Fragile Treg cells: Traitors in immune homeostasis?

Regulatory T (Treg) cells play a key role in maintaining immune tolerance and tissue homeostasis. However, in some disease microenvironments, Treg cells exhibit fragility, which manifests as preserved FoxP3 expression accompanied by inflammation and loss of immunosuppression. Fragile Treg cells are formatively, phenotypically and functionally diverse in various diseases, further complicating the role of Treg cells in the immunotherapeutic response and offering novel targets for disease treatment by modulating specific Treg subsets. In this review, we summarize findings on fragile Treg cells to provide a framework for characterizing the formation and role of fragile Treg cells in different diseases, and we discuss how this information may guide the development of more specific Treg-targeted immunotherapies.

Prognosis comparison between hepatocellular carcinoma patients with microvascular invasion who received hepatectomy alone and those who underwent early PA-TACE: a retrospective cohort study.

Background: Postoperative adjuvant transcatheter arterial chemoembolization (PA-TACE) can achieve longer overall survival (OS) and disease-free survival (DFS) in hepatocellular carcinoma (HCC) patients with microvascular invasion (MVI). We investigated whether this treatment strategy could benefit these patients by mediating the dysfunctional immunological status. Therefore, a retrospective cohort study was conducted to investigate the effect of early PA-TACE in HCC patients with MVI by measuring the levels of T helper cell 17 (Th17) and regulatory T cell (Treg). Methods: This study retrospectively included 472 patients with HCC undergoing hepatectomy between December 2015 and December 2018, and 115 patients with MVI confirmed by postoperative pathology were enrolled and divided into two groups of TACE group and non-TACE group according to whether TACE was performed. HCC patients with MVI. The proportion of Treg and Th17 cells in peripheral blood was measured one day before and four weeks after TACE. All patients in the two groups were followed up until death or until the study ended in December 2023. The rates of OS and progression-free survival (PFS) in patients with MVI were compared between those who received hepatectomy alone and those who underwent early PA-TACE. Results: Among 115 HCC patients with MVI from 472 patients, the study enrolled 51 patients with PA-TACE into the TACE group and 42 patients without TACE into the non-TACE group. There were no statistical differences in baseline data between the two groups (all P>0.05). The frequency of Treg among CD4+ T cells in HCC patients with PA-TACE was significantly lower than baseline (7.34%±3.61% vs. 5.82%±2.76%, P<0.001), and the frequency of Th17 among CD4+ T cells in these patients was significantly higher than baseline (0.49%±0.28% vs. 0.50%±0.25%, P<0.001). Among all the patients, the median OS was 61.8 months. The OS rate and PFS rate at 12, 36, and 60 months in the TACE group were significantly higher than those in the non-TACE group (all P<0.05). Conclusions: PA-TACE may have roles in improving survival outcomes, and restoring immune homeostasis in HCC patients with MVI after hepatectomy.

MEK-inhibitor treatment reduces the induction of regulatory T cells in mice after influenza A virus infection.

Regulatory T cells (Tregs) play a crucial and complex role in balancing the immune response to viral infection. Primarily, they serve to regulate the immune response by limiting the expression of proinflammatory cytokines, reducing inflammation in infected tissue, and limiting virus-specific T cell responses. But excessive activity of Tregs can also be detrimental and hinder the ability to effectively clear viral infection, leading to prolonged disease and potential worsening of disease severity. Not much is known about the impact of Tregs during severe influenza. In the present study, we show that CD4+/CD25+FoxP3+ Tregs are strongly involved in disease progression during influenza A virus (IAV) infection in mice. By comparing sublethal with lethal dose infection in vivo, we found that not the viral load but an increased number of CD4+/CD25+FoxP3+ Tregs may impair the immune response by suppressing virus specific CD8+ T cells and favors disease progression. Moreover, the transfer of induced Tregs into mice with mild disease symptoms had a negative and prolonged effect on disease outcome, emphasizing their importance for pathogenesis. Furthermore, treatment with MEK-inhibitors resulted in a significant reduction of induced Tregs in vitro and in vivo and positively influenced the progression of the disease. Our results demonstrate that CD4+/CD25+FoxP3+ Tregs are involved in the pathogenesis of severe influenza and indicate the potential of the MEK-inhibitor zapnometinib to modulate CD4+/CD25+FoxP3+ Tregs. Thus, making MEK-inhibitors even more promising for the treatment of severe influenza virus infections.

Exosomes derived from cord blood Treg cells promote diabetic wound healing by targeting monocytes.

Chronic nonhealing diabetic wounds are a critical clinical challenge. Regulatory T cells (Tregs) are immunosuppressive modulators affecting wound healing progression by controlling the inflammatory response. The current study attempted to investigate whether the exosomes derived from cord blood (CB) Tregs can accelerate the healing process. Exosomes were isolated from CB-Treg cultures using ultracentrifugation and validated with different specific markers of exosomes. The purified CB-Treg-derived exosomes were co-cultured with peripheral blood mononuclear cells (PBMCs) and CD14+ monocytes. The migration-promoting effect of CB-Treg-derived exosomes on fibroblasts and endothelial cells was investigated. We used thermosensitive Pluronic F-127 hydrogel (PF-127) loaded with CB-Treg-derived exosomes in a diabetic wound healing mouse model. CB-Treg-derived exosomes with 30-120 nm diameters revealed exosome-specific markers, such as TSG101, Alix, and CD63. CB-Treg-derived exosomes were mainly bound to the monocytes when co-cultured with PBMCs, and promoted monocyte polarization to the anti-inflammatory phenotype (M2) in vitro. CB-Treg-derived exosomes enhanced the migration of endothelial cells and fibroblasts. Furthermore, CB-Treg-derived exosomes treatment accelerated wound healing by downregulating inflammatory factor levels and upregulating the M2 macrophage ratio in vivo. Our findings indicated that CB-Treg-derived exosomes could be a promising cell-free therapeutic strategy for diabetic wound healing, partly by targeting monocytes.

Regulatory T Cell and T Helper 17 Cell Imbalance in Patients with Unexplained Infertility.

Purpose: Female infertility is a global health concern. The aim of this study was to investigate the relationship between regulatory T (Treg) cells and helper T cells 17 (Th17) in peripheral blood and unexplained infertility (UI). In addition, we explored potential valuable diagnostic biomarkers for patients with UI and ascertained whether Treg and Th17 cells are associated with primary and secondary UI. Patients and Methods: The patients underwent standard fertility evaluation test, including blood tests, ultrasound examination, fallopian tube tests, ovulation assessment, and male partner's semen analysis. According to the inclusion and exclusion criteria, this study enrolled 37 patients with UI (30 with primary UI and 7 with secondary UI) and 26 age-matched healthy volunteers as the control group. Flow cytometry was used to detect the frequency of Treg and Th17 cells. The area under the receiver operating characteristic curve (AUC) with a 95% confidence interval (CI) was used to assess the diagnostic performance. An AUC > 0.800 indicated good diagnostic performance. Results: The percentage of Treg decreased significantly, whereas the percentage and absolute count of Th17 cells increased. Moreover, the Th17/Treg ratio in patients with UI increased significantly. As a diagnostic biomarker for UI, the Th17/Treg ratio exhibited remarkable diagnostic performance (AUC: 0.813 (95% CI = 0.709-0.917)). However, the percentages and absolute counts of Treg and Th17 cells in the peripheral blood of women with primary and secondary UI, as well as their Th17/Treg ratios, did not differ significantly. Conclusion: The distribution of Treg and Th17 cells is imbalanced in patients with UI. Therefore, the Th17/Treg ratio may be a promising indicator of UI. However, there were no significant differences in the distribution of Treg and Th17 cells between women with primary and secondary UI.

Combined oral low-dose cyclophosphamide endocrine therapy may improve clinical response among patients with metastatic breast cancer via Tregs in TLSs.

Despite limited research on refractory and/or endocrine therapy failure in elderly metastatic breast cancer (MBC) patients, a prior study showed that low-dose oral cyclophosphamide (CY) can improve the overall survival rate of MBC patients, possibly through the immunoregulation of regulatory T cells (Tregs). We preliminarily investigated the combination of endocrine therapy (ET) with oral low-dose CY as salvage therapy in elderly patients via peripheral blood regulatory T-cell analyses. In addition, we evaluated the associations of tumor tertiary lymphoid structures (TLSs) with therapeutic outcomes. HR+/HER2- advanced breast cancer patients who received low-dose CY combined with ET or ET only from April 2015 to August 2021 were enrolled in this retrospective study. The primary outcome was the clinical control rate (CCR), and the secondary outcome was progression-free survival (PFS). Circulating T lymphocyte subpopulations represented by Tregs were monitored during treatment by flow cytometry methods. TLSs wereconfirmed by hematoxylin-eosin staining of pretreatment specimens, and CD3, CD4, and Foxp3 were detected using Opal multicolor immunofluorescence. A total of 85 patients who received CY + ET and 50 patients who received ET only were enrolled, the percentage of patients who received CCR was 73% (62/85) vs. 70% (45/50), and the objective response rate (ORR) was 28% (24/85) vs. 24% (12/50). No deaths occurred during the study period. The mean PFS time was 13 vs. 11 months (P = 0.03). In the CY + ET group, decreases in CD4+/CD25+/Foxp3+ T cells (P < 0.001) were favorable for both clinical control and prolonged PFS (P < 0.001). Compared with patients without TLSs, those with TLSs were more likely to have better clinical control and PFS (mean time = 6 months), and a greater number of Treg cells during TLS pretreatment correlated with longer PFS (P = 0.043). Oral low-dose CY combined with standard ET exerts immunological effects by decreasing Treg levels to achieve improved clinical responses. Moreover, patients with TLSs might benefit more from such therapy than those without TLSs, and a high Treg cell count in TLSs before treatment predicts better therapeutic efficacy.

Exploring risk factors for autoimmune diseases complicated by non-hodgkin lymphoma through regulatory T cell immune-related traits: a Mendelian randomization study.

Background: The effect of immune cells on autoimmune diseases (ADs) complicated by non-Hodgkin lymphoma (NHL) has been widely recognized, but a causal relationship between regulatory T cell (Treg) immune traits and ADs complicated by NHL remains debated. Methods: Aggregate data for 84 Treg-related immune traits were downloaded from the Genome-Wide Association Study (GWAS) catalog, and GWAS data for diffuse large B-cell lymphoma (DLBCL; n=315243), follicular lymphoma (FL; n=325831), sjögren's syndrome (SS; n=402090), rheumatoid arthritis (RA; n=276465), dermatopolymyositis (DM; n=311640), psoriasis (n=407876), atopic dermatitis (AD; n=382254), ulcerative colitis (UC; n=411317), crohn's disease(CD; n=411973) and systemic lupus erythematosus (SLE; n=307587) were downloaded from the FinnGen database. The inverse variance weighting (IVW) method was mainly used to infer any causal association between Treg-related immune traits and DLBCL, FL, SS, DM, RA, Psoriasis, AD, UC, CD and SLE, supplemented by MR-Egger, weighted median, simple mode, and weighted mode. Moreover, we performed sensitivity analyses to assess the validity of the causal relationships. Results: There was a potential genetic predisposition association identified between CD39+ CD8br AC, CD39+ CD8br % T cell, and the risk of DLBCL (OR=1.51, p<0.001; OR=1.25, p=0.001) (adjusted FDR<0.1). Genetic prediction revealed potential associations between CD25++ CD8br AC, CD28- CD25++ CD8br % T cell, CD39+ CD8br % CD8br, and the risk of FL (OR=1.13, p=0.022; OR=1.28, p=0.042; OR=0.90, p=0.016) (adjusted FDR>0.1). Furthermore, SLE and CD exhibited a genetically predicted potential association with the CD39+ CD8+ Tregs subset. SS and DM were possibly associated with an increase in the quantity of the CD4+ Tregs subset; RA may have reduced the quantity of the CD39+ CD8+ Tregs subset, although no causal relationship was identified. Sensitivity analyses supported the robustness of our findings. Conclusions: There existed a genetically predicted potential association between the CD39+ CD8+ Tregs subset and the risk of DLBCL, while SLE and CD were genetically predicted to be potentially associated with the CD39+ CD8+ Tregs subset. The CD39+ CD8+ Tregs subset potentially aided in the clinical diagnosis and treatment of SLE or CD complicated by DLBCL.

Near-infrared duocarmycin photorelease from a Treg-targeted antibody-drug conjugate improves efficacy of PD-1 blockade in syngeneic murine tumor models.

Regulatory T cells (Tregs) play a crucial role in mediating immunosuppression in the tumor microenvironment. Furthermore, Tregs contribute to the lack of efficacy and hyperprogressive disease upon Programmed cell death protein 1 (PD-1) blockade immunotherapy. Thus, Tregs are considered a promising therapeutic target, especially when combined with PD-1 blockade. However, systemic depletion of Tregs causes severe autoimmune adverse events, which poses a serious challenge to Treg-directed therapy. Here, we developed a novel treatment to locally and predominantly damage Tregs by near-infrared duocarmycin photorelease (NIR-DPR). In this technology, we prepared anti-CD25 F(ab')2 conjugates, which site-specifically uncage duocarmycin in CD25-expressing cells upon exposure to NIR light. In vitro, CD25-targeted NIR-DPR significantly increased apoptosis of CD25-expressing HT2-A5E cells. When tumors were irradiated with NIR light in vivo, intratumoral CD25+ Treg populations decreased and Ki-67 and Interleukin-10 expression was suppressed, indicating impaired functioning of intratumoral CD25+ Tregs. CD25-targeted NIR-DPR suppressed tumor growth and improved survival in syngeneic murine tumor models. Of note, CD25-targeted NIR-DPR synergistically enhanced the efficacy of PD-1 blockade, especially in tumors with higher CD8+/Treg PD-1 ratios. Furthermore, the combination therapy induced significant anti-cancer immunity including maturation of dendritic cells, extensive intratumoral infiltration of cytotoxic CD8+ T cells, and increased differentiation into CD8+ memory T cells. Altogether, CD25-targeted NIR-DPR locally and predominantly targets Tregs in the tumor microenvironment and synergistically improves the efficacy of PD-1 blockade, suggesting that this combination therapy can be a rational anti-cancer combination immunotherapy.

Transcriptional switches in melanoma T Cells: Facilitating polarizing into regulatory T cells.

Melanoma is a malignant skin tumor with a high mortality rate. Regulatory T cells (Tregs) are immune cells with immunosuppressive roles, however, the precise mechanisms governing Treg involvement in melanoma remain enigmatic. Experimental findings unveiled different transcription factor switches between normal and tumor T cell, with heightened FOXP3 and BATF in the latter. These factors induced immunosuppressive molecules and Treg maintenance genes, polarizing tumor T cells into Tregs. Spatial transcriptomics illuminated the preferential settlement of Tregs at the melanoma periphery. Within this context, FOXP3 in Tregs facilitated direct enhancement of specific ligand gene expression, fostering communication with neighboring cells. Novel functional molecules bound to FOXP3 or BATF in Tregs, such as SPOCK2, SH2D2A, and ligand molecules ITGB2, LTA, CLEC2C, CLEC2D, were discovered, which had not been previously reported in melanoma Treg studies. Furthermore, we validated our findings in a large number of clinical samples and identified the Melanoma Treg-Specific Regulatory Tag Set (Mel TregS). ELISA analysis showed that the protein levels of Mel TregS in melanoma Tregs were higher than in normal Tregs. We then utilized SERS technology to measure the signal values of Mel TregS in exosome, and successfully discriminated between healthy individuals and melanoma patients, as well as early and late-stage patients. This approach significantly enhanced detection sensitivity. In sum, our research elucidated fresh insights into the mechanisms governing Treg self-maintenance and communication with surrounding cells in melanoma. We also introduced an innovative method for clinical disease monitoring through SERS technology.

Modified spatial architecture of regulatory T cells after neoadjuvant chemotherapy in non-small cell lung cancer patients.

It is crucial to decipher the modulation of regulatory T cells (Tregs) in tumor microenvironment (TME) induced by chemotherapy, which may contribute to improving the efficacy of neoadjuvant chemoimmunotherapy in resectable non-small cell lung cancer (NSCLC). We retrospectively collected specimens from patients with II-III NSCLC, constituting two cohorts: a neoadjuvant chemotherapy (NAC) cohort (N = 141) with biopsy (N = 58) and postoperative specimens (N = 141), and a surgery-only cohort (N = 122) as the control group. Then, the cell density (Dens), infiltration score (InS), and Treg-cell proximity score (TrPS) were conducted using a panel of multiplex fluorescence staining (Foxp3, CD4, CD8, CK, CD31, ɑSMA). Subsequently, the association of Tregs with cancer microvessels (CMVs) and cancer-associated fibroblasts (CAFs) was analyzed. Patients with NAC treatment have a higher density of Tregs in both paired (P < 0.001) and unpaired analysis (P = 0.022). Additionally, patients with NAC treatment showed higher infiltration score (paired, P < 0.001; unpaired, P = 0.014) and more CD8+T cells around Tregs (paired/unpaired, both P < 0.001). Subgroup analysis indicated that tumors with a diameter of ≤ 5 cm exhibited increase in both Dens(Treg) and InS(Treg), and gemcitabine, pemetrexed and taxel enhanced Dens(Treg) and TrPS(CD8) following NAC. Multivariate analysis identified that the Dens(Tregs), InS(Tregs) and TrPS(CD8) were significantly associated with better chemotherapy response [OR = 8.54, 95%CI (1.69, 43.14), P = 0.009; OR = 7.14, 95%CI (1.70, 30.08), P = 0.024; OR = 5.50, 95%CI (1.09, 27.75), P = 0.039, respectively] and positive recurrence-free survival [HR = 3.23, 95%CI (1.47, 7.10), P = 0.004; HR = 2.70; 95%CI (1.27, 5.72); P = 0.010; HR = 2.55, 95%CI (1.21, 5.39), P = 0.014, respectively]. Moreover, TrPS(CD8) and TrPS(CD4) were negatively correlated with the CMVs and CAFs. These discoveries have deepened our comprehension of the immune-modulating impact of chemotherapy and underscored that the modified spatial landscape of Tregs after chemotherapy should be taken into account for personalized immunotherapy, aiming to ultimately improve clinical outcomes in patients with NSCLC.

Mitochondrial Oxidative Stress Regulates FOXP3+ T-Cell Activity and CD4-Mediated Inflammation in Older Adults with Frailty.

In healthy older adults, the immune system generally preserves its response and contributes to a long, healthy lifespan. However, rapid deterioration in immune regulation can lead to chronic inflammation, termed inflammaging, which accelerates pathological aging and diminishes the quality of life in older adults with frailty. A significant limitation in current aging research is the predominant focus on comparisons between young and older populations, often overlooking the differences between healthy older adults and those experiencing pathological aging. Our study elucidates the intricate immunological dynamics of the CD4/Treg axis in frail older adults compared to comparable age-matched healthy older adults. By utilizing publicly available RNA sequencing and single-cell RNA sequencing (scRNAseq) data from peripheral blood mononuclear cells (PBMCs), we identified a specific Treg cell subset and transcriptional landscape contributing to the dysregulation of CD4+ T-cell responses. We explored the molecular mechanisms underpinning Treg dysfunction, revealing that Tregs from frail older adults exhibit reduced mitochondrial protein levels, impairing mitochondrial oxidative phosphorylation. This impairment is driven by the TNF/NF-kappa B pathway, leading to cumulative inflammation. Further, we gained a deeper understanding of the CD4/Treg axis by predicting the effects of gene perturbations on cellular signaling networks. Collectively, these findings highlight the age-related relationship between mitochondrial dysfunction in the CD4/Treg axis and its role in accelerating aging and frailty in older adults. Targeting Treg dysfunction offers a critical basis for developing tailored therapeutic strategies aimed at improving the quality of life in older adults.

Update on Early-Life T Cells: Impact on Oral Rotavirus Vaccines.

Rotavirus infection continues to be a significant public health problem in developing countries, despite the availability of several vaccines. The efficacy of oral rotavirus vaccines in young children may be affected by significant immunological differences between individuals in early life and adults. Therefore, understanding the dynamics of early-life systemic and mucosal immune responses and the factors that affect them is essential to improve the current rotavirus vaccines and develop the next generation of mucosal vaccines. This review focuses on the advances in T-cell development during early life in mice and humans, discussing how immune homeostasis and response to pathogens is established in this period compared to adults. Finally, the review explores how this knowledge of early-life T-cell immunity could be utilized to enhance current and novel rotavirus vaccines.

Antimetastatic effect of intratumoral Treg antagonists in mice with renal cancer.

The interplay of regulatory T cells (Tregs) within the tumour microenvironment presents a significant challenge in anticancer immunotherapy. This study investigates the potential of Treg blockade to enhance the efficiency of effector T cells. Two distinct treatment cocktails were examined: 3p-hpRNA (5' triphosphate hairpin RNA) combined with unmethylated CpG oligonucleotide (CpG); CpG in combination with OX40 receptor-specific monoclonal antibody (anti-OX40). Treatment efficacy was assessed using a murine model of kidney adenocarcinoma.Renca cells (renal cortical cells with adenocarcinoma) were subcutaneously engrafted in 30 BALB/c mice, then animals were allocated into three treatment groups: Group 1: CpG+anti-OX40, Group 2: CpG+3p-hpRNA, Group 3: untreated control. Treatment efficacy was evaluated based on tumour growth, the occurrence of metastases and overall survival.On day 28 post-implantation, experiments had to be terminated due to tumour progression. Although comparisons of survival times and primary tumour sizes thus became inconsequential, histological examinations provided valuable insights. We observed distinct variations in primary tumour characteristics among the different groups: Groups 1 and 2 displayed demarcations, while Group 3 exhibited diffuse tumours with necrosis. Lung metastases were evident in 70% of untreated mice, whereas none were observed in either of the treated groups.Our findings instil confidence in the potential efficacy of the treatments, thereby laying a solid foundation for future investigations.

[Expression of miR-142 and its relationship with Th17/Treg imbalance in children with autoimmune thyroid disease]. / miR-142åœ¨å„¿ç«¥è‡ªèº«å ç–«æ€§ç”²çŠ¶è ºç–¾ç— ä¸­è¡¨è¾¾åŠä¸ŽTh17/Tregå¤±è¡¡å ³ç³»çš„ç ”ç©¶.

OBJECTIVES: To investigate the expression of microRNA-142 (miR-142) in children with autoimmune thyroid disease (AITD) and its relationship with the imbalance of helper T cell 17 (Th17) and regulatory T cell (Treg). METHODS: A total of 89 children hospitalized for AITD from January 2019 to December 2022 were prospectively selected as the study subjects, including 48 children with Graves' disease (GD group) and 41 children with Hashimoto's thyroiditis (HT group). Additionally, 55 healthy children undergoing physical examinations during the same period were selected as the control group. The differences in serum miR-142, antithyroglobulin antibody (TGAb), antithyroperoxidase antibody (TPOAb), Th17/Treg, and interleukin-17 (IL-17) expression were compared among the groups. RESULTS: The expression of miR-142, TPOAb, TGAb, Th17, Th17/Treg, and IL-17 in the GD group and HT group was higher than that in the control group, while Treg was lower than that in the control group (P<0.05). Pearson correlation analysis revealed that in the GD group, miR-142 was positively correlated with TPOAb, TGAb, Th17, Th17/Treg, and IL-17 (r=0.711, 0.728, 0.785, 0.716, 0.709, respectively; P<0.001) and negatively correlated with Treg (r=-0.725, P<0.001); in the HT group, miR-142 was positively correlated with TPOAb and TGAb (r=0.752, 0.717, respectively; P<0.001). CONCLUSIONS: miR-142 is highly expressed in children with AITD, and its expression may be related to the Th17/Treg imbalance in children with GD.

Graft protective effects and donor-specific antibody suppression by CD4+CD25+Foxp3+ regulatory T cell induced by HMG-CoA reductase inhibitor rosuvastatin in a murine heart transplant model.

BACKGROUND: We previously demonstrated that the hydroxymethylglutaryl-CoA (HMG-CoA) reductase inhibitor (statins) play an important role in the regulation of alloimmune responses. However, little is known regarding the effects of statin on allograft protection or donor-specific antibodies (DSA). In this study, we investigated the graft-protective and immunomodulatory effects of rosuvastatin in a model of fully major histocompatibility complex-mismatched murine cardiac allograft transplantation. METHODS: CBA mice underwent transplantation of C57BL/6 (B6) hearts and received 50 and 500 µg/kg/day of rosuvastatin from the day of transplantation until seven days after the completion of transplantation. To confirm the requirement for regulatory T cells (Tregs), we administered an anti-interleukin-2 receptor alpha antibody (PC-61) to rosuvastatin-treated CBA recipients. Additionally, histological and fluorescent staining, cell proliferation analysis, flow cytometry, and DSA measurements were performed. RESULTS: CBA recipients with no treatment rejected B6 cardiac graft acutely (median survival time [MST], 7 days). CBA mice treated with 500 µg/kg/day of rosuvastatin prolonged allograft survival (MSTs, 77 days). Fluorescent staining studies showed that rosuvastatin-treated recipients had strong aggregation of CD4+Foxp3+ cells in the myocardium and around the coronary arteries of cardiac allografts two weeks after grafting. Flow cytometry studies performed two weeks after transplantation showed an increased number of splenic CD4+CD25+Foxp3+ T cells in rosuvastatin-treated recipients. The addition of rosuvastatin to mixed leukocyte cultures suppressed cell proliferation by increasing the number of CD4+CD25+Foxp3+ Tregs. Additionally, Tregs suppressed DSA production in rosuvastatin-treated recipients. CONCLUSION: Rosuvastatin treatment may be a complementary graft-protective strategy for suppressing DSA production in the acute phase, driven by the promotion of splenic and graft-infiltrating CD4+CD25+Foxp3+ Tregs.

Soluble Fibrinogen-Like Protein 2 Downregulation and Th17/Treg Imbalance in a Taurocholate-Induced Murine Experimental Model of Severe Acute Pancreatitis.

BACKGROUND: Severe acute pancreatitis (SAP) is associated with tremendous systemic inflammation, T-helper 17 (Th17) cells, and regulatory T (Treg) cells play an essential role in the inflammatory responses. Meanwhile, soluble fibrinogen-like protein 2 (Sfgl2) is a critical immunosuppressive effector cytokine of Treg cells and modulates immune responses. However, the impact of SAP induction on Sfgl2 expression and the role of Sfgl2 in immunomodulation under SAP conditions are largely unknown. METHODS: A taurocholate-induced mouse SAP model was established. The ratios of CD4+CD25+Foxp3+ Treg cells or CD4+IL-17+ Th17 cells in blood and pancreatic tissues as well as surface expression of CD80, CD86, and major histocompatibility complex class II (MHC-II) were determined by flow cytometry. Gene mRNA expression was determined by qPCR. Serum amylase and soluble factors were quantitated by commercial kits. Bone marrow-derived dendritic cells (DCs) were generated, and NF-κB/p65 translocation was measured by immunofluorescence staining. RESULTS: SAP induction in mice decreased the Th17/Treg ratio in the pancreatic tissue and increased the Th17/Treg ratio in the peripheral blood. In addition, SAP was associated with a reduced level of Sfgl2 in the pancreatic tissue and blood: higher levels of serum IL-17, IL-2, IFN-α, and TNF-α, and lower levels of serum IL-4 and IL-10. Furthermore, the SAP-induced reduction in Sfgl2 expression was accompanied by dysregulated maturation of bone marrow-derived DCs. CONCLUSIONS: SAP causes reduced Sfgl2 expression and Th17/Treg imbalance, thus providing critical insights for the development of Sfgl2- and Th17/Treg balance-targeted immunotherapies for patients with SAP.

Wutou decoction alleviates arthritis inflammation in CIA mice by regulating Treg cell stability and Treg/Th17 balance via the JAK2/STAT3 pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Wutou Decoction (WTD) is a classic traditional Chinese medicine formula, which has shown clinical efficacy in treating rheumatoid arthritis (RA). The Treg stability and Th17/Treg imbalance is an important immunological mechanism in RA progression. Whether WTD regulates CD4+ T cell subsets has not been thoroughly investigated yet. AIM OF THE STUDY: This study aimed to explore the potential role and mechanisms of WTD in regulating the diminished stability of Treg cells and the imbalance of CD4+ T cell subsets via in vivo and in vitro experiments. MATERIALS AND METHODS: Firstly, the therapeutic effects of WTD on the collagen-induced arthritis (CIA) mouse and its potential regulatory function on CD4+ T cell subsets were evaluated in vivo. Animal specimens were collected after 31 days of treatment with WTD. The anti-arthritic and anti-inflammatory effects of WTD were assessed through arthritis scoring, body weight, spleen index, serum IL-6 levels, and micro-PET/CT imaging. Gene enrichment analysis was performed to evaluate the activation T cell-related signaling pathway. Flow cytometry was used to determine the proportions of CD4+ T cell subsets in vitro and in vitro. Additionally, ELISA was used to assess the secretion of IL-10 and TGF-ß by Treg cells under inflammatory conditions. The suppressive function of Treg cells on cell proliferation under inflammatory conditions was examined using CFSE labeling. Immunofluorescence staining was performed to detect the phosphorylation levels of STAT3 in CD4+ T cells from mouse spleen tissues. Western blotting was used to evaluate the phosphorylation levels of JAK2/STAT3 in Treg cells. RESULTS: WTD significantly alleviated joint inflammation in CIA mice. WTD reduced serum IL-6 levels in CIA mice, improved their body weight and spleen index. WTD treatment inhibited the activation of CD4+ T cell subgroup-related signaling in the joint tissues of CIA mice. In vitro and in vitro experiments showed that WTD increased the proportion of Treg cells and decreased the proportion of Th17 cells in CIA mice spleen. Furthermore, WTD promoted the secretion of IL-10 and TGF-ß by Treg cells and enhanced the inhibitory capacity of Treg cells on cell proliferation under inflammatory conditions. Immunofluorescence detected decreased STAT3 phosphorylation levels in CD4+ T cells from CIA mice spleen, while western blotting revealed a decrease in JAK2/STAT3 phosphorylation levels in Treg cells in vitro. CONCLUSIONS: Inhibiting JAK2/STAT3 phosphorylation is a potential mechanism through which WTD improves Treg cell stability, balances CD4+ T cell subsets, and attenuates RA joint inflammation.

CD4+ T cell heterogeneity in gestational age and preeclampsia using single-cell RNA sequencing.

A balance between pro-inflammatory decidual CD4+ T cells and FOXP3+ regulatory T cells (FOXP3+ Tregs) is important for maintaining fetomaternal tolerance. Using single-cell RNA-sequencing and T cell receptor repertoire analysis, we determined that diversity and clonality of decidual CD4+ T cell subsets depend on gestational age. Th1/Th2 intermediate and Th1 subsets of CD4+ T cells were clonally expanded in both early and late gestation, whereas FOXP3+ Tregs were clonally expanded in late gestation. Th1/Th2 intermediate and FOXP3+ Treg subsets showed altered gene expression in preeclampsia (PE) compared to healthy late gestation. The Th1/Th2 intermediate subset exhibited elevated levels of cytotoxicity-related gene expression in PE. Moreover, increased Treg exhaustion was observed in the PE group, and FOXP3+ Treg subcluster analysis revealed that the effector Treg like subset drove the Treg exhaustion signatures in PE. The Th1/Th2 intermediate and effector Treg like subsets are possible inflammation-driving subsets in PE.

Deubiquitinating enzymes: potential regulators of the tumor microenvironment and implications for immune evasion.

Ubiquitination and deubiquitination are important forms of posttranslational modification that govern protein homeostasis. Deubiquitinating enzymes (DUBs), a protein superfamily consisting of more than 100 members, deconjugate ubiquitin chains from client proteins to regulate cellular homeostasis. However, the dysregulation of DUBs is reportedly associated with several diseases, including cancer. The tumor microenvironment (TME) is a highly complex entity comprising diverse noncancerous cells (e.g., immune cells and stromal cells) and the extracellular matrix (ECM). Since TME heterogeneity is closely related to tumorigenesis and immune evasion, targeting TME components has recently been considered an attractive therapeutic strategy for restoring antitumor immunity. Emerging studies have revealed the involvement of DUBs in immune modulation within the TME, including the regulation of immune checkpoints and immunocyte infiltration and function, which renders DUBs promising for potent cancer immunotherapy. Nevertheless, the roles of DUBs in the crosstalk between tumors and their surrounding components have not been comprehensively reviewed. In this review, we discuss the involvement of DUBs in the dynamic interplay between tumors, immune cells, and stromal cells and illustrate how dysregulated DUBs facilitate immune evasion and promote tumor progression. We also summarize potential small molecules that target DUBs to alleviate immunosuppression and suppress tumorigenesis. Finally, we discuss the prospects and challenges regarding the targeting of DUBs in cancer immunotherapeutics and several urgent problems that warrant further investigation.