Th17 Cell Population Research Articles

Cordycepin exhibits anti-bacterial and anti-inflammatory effects against gastritis in Helicobacter pylori-infected mice.

Helicobacter pylori is one of the dominant members of gastric microbiota associated with gastritis. ChronicH. pyloricolonization may yield detrimental consequences, including mucosal layer atrophy, gastritis, and gastric cancer. The traditional antibiotic treatment might result in antibiotic resistance. To overcome this obstacle, this study aims to investigate the potential antibacterialand anti-inflammatory effects of cordycepin on mice infected withH. pylori. A mouse model ofH. pyloriinfection was established. The expression levels of target genes were evaluated by qRT-PCR, western blotting, or ELISA. The infiltrated Th17 cell population was determined by flow cytometry analysis. Our results demonstrated that the administration of cordycepin exhibited up to 3-fold antibacterial effect againstH. pyloriin vivo. Cordycepin treatment resulted in around 50% inflammatory cytokine production (e.g. IL-6 and IL-1β) and about 60% immune cell infiltration (e.g. Th17 cells) when compared to vehicle control group. Thus, we confirmed that cordycepin conferred antibacterialand anti-inflammatory effects onH. pylori-infected mice.Cordycepin may serve as a potential candidate for developing a therapeutic regimen forH. pylori-induced gastritis.

Identification of Sorafenib as a Treatment for Type 1 Diabetes.

Th1 cell activation is considered a key mediator of the pathogenesis of type 1 diabetes. Targeting IL-12-induced Th1 cell differentiation seems to be an effective way to block the development of type 1 diabetes. However, given the critical function of Th1 in the immune system, the potential side effects hinder the application of anti-Th1 therapy in the treatment of type 1 diabetes. To identify safe anti-Th1 treatment(s), we screened the FDA-approved tyrosine kinase inhibitor (TKI) drug library using an IL-12-induced Th1 differentiation cell model. We found that among the TKIs with little effect on T cell viability, sorafenib is the top contender for the inhibition of Th1 differentiation. Treatment of NOD mice with sorafenib significantly impeded the development of type 1 diabetes and ameliorated insulitis, which coincided with a specifically decreased accumulation of Th1 cell population in the pancreas but not in peripheral immune organs. Mechanistically, sorafenib indirectly inhibited janus kinase 2 (JAK2) activity and blocked IL-12-induced phosphorylations of JAK2 and signal transducer and activator of transcription 4 (STAT4). Since sorafenib is classified as an FDA-approved drug, it serves as a preliminary lead point for additional experimentation and may be a promising therapy for type 1 diabetes in humans.

Exposure to ambient air pollutants during pregnancy is linked to IL4, IL10, and IFNγ gene methylation and fewer Th1, Th2, and Th17 cell populations

Ambient air pollutants(AAPs) have been linked with adverse pregnancy outcomes, such as preeclampsia, preterm labor, and low birth weight. During pregnancy, there is an attenuation in cell-mediated immunity with shifts in the Th1/Th2/Th17/Treg environment, contributing to overall maternal susceptibility.

Endometrial Regenerative Cell-Derived Conditioned Medium Alleviates Experimental Colitis.

Background Traditional interventions can play a certain role in attenuating ulcerative colitis (UC), known as one type of inflammatory bowel diseases, but sometimes are not effective. Endometrial regenerative cells (ERCs) have been shown to exert immunosuppressive effects in different models of inflammation, and stem cell-derived conditioned media (CM) have advantages over cell therapy in terms of easy access and direct action. However, whether ERC-CM could alleviate colitis remains unclear and will be explored in this study. Methods Menstrual blood was collected from healthy female volunteers to obtain ERCs and ERC-CM. Acute colitis was induced by 3% dextran sodium sulfate (DSS), and ERC-CM was injected on days 4, 6, and 8, respectively, after induction. The disease activity index was calculated through the record of weight change, bleeding, and fecal viscosity during the treatment process. Histological features, macrophage and CD4+ T cell in the spleen and colon, and cytokine profiles in the sera and colon were measured. In addition, an in vitro lymphocyte proliferation assay was measured by using a CCK-8 kit in this study. Results ERC-CM treatment significantly improved the symptoms and histological changes in colitis mice. ERC-CM increased the percentage of Tregs in the spleen and colon but decreased the percentages of M1 macrophages and Th1 and Th17 cells in the spleen and decreased the population of Th17 cells in the colon. In addition, ERC-CM treatment decreased the local expression of TNF-α, IL-6, and iNOS in the colon. Furthermore, ERC-CM increased the levels of anti-inflammatory cytokines IL-10 and IL-27 but decreased proinflammatory cytokines IL-6 and IL-17 in the sera. In addition, ERC-CM significantly inhibited ConA-induced mouse lymphocyte proliferation in vitro. Conclusion The results suggest that ERC-CM can exert similar therapeutic effects as ERCs and could be explored for future application of cell-free therapy in the treatment of colitis.

A bioinformatics approach to introduce novel multi-epitope vaccines against Acinetobacter baumannii retrieved from immunogenic extracellular loops of outer membrane proteins

Increasing antibiotic resistance of Acinetobacter baumannii highlights the urgent need to develop an effective vaccine to combat it. Despite all efforts, no approved vaccine against this bacterium is on the market, and attempts are underway to find effective vaccines. Due to their surface exposure, outer membrane proteins (OMPs) are excellent vaccine targets. The extracellular loops of OMPs are highly immunogenic. Therefore, in the present study, we aimed to detect the conserved, exposed, and immunogenic linear B-cell epitopes among sixty-five OMPs available in 35 A. baumannii genomes deposited in the Vaxign database. According to defined criteria (allergenicity, antigenicity, and non-similarity to host), four linear B-cell epitopes were selected as highly immunogenic peptides. Epitope shuffling was performed to select the best arrangements of these epitopes. We proposed six different multi-epitope vaccines consisting of two Naked arrangements, two loop-less C-lobe (LCL) structures, and two FliC-based constructs. An LCL-based multi-epitope vaccine showed the best molecular docking score with human TLRs (TLR 1, 2, 4, and 5) and immune simulation results (Th1 cell population, IFN-γ, IL-12, IgM, IgG1, and IgG2 levels). This study provided a valuable library of exposed epitopes including 26 conserved, linear and immunogenic B-cell epitopes (belonged to 14 OMPs), and three optimized multi-epitope vaccines. However, more experimental studies are needed to confirm the actual immunoreactivity of these vaccine candidates.

The CTRP3-AdipoR2 Axis Regulates the Development of Experimental Autoimmune Encephalomyelitis by Suppressing Th17 Cell Differentiation.

C1q/TNF-related proteins (CTRP) including CTRP3 are a group of secreted proteins which have a complement C1q-like domain in common, and play versatile roles in lipid metabolism, inflammation, tumor metastasis and bone metabolism. Previously, we showed that the expression of C1qtnf3, encoding CTRP3, is highly augmented in joints of autoimmune arthritis models and CTRP3-deficiency exacerbates collagen-induced arthritis in mice. However, the mechanisms how CTRP3-deficiency exacerbates arthritis still remain to be elucidated. In this study, we showed that CTRP3 was highly expressed in Th17 cell, a key player for the development of autoimmune diseases, and Th17 cell differentiation was augmented in C1qtnf3–/– mice. Th17 cell differentiation, but not Th1 cell differentiation, was suppressed by CTRP3 and this suppression was abolished by the treatment with a receptor antagonist against AdipoR2, but not AdipoR1, associated with suppression of Rorc and Stat3 expression. Furthermore, AdipoR1 and AdipoR2 agonist, AdipoRon suppressed Th17 cell differentiation via AdipoR2, but not AdipoR1. The development of myelin oligodendrocyte glycoprotein (MOG)-induced experimental autoimmune encephalomyelitis was enhanced in C1qtnf3 –/– mice associated with increase of Th17 cell population. CTRP3 inhibited MOG-induced IL-17 production from T cells by affecting both T cells and dendritic cells. These results show that CTRP3 is an endogenous regulator of Th17 differentiation, suggesting that the CTRP3-AdipoR2 axis is a good target for the treatment of Th17 cell-mediated diseases.

Gomisin M2 alleviates psoriasis‑like skin inflammation by inhibiting inflammatory signaling pathways.

Psoriasis, a chronic inflammatory skin disease, is characterized by the excessive proliferation and impaired differentiation of epidermal keratinocytes and is accompanied by the increased infiltration of inflammatory cells. The condition requires long-term treatment and has no definitive cure. Hence, supplements and therapeutic agents have been intensely investigated. Gomisin M2 (GM2), a lignan extracted from Schisandra chinensis (Turcz). Baill. (Schisandraceae; S. chinensis), has demonstrated diverse pharmacological properties, including anticancer, anti-inflammatory and antiallergic effects. Based on these findings, the present study examined the effects of GM2 on an imiquimod (IMQ)-induced psoriasis mouse model and on keratinocytes stimulated by tumor necrosis factor (TNF)-α and interferon-γ. IMQ was topically applied to the back skin of mice for 7 consecutive days, and the mice were orally administered CD. These results showed that the oral administration of GM2 suppressed the symptoms of psoriasis, as evidenced by reductions in skin thickness, psoriasis area severity index scores for psoriasis lesions, transepidermal water loss and myeloperoxidase (MPO)-associated cell infiltration. Furthermore, GM2 reduced the pathologically increased levels of immunoglobulin G2a, MPO and TNF-α in the serum and T helper (Th)1 and Th17 cell populations in the spleen. GM2 decreased the gene expression of inflammatory-related cytokines and chemokines and inhibited the expression of signal transducer and activator of transcription 1 and nuclear factor-κB in the activated keratinocytes. These results suggested that GM2 from S. chinensis is a potential therapeutic candidate to alleviate psoriasis-like skin inflammation.

IL2-inducible T-cell kinase inhibitor ibrutinib reduces symptoms and Th2 differentiation in mouse allergic-rhinitis model.

Th2 and Th17 immune response contribute to allergic rhinitis (AR) development. Targeting Th2 and Th17 response has been shown to ameliorate AR. Ibrutinib is an inhibitor for IL2-inducible T-cell kinase, which can promote Th2 and Th17 immune response. We sought to investigate the effect of ibrutinib on AR and the underlying mechanisms. We established house dust mite-induced AR mouse model and treated AR mice with ibrutinib. The symptoms of AR, serum level of immunoglobulin E, percentage of Th1, Th2, Th17, and Treg in nasal lymphoid tissues were monitored. We also established in vitro T cell differentiation cell culture model. The T cells were treated with ibrutinib and the expression of specific transcriptional factors and cytokines was measured. The activation of PLC-γ1/calcium/NFAT2 signaling pathway was detected. Ibrutinib treatment had no effects on the development of lymphocytes and myeloid cells, but alleviated AR symptoms and decreased Th2 cell population in nasal lymphoid tissue. Meanwhile, iburitnib suppressed Th2 and Th17 differentiation in vitro. Moreover, iburitnib prevented phosphorylation of PLC-γ1and nuclear translocation of NFAT2 in Th2 cells. Our results suggested that ibrutinib could ameliorate AR symptoms through suppression of Th2 differentiation in AR mouse model.

HIF-1α modulates sex-specific Th17/Treg responses during hepatic amoebiasis

An invasive form of intestinal Entamoeba (E.) histolytica infection, which causes amoebic liver abscess, is more common in men than in women. Immunopathological mechanisms are responsible for the more severe outcome in males. Here, we used a mouse model of hepatic amoebiasis to investigate the contribution of hepatic hypoxia-inducible factor (HIF)-1α to T helper 17 (Th17)/regulatory T cell (Treg) responses in the context of the sex-specific outcome of liver damage. C57BL/6J mice were infected intrahepatically with E.histolytica trophozoites. HIF-1α expression was determined by qPCR, flow cytometry and immunohistochemistry. Tregs and Th17 cells were analysed by immunohistochemistry and flow cytometry. Finally, male and female hepatocyte-specific Hif1α knockout mice were generated, and the effect of HIF-1α on abscess development, the cytokine milieu, and Th17/Treg differentiation was examined. E.histolytica infection increased hepatic HIF-1α levels, along with the elevated frequencies of hepatic Th17 and Treg cells. While the Th17 cell population was larger in male mice, Tregs characterised by increased expression of Foxp3 in female mice. Male mice displayed increased IL-6 expression, contributing to immunopathology; this increase in IL-6 expression declined upon deletion of hepatic HIF-1α. In both sexes, hepatic deletion of HIF-1α reduced the Th17 cell frequency; however, the percentage of Tregs was reduced in female mice only. Hepatic HIF-1α modulates the sex-specific outcome of murine E.histolytica infection. Our results suggest that in male mice, Th17 cells can be modulated by hepatic HIF-1α via IL-6, indicating marked involvement in the immunopathology underlying abscess development. Strong expression of Foxp3 by hepatic Tregs from female mice suggests a potent immunosuppressive function, leading to initiation of liver regeneration. Infection with the parasite Entamoeba histolytica activates immunopathological mechanisms in male mice, which lead to liver abscesses that are larger than those in female mice. In the absence of the protein HIF-1α in hepatocytes, abscess formation is reduced; moreover, the sex difference in abscess size is abolished. These results suggest that HIF-1α modulates the immune response involved in the induction of immunopathology, resulting in differential disease susceptibility in males and females.

KLF4 Promotes Diabetic Chronic Wound Healing by Suppressing Th17 Cell Differentiation in an MDSC-Dependent Manner.

Objectives Diabetic wound inflammation deficiencies lead to ulcer development and eventual amputation and disability. Our previous research demonstrates that myeloid-derived suppressor cells (MDSCs) accumulate during inflammation and promote chronic wound healing via the regulation of Kruppel-like factor 4 (KLF4). In this study, we aimed to investigate the potential roles of MDSCs and KLF4 in diabetic wound healing. Methods An ob/ob mouse pressure ulcer (PU) model was used to evaluate the process of wound healing. The expression levels of KLF4 and IL-17A were measured by real-time PCR, and the population of MDSCs and Th17 cells was measured by flow cytometry. The levels of cytokines were determined by an immunosuppression assay. Results KLF4 deficiency in the diabetic PU model resulted in decreased accumulation of MDSCs, increased expansion of Th17 cells, and significantly delayed wound healing. Conversely, KLF4 activation by APTO-253 accelerated wound healing accompanied by increased MDSC populations and decreased numbers of Th17 cells. MDSCs have been proven to mediate Th17 differentiation via cytokines, and our in vitro data showed that elevated KLF4 expression in MDSCs resulted in reduced Th17 cell numbers and, thus, decreased levels of cytokines indispensable for Th17 differentiation. Conclusions Our study revealed a previously unreported function of KLF4-regulated MDSCs in diabetic wound healing and identified APTO-253 as a potential agent to improve the healing of pressure ulcers.

In-silico immunoinformatic analysis of SARS-CoV-2 virus for the development of putative vaccine construct

COVID-19 (CoronaVirus disease 2019) is caused by the SARS-CoV-2 virus (severe acute respiratory syndrome corona virus 2). SARS-CoV-2 virus is highly contagious and affects the human respiratory tract resulting in symptoms such as high fever, body ache, cough, dysfunctions of tastebuds and smelling sense of body. The objective of the present study involves immunoinformatic analysis to predict COVID-19 protein for vaccine construct based on the genomic information SARS-CoV-2 virus. At present, as per WHO estimates, around 133 COVID-19 novel vaccines under development. Three amino acid sequences of SARS-CoV-2 were retrieved from the NCBI database for the analysis of vaccine construct. This study involves computational and immunoinformatic methods. The Immunoinformatic tools used in the present study are NetCTL server, IFN epitope server, Toxin PRED, BCPred, CTL + HTL + ADJUVANTS + LINKERS, AlgPredserver, VaxiJenserver, ProtParam to predict vaccine construct. The secondary and tertiary structure prediction is done by PSIPRED, I-TASSER, Galaxy refine, prosA + Ramachandran. Finally, docking of the vaccine constructs and ligand was done with the help of Cluspro 2.0.C-ImmSimm webserver to simulate the potential vaccine construct. The present study demonstrated three potential Vaccine constructs for the SARS-CoV-2 virus, which were docked with TLR8 (Toll-likereceptor8). Interestingly from these, all constructs one having a high potential for the inhibition effect of the SARS-CoV-2virus. Immunological simulation data shows significant elevated amount of memory B cell; also, the high response was seen in TH(Helper) and TC(cytotoxic) cell population from the vaccine construct proposed in the current study. Hence, these constructs are suitable vaccine candidates that might be useful in developing a novel vaccine.

Effector functions of Th17 cells are regulated by IL-35 and TGF-β in visceral leishmaniasis.

Visceral leishmaniasis (VL) is a debilitating human pathogenesis in which the body's immune functions are severely compromised. Various subsets of T cells, including Th17 cells are important regulators of immune responses observed in various pathologies. The role of Th17 cells and its correlation with immuno-regulatory cytokines are however not well understood in human VL. Herein we studied how IL-17 is associated with the progression of Leishmania donovani infection using murine model of VL. We found induction of a strong IL-17 response at the early phase of infection which progressively reduced to basal level during chronic VL. The mechanistic study of this behavior was found to be linked with the role of regulatory T cells (CD4+ CD25+ T cells) that suppresses the proliferation of the Th17 cell population. Moreover, TGF-β and IL-35 derived from CD4+ CD25+ T cells are the key mediators for the downregulation of IL-17 during chronic VL. Thus, this study points to an antagonistic effect of Tregs and Th17 cells that can be used for designing better therapeutic and preventive strategies against leishmaniasis.

IL-37 Targets TSLP-Primed Basophils to Alleviate Atopic Dermatitis

Atopic dermatitis (AD) represents a severe global burden on physical, physiological and mental health. Innate immune cell basophils are essential for provoking allergic inflammation in AD. However, the roles of novel immunoregulatory cytokine IL-37 in basophils remain elusive. We employed in vitro co-culture of human basophils and human keratinocyte HaCaT cells and an in vivo MC903-induced AD murine model to investigate the anti-inflammatory mechanism of IL-37. In the in vitro model, IL-37b significantly decreased Der p1-induced thymic stromal lymphopoietin (TSLP) overexpression in HaCaT cells and decreased the expression of TSLP receptor as well as basophil activation marker CD203c on basophils. IL-37 could also reduce Th2 cytokine IL-4 release from TSLP-primed basophils ex vivo. In the in vivo model, alternative depletion of basophils ameliorated AD symptoms and significantly lowered the Th2 cell and eosinophil populations in the ear and spleen of the mice. Blocking TSLP alleviated the AD-like symptoms and reduced the infiltration of basophils in the spleen. In CRISPR/Cas9 human IL-37b knock-in mice or mice with direct treatment by human IL-37b antibody, AD symptoms including ear swelling and itching were significantly alleviated upon MC903 challenge. Notably, IL-37b presence significantly reduced the basophil infiltration in ear lesions. In summary, IL-37b could regulate the TSLP-mediated activation of basophils and reduce the release of IL-4. The results, therefore, suggest that IL-37 may target TSLP-primed basophils to alleviate AD.

MicroRNA-3100 from Bone Marrow Mesenchymal Stem Cells (BMSCs) Exosomes Alleviates Autoimmune Arthritis in Mice via Regulating Forkhead Box P3 Acetylase

To study the effect and mechanism of microRNA-3100 from exosome of BMSCs on autoimmune arthritis in mice. CIA mice model was established and treated with BMSCs exosomes or DMSO. The expression of Foxp3 was detected by PCR and Western blot. The Treg/Th17 population of T cells and the expression of related inflammatory factors were analyzed by flow cytometry. BMSCs trans-fected with MicroRNA-3100 inhibitor or MicroRNA-3100 was used to treat CIA mice through intravenous injection. The levels of Foxp3 and its acetylation, Treg/Th17 differentiation and inflammatory factors were detected, and the synovial tissues of mice were collected for analysis of its inflammatory infiltration. RA rats exhibited down-regulated Treg population and increased Th17 population in PBMC and SF, accompanied by reduced Foxp3 and acetylated Foxp3 expression, suppressed Tip60 expression in SF, in relative to normal control. However, exosomes of BMSCs administration recovered the changes. Further result showed that exosomes from BMSCs was enriched for miR-3100. miR-3100 inhibitor exposed CD4+ T cell exhibited reduced Tip60 and acetylated-Foxp3 expression, decreased Tregs population and increased Th17 cell population, vice versa as the miR-3100 mimics does. In addition, Tip60 inhibitors NU905 exerted the same effect as miR-3100 inhibitor transfection dose. Finally, miR-3100 overexpression could significantly reduce inflammation and immune cell infiltration, correct the dysfunction between Th17 and Tregs, recovered Tip60 and acetylation Foxp3 expression. MiRNA-3100 is enriched in exosomes of BMSCs, and can up-regulatethe expression of Foxp3, acetylated Foxp3 and Tip60 level, improving the imbalance of Treg/Th17 ratio, and improve autoimmune arthritis in mice.

113-OR: Identification of Sorafenib for the Treatment of Type 1 Diabetes

Type 1 diabetes (T1D) is an immune-mediated disease characterized by impairment of pancreatic beta cells. To date, a comprehensive cure is not available. Th1 cell activation is considered a key mediator of the pathogenesis of T1D. IL-12-induced STAT4 activation is a paramount signaling pathway to instigate Th1 cell differentiation. Targeting IL-12-induced Th1 cell differentiation seems to be an effective way to block the development of T1D. However, given the critical function of Th1 in the immune system, the potential side effects hinder the application of anti-Th1 therapy in the treatment of T1D. To identify safe anti-Th1 treatment(s), we screened the FDA-approved tyrosine kinase inhibitor (TKI) drug library using an IL-12-induced Th1 differentiation cell model. We found that sorafenib, an oral anti-cancer drug, is a top contender for inhibition of Th1 differentiation relative to induced toxicity. Sorafenib inhibited IL-12-induced Th1 cell differentiation in a dose-dependent manner without any change of CD4+ T-cell viability. Treatment of NOD mice with sorafenib significantly impeded the development of T1D and ameliorated insulitis, which coincided with a specifically decreased aggregate of Th1 cell population in pancreas but not in peripheral immune organs. Inflammatory cytokines and Th1 signature transcription factor Tbet mRNA expression in pancreas were decreased in sorafenib-treated NOD mice compared to vehicle-treated NOD mice. Mechanistically, sorafenib directly inhibited JAK2 activity and blocked IL-12-induced phosphorylations of JAK2 and STAT4. In conclusion, sorafenib administration inhibits the development of T1D by suppressing IL-12b/JAK2/STAT4 signaling-mediated Th1 cell differentiation specifically in pancreas in NOD mice. Since sorafenib is classified as an FDA-approved drug, it serves as a preliminary lead point for additional experimentation and may be a promising therapeutic for T1D in humans. Disclosure Q. Zeng: None. J. Song: None. X. Sun: None. D. Wang: None. Y. Xiao: None. Z. Zhou: None. T. Deng: None.

HMSC-Derived Exosome Inhibited Th2 Cell Differentiation via Regulating miR-146a-5p/SERPINB2 Pathway.

Background Allergic rhinitis (AR) is a global disease without specific treatment. Human mesenchymal stem cell- (HMSC-) derived exosomes (HMSC-exos) have been implicated for the amelioration of allergic inflammation by delivering miR-146a-5p in a mouse asthma model. However, the antiallergic activity and the underlying mechanism of HMSC-exos in AR remain unclear. The present study aimed to investigate the role of HMSC-exos in the pathogenesis of AR. Materials and Methods Blood specimens were collected from AR patients and healthy donators for investigation. HMSC and CD4+ T cells were used in the present study. Flow cytometry was used to characterize the population of Type 1 helper T (Th1) and Th2 cells. Specific siRNA and overexpressed plasmids were designed to silence or overexpress the expressions of miR-146a-5p and SERPINB2. Luciferase reporter assay was adopted to explore the binding site of miR-146a-5p and SERPINB2. Quantitative real-time PCR and immunoblots were performed to estimate the expression of target genes. Results The population of Th2 cells was significantly elevated in AR patients as compared with that in healthy donators. HMSC-exos could decrease the expression of SERPINB2 and the differentiation of Th2 cells. miR-146a-5p in HMSC-exos exhibited consistent effects and lowered the expression of SERPINB2 by binding on its 3′UTR region. Moreover, the differentiation of Th2 cells was promoted by SERPINB2 that could be reversed by HMSC-exos. Additionally, the miR-146a-5p expression was negatively associated with the SERPINB2 expression in the serum of AR patients. Conclusion HMSC-exos could inhibit the differentiation of Th2 cells via the regulation of the miR-146a-5p/SERPINB2 pathway. miR-146a-5p and SERPINB2 could be applied as potential targets for AR treatment.

Lactobacillus rhamnosus HDB1258 modulates gut microbiota-mediated immune response in mice with or without lipopolysaccharide-induced systemic inflammation

BackgroundGut microbiota closely communicate in the immune system to maintain a balanced immune homeostasis in the gastrointestinal tract of the host. Oral administration of probiotics modulates gut microbiota composition. In the present study, we isolated Lactobacillus rhamnosus HDB1258, which induced tumor necrosis factor (TNF)-α and interleukin (IL)-10 expression in macrophages, from the feces of breastfeeding infants and examined how HDB1258 could regulate the homeostatic immune response in mice with or without lipopolysaccharide (LPS)-induced systemic inflammation.ResultsOral administration of HDB1258 significantly increased splenic NK cell cytotoxicity, peritoneal macrophage phagocytosis, splenic and colonic TNF-α expression, TNF-α to IL-10 expression ratio, and fecal IgA level in control mice, while Th1 and Treg cell differentiation was not affected in the spleen. However, HDB1258 treatment significantly suppressed peritoneal macrophage phagocytosis and blood prostaglandin E2 level in mice with LPS-induced systemic inflammation. Its treatment increased LPS-suppressed ratios of Treg to Th1 cell population, Foxp3 to T-bet expression, and IL-10 to TNF-α expression. Oral administration of HDB1258 significantly decreased LPS-induced colon shortening, myeloperoxidase activity and NF-κB+/CD11c+ cell population in the colon, while the ratio of IL-10 to TNF-α expression increased. Moreover, HDB1258 treatment shifted gut microbiota composition in mice with and without LPS-induced systemic inflammation: it increased the Cyanobacteria and PAC000664_g (belonging to Bacteroidetes) populations and reduced Deferribacteres and EU622763_s group (belonging to Bacteroidetes) populations. In particular, PAC001066_g and PAC001072_s populations were negatively correlated with the ratio of IL-10 to TNF-α expression in the colon, while the PAC001070_s group population was positively correlated.ConclusionsOral administered HDB1258 may enhance the immune response by activating innate immunity including to macrophage phagocytosis and NK cell cytotoxicity in the healthy host and suppress systemic inflammation in the host with inflammation by the modulation of gut microbiota and IL-10 to TNF-α expression ratio in immune cells.

Single-cell clonotype tracing of tissue Th17 cells during CNS autoimmunity reveals the conversion of intestinal homeostatic cells to pathogenic cells

Abstract At homeostasis, most Th17 cells are found in the lamina propria of the intestine, where they contribute to tissue homeostasis by inhibiting microbiota from tissue invasion and promote barrier functions. Recent studies in humans and mice have implicated intestinal Th17 cells in extra-intestinal autoimmune diseases, including multiple sclerosis, but the mechanism in which intestinal Th17 cells mediate these dichotomous functions remains unknown. Here, we combined single-cell RNA- and TCR-seq (scRNA/TCR-seq) with fate mapping of tissue Th17 cells to profile over 84,000 tissue Th17 cells and characterize their heterogeneity, plasticity, and migration at homeostasis and during CNS autoimmunity. We discovered a homeostatic Th17 cell population, that is induced by the intestinal microbiota, is present in both lymphoid organs and the intestine, and expresses IL-17. Joint scRNA- and TCR-seq showed that during EAE this homeostatic population gives rise to a pathogenic Th17 cell population, that migrates specifically through the draining lymph nodes and the spleen to the CNS, and highly expresses GM-CSF and IFNγ, but expresses little or low levels of IL-17. This conversion of homeostatic Th17 cells into encephalitogenic Th17 cells depended on IL-23R signaling. Our work characterizes Th17 heterogeneity, plasticity, and migration during homeostasis and CNS autoimmune disease, identifies a mechanism by which intestinal Th17 cells influence the generation of an autoimmune disease-inducing population, and provides further evidence for a direct relationship between the IL-17-producing and GM-CSF- and IFNγ- producing pathogenic T cells.

Genetically engineered CD4 T cells as a novel immunotherapy against drug resistant Klebsiella pneumoniae infections.

Abstract With mortality rates ranging from 30% to 70%, carbapenem-resistant Klebsiella pneumoniae (Kp) is a significant public health threat. With growing incidence of drug resistance, it is imperative to develop alternative strategies to combat these severe infections. One novel immunotherapy is adoptive T cell transfer therapy using T cells genetically modified to express a T cell receptor (TCR) specific to an antigen of interest. Though successful against cancers and viral infections, this strategy has yet to be utilized for bacterial infections. Our study’s objective was to validate adoptive transfer as an immunotherapy against Kp infections using CD4 T cells from immunized mice. Further, we sought to identify TCRs specific to Kp epitopes for the generation of engineered Kp specific T cells. CD45.1 WT mice were intratracheally vaccinated with outer membrane protein X (OmpX) adjuvanted with heat-labile enterotoxin A1 twice at three weeks apart. Lung and splenic CD4 T cells from immunized mice were transferred into CD45.2 WT mice, followed by a challenge with Kp ST258 C4 strain. Immunized CD4 T cells were sequenced by single cell TCRseq to identify Kp specific TCR sequences. OmpX immunization resulted in a robust population of Kp specific Th17 cells in the lungs. Adoptive transfer of lung, but not splenic T cells from immunized mice significantly reduced bacterial burdens in challenged mice. Additionally, we identified 7 enriched TCR sequences that may be Omp specific. We have confirmed the efficacy of adoptive T cell transfer therapy against Kp infections using T cells from immunized mice. Moving forward we will validate our TCR sequences as Kp specific to genetically engineer both primary and cell line T cells for future therapeutic studies.

Melatonin Synergizes With Mesenchymal Stromal Cells Attenuates Chronic Allograft Vasculopathy.

BackgroundChronic rejection characterized by chronic allograft vasculopathy (CAV) remains a major obstacle to long-term graft survival. Due to multiple complicated mechanisms involved, a novel therapy for CAV remains exploration. Although mesenchymal stromal cells (MSCs) have been ubiquitously applied to various refractory immune-related diseases, rare research makes a thorough inquiry in CAV. Meanwhile, melatonin (MT), a wide spectrum of immunomodulator, plays a non-negligible role in transplantation immunity. Here, we have investigated the synergistic effects of MT in combination with MSCs in attenuation of CAV.MethodsC57BL/6 (B6) mouse recipients receiving BALB/c mouse donor aorta transplantation have been treated with MT and/or adipose-derived MSCs. Graft pathological changes, intragraft immunocyte infiltration, splenic immune cell populations, circulating donor-specific antibodies levels, cytokine profiles were detected on post-operative day 40. The proliferation capacity of CD4+ and CD8+ T cells, populations of Th1, Th17, and Tregs were also assessed in vitro.ResultsGrafts in untreated recipients developed a typical pathological feature of CAV characterized by intimal thickening 40 days after transplantation. Compared to untreated and monotherapy groups, MT in combination with MSCs effectively ameliorated pathological changes of aorta grafts indicated by markedly decreased levels of intimal hyperplasia and the infiltration of CD4+ cells, CD8+ cells, and macrophages, but elevated infiltration of Foxp3+ cells. MT either alone or in combination with MSCs effectively inhibited the proliferation of T cells, decreased populations of Th1 and Th17 cells, but increased the proportion of Tregs in vitro. MT synergized with MSCs displayed much fewer splenic populations of CD4+ and CD8+ T cells, Th1 cells, Th17 cells, CD4+ central memory T cells (Tcm), as well as effector memory T cells (Tem) in aorta transplant recipients. In addition, the percentage of splenic Tregs was substantially increased in the combination therapy group. Furthermore, MT combined with MSCs markedly reduced serum levels of circulating allospecific IgG and IgM, as well as decreased the levels of pro-inflammatory IFN-γ, TNF-α, IL-1β, IL-6, IL-17A, and MCP-1, but increased the level of IL-10 in the recipients.ConclusionsThese data suggest that MT has synergy with MSCs to markedly attenuate CAV and provide a novel therapeutic strategy to improve the long-term allograft acceptance in transplant recipients.