IL-2 Receptor Expression Research Articles

T cells Instruct Immune Checkpoint Inhibitor Therapy Resistance in Tumors Responsive to IL-1 and TNFα Inflammation.

Resistance to immune checkpoint inhibitors (ICIs) is common, even in tumors with T cell infiltration. We thus investigated consequences of ICI-induced T cell infiltration in the microenvironment of resistant tumors. T cells and neutrophil numbers increased in ICI-resistant tumors following treatment, in contrast to ICI-responsive tumors. Resistant tumors were distinguished by high expression of IL-1 Receptor 1 (IL1R1), enabling a synergistic response to IL-1 and TNFα to induce G-CSF, CXCL1, and CXCL2 via NF-κB signaling, supporting immunosuppressive neutrophil accumulation in tumor. Perturbation of this inflammatory resistance circuit sensitized tumors to ICIs. Paradoxically, T cells drove this resistance circuit via TNF both in vitro and in vivo. Evidence of this inflammatory resistance circuit and its impact also translated to human cancers. These data support a mechanism of ICI resistance, wherein treatment-induced T cell activity can drive resistance in tumors responsive to IL-1 and TNFα, with important therapeutic implications.

Dupilumab Reduces Urticaria Activity, Itch, and Hives in Patients with Chronic Spontaneous Urticaria Regardless of Baseline Serum Immunoglobulin E Levels.

In chronic spontaneous urticaria (CSU), interleukin (IL)-4 and IL-13 may promote mast cell activation directly via IL-4 receptor expression, or indirectly via upregulated immunoglobulin E (IgE) production. Dupilumab significantly improved CSU signs and symptoms in the phase 3, randomized, placebo-controlled LIBERTY-CSU CUPID Study A. This analysis explores the impact of dupilumab on CSU signs and symptoms and serum IgE levels in patients from LIBERTY-CSU CUPID Study A with serum total IgE above and below 100IU/mL at baseline. Patients with H1-antihistamine-refractory CSU received dupilumab (n = 70) or placebo (n = 68) for 24 weeks. Efficacy endpoints were change from baseline to weeks 12 and 24 in serum total IgE levels, Itch Severity Score over 7 days (ISS7), Urticaria Activity Score over 7 days (UAS7), and Hives Severity Score over 7days (HSS7) in dupilumab- or placebo-treated patients with serum total IgE above and below 100IU/mL at baseline. Dupilumab treatment significantly reduced median (interquartile range) IgE levels at week 12 [dupilumab: -31.9% (-41.9; -22.6); placebo: -6.3% (-21.3; 14.9)] and week 24 [dupilumab: -48.2% (-56.8; - 39.5); placebo: - 6.3% (-34.5; 14.8)]. Similar IgE reductions relative to baseline were observed in dupilumab-treated patients regardless of baseline IgE level. Dupilumab treatment improved ISS7, UAS7, and HSS7 over 12 and 24 weeks, regardless of baseline serum IgE level (interaction p ≥ 0.59 for all treatment by subgroup comparisons), with weak correlations (r < 0.2) observed between IgE level changes and ISS7, UAS7, and HSS7 outcomes. Dupilumab significantly improved CSU signs and symptoms and reduced serum IgE, regardless of baseline IgE levels. In the current analysis, baseline total IgE had no predictive value as a dupilumab treatment response biomarker in CSU. Downregulation of IgE, a key mediator of mast cell activation and histamine release, may at least partially explain the effectiveness of dupilumab in reducing CSU signs and symptoms. ClinicalTrials.gov Identifier: NCT04180488.

Profound T Lymphocyte and DNA Repair Defect Characterizes Schimke Immuno-Osseous Dysplasia

Schimke immuno-osseous dysplasia is a rare multisystemic disorder caused by biallelic loss of function of the SMARCAL1 gene that plays a pivotal role in replication fork stabilization and thus DNA repair. Individuals affected from this disease suffer from disproportionate growth failure, steroid resistant nephrotic syndrome leading to renal failure and primary immunodeficiency mediated by T cell lymphopenia. With infectious complications being the leading cause of death in this disease, researching the nature of the immunodeficiency is crucial, particularly as the state is exacerbated by loss of antibodies due to nephrotic syndrome or immunosuppressive treatment. Building on previous findings that identified the loss of IL-7 receptor expression as a possible cause of the immunodeficiency and increased sensitivity to radiation-induced damage, we have employed spectral cytometry and multiplex RNA-sequencing to assess the phenotype and function of T cells ex-vivo and to study changes induced by in-vitro UV irradiation and reaction of cells to the presence of IL-7. Our findings highlight the mature phenotype of T cells with proinflammatory Th1 skew and signs of exhaustion and lack of response to IL-7. UV light irradiation caused a severe increase in the apoptosis of T cells, however the expression of the genes related to immune response and regulation remained surprisingly similar to healthy cells. Due to the disease’s rarity, more studies will be necessary for complete understanding of this unique immunodeficiency.

Bipotential B-neutrophil progenitors are present in human and mouse bone marrow and emerge in the periphery upon stress hematopoiesis.

This study investigates the dynamics of hematopoiesis in COVID-19, focusing on neutrophil responses. Through RNA sequencing of neutrophils from healthy controls and COVID-19 patients, distinct gene expression alterations are identified, particularly in ICU patients. Notably, neutrophils from COVID-19 patients, especially in the ICU, exhibit enrichment of immunoglobulin and B cell lineage-associated genes, suggesting potential lineage plasticity. Validation in a larger patient cohort and single-cell analysis of bone marrow granulocytes support the presence of granulocyte-monocyte progenitors with B cell lineage-associated genes. The findings propose a link between B-neutrophil lineages during severe infection, implicating a potential role for these cells in altered hematopoiesis favoring myeloid cells over B cells. Elevated GM-CSF and reduced IL-7 receptor expression in stress hematopoiesis suggest cytokine involvement in these dynamics, providing novel insights into disease pathogenesis.

Gut Microbiota Disorder Contributes to the Production of IL-17A That Exerts Chemotaxis via Binding to IL-17RA in Endometriosis.

Endometriosis (EM) is a chronic estrogen-dependent condition characterized by the growth of endometrial-like tissue outside the uterus, posing a significant burden on reproductive-aged women. Previous research has shown a correlation between gut microbiota dysbiosis and interleukin-17A (IL-17A) in EM patients. IL-17A, a promising immunomodulatory molecule, exerts dual roles in human physiology, driving inflammatory diseases. However, the functions and origins of IL-17A in EM remain poorly characterized. Single-cell data analysis was employed to characterize IL-17A activity in EM lesions. Fecal microbiota transplantation was conducted to explore the impact of gut microbiota on EM. Gut microbiota and bile acid metabolism were assessed via 16S rRNA sequencing and targeted metabolomics. Th17 cell proportions were measured using flow cytometry. High expression of IL-17 receptor A (IL-17RA) was observed in myeloid cell subpopulations within EM lesions and may be involved in the migration and recruitment of inflammatory cells in lesions. Elevated IL-17A levels were further validated in peritoneal and follicular fluids of EM patients. Dysregulated bile acid levels, particularly elevated chenodeoxycholic acid (CDCA) and ursodeoxycholic acid (UDCA), were found in the gut and peritoneal fluid of EM mouse models. Additional CDCA administration reduced EM lesions and modulated Th17 cell proportions, while UDCA showed no significant effects. Our findings shed light on the origins and functions of IL-17A in EM, implicating its involvement in lesion migration and recruitment. Dysregulated bile acid metabolism may contribute to EM pathogenesis, with CDCA exhibiting therapeutic potential.

RhIL-7-hyFc, a long-acting interleukin-7, improves efficacy of CAR-T cell therapy in solid tumors

BackgroundChimeric antigen receptor T-cell (CAR-T) therapy has achieved remarkable remission in patients with B-cell malignancies. However, its efficacy in treating solid tumors remains limited. Here, we investigated a combination therapy...

T cell responsiveness to IL-10 defines the immunomodulatory effect of costimulation blockade via anti-CD154 and impacts transplant survival.

Costimulation blockade (CoB)-based immunotherapy is a promising alternative to immunosuppression for transplant recipients; however, the current limited understanding of the factors that impact its efficacy restrains its clinical applicability. In this context, pro- and anti-inflammatory cytokines are being recognized as having an impact on T cell activation beyond effector differentiation. This study aims at elucidating the impact of direct IL-10 signaling in T cells on CoB outcomes. We used a full-mismatch skin transplantation model where recipients had a T cell-restricted expression of a dominant negative IL-10 receptor (10R-DN), alongside anti-CD154 as CoB therapy. Unlike wild-type recipients, 10R-DN mice failed to benefit from CoB. This accelerated graft rejection correlated with increased accumulation of T cells producing TNF-α, IFN-γ, and IL-17. In vitro experiments indicated that while lack of IL-10 signaling did not change the ability of anti-CD154 to modulate alloreactive T cell proliferation, the absence of this pathway heightened T H 1 effector cell differentiation. Furthermore, deficiency of IL-10 signaling in T cells impaired Treg induction, a hallmark of anti-CD154 therapy. Overall, these findings unveil an important and novel role of IL-10 signaling in T cells that defines the success of CoB therapies and identifies a target pathway for obtaining robust immunoregulation.

Steady-state, therapeutic, and helminth-induced IL-4 compromise protective CD8 T cell bystander activation.

Memory CD8 T cells (Tmem) can be activated into innate-like killers by cytokines like IL-12, IL-15, and/or IL-18; but mechanisms regulating this phenomenon (termed bystander activation) are not fully resolved. We found strain-intrinsic deficiencies in bystander activation using specific pathogen-free mice, whereby basal IL-4 signals antagonize IL-18 sensing. We show that therapeutic and helminth-induced IL-4 impairs protective bystander-mediated responses against pathogens. However, this IL-4/IL-18 axis does not completely abolish bystander activation but rather tunes the expression of direct versus indirect mediators of cytotoxicity (granzymes and interferon-γ, respectively). We show that antigen-experience overrides strain-specific deficiencies in bystander activation, leading to uniform IL-18 receptor expression and enhanced capacity for bystander activation/cytotoxicity. Our data highlight that bystander activation is not a binary process but tuned/deregulated by other cytokines that are elevated by contemporaneous infections. Further, our findings underscore the importance of antigen-experienced Tmem to dissect the contributions of bystander Tmem in health and disease.

Phase 2 study of bizaxofusp, an IL-4R targeted toxin payload, in nonresectable recurrent GBM: Comparison of overall survival with contemporaneous eligibility-matched and propensity score balanced external control arm.

2079 Background: Bizaxofusp (MDNA55) is designed to selectively deliver a potent toxin payload to tumor cells by targeting the IL-4 receptor (IL-4R) overexpressed by GBM, but not normal brain. Localized delivery of bizaxofusp minimizes risk of off-target toxicities and systemic exposure while eliciting effective tumor cell killing. Bizaxofusp was evaluated using convection-enhanced delivery in MDNA55-05 (NCT02858895), a single-arm, open-label, multi-center Ph2 study in nonresectable recurrent GBM (rGBM). Methods: The study population comprised of de novo IDH wild-type GBM patients with 1st or 2nd relapse that were non-resectable. Forty-four patients were administered a single treatment of bizaxofusp (range: 18-240 μg). The primary endpoint was OS; secondary endpoints were safety and tumor response. IL-4R expression in tumor biopsies was determined using a validated assay. OS was compared to an eligibility matched external control arm (emECA) of 81 contemporaneous rGBM subjects receiving standard of care. The emECA was further refined by propensity-score balancing (pbECA) to the bizaxofusp arm based on known prognostic factors, resulting in a weighted sample size of 40.8. OS was defined as time from relapse (to unify index date in both arms) or treatment start (for analysis of tumor response) to death/censor. Tumor response was assessed following RANO and mRANO criteria. Results: Bizaxofusp showed an acceptable safety profile at doses up to 240 μg. TRAEs were primarily neurological or aggravation of pre-existing neurological deficits associated with GBM and were manageable with standard measures. Results showed that IL-4R expression did not impact mOS except in patients receiving low doses of bizaxofusp. The bizaxofusp arm had significantly longer median OS (mOS) than contemporaneous emECA (12.4 vs 7.7 months; HR: 0.64, 95% CI 0.46-0.93; p = 0.02). Survival benefit was also evident when compared to the pbECA: (12.4 vs 7.2 months; HR: 0.72, 95% CI 0.46-1.1; p = 0.27). Patients with tumor control (SD or PR/CR, n=21) had significantly longer mOS than patients with tumor progression (n=23) (16.7 vs 8.5 months; HR = 0.5, 95% CI 0.27-0.9; p = 0.01). Among patients with tumor control who had pseudoprogression (PsP) per mRANO, mOS (22.8 months) was significantly longer than patients with tumor progression (HR: 0.49, 95% CI 0.25-0.98; p = 0.049). Conclusions: Bizaxofusp achieved significant OS benefit in patients with nonresectable rGBM compared to contemporaneous emECA. Patients who experienced tumor control, including those with PsP, showed significantly longer survival than patients with tumor progression. Phase 3 ready registrational trial will comprise a high dose bizaxofusp arm and a hybrid control arm with 1/3 randomized subjects and 2/3 pmECA. Clinical trial information: NCT02858895 .

Interleukin (IL)-21 and IL-21 receptor expression in peripheral T and B cells of patients with breast cancer.

IL-21 is a cytokine with versatile antitumor and pro-tumorigenic activities. It is mainly produced by CD4+ T cells and B cells are one of its pivotal targets. In this study, we assessed and compared the expression of IL-21 by CD4+ T cells and the IL-21 receptor (IL-21R) on B cells in the peripheral blood of women with breast cancer and healthy individuals. Blood samples were taken from both patients and controls. Mononuclear cells were seperated using Ficoll-Hypaque density gradient centrifugation. These isolated cells were then stained with either anti-CD19/anti-IL-21R or anti-CD4/anti-IL-21 antibodies and analyzed using flow cytometry. The results showed that there was no significant difference in the percentage of IL-21R+ B cells and IL-21+CD4+ T cells between patients and controls. However, the percentage of CD4+ T cells decreased significantly in patients with breast cancer (P=0.003). This decline was observed from the early stage and before lymph node (LN) involvement. In comparison to the control group, IL-21R+ B cells were relatively lower in patients with stages I+II and those with fewer than 4 involved LNs. The intensity of IL-21 expression in T cells was associated with HER2 expression (P=0.029). Furthermore, we found that the majority of IL-21R+ B cells exhibited a naïve phenotype and most of IL-21+CD4+ T cells did not produce IFN-γ or IL-17. In conclusion, breast cancer from the early stages leads to a significant reduction in the proportion of peripheral CD4+ T cells. However, we did not find a significant change in IL-21 and its receptor expression during disease progression.

Tumour-associated myeloid cells expressing IL-10R2/IL-22R1 as a potential biomarker for diagnosis and recurrence of pancreatic ductal adenocarcinoma

BackgroundPancreatic ductal adenocarcinoma (PDAC) is an aggressive malignancy with a poor survival rate, largely due to the lack of early diagnosis. Although myeloid cells are crucial in the tumour microenvironment, whether their specific subset can be a biomarker of PDAC progression is unclear.MethodsWe analysed IL-22 receptor expression in PDAC and peripheral blood. Additionally, we analysed gene expression profiles of IL-10R2+/IL-22R1+ myeloid cells and the presence of these cells using single-cell RNA sequencing and murine orthotropic PDAC models, respectively, followed by examining the immunosuppressive function of IL-10R2+/IL-22R1+ myeloid cells. Finally, the correlation between IL-10R2 expression and PDAC progression was evaluated.ResultsIL-10R2+/IL-22R1+ myeloid cells were present in PDAC and peripheral blood. Blood IL-10R2+ myeloid cells displayed a gene expression signature associated with tumour-educated circulating monocytes. IL-10R2+/IL-22R1+ myeloid cells from human myeloid cell culture inhibited T cell proliferation. By mouse models for PDAC, we found a positive correlation between pancreatic tumour growth and increased blood IL-10R2+/IL-22R1+ myeloid cells. IL-10R2+/IL-22R1+ myeloid cells from an early phase of the PDAC model suppressed T cell proliferation and cytotoxicity. IL-10R2+ myeloid cells indicated tumour recurrence 130 days sooner than CA19-9 in post-pancreatectomy patients.ConclusionsIL-10R2+/IL-22R1+ myeloid cells in the peripheral blood might be an early marker of PDAC prognosis.

Determination of IL-23 receptor expression and gene polymorphism (rs1884444) in Iranian patients with ankylosing spondylitis

BackgroundThrough investigating genetic variations, it has been demonstrated that single nucleotide polymorphisms (SNPs) in the IL-23 receptor (IL23R) gene have a critical role in the pathophysiology of ankylosing spondylitis (AS). Here, we investigated whether the IL23R variant (rs1884444) is associated with AS in the Iranian population.Methods and materialIn this research, we analyzed rs1884444 in a group of 425 patients with AS and 400 matched controls. For DNA extraction, the phenol/chloroform technique was utilized. Peripheral blood mononuclear cells (PBMCs) were obtained from the whole blood of 39 patients and 43 healthy controls and total RNA was extracted. Genotyping was performed by amplification-refractory mutation system (ARMS)–PCR method. Afterward, the expression level of IL23R was analyzed by the real-time quantitative (Q)-PCR method.ResultsWe observed no significant association between the distribution of alleles and genotypes of rs1884444 and susceptibility to AS. In addition, the expression level of IL23R did not differ between PBMCs from AS patients compared to the control group (P = 0.167). Furthermore, the relative expression level of IL23R was positively correlated with the BASDAI (P < 0.01) and BASFI (P < 0.05) scores of the patients.ConclusionIt appears that IL23R polymorphism (rs1884444) and the level of gene expression might not contribute to the susceptibility to AS in the Iranian population. The correlation of IL23R expression with the level of BASDAI and BASFI scores in patients may be due to the role of the IL-23/IL-23R signaling cascade in inflammation and exert a critical role in the development of AS.

Early dynamic changes to monocytes following major surgery are associated with subsequent infections.

Post-operative infections are a common cause of morbidity following major surgery. Little is understood about how major surgery perturbs immune function leading to heightened risk of subsequent infection. Through analysis of paired blood samples obtained immediately before and 24 h following surgery, we evaluated changes in circulating immune cell phenotype and function across the first 24 h, to identify early immune changes associated with subsequent infection. We conducted a prospective observational study of adult patients undergoing major elective gastrointestinal, gynecological, or maxillofacial surgery requiring planned admission to the post-anesthetic care unit. Patients were followed up to hospital discharge or death. Outcome data collected included mortality, length of stay, unplanned intensive care unit admission, and post-operative infections (using the standardized endpoints in perioperative medicine-core outcome measures for perioperative and anesthetic care criteria). Peripheral blood mononuclear cells were isolated prior to and 24 h following surgery from which cellular immune traits including activation and functional status were assessed by multi-parameter flow cytometry and serum immune analytes compared by enzyme-linked immunosorbent assay (ELISA). Forty-eight patients were recruited, 26 (54%) of whom developed a post-operative infection. We observed reduced baseline pre- and post-operative monocyte CXCR4 and CD80 expression (chemokine receptors and co-stimulation markers, respectively) in patients who subsequently developed an infection as well as a profound and selective post-operative increase in CD4+ lymphocyte IL-7 receptor expression in the infection group only. Higher post-operative monocyte count was significantly associated with the development of post-operative infection (false discovery rate < 1%; adjusted p-value = 0.001) with an area under the receiver operating characteristic curve of 0.84 (p < 0.0001). Lower monocyte chemotaxis markers, higher post-operative circulating monocyte counts, and reduced co-stimulatory signals are associated with subsequent post-operative infections. Identifying the underlying mechanisms and therapeutics to reverse defects in immune cell function requires further exploration.

P470 Crohn’s disease patients with Ustekinumab-induced remission are characterized by high baseline p35 cytokines subunits in LPMC and high IL-23 receptor expression in the lamina propria Th1 cells

Abstract Background Crohn’s disease (CD) is a chronic relapsing disorder of the gastrointestinal tract characterized by excessive inflammation sustained by the induction and expansion of Th1 and Th17 cells. Ustekinumab (UST), a fully humanized monoclonal antibody, is currently used for the therapy of CD patients. It acts by neutralizing both IL12 and IL23, essential for the maturation and expansion of Th1 and Th17 cells, by binding the common subunit p40. To be effective UST needs, in addition to the expression of target cytokines, the presence of cells responsive to IL12 and IL23, thus cells with IL12 and IL23 receptors. Methods This is a prospective, observational, single-center open-label study. All CD patients with clinical indications to receive UST for active luminal disease were enrolled. Patients had baseline endoscopy to confirm disease activity by SES-CD and to collect biopsies to isolate LPMC. Real-time pcr was performed to evaluate IL12 (p40 and p35) and IL23 (p40 and p19) cytokines subunits and IL12 (IL12Rb1 and IL12Rb2) and IL23 (IL12Rb1 and IL23R) receptors subunits expression in remitters (R) and not remitters (NR). Moreover, LPMCs were analyzed by flow cytometry, and IL12Rb2 and IL23R expression in Th1, Th17, and type 1 Innate lymphoid cells (iLC) cells were evaluated. Results were analyzed for clinical remission status (HBI&amp;lt;5) at week 16. Differences between groups were analyzed by Mann-Whitney test and statistical significance was considered for p&amp;lt;0.05. Results From April 2019 to November 2022, 13 CD patients were enrolled, whose clinical and demographic characteristics are enclosed in Table 1. At baseline there was no difference in IL12Rb2, ILRb1, and IL23R expression between R and NR (Fig.1). Regarding cytokines subunits, there was no difference in p19 but there was a significantly higher expression of p35 subunits in R than in NR (mean 1,52 vs 0,34; p=0,001) (Fig.1). Flow cytometry showed no difference in Th and ILCs subgroups frequency between R and NR. IL12Rb2 and IL23R were also equally expressed by Th17, Th1/Th17 and ILC1 cells in the two groups. In contrast, Th1 cells, IL23R expression was higher in R group as compared to NR (mean 20,6% vs 6,8; p=0.009) (Fig.2). Conclusion In CD patients treated with UST, higher expression p35 in LPMC and higher expression of IL23R in the lamina propria Th1 cells was associated with remission at week 16.

Assessment of IL-9/IL-9R Expression and Therapeutic Effect of IL-9 Blockade in a Mouse Breast Cancer Model

Background and Hypothesis:IL-9 is a cytokine produced by TH9, mast cells, and innate lymphoid cells. IL-9 acts on the IL-9 receptor (IL-9R) and is primarily involved in parasitic immunity and allergic inflammation. IL-9 has anti-tumor effects in solid tumors such as melanoma. IL-9 also has pro-tumor effects in various cancer types, including lung cancer. Previous studies have shown that increased serum IL-9 in breast cancer patients correlates with tumor metastasis. Therefore, we hypothesize that IL9 signaling contributes to breast tumor metastasis and blocking IL-9 may have a therapeutic benefit in reducing metastasis.&#x0D; Methods:Mice were orthotopically implanted with 4T1 breast tumor cells. Organs were harvested after 28 days. Total RNA was extracted and expression of IL-9/IL-9R and leukocyte markers were assessed. To test therapeutic effect of IL-9 blockade, tumor-bearing mice were treated with anti-IL-9 monoclonal antibody (αIL-9). Tumor sizes were monitored every 4 days. On day 28, tumor tissue was harvested and weighed. Lung was also harvested and stained with H&amp;E for metastasis analysis.&#x0D; Results:Primary tumor growth was not altered by αIL-9. The effect of αIL-9 treatment on lung metastasis is pending pathological analysis. We further examined IL-9R to define how tumor burden alters expression of IL-9 and IL-9R across tissues. Lymphoid organs, such as the thymus, spleen, and inguinal lymph node (inguinal LN), exhibit high levels of IL-9R. Additionally, the small intestine is notably enriched with IL-9R. The thymus exhibited the highest IL-9 expression.&#x0D; Conclusion and Future Directions:Although blockade of IL-9 did not impact tumor growth in this model, this will be examined in other models to confirm the findings. Further research is needed to investigate the potential therapeutic benefits of αIL-9 with other established therapies. We observed that there is altered IL-9 and IL-9R expression in tumor bearing mice and the physiological significance of that finding remains to be determined.

INCREASED CD4+ TO CD8+ T-CELL RATIO AS A DRIVER OF IMPAIRED ACHILLES TENDON HEALING IN HUMAN PATIENTS

Early identification of patients at risk for impaired tendon healing and corresponding novel therapeutic approaches are urgent medical needs. This study aimed to clarify the role of CD3+ T-cells during acute Achilles tendon (AT) healing. Blood and hematoma aspirate were taken from 26 patients during AT reconstruction, and additional blood samples were obtained during clinical follow-up at 6, 26 and 52 weeks after surgery. T-cell subsets were analyzed by flow cytometry using CD3, CD4, CD8, CD11a, CD57 and CD28 antibodies. Clinical follow-up included functional tests, MRI assessments, and subjective questionnaires. In vitro, the functional behavior of patient-derived tenocytes was investigated in co-cultures with autologous unpolarized CD4+ or CD8+ T-cells, or IFNy-polarized CD8+ or IL17-polarized CD4+ Tcells (n=5-6). This included alterations in gene expression (qPCR), MMP secretion (ELISA), migration rate (scratch wound healing assay) or contractility (collagen gels). Analysis revealed that elevated CD4+ T-cell levels and reduced CD8+ T-cell levels (increased CD4/CD8 ratio) in hematoma aspirate and pre-operative blood were associated with inferior clinical outcomes regarding pain and function at 26 and 52 weeks. Increased levels of CD8+ -memory T-cell subpopulations in blood 6 weeks after surgery were associated with less tendon elongation. In vitro, tenocytes showed increased MMP1/2/3 levels and collagen III/I ratio in co-culture with unpolarized and/or IL17-polarized CD4+ T-cells compared to unpolarized CD8+ T-cells. This coincided with increased IL17 receptor expression in tenocytes co-cultured with CD4+ T-cells. Exposure of tenocytes to IL17-polarized CD4+ T-cells decreased their migration rate and increased their matrix contractility, especially compared to IFNy-polarized CD8+ T-cells. The CD4+ /CD8+ T-cell ratio could serve as prognostic marker for early identification of patients with impaired AT healing potential. Local reduction of CD4+ T-cell levels or their IL17 secretion represent a potential therapeutic approach to improve AT healing and to prevent weakening of the tendon ECM.

IL-6 signaling accelerates iron overload by upregulating DMT1 in endothelial cells to promote aortic dissection.

Aortic dissection (AD), caused by tearing of the intima and avulsion of the aortic media, is a severe threat to patient life and organ function. Iron is closely related to dissection formation and organ injury, but the mechanism of iron ion transport disorder in endothelial cells (ECs) remains unclear. We identified the characteristic EC of dissection with iron overload by single-cell RNA sequencing data. After intersecting iron homeostasis and differentially expressed genes, it was found that hypoxia-inducible factor-1α (HIF-1α) and divalent metal transporter 1 (DMT1) are key genes for iron ion disorder. Subsequently, IL-6R was identified as an essential reason for the JAK-STAT activation, a classical iron regulation pathway, through further intersection and validation. In in vivo and in vitro, both high IL-6 receptor expression and elevated IL-6 levels promote JAK1-STAT3 phosphorylation, leading to increased HIF-1α protein levels. Elevated HIF-1α binds explicitly to the 5'-UTR sequence of the DMT1 gene and transcriptionally promotes DMT1 expression, thereby increasing Fe2+ accumulation and endoplasmic reticulum stress (ERS). Blocking IL-6R and free iron with deferoxamine and tocilizumab significantly prolonged survival and reduced aortic and organ damage in dissection mice. A comparison of perioperative data between AD patients and others revealed that high free iron, IL-6, and ERS levels are characteristics of AD patients and are correlated with prognosis. In conclusion, activated IL-6/JAK1/STAT3 signaling axis up-regulates DMT1 expression by increasing HIF-1α, thereby increasing intracellular Fe2+ accumulation and tissue injury, which suggests a potential therapeutic target for AD.

IL-21 promoting angiogenesis contributes to the development of psoriasis.

Elevated IL-21 expression which can effectively induce Th17 cell differentiation has been implicated in the pathogenesis of psoriasis, but its role in angiogenesis remains poorly understood. PASI and PSI score assessment was applied to evaluate the severity of psoriatic lesions. The expression of IL-21, IL-21 receptor (IL-21R), CD31, VEGFA, MMP-9, and ICAM-1 in skin was determined by immunohistochemistry or quantitative real-time polymerase chain reaction. The serum level of IL-21 was measured by enzyme-linked immunosorbent assay (ELISA). Then, their correlation was analyzed statistically. Human umbilical vein endothelial cells (HUVECs) cocultured with conditional medium from normal human epidermal keratinocytes (NHEKs) were treated with IL-21 and/or M5 cocktail (mixture of IL-1α, IL-17A, IL-22, TNF-α, and oncostatin M). The migration and tube formation of HUVECs were detected, and the levels of VEGFA, MMP-9, and ICAM-1 in NHEKs were measured by Western blotting or ELISA. Increased IL-21 and IL-21R expression was observed in psoriatic sera or skin specimens, with IL-21R mainly locating in keratinocytes and IL-21 in immune cells. Pearson analysis showed significantly positive correlation between IL-21/IL-21R and erythema scores/microvessel density in psoriatic lesions. Moreover, the expression of proangiogenic genes, VEGFA, ICAM-1, and MMP-9 was upregulated in skins of psoriasis. Additionally, in M5 microenvironment, migration and tube formation could be magnified in HUVECs using IL-21 pre-treated NHEK medium. Mechanically, the co-stimulation of IL-21 and M5 to NEHKs increased the expression of ICAM-1. IL-21 could regulate keratinocytes to secrete ICAM-1, thereby promoting angiogenesis in psoriasis.

Preclinical evidence for preventive and curative effects of resveratrol on xenograft cholangiocarcinogenesis

Cholangiocarcinoma (CCA), the malignant tumor of bile duct epithelial cells, is a relatively rare yet highly lethal cancer. In this work, we tested the ability of Resveratrol (RV) to prevent and cure CCA xenograft in nude mice and investigated molecular mechanisms underpinning such anticancer effect. Human CCA cells were xenografted in mice that were or not treated prior to or after to transplantation with RV. Tumor growth was monitored and analyzed for the markers of cell proliferation, apoptosis, and autophagy. TCGA was interrogated for the molecules possibly targeted by RV. RV could inhibit the growth of human CCA xenograft when administered after implantation and could reduce the growth or even impair the implantation of the tumors when administered prior the transplantation. RV inhibited CCA cell proliferation, induced apoptosis with autophagy, and strongly reduced the presence of CAFs and production of IL-6. Interrogation of CCA dataset in TCGA database revealed that the expression of IL-6 Receptor (IL-6R) inversely correlated with that of MAP-LC3 and BECLIN-1, and that low expression of IL-6R and of MIK67, two pathways downregulated by RV, associated with better survival of CCA patients. Our data demonstrate that RV elicits a strong preventive and curative anticancer effect in CCA by limiting the formation of CAFs and their release of IL-6, and this results in up-regulation of autophagy and apoptosis in the cancer cells. These findings support the clinical use of RV as a primary line of prevention in patients exposed at risk and as an adjuvant therapeutics in CCA patients.

P2Y1 and P2Y2 receptors differ in their role in the regulation of signaling pathways during unloading-induced rat soleus muscle atrophy

The current study aimed to investigate the hypothesis that purinergic receptors P2Y1 and P2Y2 play a regulatory role in gene expression in unloaded muscle. ATP is released from cells through pannexin channels, and it interacts with P2Y1 and P2Y2 receptors, leading to the activation of markers of protein catabolism and a reduction in protein synthesis. To test this hypothesis thirty-two rats were randomly divided into four groups (8 per group): a non-treated control group (C), a group subjected to three days of hindlimb unloading with a placebo (HS), a group subjected to three days of hindlimb unloading treated with a P2Y1 receptor inhibitor, MRS2179 (HSM), and a group subjected to three days of hindlimb unloading treated with a P2Y2 receptor inhibitor, AR-C 118925XX (HSA). This study revealed several key findings following three days of soleus muscle unloading: 1Inhibition of P2Y1 or P2Y2 receptors prevented the accumulation of ATP, the increase in IP3 receptor content, and the decrease in the phosphorylation of GSK-3beta. This inhibition also mitigated the reduction in the rate of protein synthesis. However, it had no significant effect on the markers of mTORC1-dependent signaling. 2Blocking P2Y1 receptors prevented the unloading-induced upregulation of phosphorylated p38MAPK and partially reduced the increase in MuRF1mRNA expression. 3Blocking P2Y2 receptors prevented muscle atrophy during unloading, partially maintained the levels of phosphorylated ERK1/2, reduced the increase in mRNA expression of MAFbx, ubiquitin, and IL-6 receptor, prevented the decrease in phosphorylated AMPK, and attenuated the increase in phosphorylated p70S6K.Taken together, these results suggest that the prevention of muscle atrophy during unloading, as achieved by the P2Y2 receptor inhibitor, is likely mediated through a reduction in catabolic processes and maintenance of energy homeostasis. In contrast, the P2Y1 receptor appears to play a relatively minor role in muscle atrophy during unloading.