Tregs In Tissues Research Articles

Amphiregulin promotes activated regulatory T cell-suppressive function via the AREG/EGFR pathway in laryngeal squamous cell carcinoma

BackgroundActivated regulatory T cells (aTregs) play a vital role in promoting a tumor immunosuppressive microenvironment in laryngeal squamous cell carcinoma (LSCC). However, the regulatory factors that induce the generation of aTregs are not clear. Herein, we investigated the effect of amphiregulin (AREG) on the production of aTregs in the tumor microenvironment of LSCC.MethodsImmunohistochemical (IHC) analysis was conducted to examine the expression of AREG and FOXP3, and their association with clinical parameters and patient outcomes was demonstrated. The expression level of EGFRs in three functional subsets of Tregs was assessed, and the induction of CD4+ T cells into aTregs in the presence or absence of AREG or Gefitinib was analyzed using flow cytometry.ResultsOur results showed a higher expression level of AREG was significantly related to advanced clinical stage and worse survival, particularly with increased infiltration of Tregs in LSCC tumor tissue. The in vitro study showed that AREG significantly promoted the differentiation of aTregs, and enhanced the inhibitory effect of Tregs on T cell proliferation, which could be reversed by epidermal growth factor receptor (EGFR) inhibitors. In addition, we found that EGFR was highly expressed in aTregs, but not in other subsets of Tregs. It is suggested that AREG might induce aTregs, and enhance the immunosuppressive function of Tregs via the AREG/EGFR signal pathway.ConclusionsCollectively, this study revealed the role and mechanism of AREG in negative immune regulation, and targeting AREG might be a novel immunotherapy for LSCC.

IL-35 promotes IL-35+IL-10+ Bregs and Conventional LAG3+ Tregs in the lung tissue of OVA-Induced Asthmatic Mice.

This study aimed to investigate the effect of interleukin-35 (IL-35) on inflamed lung tissue in a murine model of asthma. IL-35 was examined for its potential to induce regulatory lymphocytes during ovalbumin (OVA)-induced acute lung injury. Female BALB/c mice sensitized with OVA and were treated with recombinant IL-35 (rIL-35) via intranasal or intraperitoneal routes and were administered 4h before OVA challenge. The effects of rIL-35 treatment on the lung and blood levels of regulatory B cells (Bregs) and regulatory T cells (Tregs), as well as their production of immunosuppressive cytokines, were determined using flow cytometry and enzyme-linked immunosorbent assay (ELISA), respectively. Treatment of OVA-sensitized asthmatic mice with rIL-35, whether administered intranasally or intraperitoneally, resulted in reduced lung inflammation and injury. This reduction was accompanied by an increase in the frequency of IL-35 producing Bregs, IL-35 and IL-10 producing Bregs, and conventional LAG3+ Tregs in the lung tissues and blood. This increase was more pronounced with intranasal rIL-35. Furthermore, there was a positive correlation between the levels of these regulatory cells and lung gene expression of IL-35 and IL-10, and an inverse correlation with both lung gene expression and plasma level of IL-17. The results of this study suggest that IL-35, through its ability to increase Bregs and Tregs, is effective in reversing lung inflammation in the context of asthma. Since the increase was more pronounced with intranasal administration, this highlights the therapeutic potential of its local intrapulmonary application in managing asthma-related inflammation.

Single-cell chromatin accessibility and transposable element landscapes reveal shared features of tissue-residing immune cells

Tissue adaptation is required for regulatory T (Treg) cell function within organs. Whether this program shares aspects with other tissue-localized immune populations is unclear. Here, we analyzed single-cell chromatin accessibility data, including the transposable element (TE) landscape of CD45+ immune cells from colon, skin, adipose tissue, and spleen. We identified features of organ-specific tissue adaptation across different immune cells. Focusing on tissue Treg cells, we found conservation of the Treg tissue adaptation program in other tissue-localized immune cells, such as amphiregulin-producing T helper (Th)17 cells. Accessible TEs can act as regulatory elements, but their contribution to tissue adaptation is not understood. TE landscape analysis revealed an enrichment of specific transcription factor binding motifs in TE regions within accessible chromatin peaks. TEs, specifically from the LTR family, were located in enhancer regions and associated with tissue adaptation. These findings broaden our understanding of immune tissue residency and provide an important step toward organ-specific immune interventions.

Correlation between Tregs and ICOS-induced M2 macrophages polarization in colorectal cancer progression.

To explore the mechanism by which Tregs promote the progression of colorectal cancer by inducing tumor-associated macrophages to polarize into M2 type via ICOS. Postoperative pathological tissues and clinical pathological data of 268 colorectal cancer patients who underwent initial surgery were collected. Immunohistochemistry (IHC) was used to detect the expression levels of ICOS, CD163 (a marker for M2 macrophages), and Foxp3 (a marker for Tregs) in cancerous, adjacent non-tumorous, and normal tissues. The relationship of ICOS, M2 macrophages, and Tregs in CRC with clinical pathological characteristics and pre-surgical tumor markers (such as CEA and CA199) was explored. The expression levels of M2 macrophages and Tregs increased with tumor progression, while ICOS expression showed a decreasing trend. Compared to adjacent and normal tissues, the expression levels of ICOS, M2 macrophages, and Tregs were higher in CRC tissues. The expression levels of M2 macrophages and Tregs were significantly positively correlated with tumor markers, while ICOS expression was significantly negatively correlated. Tumor-associated m2 macrophages induced by Tregs and ICOS participate in the dynamic balance of the colorectal cancer tumor microenvironment, and their interaction affects colorectal carcinogenesis and progression. High levels of ICOS are associated with better long-term survival rates.

Distinct Localization, Transcriptional Profiles, and Functionality in Early Life Tonsil Regulatory T Cells.

CD4+ regulatory T cells (Tregs) are key orchestrators of the immune system, fostering the establishment of protective immunity while preventing deleterious responses. Infancy and childhood are crucial periods of rapid immunologic development, but how Tregs mediate immune responses at these earliest timepoints of human life is poorly understood. In this study, we compare blood and tissue (tonsil) Tregs across pediatric and adult subjects to investigate age-related differences in Treg biology. We observed increased FOXP3 expression and proportions of Tregs in tonsil compared with paired blood samples in children. Within tonsil, early life Tregs accumulated in extrafollicular regions with cellular interactions biased toward CD8+ T cells. Tonsil Tregs in both children and adults expressed transcriptional profiles enriched for lineage defining signatures and canonical functionality compared with blood, suggesting tissue as the primary site of Treg activity. Early life tonsil Tregs transcriptional profiles were further defined by pathways associated with activation, proliferation, and polyfunctionality. Observed differences in pediatric tonsil Treg transcriptional signatures were associated with phenotypic differences, high proliferative capacity, and robust production of IL-10 compared with adult Tregs. These results identify tissue as a major driver of Treg identity, provide new insights into developmental differences in Treg biology across the human lifespan, and demonstrate unique functional properties of early life Tregs.

The tissue-resident regulatory T cell pool is shaped by transient multi-tissue migration and a conserved residency program

The tissues are the site of many important immunological reactions, yet how the immune system is controlled at these sites remains opaque. Recent studies have identified Foxp3+ regulatory T (Treg) cells in non-lymphoid tissues with unique characteristics compared with lymphoid Treg cells. However, tissue Treg cells have not been considered holistically across tissues. Here, we performed a systematic analysis of the Treg cell population residing in non-lymphoid organs throughout the body, revealing shared phenotypes, transient residency, and common molecular dependencies. Tissue Treg cells from different non-lymphoid organs shared Tcell receptor (TCR) sequences, with functional capacity to drive multi-tissue Treg cell entry and were tissue-agnostic on tissue homing. Together, these results demonstrate that the tissue-resident Treg cell pool in most non-lymphoid organs, other than the gut, is largely constituted by broadly self-reactive Treg cells, characterized by transient multi-tissue migration. This work suggests common regulatory mechanisms may allow pan-tissue Treg cells to safeguard homeostasis across the body.

Differences in the characteristics and functions of brain and spinal cord regulatory T cells

T cells play an important role in the acquired immune response, with regulatory T cells (Tregs) serving as key players in immune tolerance. Tregs are found in nonlymphoid and damaged tissues and are referred to as “tissue Tregs”. They have tissue-specific characteristics and contribute to immunomodulation, homeostasis, and tissue repair through interactions with tissue cells. However, important determinants of Treg tissue specificity, such as antigen specificity, tissue environment, and pathology, remain unclear. In this study, we analyzed Tregs in the central nervous system of mice with ischemic stroke and experimental autoimmune encephalomyelitis (EAE), a mouse model of multiple sclerosis. The gene expression pattern of brain Tregs in the EAE model was more similar to that of ischemic stroke Tregs in the brain than to that of spinal cord Tregs. In addition, most T-cell receptors (TCRs) with high clonality were present in both the brain and spinal cord. Furthermore, Gata3+ and Rorc+ Tregs expressed TCRs recognizing MOG in the spinal cord, suggesting a tissue environment conducive to Rorc expression. Tissue-specific chemokine/chemokine receptor interactions in the spinal cord and brain influenced Treg localization. Finally, spinal cord- or brain-derived Tregs had greater anti-inflammatory capacities in EAE mice, respectively. Taken together, these findings suggest that the tissue environment, rather than pathogenesis or antigen specificity, is the primary determinant of the tissue-specific properties of Tregs. These findings may contribute to the development of novel therapies to suppress inflammation through tissue-specific Treg regulation.

IKK2 controls the inflammatory potential of tissue-resident regulatory T cells in a murine gain of function model

Loss-of-function mutations have provided crucial insights into the immunoregulatory actions of Foxp3+ regulatory T cells (Tregs). By contrast, we know very little about the consequences of defects that amplify aspects of Treg function or differentiation. Here we show that mice heterozygous for an Ikbkb gain-of-function mutation develop psoriasis. Doubling the gene dose (IkbkbGoF/GoF) results in dactylitis, spondylitis, and characteristic nail changes, which are features of psoriatic arthritis. IkbkbGoF mice exhibit a selective expansion of Foxp3 + CD25+ Tregs of which a subset express IL-17. These modified Tregs are enriched in both inflamed tissues, blood and spleen, and their transfer is sufficient to induce disease without conventional T cells. Single-cell transcriptional and phenotyping analyses of isolated Tregs reveal expansion of non-lymphoid tissue (tissue-resident) Tregs expressing Th17-related genes, Helios, tissue-resident markers including CD103 and CD69, and a prominent NF-κB transcriptome. Thus, IKK2 regulates tissue-resident Treg differentiation, and overactivity drives dose-dependent skin and systemic inflammation.

Abstract 4280: Regulatory T cells contribute to tumor cell dissemination by extracellular matrix remodeling

Abstract Regulatory T (Treg) cells function to enforce peripheral tolerance and are potent suppressors of tumor immunity. In breast cancer, we have shown that Treg cells promote tumor growth by favoring alternative activation of macrophages via suppression of IFN-γ. The tumor-associated extracellular matrix (ECM) is a key regulator of metastatic dissemination and is emerging as a critical regulator of tumor immunity. However, the reciprocal effects of the immune system on the ECM remain elusive. Using murine in vivo, ex vivo and bioengineering models we describe Treg cell-dependent changes in the tumor ECM that facilitate tumor cell movement and metastatic dissemination. Further, Treg cell-dependent matrisome signatures correlate with survival in human breast cancer samples. This work underscores a previously unrecognized role of Treg cells on the ECM that facilitates metastasis and is consistent with tissue Treg cell emergent function as critical regulators of tissue repair and cancer. Citation Format: Ailen Garcia-Santillan, Jessanne Y. Lichtenberg, He Shen, Mikhail G. Dozmorov, Amy L. Olex, Priscilla Y. Hwang, Paula D. Bos. Regulatory T cells contribute to tumor cell dissemination by extracellular matrix remodeling [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4280.

Intragraft memory-like CD127hiCD4+Foxp3+ Tregs maintain transplant tolerance.

CD4+Foxp3+ regulatory T cells (Tregs) play an essential role in suppressing transplant rejection, but their role within the graft and heterogeneity in tolerance are poorly understood. Here, we compared phenotypic and transcriptomic characteristics of Treg populations within lymphoid organs and grafts in an islet xenotransplant model of tolerance. We showed Tregs were essential for tolerance induction and maintenance. Tregs demonstrated heterogeneity within the graft and lymphoid organs of tolerant mice. A subpopulation of CD127hi Tregs with memory features were found in lymphoid organs, presented in high proportions within long-surviving islet grafts, and had a transcriptomic and phenotypic profile similar to tissue Tregs. Importantly, these memory-like CD127hi Tregs were better able to prevent rejection by effector T cells, after adoptive transfer into secondary Rag-/- hosts, than naive Tregs or unselected Tregs from tolerant mice. Administration of IL-7 to the CD127hi Treg subset was associated with a strong activation of phosphorylation of STAT5. We proposed that memory-like CD127hi Tregs developed within the draining lymph node and underwent further genetic reprogramming within the graft toward a phenotype that had shared characteristics with other tissue or tumor Tregs. These findings suggested that engineering Tregs with these characteristics either in vivo or for adoptive transfer could enhance transplant tolerance.

Calcitriol restrains microglial M1 polarization and alleviates dopaminergic degeneration in hemiparkinsonian mice by boosting regulatory T-cell expansion.

Vitamin D deficiency is a risk factor for Parkinson's disease (PD) and vitamin D supplementation robustly alleviates neurodegeneration in PD models. However, the mechanisms underlying this effect require further clarification. Current evidence suggests that harnessing regulatory T cells (Treg) may mitigate neuronal degeneration. In this study, we investigated the therapeutic effects of vitamin D receptor activation by calcitriol on PD, specifically focusing on its role in Treg. Hemiparkinsonian mice model was established through the injection of 6-OHDA into the striatum. Mice were pretreated with calcitriol before 6-OHDA injection. The motor performance, dopaminergic neuronal survival, contents of dopamine, and dopamine metabolites were evaluated. The pro-inflammatory cytokines levels, T-cell infiltration, mRNA expression of indicated microglial M1/M2 phenotypic markers, and microglial marker in the midbrain were detected. Populations of Treg in the splenic tissues were assessed using a flow cytometry assay. PC61 monoclonal antibody was applied to deplete Treg in vivo. We show that calcitriol supplementation notably improved motor performance and reduced dopaminergic degeneration in the 6-OHDA-induced PD model. Mechanistically, calcitriol promoted anti-inflammatory/neuroprotective Treg and inhibited pro-inflammatory/neurodestructive effector T-cell generation in this model. This process significantly inhibited T-cell infiltration in the midbrain, restrained microglial activation, microglial M1 polarization, and decreased pro-inflammatory cytokines release. This more favorable inflammatory microenvironment rescued dopaminergic degeneration. To further verify that the anti-inflammatory effects of calcitriol are associated with Treg expansion, we applied an antibody-mediated Treg depletion assay. As predicted, the anti-inflammatory effects of calcitriol in the PD model were diminished following Treg depletion. These findings suggest that calcitriol's anti-inflammatory and neuroprotective effects in PD are associated with its potential to boost Treg expansion.

Adipose-tissue regulatory T cells are a consortium of subtypes that evolves with age and diet

Foxp3+CD4+ regulatory T (Treg) cells found within tissues regulate local immunity, inflammation, and homeostasis. Tregs in epididymal visceral adipose tissue (eVAT) are critical regulators of local and systemic inflammation and metabolism. During aging and under obesogenic conditions, eVAT Tregs undergo transcriptional and phenotypic changes and are important for containing inflammation and normalizing metabolic indices. We have employed single-cell RNA sequencing, single-cell Tra and Trb sequencing, adoptive transfers, photoconvertible mice, cellular interaction analyses, and in vitro cultures to dissect the evolving heterogeneity of eVAT Tregs with aging and obesity. Distinct Treg subtypes with distinguishable gene expression profiles and functional roles were enriched at differing ages and with differing diets. Like those in lean mice, eVAT Tregs in obese mice were not primarily recruited from the circulation but instead underwent local expansion and had a distinct and diversified T cell receptor repertoire. The different eVAT-Treg subtypes were specialized in different functions; for example, the subtypes enriched in lean, but not obese, mice suppressed adipogenesis. The existence of functionally divergent eVAT-Treg subtypes in response to obesogenic conditions presents possibilities for precision therapeutics in the context of obesity.

Upregulation of IDO gene expression reduces the immunogenicity of epidermal cells and strengthens the immune protection of epidermal cells during transplantation treatment of wounds

BackgroundEpidermal cell transplantation is a feasible treatment option for large wounds; however, sources of autologous epidermal cells are often limited. Allogeneic epidermal cells can be cultured conveniently; however, related immune rejection needs to be addressed. Herein, we hypothesized that the immunogenicity of epidermal cells with high indoleamine 2,3-dioxygenase (IDO) expression may be reduced by gene transfection. Methods/ResultsTo test this hypothesis, we obtained stable transfectants by transfecting epidermal stem cells with a lentiviral vector encoding the IDO gene and screening them for puromycin resistance (a marker for successful transfection). The phenotype tested using cell counting kit −8 and Transwell assays confirmed that IDO-transfected epidermal cells maintained their characteristics. Co-culture of IDO-transfected epidermal cells with allogeneic CD4+ T cells in vitro showed that the upregulation of IDO expression in epidermal cells inhibited the proliferation of CD4+ T cells (P < 0.001, P < 0.001, and P < 0.001, respectively) and promoted their apoptosis (P = 0.00028, P = 0.0006, and P = 0.00247, respectively) and transformation into functional regulatory T cells (Tregs) (P = 0.0051, P = 0.0132, and P = 0.0248, respectively) compared with Con, NC, and 1-MT groups. The increased proportion of Tregs may be related to the overexpression of IDO, which promoted the expression of transforming growth factor beta (TGF-β) (P = 0.0001, P = 0.0013, and, P = 0.0009) and interleukin (IL) 10 (IL-10) (P = 0.0062, P = 0.0058, and P = 0.0119) while inhibited the expression of IL-2 (P = 0.0012, P = 0.0126, and P = 0.0066). We further verified these effects in vivo as transplanted IDO-transfected epidermal stem cells were effective in treating wounds in mice. On days 5 and 7, wounds treated with IDO cells healed faster than those in the other groups (day 5: P = 0.012 and P = 0.0136; day 7: P = 0.0242 and P = 0.0187, respectively), whereas this effect was significantly inhibited by 1-methyltryptophan (1-MT) (day 5: P = 0.0303; day 7: P = 0.0105). Immunofluorescence staining detected IDO and CD4+ Foxp3+ Tregs in the transplanted wounds, which may promote Foxp3+ Tregs in the wound tissue (day 5: P < 0.0001, P < 0.0001, and P < 0.0001; day 7: P < 0.0001, P < 0.0001, and P < 0.0001), respectively) and decrease CD4+ T cells (day 5: P < 0.0001, P < 0.0001, and P < 0.0001; day 7: P < 0.0001, P < 0.0001, and P < 0.0001). ConclusionOur results suggest that the upregulation of IDO expression in epidermal stem cells can reduce their immunogenicity by promoting Tregs, thus inducing the immune protection of epidermal stem cells.

Plexin B1 controls Treg numbers, limits allergic airway inflammation, and regulates mucins.

We investigated the effect of global Plexin B1 deficiency on allergic airway responses to house dust mite (HDM) or ovalbumin (OVA). In the HDM model, there were higher Th2 cytokine levels in the BALF of Plexin B1 knock-out (KO) mice compared to wild type (WT), and tissue inflammation and mucus production were modestly enhanced. In the OVA model, Plexin B1 deficiency led to increases in lung inflammation, mucus production, and lung Th2 cytokines accompanied by dysregulated mucin gene expression without affecting anti-OVA IgE/IgG1 levels. Spleen cells from Plexin B1 KO mice proliferated more robustly than WT cells in vitro to a variety of stimuli. Plexin B1 KO CD4+ T cells from spleens expressed higher levels of Ki-67 and CD69 compared to WT cells. Spleen cells from naïve Plexin B1 KO mice secreted increased amounts of IL-4 and IL-6 when pulsed in vitro with OVA whereas in vivo OVA-primed spleen cells produced IL-4/IL-5 when subjected to in vitro OVA restimulation. The upregulated allergic inflammatory response in Plexin B1 KO mice was associated with a lower number of Tregs in the lung tissues. Moreover, these mice displayed lower numbers of Treg cells in the lymphoid tissues at the baseline. These results demonstrate a previously unrecognized link between Plexin B1, Treg cells, and mucus in allergic lung inflammation.

Genistein Alleviates Dextran Sulfate Sodium-induced Ulcerative Colitis in Mice by Regulating Th17/Treg Cell Balance: Implication for the G Protein-coupled Estrogen Receptor

Objectives This study aimed to evaluate the therapeutic effects and mechanism of genistein on dextran sulfate sodium-induced mouse model of ulcerative colitis (UC). Materials and Methods A DSS-induced mouse model for UC was used in this study. Mice were then treated with genistein or a combination of genistein and the G protein-coupled estrogen receptor (GPER) antagonist G15. Colon length, disease activity index (DAI), spleen index (SI), histopathological alterations, and integrity of the colonic barrier were evaluated. The expression of inflammatory cytokines and antioxidant capacity were detected. Populations of T helper 17 cells (Th17) and Regulatory T cells (Treg) in the mesenteric lymph nodes (MLN) were analyzed. The expression of a transcription factor for Th17 and Treg in the colonic tissues was detected. Results Genistein treatment significantly alleviated DSS-induced colitis, summarized as increased colonic length, decreased DAI, SI, improved histopathological alterations, and colonic barrier integrity. Genistein treatment restrained pro-inflammatory cytokines and oxidative enzyme release while promoting anti-inflammatory and anti-oxidative enzyme release. Flow cytometry indicated that genistein significantly reduced the Th17 population while boosting Treg populations. Furthermore, genistein inhibited the expression of transcription factors associated with Th17 and promoted the expression of transcription factors associated with Treg in the colonic tissue. Intriguingly, these observed effects of genistein were abolished when UC mice were treated with a combination of genistein and GPER antagonist G15. Conclusion This study suggests that genistein is potent in protecting against DSS-induced colonic injuries by rebalancing Th17/Treg and that GPER may be a vital target for genistein-mediated immunomodulatory effects in UC.

Immunohistochemical analyses reveal FoxP3 expressions in spleen and colorectal cancer in mice treated with AOM/DSS, and their suppression by glycyrrhizin.

We previously demonstrated that glycyrrhizin (GL) suppressed inflammation and carcinogenesis in an azoxymethane (AOM)/dextran sodium sulfate (DSS)-induced murine model of colorectal cancer (CC). In this study, we found an accumulation of regulatory T cells (Tregs) in the spleen and suppression by GL in model mice. ICR mice were divided into four groups: Control, GL, CC, and GL-treated CC (CC+GL), and were sacrificed 20 weeks after AOM/DSS treatment. We measured spleen weight, areas of white and red pulp, and CD8+ T cells (cytotoxic T lymphocytes, CTL), and CD11c-positive cells (dendritic cells) in splenic tissues and forkhead box protein 3 (FoxP3)-positive cells (Tregs) in colorectal and splenic tissues. In all cases, the CC group showed a significant increase compared with those in Control group, and GL administration significantly attenuated this increase. These results indicate that Tregs accumulated in the spleen may participate in inflammation-related carcinogenesis by suppressing CTL. We also suggest that GL which binds to high-mobility group box 1 (HMGB1), suppresses carcinogenesis with decreasing Tregs in the spleen. Furthermore, there was an expression of FoxP3 in cancer cells, indicating that it may be involved in the malignant transformation of cancer cells.

The Regulation of FOXP3 Expression and Its Therapeutic Potential in Non-Small Lung Cancer

Non-small cell cancer (NSCLC) is today one of the most threatening malignant tumors, and it is still rarely cured by current standard therapies with a high mortality rate. The transcription factor Forkhead box p3 (FOXP3) is a potential molecular target for NSCLC since it has been identified with an oncogenic effect in the tumor microenvironment (TME). It is well known that FOXP3 expression could characterize the regulatory T cells (Treg), which plays a critical role in regulating anti-tumor immune responses in TME. Through analyzing the basic mechanisms of FOXP3 and Treg in distinct tumor tissues, they have been found with contradictory expression and the frequency in different TME. In NSCLC, the suppressive function of Treg to immune systems and frequency of FOXP3 is accumulated to promote the development of tumors. Most recently, accumulating evidence reveals the therapeutic potential of targeting FOXP3 and the related signaling pathways in the treatment of NSCLC patients. Although unsolved side effects and limitations that still must be considered when targeting FOXP3 and FOXP3-related proteins, there is no doubt that they are potential therapeutic applications for NSCLC treatment. The main objective of this review is to mention the structure, function and recent progress in treatment targeting FOXP3 and FOXP3-related signaling pathways, especially to therapies with immune checkpoint inhibitors.

F. prausnitzii-derived extracellular vesicles attenuate experimental colitis by regulating intestinal homeostasis in mice

BackgroundEmerging evidence has shown that extracellular vesicles (EVs) derived from gut bacteria play a crucial role in microbiota-host interactions. Here, we aimed to evaluate the attenuating effect of EVs derived from a reduced commensal bacterium, F. prausnitzii (Fp-EVs), in inflammatory bowel disease (IBD) on dextran sulfate sodium (DSS)-induced colitis in mice.ResultsFp-EVs isolated by ultracentrifugation and typically exhibited a double concave disc shape with an average diameter of 172 nm. Fp-EVs treatment reduced DSS-induced weight loss, disease activity index (DAI) score, colon length shortening, histological damage, neutrophil infiltration and increased intestinal epithelial apoptotic cells in DSS-induced colitis mice. Fp-EVs upregulated the protein expression of zona occludens (ZO)-1 and Occludin and increased the ratio of Tregs in the colon tissue of colitis mice. Furthermore, Fp-EVs downregulated the expression of the proinflammatory cytokines interleukin-1β (IL-1β), IL-2, IL-6, IL-12a, IL-17a, Interferon-γ (IFN-γ), tumor necrosis factor - α (TNF-α), granulocyte-macrophage colony stimulating factor (GM-CSF) and upregulated the anti-inflammatory cytokines IL-4, IL-10, and transforming growth factor β (TGF-β) in DSS-treated mice. Moreover, Fp-EV treatment markedly reduced the phosphorylation of these proteins Nuclear factor-κB (NF-κB) and Mitogen activated protein kinase (MAPK), and regulated the expression of nuclear factor erythroid 2-related factor (Nrf2) and heme oxygenase-1 (HO-1).ConclusionOur findings revealed that Fp-EVs attenuated DSS-induced colitis by modulating the intestinal mucosal barrier function and immunological profile. Our findings reveal that Fp-EVs attenuate DSS-induced colitis by modulating intestinal mucosal barrier function and the immunological profile.

CD4+CCR8+ Tregs in ovarian cancer: a potential effector Tregs for immune regulation

BackgroundTregs are key drivers of immunosuppression in solid tumors. As an important chemokine receptor on Tregs, the regulatory effect of CCR8 on tumor immunity has received more and more attention. However, the current research on CCR8 in the immune microenvironment of ovarian cancer has not been clear.MethodsBioinformatics analysis was used to compare the transcriptome differences between CD4+ T cells in the peripheral circulation and infiltrated in ovarian tumor tissues. RT-PCR was used to detect the expression levels of chemokine receptor-related differential genes on CD4+ T cells in peripheral blood and ovarian tumor tissues. Multiparameter flow cytometry was used to detect the proportion and phenotypic characteristics of CD4+CCR8+ Tregs and CD4+CCR8− Tregs in different sample types. The expression level of CCR8 ligands was detected at multiple levels. To explore the important role of CCR8-CCL1 and CCR8-CCL18 axis in the migration and invasion of CD4+CCR8+ Tregs into ovarian tumor tissues by establishing a chemotaxis system in vitro.ResultsIn this study, significantly different gene expression profiles were found between peripheral circulating CD4+ T cells and infiltrating CD4+ T cells in ovarian tumor tissues, in which chemokine-chemokine receptor signaling pathway was significantly enriched in all three groups of differential genes. The expression level of CCR8 in infiltrating CD4+ T cells of ovarian cancer tissue was significantly higher than that in peripheral blood of healthy controls and ovarian cancer patients, and high expression of CCR8 was significantly correlated with advanced tumor stage and poor differentiation. CD4+CCR8+ Tregs are the main type of infiltrating CD4+ Tregs in ovarian tumor tissues, which have stronger immunosuppressive phenotypes, secrete more inhibitory cytokines and have stronger proliferation ability. The ligands CCL1 and CCL18 corresponding to CCR8 were significantly overexpressed in ovarian tumor tissues, and the CCR8-CCL1 and CCR8-CCL18 axis played a key role in the migration and infiltration of CD4+CCR8+ Tregs into ovarian tumor tissues.ConclusionsThe results of this study may help to understand the phenotypic characteristics and recruitment process of Tregs in the tumor, and provide new ideas for improving the immunosuppressive status of the ovarian cancer microenvironment.

Jiawei Maxing Shigan Tang alleviates radiation-induced lung injury via TGF-β1/Smad signaling pathway mediated by regulatory T cells

Ethnopharmacological relevanceRadiation-induced lung injury (RILI) is a common complication during thoracic radiotherapy which impairs the quality of life in patients and limits radiation doses. Jiawei Maxing Shigan Tang (JMST), which is a modified decoction made of Ephedra, Apricot Kernel, Gypsum, and Licorice, can alleviate the symptoms of RILI in patients. Previous studies and preliminary findings suggested a potential molecular mechanism of JMST in the treatment of RILI. Further studies are needed. Aim of the studyTo elucidate the mechanisms of how regulatory T cells (Tregs) promote RILI and the effect of JMST on Tregs, as well as the corresponding pathway. Materials and methodsCD4+CD25+ Tregs were isolated from rats, and the supernatant's TGF-β1 level was examined by using enzyme-linked immunosorbent assay (ELISA). Type II alveolar epithelial cells (AECs) were co-cultured with the supernatant of Tregs, and the expression levels of epithelial-to-mesenchymal transition (EMT)-related and TGF-β1/Smad signaling pathway-related proteins were analyzed by using western blotting (WB). Afterward, the Tregs were incubated with different concentrations of JMST. The cell viability and TGF-β1 concentration were confirmed by cell counting kit-8 (CCK-8) assay and ELISA, respectively. The optimized concentration of JMST was applied in vitro and vivo experiments. The specific mechanism was investigated through the combination of using flow cytometry, lung histopathology analysis, ELISA, and WB. ResultsRadiation could promote Tregs to secrete TGF-β1. After radiation, the expression levels of Smad2/3, phosphorylated Smad2/3 (p-Smad2/3), Smad4 and mesenchymal markers Vimentin and α-SMA were all increased, while the expression level of epithelial markers E-cadherin was decreased. The expression levels of these proteins were reversed after interventions involving Treg cell activation inhibition or TGF-β1 receptor inhibitor. JMST reduced the number of Tregs in lung tissue and alleviated the degree of pulmonary fibrosis. The expression of Smad2/3, p-Smad2/3, Smad4, TGF-β1, Vimentin, and α-SMA were significantly downregulated, while the E-cadherin was upregulated, through the intervention of JMST. ConclusionTregs could mediate EMT through TGF-β1/Smad pathway. JMST inhibits EMT via TGF-β1/Smad pathway by regulating Tregs, therefore alleviating RILI.