17 cells in a time and dose-dependent manner. Treatment with Th17 signature cytokine IL-21, but not IL-17, upregulated T cell expression of Areg. Consistently, IL-21R-/- T cells produced lower levels of Areg. Mechanically, STAT3 mediates Areg expression in Th17 cells, in that IL-6 promoted STAT3 activation, and IL-6 did not induce AREG expression in STAT3-/- T cells. Areg expression was lower in STAT3-/- T cells compared with WT T cells under Th17 conditions. Treatment with RORgt inhibitor, which inhibits Th17 differentiation, suppressed Th17 cell Areg expression, indicating that higher Areg expression requires fully Th17 differentiation. Areg promoted human intestinal fibroblast proliferation and motility, which was mediated by activation of mTOR, and mTOR inhibitors inhibited Areg-induced human intestinal fibroblast proliferation and motility. Finally, Areg expression is increased in peripheral blood CD4+ T cells, intestinal biopsy tissues, and lamina propria lymphocytes of CD patients with fibrosis. Collectively, these findings reveal that Th17-derived Areg promotes fibrotic responses in both experimental colitis and human CD patients. Thereby, Areg could serve as a potential therapeutic target for fibrosis in CD.

Periodontal ligament cells under mechanical force regulate local immune homeostasis by modulating Th17/Treg cell differentiation.

Improper orthodontic force often causes root resorption or destructive bone resorption. There is evidence that T helper 17 (Th17) cells and regulatory T (Treg) cells may be actively involved in bone remodeling during tooth movement. In a combination of in vitro and in vivo studies, we investigated the effect of human periodontal ligament cells (hPDLCs) on Th17/Treg cells under different orthodontic forces and corticotomy. hPDLCs were cultured in vitro and subjected to different mechanical forces. The expression of interleukin (IL)-6 and transforming growth factor (TGF)-β in the supernatant and the mRNA levels of hypoxia inducible factor (HIF)-1α, Notch1, and TGF-β in hPDLCs were investigated. Supernatants were collected and co-cultured with activated CD4+T cells, and the differentiation of Th17/Treg cells was analyzed by flow cytometry. We also established an animal model of tooth movement with or without corticotomy. The tooth movement distance, alveolar bone height, and root resorption were analyzed using micro-computed tomography. Expression of interleukin (IL)-17A, forkhead Box P3 (Foxp3), and IL-6 were analyzed using immunohistochemistry, while osteoclasts were evaluated by tartrate-resistant acid phosphatase (TRAP) staining. The mRNA levels of IL-17A, IL-6, Foxp3, IL-10, HIF-1α, notch1, and C-X-C motif chemokine ligand 12 (CXCL12) in alveolar bone and gingiva were investigated. Heavy force repressed cell viability and increased the mortality rate of hPDLCs; it also improved the expression of IL-6, declined the expression of TGF-β, and promoted the mRNA expression level of HIF-1α. The expression of TGF-β and Notch1 mRNA decreased and then increased. The supernatant of hPDLCs under heavy force promotes the polarization of Th17 cells. The heavy force caused root resorption and decreased alveolar bone height and increased the positive area of IL-17A immunohistochemical staining and the expression of IL-17A, IL-6, HIF-1α, and Notch1 mRNA. Corticotomy accelerated tooth movement, increased the proportion of Foxp3-positive cells, and up-regulated the expression of Foxp3, IL-10, and CXCL12 mRNA. During orthodontic tooth movement, the heavy force causes root resorption and inflammatory bone destruction, which could be associated with increased expression of Th17 cells and IL-6. Corticotomy can accelerate tooth movement without causing root resorption and periodontal bone loss, which may be related to the increased expression of Treg cells. Altogether, this report provides a new perspective on the prevention of inflammatory injury via the regulation of Th17/Treg cells in orthodontics.

Forsythoside A Alleviates Imiquimod-Induced Psoriasis-like Dermatitis in Mice by Regulating Th17 Cells and IL-17A Expression.

Psoriasis is a recurrent inflammatory skin disease characterized by redness and scaly skin lesions with itchy or painful sensations. Forsythoside A, one of the main active compounds isolated from the fruit of Forsythia suspensa, has been widely applied to treat inflammatory diseases in the clinical use of traditional oriental medicine. However, the effect of forsythoside A on psoriasis remains unclear. This study aimed to explore the therapeutic effects and immune regulation of forsythoside A on psoriasis. C57BL/6 mice were divided into six groups and treated with imiquimod cream on their shaved back skin to induce psoriasis-like dermatitis. Different doses of forsythoside A (5 mg/kg, 10 mg/kg, or 20 mg/kg) were administered to the respective treatment groups. Skin redness, scaling, and ear thickness were measured; keratinocyte proliferation and inflammatory cytokine expression were detected by hematoxylin–eosin and immunohistochemical staining. Th17 cells in the inguinal lymph nodes were detected by flow cytometric analysis. IL-17A levels were measured using ELISA. The results showed that forsythoside A relieved psoriatic skin symptoms such as skin redness, thickness, scaling, and reduced epidermal thickening. The expression of IL-6, IL-17, and Ki-67 was downregulated in the forsythoside-A-treated groups. Th17 cell expression in inguinal lymph nodes and IL-17A secretion was suppressed by forsythoside A. In conclusion, forsythoside A was found to alleviate imiquimod-induced psoriasis-like dermatitis in mice by suppressing Th17 development and IL-17A secretion. These findings demonstrate the feasibility of forsythoside A in treating human psoriasis.

Therapeutic Effect of IL-38 on Experimental Autoimmune Uveitis: Reprogrammed Immune Cell Landscape and Reduced Th17 Cell Pathogenicity.

PurposeThe purpose of this study was to elucidate the effects of interleukin (IL)-38 on experimental autoimmune uveitis (EAU) and its underlying mechanisms.MethodsMice with EAU were treated with IL-38, and the retinas and cervical draining lymph nodes (CDLNs) were analyzed by flow cytometry. Single-cell RNA sequencing (scRNA-seq) was conducted to analyze the immune cell profiles of CDLNs from normal, EAU, and IL-38-treated mice.ResultsAdministration of IL-38 attenuated EAU symptoms and reduced the proportion of T helper 17 (Th17) and T helper 1 (Th1) cells in the retinas and CDLNs. In scRNA-seq analysis, IL-38 downregulated the IL-17 signaling pathway and reduced the expression of Th17 cell pathogenicity-related genes (Csf2 and Il23r), findings which were also confirmed by flow cytometry. In vitro, IL-38 reduced the granulocyte-macrophage colony-stimulating factor (GM-CSF) stimulation function of IL-23 and inhibited IL-23R expression in Th17 cells. Moreover, when co-cultured with Th17 cells, IL-38 prevented IL-23 production in antigen-presenting cells (APCs).ConclusionsOur data demonstrate the therapeutic effect of IL-38 on EAU, and suggest that the effect of IL-38 may be caused by dampening of the GM-CSF/IL-23R/IL-23 feedback loop between Th17 cells and APCs.