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PURPOSEPostconsolidation immunotherapy including dinutuximab, granulocyte-macrophage colony-stimulating factor, and interleukin-2 improved outcomes for patients with high-risk neuroblastoma enrolled on the randomized portion of Children's Oncology Group study ANBL0032. After random assignment ended, all patients were assigned to immunotherapy. Survival and toxicities were assessed.PATIENTS AND METHODSPatients with a pre-autologous stem cell transplant (ASCT) response (excluding bone marrow) of partial response or better were eligible. Demographics, stage, tumor biology, pre-ASCT response, and adverse events were summarized using descriptive statistics. Event-free survival (EFS) and overall survival (OS) from time of enrollment (up to day +200 from last ASCT) were evaluated.RESULTSFrom 2009 to 2015, 1,183 patients were treated. Five-year EFS and OS for the entire cohort were 61.1 ± 1.9% and 71.9 ± 1.7%, respectively. For patients ≥ 18 months old at diagnosis with International Neuroblastoma Staging System stage 4 disease (n = 662) 5-year EFS and OS were 57.0 ± 2.4% and 70.9 ± 2.2%, respectively. EFS was superior for patients with complete response/very good partial response pre-ASCT compared with those with PR (5-year EFS: 64.2 ± 2.2% v 55.4 ± 3.2%, P = .0133); however, OS was not significantly different. Allergic reactions, capillary leak, fever, and hypotension were more frequent during interleukin-2–containing cycles than granulocyte-macrophage colony-stimulating factor–containing cycles ( P < .0001). EFS was superior in patients with higher peak dinutuximab levels during cycle 1 ( P = .034) and those with a high affinity FCGR3A genotype ( P = .0418). Human antichimeric antibody status did not correlate with survival.CONCLUSIONAnalysis of a cohort assigned to immunotherapy after cessation of random assignment on ANBL0032 confirmed previously described survival and toxicity outcomes. EFS was highest among patients with end-induction complete response/very good partial response. Among patients with available data, higher dinutuximab levels and FCGR3A genotype were associated with superior EFS. These may be predictive biomarkers for dinutuximab therapy.

Background & Aims:CD4+CD25+Foxp3+regulatory T cells (Tregs) are essential to maintain immunological tolerance and have been shown to promote liver allograft tolerance in both rodents and humans. Low-dose IL-2 (LDIL-2) can expand human endogenous circulating Tregsinvivo, but its role in suppressing antigen-specific responses and promoting Treg trafficking to the sites of inflammation is unknown. Likewise, whether LDIL-2 facilitates the induction of allograft tolerance has not been investigated in humans. Methods:We conducted a clinical trial in stable liver transplant recipients 2–6 years post-transplant to determine the capacity of LDIL-2 to suppress allospecific immune responses and allow for the complete discontinuation of maintenance immunosuppression (ClinicalTrials.govNCT02949492). One month after LDIL-2 was initiated, those exhibiting at least a 2-fold increase in circulating Tregs gradually discontinued immunosuppression over a 4-month period while continuing LDIL-2 for a total treatment duration of 6 months. Results:All participants achieved a marked and sustained increase in circulating Tregs. However, this was not associated with the preferential expansion of donor-reactive Tregs and did not promote the accumulation of intrahepatic Tregs. Furthermore, LDIL-2 induced a marked IFNγ-orchestrated transcriptional response in the liver even before immunosuppression weaning was initiated. The trial was terminated after the first 6 participants failed to reach the primary endpoint owing to rejection requiring reinstitution of immunosuppression. Conclusions:The expansion of circulating Tregs in response to LDIL-2 is not sufficient to control alloimmunity and to promote liver allograft tolerance, due, at least in part, to off-target effects that increase liver immunogenicity. Our trial provides unique insight into the mechanisms of action of immunomodulatory therapies such as LDIL-2 and their limitations in promoting alloantigen-specific effects and immunological tolerance. Clinical Trials Registration:The study is registered at ClinicalTrials.gov (NCT02949492). Impact and implications:The administration of low-dose IL-2 is an effective way of increasing the number of circulating regulatory T cells (Tregs), an immunosuppressive lymphocyte subset that is key for the establishment of immunological tolerance, but its use to promote allograft tolerance in the setting of clinical liver transplantation had not been explored before. In liver transplant recipients on tacrolimus monotherapy, low-dose IL-2 effectively expanded circulating Tregs but did not increase the number of Tregs with donor specificity, nor did it promote their trafficking to the transplanted liver. Low-dose IL-2 did not facilitate the discontinuation of tacrolimus and elicited, as an off-target effect, an IFNγ-orchestrated inflammatory response in the liver that resembled T cell-mediated rejection. These results, supporting an unexpected role for IL-2 in regulating the immunogenicity of the liver, highlight the need to carefully evaluate systemic immunoregulatory strategies with investigations that are not restricted to the blood compartment and involve target tissues such as the liver.

This pilot study aims to assess the effect of Cognitive Behavioral Therapy for insomnia (CBTi) in individuals with cannabis use disorder and insomnia. It also aims to investigate the effect of CBTi on levels of serum inflammatory markers in relation to insomnia symptoms.Individuals with cannabis use disorder and insomnia symptoms were recruited over 18 months. Data collected included demographics, self-reported sleep parameters, and cannabis use. Blood samples were drawn to measure IL-2, IL-6, CRP, and cortisol. Participants completed the Insomnia Severity Index questionnaire (ISI) and the Patient Health Questionnaire-4 (PHQ-4), and they were provided with an actigraphy (wrist) device for 1 week before CBTi and a subsequent week after completing the 4 CBTi sessions.Nineteen participants were enrolled in the study. The mean ISI score decreased from moderately severe insomnia at baseline to no clinically significant insomnia after CBTi with a sustained decrease at 3- and 6-months follow-up. Actigraphy showed a significant decrease in sleep onset latency (SOL) after CBTi. Three months after CBTi, 80% of participants reported a decrease in their cannabis use. There was also a significant and sustained decrease in mean PHQ-4 scores after CBTi. Although only trending towards significance, the levels of three out of four biomarkers (IL-2, IL-6, CRP) were decreased 6 months after CBTi.CBTi is effective as a short- and long-term treatment of insomnia and comorbid anxiety/depression in individuals who regularly use cannabis. A potential added benefit is a reduction in cannabis consumption and inflammatory serum biomarkers.

To determine if Exendin-4 could be a therapeutic agent for burn-induced hyperglycemia.Male Balb/c mice received a bolus of Exendin-4 intraperitoneally immediately after 15% total body surface area scald injury. Tail glucose levels were recorded and T-cell functions were analyzed at 4 h and 24 h postburn (pb). Pancreatic pathology was observed consecutively. The secretions of cytokines were detected in serum, spleen, and lung. Apoptosis of splenic CD3+ T-cells was examined by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling and flow cytometry.Although Exendin-4 could attenuate burn-induced hyperglycemia in mice at 4 h pb, it accelerated their survival dose dependently with progressive depletion of splenocyte number. T-cell function underwent two-phasic changes following Exendin-4 treatment. Compared to placebo mice, T-cell from Exendin-4-treated mice was manifested with increased proliferation, while decreased IL-2 secretion and lower ratio of IL-4/IFN-γ at 4 h pb. However, at 24 h pb, it showed decreased proliferation, while increased IL-2 secretion and higher ratio of IL-4/IFN-γ. Exendin-4 could elicit higher circulating IL-6 and IL-10 levels at 4 h pb, which were pronounced in the lung at 24 h pb. In the meanwhile, severe inflammation could be found in the pancreas. At 24 h pb, the numbers of terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling or caspase-3 positive cells and the apoptosis of CD3+ T-cells were significantly increased in the spleens of Exendin-4 mice relative to placebo mice.These data support a pathogenic role of Exendin-4 signaling during thermal injury, warning against its clinical application in acute insults.

This study investigated the impact of colitis induced by dextran sulphate sodium (DSS)-induced colitis (DIC) on histopathological and immunological outcomes in the periodontal tissues of Wistar rats.Inflammatory bowel diseases (IBD) and periodontitis have been reported to present a bidirectional relationship. However, the inflammatory pathway that connects both diseases needs further investigation.Twenty-five male Wistar rats were allocated in four groups: unilateral ligature-induced periodontitis for 14 days: LIP (n = 7); dextran sulphate sodium-induced colitis only: DIC (n = 6); DIC + LIP (n = 6) and controls (n = 6). Digital images were obtained from the histological sections. In order to assess the attachment loss (AL), the linear distance between the cementoenamel junction (CEJ) and the alveolar bone crest was measured on the mesial root using histological photomicrography's ImageJ software. Immunological analyses of gingival tissues and plasma were performed by Bio-Plex Th1/Th2 Assay.The DIC group showed inflammatory cells extending to the periodontal connective tissues, which contained significantly elevated expressions of IL-1α, IL-1β, IL-2, IL-6, IL-12, IL-13, GM-CSF, IFN-γ and TNF-α compared to controls. There was no significant difference in bone loss between controls and DIC. There were no significant histopathological differences between DIC + LIP and LIP. However, DIC + LIP presented a significantly lower IL-2 and IL-5 than the LIP group. There was no bone loss difference between LIP+DIC and LIP groups. DIC + LIP group presented significantly higher levels of GM-CSF in plasma.DSS-induced colitis was associated with an overexpression of Th1/Th2- related cytokines in the gingival tissue.

Many studies have explored new methods to cure acute lung injury (ALI); however, none of those methods could significantly change the high mortality rate of ALI. Shenfu is a Chinese traditional medicine that might be effective against ALI.Our study explores the therapeutic potential of Shenfu in ALI.Male C57BL/6 mice were assigned to control, lipopolysaccharide (LPS) (500 µg/100 μL per mouse), and LPS + Shenfu (30 mL/kg) groups. Shenfu (10 µL/mL) was added to LPS (10 µg/mL) treated MLE-12 cells for 48 hCompared with the ALI group, Shenfu reduced wet/dry weight ratio (19.8%, 36.2%), and reduced the IL-2 (40.9%, 61.6%), IFN-γ (43.5%, 53.3%) TNF-α (54.1%, 42.1%), IL-6 (54.8%,70%), and IL-1β (39.9%, 65.1%), reduced serum uric acid (18.8%, 48.7%) and creatinine (17.4%, 41.1%). Moreover, Shenfu enhanced cell viability (17.2%, 59.9%) and inhibited cell apoptosis (63.0%) and p38/ERK phosphorylation inTaken together, the results revealed the therapeutic effect and the underlying mechanism of Shenfu injection in an ALI in mouse model, indicating its clinical potential to treat patients with ALI.

Despite recent changes in the treatment landscape, there remains an unmet need for effective, tolerable, chemotherapy-free treatments for patients with advanced/metastatic urothelial carcinoma (mUC), especially cisplatin-ineligible patients.To evaluate the immunostimulatory interleukin-2 cytokine prodrug bempegaldesleukin (BEMPEG) plus nivolumab in patients with advanced/mUC from the phase 2 multicenter PIVOT-02 study.This open-label, multicohort phase 1/2 study enrolled patients with previously untreated locally advanced/surgically unresectable or mUC (N = 41).Patients received BEMPEG 0.006 mg/kg plus nivolumab 360 mg intravenously every 3 wk.The primary objectives were safety and the objective response rate (ORR) in patients with measurable disease at baseline and at least one postbaseline tumor response assessment (response-evaluable). Secondary objectives were overall survival (OS) and progression-free survival (PFS). Exploratory biomarker analyses via univariate logistic regression were performed to test the association between potential biomarkers (CD8The ORR was 35% (13/37 evaluable patients) and the complete response rate was 19% (7/37 patients); the median duration of response was not reached. Median PFS was 4.1 mo (95% confidence interval [CI] 2.1-8.7) and median OS was 23.7 mo (95% CI 15.8-not reached). Overall, 40/41 patients (98%) experienced at least one treatment-related adverse event (TRAE); grade 3/4 TRAEs occurred in 11 patients (27%), most commonly pyrexia (4.9%; 2 patients). Exploratory biomarker analyses showed no association between biomarkers and response. Limitations include the small sample size and single-arm design.BEMPEG plus nivolumab was well tolerated and showed antitumor activity as first-line treatment in patients with locally advanced/mUC.We investigated an immune-stimulating prodrug called bempegaldesleukin plus the antibody nivolumab as the first therapy for patients with advanced or metastatic cancer of the urinary tract. This combination had manageable treatment-related side effects and was effective in a subset of patients. This trial is registered at ClinicalTrials.gov as NCT02983045.

Interleukin 2 (IL-2) is a pleiotropic cytokine that elicits both immunogenic and tolerogenic immune response essential for homeostasis. Initial clinical application of IL-2 based therapy was hampered by its function on broad spectrum of cells expressing corresponding receptors, which exerts uncontrolled side effects in clinical trials. While recent progress on structural modification or adjusted regimen of IL-2, revolutionized the use of IL-2 in immunomodulation process that requires stringent selection of effector cells, such as promotion of T

The coronavirus disease 2019 (COVID-19) pandemic has affected the entire world, and has a variety of clinical presentations. The aim of this study is to determine the relationships of fecal cytokines and markers with the symptoms and prognosis of children with COVID-19 infection, and to identify noninvasive markers during follow-up. In a cohort of 40 COVID-19-positive children and 40 healthy controls, fecal cytokines and markers were examined in stool samples. A binary logistic model was used to assess the potential of cytokines as risk factors for hospitalization. Odds ratios (ORs) with 95% confidence intervals (CIs) were reported. A

Inflammation is at the core of many chronic conditions and exacerbates infectious conditions, including the severity of coronavirus disease 2019 (COVID-19) infections.This study aimed to examine the effects of a novel food supplement, palmitoylethanolamide (PEA), specifically Levagen+, as compared with a placebo on proinflammatory biomarkers in adults recently diagnosed with COVID-19 who were unvaccinated and nonhospitalized.This study was a double-blind randomized placebo-controlled trial conducted October 2020-March 2021 (clinicaltrials.gov: NCT04912921). Participants aged 19-53 y were unvaccinated and recently infected with COVID-19 as indicated by a positive test result per RT-PCR or antigen test, and they reported to the test site following diagnosis as allowed by the CDC's return-to-work policy. Participants were stratified by age, sex, and BMI and randomly assigned by coin toss to receive 600 mg Levagen+ twice daily (LEV) or placebo tablets twice daily (CON) for 4 wk. At baseline and week 4, participants completed health histories, 24-h dietary recalls, anthropometrics, and nonfasting blood sampling. The primary outcomes were the 4-wk change between groups for IL-6, C-reactive protein, ferritin, intercellular adhesion molecule 1, soluble P-selectin (sP-selectin), and neutrophil/lymphocyte ratio. Multiple linear regression models were utilized to assess treatment effects on outcomes, adjusting for covariates.A total of 60 participants completed the study (LEV: n = 30; CON: n = 30). After 4 wk of supplementation, sP-selectin (β = -11.5; 95% CI: -19.8, -3.15; P = 0.0078), IL-1β (β = -22.9; 95% CI: -42.4, -3.40; P = 0.0222), and IL-2 (β = -1.73; 95% CI: -3.45, -0.065; P = 0.0492) concentrations were significantly reduced in the LEV group compared with the CON group.Inflammatory mechanisms are crucial to optimal resolution of infectious conditions, yet unchecked secretion of inflammatory mediators can promote the dysregulated immune response implicated in COVID-19 complications. Overall, PEA supplementation produced anti-inflammatory effects in individuals recently diagnosed with COVID-19 who were nonhospitalized.

Recent studies have demonstrated that regulatory T cells (Tregs) develop in the thymus via two pathways involving distinct Treg progenitors (TregP): CD25+FOXP3− (CD25+ TregP) and CD25−FOXP3lo (FOXP3lo TregP) Treg progenitors. To examine this process in more detail, we carried out single-cell RNA sequencing (scRNA-Seq) and TCR-Seq on sorted murine CD4+CD8+ double-positive (DP) thymocytes, CD4+ single-positive (CD4SP) thymocytes, CD25+FOXP3−CD73− TregP, CD25−FOXP3loCD73− TregP, newly generated mature CD25+FOXP3+CD73− Tregs, and FOXP3+CD73+ recirculating/long-term resident Tregs (RT-Tregs). Sorted populations were individually hashtagged and then combined into one scRNA-Seq/TCR-Seq library before sequencing and subsequent analysis. We found that both CD25+ TregP and FOXP3lo TregP arise via an initial agonist-activated state that gives rise to a second transitional stage before differentiating into mature Tregs. Using both scRNA-Seq and bulk RNA-Seq on sorted thymocyte subsets, we demonstrate that CD25+ TregP are significantly enriched for Il2 production, suggesting that they are the major source of IL-2 needed to convert TregP into mature Tregs. Using TCR-Seq, we found that several TCRs were clearly biased in favor of the conventional or Treg lineages, but that a large fraction of TCRs were found in both these lineages. Finally, we found that RT-Tregs in the thymus are not monomorphic but are composed of multiple distinct subsets and that these RT-Tregs express the most diverse TCR repertoire of all CD4SP thymocytes. Thus, our studies define multiple stages of Treg differentiation within the murine thymus and serve as a resource for future studies on CD4+ thymocyte development and Treg differentiation.

Murine myeloid cells are developed from hemopoietic stem/progenitor cells. Different types of progenitor cells have variable differentiation potentials. Among the ten main types of cells differentiated from lymphoid progenitor cells, regulatory T cells (Tregs), an important cell subpopulation regulating immune and inflammatory responses, arise from the hematopoietic stem cells in the bone marrow. Tregs then differentiate into T lymphocytes and migrate to the thymus and finally generate Treg subsets, which are subsequently activated and regulated by inflammatory cytokines in the peripheral blood. Tregs also have different phenotypes and immunomodulatory functions. The cytokine interleukin-2/interleukin-2 receptor (IL-2/IL-2R) pathway is an important regulatory signaling pathway of Tregs. Besides, different types of CD4

Since most orthopedic patients’ wounds are open in clinical practice, postoperative wound infection and other conditions are prone to occur, which pose varying degrees of threat to patients’ prognosis and life and health. In this paper, a retrospective study is conducted on orthopedic trauma patients in our hospital from January 2020 to January 2022, including 98 patients with postoperative infection and 98 patients without infection, to detect and compare the levels of inflammatory factors and the level of T cell lymphatic group index difference. The ROC curve is drawn to analyze the diagnostic efficacy of postoperative infection indicators for patients with orthopedic trauma. The differences in baseline data between the two groups are compared. Multivariate Logistic regression analysis is performed on infection status. The experimental results show that the IL-6, IL-2, and CRP of the infection group of patients are significantly higher than uninfected group, and the CD3, CD4, and CD8 are significantly lower than uninfected group, which means that patients with infection after orthopedic trauma are in a disordered state of immune cytokines and function. Therefore, postoperative infection can be effectively assessed by early combined detection of the above indicators. In addition, the analysis of other clinical data showed that the operation time, the number of underlying diseases, and the surgical method are also risk factors for postoperative infection.

Selenium-enriched egg white peptides (Se-EWP) were prepared by pre-heat treatment and enzymatic hydrolysis in this study. In addition, their selenopeptide sequence identification and immunomodulatory effect were investigated. Results showed that the yield of Se-EWP obtained from alkaline-neutral protease treatment reached 76.90%, and peptides with a molecular weight of 200-1000 Da accounted for 98.33%. Four characteristic selenopeptides, including SeCys-Trp-Leu-Glu, Trp-Ser-SeCys, SeMet-Ala-Pro, and SeMet-Leu, were identified by HPLC-ESI-MS/MS, which were rich in hydrophobic and branched-chain amino acids. Se-EWP (750 mg/kg/d) could effectively retard the decrease of immune organ index in immunosuppressed mice induced by cyclophosphamide. Moreover, supplementation of Se-EWP could promote a higher content of Se in liver, the number of white blood cells, and the levels of serum cytokines (IL-6, IL-2, and TNF-α) as compared with EWP groups, indicating that Se-EWP could effectively alleviate immunosuppression induced by cyclophosphamide. These findings suggested that Se-EWP exhibited great potential as functional foods for immunomodulatory effect.

To determine the metabolic activity, and cytokine expression over time from immortalized human corneal epithelial cells (HCECs) exposed to hyperosmotic stress.HCECs were cultured and expanded in DMEM/F-12 with 10% FBS. The cells were exposed to either normal media (295 mmol/kg) or hyperosmolar media (500 mmol/kg) for 0.25, 3, 6, and 12 hours. After each exposure duration, metabolic activity was quantified using alamarBlue, and a panel of pro-inflammatory cytokines (IL-1β, IL-2, IL-4, IL-6, IL-8, IL-10, IL-12p70, IL-13, TNF-α, IFN-γ, and IL-17A) was quantified using multiplexed electrochemiluminescence (Meso Scale Diagnostics, Rockville, MD).Metabolic activity of the HCEC exposed to hyperosmolar conditions was significantly reduced at the 3-, 6-, and 12-hour mark compared to the control (allHyperosmolar stress reduced cell metabolic activity and increased expression of IL-1β, IL-4, IL6, IL8, IL-12p70, IL-13, and TNF-α over a 6-hour period in an immortalized HCEC line.

Billions of individuals worldwide suffer from periodontal disease, an inflammatory disease that results in hard-tissue and soft-tissue destruction. A viable therapeutic option to treat periodontal disease may be via cannabinoids that exert immunomodulatory effects, and the endocannabinoid system (ECS) is readily present in periodontal tissues that exhibit cannabinoid type 1 and 2 receptors (CB1R and CB2R). Phytocannabinoids (pCBs), which are a part of a heterogeneous group of molecules acting on cannabinoid receptors (CBR) derived from the cannabis plants, have been attributed to a wide variety of effects including anti-inflammatory activity and some pro-inflammatory effects depending on the cell type. Thus, this study aims to examine the effects of pCBs on primary human gingival fibroblasts (HGFs) in IL-1β stimulated (simulated periodontal disease) HGFs.Human gingival fibroblasts (HGFs) obtained from ATCC were cultured per the manufacturer's recommendation. The functional activity of cannabinoid receptors was measured using ACTOne (cAMP)-based CB1R and CB2R assay. The effects of three pCBs (0.1-10 μg/ml or 10Cannabidivarin (CBVN or CBDV) (ECThe effective inhibition of IL-1β-stimulated production of PGE2 and cytokines by the pCB in HGFs suggests that targeting the endocannabinoid system may lead to the development of therapeutic strategies for periodontal therapy. However, each pCB has its unique anti-inflammatory profile, in which certain pro-inflammatory activities are also exhibited. The pCBs alone or in combination may benefit and aid in improving public oral health.

Asthma is a common chronic lung disease without absolute treatment, and hypersensitivity reactions and type 2 immune responses are responsible for asthma pathophysiology. ADAM10 as a metalloproteinase transmembrane protein is critical for development of Th2 responses, and levamisole as an anthelmintic drug has immunomodulatory effects, which not only regulates ADAM10 activity but also can suppress the bone marrow and neutrophil production. Therefore, in the present study, nanoparticles were used as a levamisole delivery system to reduce bone marrow suppression, and the immunomodulatory and ADAM10 inhibitory effects of levamisole were studied in allergic asthma. Asthmatic mice were treated with PLGA-levamisole nanoparticles. Then, AHR, BALF, and blood cell counts, levels of the IgG1 subclass, total and OVA-specific IgE, IL2, IL-4, IL-5, IL-10, IL-13, IL-17, IL-25, IL-33, INF-γ, and TNF-α, gene expression of FoxP3, T-bet, RORγt, PU.1, GATA3, FcεRII, CysLT1R, eotaxin, and ADAM10, and lung histopathology were evaluated. PLGA-LMHCl with considered characteristics could control airway hyper-responsiveness, eosinophils in the BALF, levels of immunoglobulins, Th2-, Th9-, and Th17-derived cytokines and pivotal genes, eosinophilic inflammation, hyperplasia of the goblet cell, and hyperproduction of mucus and could increase Th1- and Treg-derived cytokines and also pivotal genes. It could also modulate the ADAM10 activity and had no effect on the number of neutrophils in the bloodstream. The novel safe nanodrug had no side effect on the bone marrow to produce neutrophils and could control the allegro-immuno-inflammatory response of asthma.

To determine the effects and immunological mechanisms of low-dose interleukin-2 (IL-2) in a murine model of chronic cardiac allograft rejection (BALB/c to C57BL/6) after costimulatory blockade consisting of MR1 (250 μg/ip day 0) and CTLA4-Ig (200 μg/ip day 2), we administered low-dose IL-2 (2000 IU/day) starting on posttransplant day 14 for 3 weeks. T regulatory (Treg) cell infiltration of the grafts was determined by immunohistochemistry; circulating exosomes by western blot and aldehyde bead flow cytometry; antibodies to donor MHC by immunofluorescent staining of donor cells; and antibodies to cardiac self-antigens (myosin, vimentin) by ELISA. We demonstrated that costimulation blockade after allogeneic heart transplantation induced circulating exosomes containing cardiac self-antigens and antibodies to both donor MHC and self-antigens, leading to chronic rejection by day 45. Treatment with low-dose IL-2 prolonged allograft survival (100 days), prevented chronic rejection, and induced splenic and graft-infiltrating CD4+ CD25+ Foxp3 Treg cells by day 45 and circulating exosomes (Foxp3+) with PD-L1 and CD73. MicroRNA 142, associated with the TGFβ pathway, was significantly downregulated in exosomes from IL-2-treated mice. In conclusion, low-dose IL-2 delays rejection in a murine model of chronic cardiac allograft rejection and also induces graft-infiltrating Tregs and circulating exosomes with immunoregulatory molecules.

Inflammation plays an important role in the pathogenesis of stroke. The differential expression of inflammatory and angiogenic factors in thrombi and plasma remain undefined. In this observational cohort study, we evaluated angiogenic factors and inflammatory cytokines, in cerebral thrombi, local cerebral plasma (CP), and peripheral plasma (PP) in patients with acute ischemic stroke. Protein analysis of thrombi, CP and PP were used to measure angiogenic and inflammatory proteins using electrochemiluminescence. Our data indicate that VEGF-A, VEGF-C, bFGF, IL-4, IL-13, IL-1β, IL-2, IL-8, IL-16, IL-6 and IL-12p70 were higher in the thrombi of acute ischemic stroke patients than in the CP and PP of stroke patients. Moreover, the protein levels of GM-CSF were lower in the PP than in the CP and the clot. Moreover, VEGF-D, Flt-1, PIGF, TIE-2, IL-5, TNF-β, IL-15, IL-12/IL-23p40, IFN-γ and IL-17A were higher in PP and CP than in thrombi. Our results show that cytokines mediating the inflammatory response and proteins involved in angiogenesis are differentially expressed in thrombi within the cerebral and peripheral circulations. These data highlight the importance of identifying new biomarkers in different compartments of the circulatory system and in thrombi that may be used for the diagnosis and treatment of stroke patients.

PurposeBiochemical biomarkers to determine the injury severity and the potential for functional recovery of traumatic spinal cord injury (TSCI) are highly warranted; however, it remains to be clarified whether cerebrospinal fluid (CSF) or peripheral blood (PB) is the ideal sample media. This study aims to measure and compare biomarker concentrations in CSF and PB and to explore associations between biomarker concentrations and injury severity, i.e., American Spinal Injury Association (ASIA) Impairment Scale (AIS) grade, and biomarker concentrations and clinical outcome, i.e., AIS grade improvement and Spinal Cord Independent Measure version III (SCIM-III) score.MethodsFrom 2018 to 2020, we conducted a single-center prospective pilot study of TSCI patients (n=15) and healthy controls (n=15). Sample collection and clinical outcome assessment were performed at median 13 h [IQR: 19], 9 days [IQR: 2], and 148 days [IQR: 49] after TSCI. Concentrations of neuron-specific enolase (NSE); glial fibrillary acid protein (GFAP); neurofilament light chain (NfL); interferon-γ (IFN-γ); interleukin (IL)-1ß, IL-2, IL-4, IL-6, IL-8, IL-10, IL-12p70, and IL-13; and tumor necrosis factor α (TNF-α) were measured and associated to clinical outcomes.ResultsThe biomarker concentrations were higher in CSF than PB. CSF concentrations of GFAP, NSE, IFN-y, TNF-a, IL-2, IL-12p70, IL-4, IL-10, and IL-13 and PB concentrations of GFAP and IFN-y were significantly associated with AIS grade, but not with AIS grade improvement or SCIM-III score.ConclusionsOur results support GFAP as a potential diagnostic biomarker that may be measured in CSF as well as PB. PURPOSE: Biochemical biomarkers to determine the injury severity and the potential for functional recovery of traumatic spinal cord injury (TSCI) are highly warranted; however, it remains to be clarified whether cerebrospinal fluid (CSF) or peripheral blood (PB) is the ideal sample media. This study aims to measure and compare biomarker concentrations in CSF and PB and to explore associations between biomarker concentrations and injury severity, i.e., American Spinal Injury Association (ASIA) Impairment Scale (AIS) grade, and biomarker concentrations and clinical outcome, i.e., AIS grade improvement and Spinal Cord Independent Measure version III (SCIM-III) score.METHODS: From 2018 to 2020, we conducted a single-center prospective pilot study of TSCI patients (n=15) and healthy controls (n=15). Sample collection and clinical outcome assessment were performed at median 13 h [IQR: 19], 9 days [IQR: 2], and 148 days [IQR: 49] after TSCI. Concentrations of neuron-specific enolase (NSE); glial fibrillary acid protein (GFAP); neurofilament light chain (NfL); interferon-γ (IFN-γ); interleukin (IL)-1ß, IL-2, IL-4, IL-6, IL-8, IL-10, IL-12p70, and IL-13; and tumor necrosis factor α (TNF-α) were measured and associated to clinical outcomes.RESULTS: The biomarker concentrations were higher in CSF than PB. CSF concentrations of GFAP, NSE, IFN-y, TNF-a, IL-2, IL-12p70, IL-4, IL-10, and IL-13 and PB concentrations of GFAP and IFN-y were significantly associated with AIS grade, but not with AIS grade improvement or SCIM-III score.CONCLUSIONS: Our results support GFAP as a potential diagnostic biomarker that may be measured in CSF as well as PB.

T cell antigen receptor stimulation induces tyrosine phosphorylation of downstream signaling molecules and the phosphatidylinositol, Ras, MAPK, and PI3 kinase pathways, leading to T cell activation. Previously, we reported that the G-protein-coupled human muscarinic receptor could bypass tyrosine kinases to activate the phosphatidylinositol pathway and induce interleukin-2 production in Jurkat leukemic T cells. Here, we demonstrate that stimulating G-protein-coupled muscarinic receptors (M1 and synthetic hM3Dq) can activate primary mouse T cells if PLCβ1 is coexpressed. Resting peripheral hM3Dq+PLCβ1 (hM3Dq/β1) T cells did not respond to clozapine, an hM3Dq agonist, unless they were preactivated by TCR and CD28 stimulation which increased hM3Dq and PLCβ1 expression. This permitted large calcium and phosphorylated ERK responses to clozapine. Clozapine treatment induced high IFN-γ, CD69, and CD25 expression, but surprisingly did not induce substantial IL-2 in hM3Dq/β1 T cells. Importantly, costimulation of both muscarinic receptors plus the TCR even led to reduced IL-2 expression, suggesting a selective inhibitory effect of muscarinic receptor costimulation. Stimulation of muscarinic receptors induced strong nuclear translocation of NFAT and NFκB and activated AP-1. However, stimulation of hM3Dq led to reduced IL-2 mRNA stability which correlated with an effect on the IL-2 3'UTR activity. Interestingly, stimulation of hM3Dq resulted in reduced pAKT and its downstream pathway. This may explain the inhibitory impact on IL-2 production in hM3Dq/β1T cells. Moreover, an inhibitor of PI3K reduced IL-2 production in TCR-stimulated hM3Dq/β1 CD4 T cells, suggesting that activating the pAKT pathway is critical for IL-2 production in T cells.

Background: Acute lung injury (ALI)/acute respiratory distress syndrome (ARDS) remains a serious clinical problem but has no approved pharmacotherapy. Mesenchymal stem cells (MSCs) represent an attractive therapeutic tool for tissue damage and inflammation owing to their unique immunomodulatory properties. The present study aims to explore the therapeutic effect and underlying mechanisms of human umbilical cord MSCs (UC-MSCs) in ALI mice. Objective: In this study, we identify a novel mechanism for human umbilical cord-derived MSCs (UC-MSCs)-mediated immunomodulation through PGE2-dependent reprogramming of host macrophages to promote their PD-L1 expression. Our study suggests that UC-MSCs or primed- UC-MSCs offer new therapeutic approaches for lung inflammatory diseases. Methods: Lipopolysaccharide (LPS)-induced ALI mice were injected with 5×105 UC-MSCs via the tail vein after 4 hours of LPS exposure. After 24 hours of UC-MSC administration, the total protein concentration and cell number in the bronchoalveolar lavage fluid (BALF) and cytokine levels in the lung tissue were measured. Lung pathological changes and macrophage infiltration after UCMSC treatment were analyzed. Moreover, in vitro co-culture experiments were performed to analyze cytokine levels of RAW264.7 cells and Jurkat T cells. Results: UC-MSC treatment significantly improved LPS-induced ALI, as indicated by decreased total protein exudation concentration and cell number in BALF and reduced pathological damage in ALI mice. UC-MSCs could inhibit pro-inflammatory cytokine levels (IL-1β, TNF-α, MCP-1, IL-2, and IFN-γ), while enhancing anti-inflammatory cytokine IL-10 expression, as well as reducing macrophage infiltration into the injured lung tissue. Importantly, UC-MSC administration increased programmed cell death protein ligand 1 (PD-L1) expression in the lung macrophages. Mechanistically, UC-MSCs upregulated cyclooxygenase-2 (COX2) expression and prostaglandin E2 (PGE2) secretion in response to LPS stimulation. UC-MSCs reduced the inflammatory cytokine levels in murine macrophage Raw264.7 through the COX2/PGE2 axis. Furthermore, UC-MSC- derived PGE2 enhanced PD-L1 expression in RAW264.7 cells, which in turn promoted programmed cell death protein 1 (PD-1) expression and reduced IL-2 and IFN-γ production in Jurkat T cells. Conclusion: Our results suggest that UC-MSCs attenuate ALI via PGE2-dependent reprogramming of macrophages to promote their PD-L1 expression.

Cancer immunotherapy approaches have progressed significantly during the last decade due to the significant improvement of our understanding of immunologic evasion of malignant cells. Depending on the type, stage, and grade of cancer, distinct immunotherapy approaches are being designed and recommended; each is different in efficacy and adverse effects. Malignant cells can adopt multiple strategies to alter the normal functioning of the immune system in recognizing and eliminating them. These strategies include secreting different immunosuppressive factors, polarizing tumor microenvironment cells to immunosuppressive ones, and interfering with the normal function of the antigen processing machinery (APM). In this context, careful evaluation of immune surveillance has led to a better understanding of the roles of cytokines, including IL-2, IL-12, IL-15, interferon-α (IFN-α), tumor necrosis factor-α (TNF-α), and transforming growth factor-β (TGF-β) in cancer formation and their potential application in cancer immunotherapy. Additionally, monoclonal antibodies (mAbs), adoptive cell therapy approaches, immune checkpoint blockade, and cancer vaccines also play significant roles in cancer immunotherapy. Moreover, the development of clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9 (CRISPR/CAS9) as an outstanding genome editing tool resolved many obstacles in cancer immunotherapy. In this regard, this review aimed to investigate the impacts of different immunotherapy approaches and their potential roles in the current and future roads of cancer treatment. Whatever the underlying solution for treating highly malignant cancers is, it seems that solving the question is nowhere near an achievement unless the precise cooperation of basic science knowledge with our translational experience.

Interleukin-2 inducible T cell kinase (ITK) plays a predominant role in the T-cell receptor (TCR) signaling cascade to ensure valid T-cell activation and function. Nevertheless, whether it regulates T-cell response of early vertebrates remains unknown. Herein, we investigated the involvement of ITK in the lymphocyte-mediated adaptive immune response, and its regulation to T-cell activation in the Nile tilapia Oreochromis niloticus. Both sequence and structure of O. niloticus ITK (OnITK) were remarkably conserved with its homologues from other vertebrates, implying its potential conserved function. OnITK mRNA was extensively expressed in lymphoid-related tissues, and with the relative highest level in peripheral blood. Once Nile tilapia was infected by Edwardsiella piscicida, OnITK in splenic lymphocytes was significantly up-regulated on 7-day post infection at both transcription and translation levels, suggesting that OnITK might involve in the primary adaptive immune response of teleost. Furthermore, upon splenic lymphocytes were stimulated by T-cell specific mitogen PHA, OnITK mRNA and protein levels were dramatically elevated. More importantly, treatment of splenic lymphocytes with specific inhibitor significantly crippled OnITK expression, which in turn impaired the inducible expression of T-cell activation markers IFN-γ, IL-2 and CD122, indicating the critical roles of ITK in regulating T-cell activation of Nile tilapia. Taken together, our results suggest that ITK takes part in the lymphocyte-mediated adaptive immunity of tilapia, and is indispensable for T-cell activation of teleost. Our findings thus provide novel evidences for understanding the mechanism regulating T-cell immunity of early vertebrates, as well as the evolution of adaptive immune system.

Despite new therapeutic options, treatment of steroid-refractory chronic graft-versus-host disease (SR-cGVHD) remains challenging as organ involvement and clinical manifestations are highly variable. In previous trials of low-dose interleukin-2 (LD IL-2), we established the safety and efficacy of LD IL-2 for the treatment of SR-cGVHD. In the present report, we combined five phase 1 or 2 clinical trials conducted at our center to investigate organ-specific response rate, coinvolvement of organs, predictors of organ-specific response, and its possible association with immune response. For the 105 adult patients included in this report, the overall response rate after 8 or 12 weeks of LD IL-2 was 48.6% and 53.3%, including late responses in patients who continued treatment for extended periods. Skin was the most frequent organ involved (84%). The organ-specific response rate was highest in liver (66.7%) followed by the gastrointestinal tract (62.5%), skin (36.4%), joint/muscle/fascia (34.2%), and lung (19.2%). In multivariable analysis, shorter time from diagnosis of cGVHD to IL-2 initiation, shorter time from transplant to IL-2 initiation, and fewer prior therapies were associated with overall response as well as skin response. For immunologic correlates, the ratio of regulatory T cells:conventional T cells (ie, CD4Treg:CD4Tcon) ratio at 1 week was significantly higher in patients with overall and skin response; skin response was significantly associated with lower number of total CD3 T cells, CD4Tcon cells, and CD8 T cells and a higher number of B cells. For lung responders, terminal effector memory cell counts were lower within all T-cell populations compared with nonresponders. Organ-specific mechanisms of injury should be investigated, and organ-specific targeted therapies need to be developed.

Type 1 diabetes (T1D), an autoimmune disorder in which the insulin-producing β-cells in the islets of Langerhans in the pancreas are destroyed, afflicts over 1.6 million Americans. Although pancreatic islet transplantation has shown promise in treating T1D, continuous use of required immunosuppression regimens limits clinical islet transplantation as it poses significant adverse effects on graft recipients and does not achieve consistent long-term graft survival with 50%-70% of recipients maintaining insulin independence at 5 years. T cells play a key role in graft rejection, and rebalancing pathogenic T effector and protective T regulatory cells can regulate autoimmune disorders and transplant rejection. The synergy of the interleukin-2 (IL-2) and Fas immunomodulatory pathways presents an avenue for eliminating the need for systemic immune suppression by exploiting IL-2's role in expanding regulatory T cells and leveraging Fas ligand (FasL) activity on antigen-induced cell death of effector T cells. Herein, we developed a hydrogel platform for co-delivering an analog of IL-2, IL-2D, and FasL-presenting microgels to achieve localized immunotolerance to pancreatic islets by targeting the upregulation of regulatory T cells and effector T cells simultaneously. Although this hydrogel provided for sustained, local delivery of active immunomodulatory proteins, indefinite allograft survival was not achieved. Immune profiling analysis revealed upregulation of target regulatory T cells but also increases in Granzyme B-expressing CD8sup+/supT cells at the graft site. We attribute the failed establishment of allograft survival to these Granzyme B-expressing T cells. This study underscores the delicate balance of immunomodulatory components important for allograft survival - whose outcome can be dependent on timing, duration, modality of delivery, and disease model.

Purpose:Half of the patients with high-risk neuroblastoma who receive GD2-targeted mAb do not achieve long-term remissions. Recently, the antibody hu14.18 has been linked to IL2 (hu14.18-IL2) to enhance its efficacy and shown promising preclinical and clinical activity. We developed two new immunocytokines (IC) by linking two other γc cytokines, IL15 and IL21, to hu14.18. The purpose of this study was to compare hu14.18-IL15 and -IL21 with hu14.18-IL2 in their ability to induce antibody-dependent cell-mediated cytotoxicity (ADCC) against neuroblastoma.Experimental Design:We assessed ADCC of hu14.18-IL15 and -IL2 (human cytokines, cross-reactive to mouse) against GD2low and GD2high neuroblastoma cell lines in vitro. T-cell–deficient mice with orthotopic patient-derived xenografts (PDX) and immunocompetent mice with transplantable orthotopic neuroblastoma were used to test all three ICs, including hu14.18-IL21 (murine IL21, not cross-reactive to human). Mechanistic studies were performed using single-cell RNA-sequencing (scRNA-seq).Results:hu14.18-IL15 and hu14.18-IL2 mediated equivalent in vitro ADCC by human NK cells. When combined with chemotherapy, all three ICs similarly controlled the growth of PDXs in nude mice with murine NK effector cells. However, hu14.18-IL15 and -IL21 outperformed hu14.18-IL2 in immunocompetent mice with syngeneic neuroblastoma, inducing complete tumor regressions and extending survival. scRNA-seq data revealed an increase in CD8+ T cells and M1 tumor-associated macrophages and decreased regulatory T cells and myeloid-derived suppressor cells in the tumor microenvironment.Conclusions:Hu14.18-IL15 and Hu14.18-IL21 exhibit robust preclinical activity, warranting further consideration for clinical testing in patients with GD2-expressing neuroblastoma.

Enteric glial cells (EGCs) are involved in intestinal inflammation. In this study, we will investigate how Bifidobacterium bifidum (B.b.) and Bacteroides fragilis (B.f.) influence EGC regulation. After pretreatment with lipopolysaccharide (LPS) and interferon-γ (IFN-γ), the expressions of major histocompatibility complex class II (MHC-II), CD80, CD86, glial cell line-derived neurotrophic factor (GDNF), toll-like receptor 2 (TLR-2), and tumor necrosis factor-α (TNF-α) in EGCs were detected using polymerase chain reaction and western blot after co-culture with the supernatants of B.b. or B.f. (multiplicity of infection, 40:1 or 80:1). Finally, EGCs were co-cultured with naive CD

Cytokines are powerful mediators of immune responses and some, such as interleukin-2 (IL-2), have achieved dramatic responses as cancer immunotherapies. Unfortunately, systemic administration often results in deleterious side effects, prompting exploration of strategies to localize cytokine activity to the tumor microenvironment (TME). To this end, we constructed an IL-2/IL2Ra fusion protein (IL-2FP) with an MMP2/9-specific cleavage site, designed to exploit the dysregulated protease activity in the TME to selectively activate IL-2 in the tumor. To determine if TME protease activity is sufficient to cleave the FP and if FP activity is due to specific cleavage, we created Colon 38 tumor cell lines expressing similar levels of IL-2FPs with either a functional cleavage site [H11(cs-1FP)] or a scrambled, noncleavable sequence [H2(scramFP)]. H11(cs-1FP) tumors demonstrated reduced tumor growth, characterized by regressions not observed in H2(scramFP) tumors. Analysis through qRT-PCR, flow cytometry, and immunohistochemistry indicate robust CD8 responses in the H11(cs-1FP) tumors. Interferon gamma (IFNg) knockout mice revealed that the immune effects of the cleavable FP are mediated through both IFNg-dependent and IFNg-independent mechanisms. Collectively, these data suggest that matrix metalloproteinases (MMPs) in the TME can cleave the IL-2FP specifically, thus enhancing an antitumor response, and provide a rationale for further developing this approach.

Macrolide antibiotics have immunomodulatory properties which may be beneficial in viral infections. However, the precise effects of macrolides on T cell responses to COVID, differences between different macrolides, and synergistic effects with other antibiotics have not been explored.We investigated the effect of antibiotics (amoxicillin, azithromycin, clarithromycin, and combined amoxicillin with clarithromycin) on lymphocyte intracellular cytokine levels and monocyte phagocytosis in healthy volunteer PBMCs stimulated ex vivo with SARS-CoV-2 S1+2 spike protein. A retrospective cohort study was performed on intensive care COVID-19 patients.Co-incubation of clarithromycin with spike protein-stimulated healthy volunteer PBMCs ex vivo resulted in an increase in CD8Clarithromycin has immunomodulatory properties over and above azithromycin. Amoxicillin in addition to clarithromycin is associated with synergistic ex vivo immunomodulatory properties. The potential benefit of clarithromycin in critically ill patients with COVID-19 and other viral pneumonitis merits further exploration.

Regulatory B cells (Bregs) are a subset of B cells that secrete interleukin 10 (IL-10) and play a vital role in suppressing the immune response. The aim of this study was to evaluate the proportion of Bregs in patients with thymoma.The proportions of subgroups of Bregs in 23 patients with thymoma and 15 healthy controls were detected by flow cytometry. The serum IL-2, IL-4, IL-6, IL-10, IL-17A, IFN-γ, and TNF-α levels of the subjects were measured using a cytometric bead array (CBA).The proportions of circulating IL-10The expression of IL-10 produced by Bregs is increased in patients with thymoma, particularly those with advanced-stage disease, which may suggest that Bregs are involved in the pathogenesis and progression of thymoma.

Systemic sclerosis (SSc) is a debilitating autoimmune disease characterized by severe lung outcomes resulting in reduced life expectancy. Fra-2-transgenic mice offer the opportunity to decipher the relationships between the immune system and lung fibrosis. This study was undertaken to investigate whether the Fra-2-transgenic mouse lung phenotype may result from an imbalance between the effector and regulatory arms in the CD4+ T cell compartment.We first used multicolor flow cytometry to extensively characterize homeostasis and the phenotype of peripheral CD4+ T cells from Fra-2-transgenic mice and control mice. We then tested different treatments for their effectiveness in restoring CD4+ Treg cell homeostasis, including adoptive transfer of Treg cells and treatment with low-dose interleukin-2 (IL-2).Fra-2-transgenic mice demonstrated a marked decrease in the proportion and absolute number of peripheral Treg cells that preceded accumulation of activated, T helper cell type 2-polarized, CD4+ T cells. This defect in Treg cell homeostasis was derived from a combination of mechanisms including impaired generation of these cells in both the thymus and the periphery. The impaired ability of peripheral conventional CD4+ T cells to produce IL-2 may greatly contribute to Treg cell deficiency in Fra-2-transgenic mice. Notably, adoptive transfer of Treg cells, low-dose IL-2 therapy, or combination therapy changed the phenotype of Fra-2-transgenic mice, resulting in a significant reduction in pulmonary parenchymal fibrosis and vascular remodeling in the lungs.Immunotherapies for restoring Treg cell homeostasis could be relevant in SSc. An intervention based on low-dose IL-2 injections, as is already proposed in other autoimmune diseases, could be the most suitable treatment modality for restoring Treg cell homeostasis for future research.

To improve the adjuvant activity of polysaccharides from Eucommia ulmoides leaves (PsEUL) in inducing an effective immune response against ovalbumin (OVA), PsEUL were conjugated to OVA using the N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride (EDC) method. The synthesized PsEUL-OVA was encapsulated using phytantriol and F127 to produce PsEUL-OVA cubosomes (Cubs), a novel delivery system. The physicochemical properties and immune modulation effects of this novel delivery system were explored.In vitro, PsEUL-OVA/Cubs carrying large amounts of OVA were rapidly phagocytized by macrophages and upregulated macrophage proliferation, thereby stimulating cytokine production (interleukin (IL)-6 and IL-4). In vivo, PsEUL-OVA/Cubs increased the titer of OVA-specific antibodies (immunoglobulin (Ig)G, IgG2b, IgG2a and IgG1) and cytokine levels (IL-2, IL-6, IL-4 and interferon-γ). In addition, the cubosomes promoted the differentiation of CD8Collectively, the findings demonstrate that PsEUL-antigen/Cubs can be a useful delivery vehicle with immune response-promoting effects. Therefore, this study lays the foundation for the development of novel adjuvant-antigen delivery systems with potential applications in vaccine design. © 2022 Society of Chemical Industry.

Chimeric-antigen receptor (CAR) T-cell therapy has shown remarkable efficacy against hematologic tumors. Yet, CAR T-cell therapy has had little success against solid tumors due to obstacles presented by the tumor microenvironment (TME) of these cancers. Here, we show that CAR T cells armored with the engineered IL-2 superkine Super2 and IL-33 were able to promote tumor control as a single-agent therapy. IFNγ and perforin were dispensable for the effects of Super2- and IL-33-armored CAR T cells. Super2 and IL-33 synergized to shift leukocyte proportions in the TME and to recruit and activate a broad repertoire of endogenous innate and adaptive immune cells including tumor-specific T cells. However, depletion of CD8+ T cells or NK cells did not disrupt tumor control, suggesting that broad immune activation compensated for loss of individual cell subsets. Thus, we have shown that Super2 and IL-33 CAR T cells can promote antitumor immunity in multiple solid tumor models and can potentially overcome antigen loss, highlighting the potential of this universal CAR T-cell platform for the treatment of solid tumors.

The delivery of specific cytokine payloads to a neoplastic environment employing antibodies able to selectively accumulate at the tumor site represents an attractive strategy to stimulate an immune response to cancer. Whilst conventional antibody–cytokine fusions based on a single payload have shown potent anticancer activity, the concomitant delivery of two cytokine payloads may further improve the therapeutic outcome as the immune system typically adopts multiple signals to reinforce an antitumor strategy. We here describe a potency-matched dual-cytokine antibody fusion protein containing a tumor-targeting antibody fragment specific to human fibroblast activation protein (FAP), simultaneously linked to both interleukin-2 (IL2) and a tumor necrosis factor (TNF) mutant. The resulting fusion protein, termed IL2-7NP2-TNFmut, formed stable non-covalent trimers driven by the interaction of the tumor necrosis factor subunits. Both cytokine payloads retained their biological activity within the fusion protein, as shown by in vitro cellular assays. The tumor-targeting properties and the anticancer activity of IL2-7NP2-TNFmut were investigated in vivo in immunocompromised mice bearing SKRC52 cells transduced with human FAP. The fusion protein preferentially localized to the cancer site and induced partial tumor retardation.

Hepatic CD4 tissue-resident memory T cells (TRM) are required for robust protection against Salmonella infection; however, the generation of this T cell population is poorly understood. To interrogate the contribution of inflammation, we developed a simple Salmonella -specific T cell transfer system that allowed direct visualization of hepatic TRM formation. Salmonella -specific (SM1) T cell receptor (TCR) transgenic CD4 T cells were activated in vitro and adoptively transferred into C57BL/6 mice while hepatic inflammation was induced by acetaminophen overdose or L. monocytogenes infection. In both model systems, hepatic CD4 TRM formation was accentuated by local tissue responses. Liver inflammation also enhanced the suboptimal protection provided by a subunit Salmonella vaccine which typically induces circulating memory CD4 T cells. To further elucidate the mechanism of CD4 TRM formation in response to liver inflammation, various cytokines were examined by RNAseq, bone marrow chimeras, and in vivo neutralization. Surprisingly, IL-2 and IL-1 were found to enhance CD4 TRM formation. Thus, local inflammatory mediators enhance CD4 TRM populations and can boost the protective immunity provided by a suboptimal vaccine. This knowledge will be foundational for the development of a more effective vaccine against invasive nontyphoidal salmonellosis (iNTS).

Mulisan decoction (MLS) is a classic formula of traditional Chinese medicine for treating hyperhidrosis. The mechanism remains unclear.To investigate the antiperspirant effect and underlying mechanisms of MLS.Fifty rats were divided into control, model, and three doses of MLS intervention groups (Rats treated with MLS presented a significant decrease in sweat point numbers (13.5%), increase in body weight (13.2%), and promotion in the balance of Th1/Th2 cytokine ratioMLS demonstrated a good antiperspirant effect and metabolism improvement. These findings inspire more clinical study validation on immune improvement and antiperspirant effect.

The influence of peripheral inflammation on nonmotor symptoms (NMSs) in Parkinson's disease (PD) remains unclear.The aim of this study was to explore whether serum inflammatory marker profiles are associated with the progression of NMSs in early PD.We included 45 patients with early PD and 20 healthy control subjects. Six inflammatory markers, including interleukin (IL)-1β, IL-2, IL-6, IL-10, tumor necrosis factor-α, and high-sensitivity C-reactive protein, were measured. NMSs were assessed using the Non-Motor Symptoms Scale, Montreal Cognitive Assessment, and Composite Autonomic Symptom Score-31 at baseline and after 3 years.Principal component (PC) analysis showed that only PC3 scores, mainly loaded by IL-2 and IL-6, were significantly elevated in the PD group compared with the control group. Higher PC3 scores in the PD group were associated with faster progression of Non-Motor Symptoms Scale total and mood/apathy domain scores. There were no significant associations of PC scores with Montreal Cognitive Assessment and Composite Autonomic Symptom Score-31 score changes.Peripheral inflammation may be related to the evolution of NMSs, particularly mood symptoms, in the early stages of PD. © 2022 International Parkinson and Movement Disorder Society.

Glutaminase (GLS) isoenzymes GLS1 and GLS2 catalyze the first step of glutaminolysis. GLS1 is requisite for Th17 cell differentiation, and its inhibition suppresses autoimmune disease in animals, but the function of GLS2 is not known. The aim of this study was to investigate the role of GLS2 in CD4+ T cell function and systemic lupus erythematosus (SLE) pathogenesis.We measured reactive oxygen species (ROS) levels, lipid peroxidation, and mitochondrial mass and polarization by flow cytometry, interleukin-2 (IL-2) production by a dual luciferase assay, and CpG DNA methylation of Il2 by a real-time polymerase chain reaction system. The impact of the overexpression of wild-type GLS1, wild-type GLS2, or mutated GLS2 at the PDZ domain-binding motif in CD4+ T cells was examined. Furthermore, GLS2 expression in CD4+ T cells from lupus-prone mice and patients with SLE was analyzed by Western blotting.GLS2, but not GLS1, reduced ROS levels and lipid peroxidation and restored mitochondrial function in T cells. GLS2 promoted IL-2 production through the demethylation of the Il2 promoter. Mutation of the PDZ domain-binding motif abated the ability of GLS2 to regulate IL-2 and ROS levels. In lupus-prone mice and patients with SLE, the expression of GLS2 was decreased in CD4+ T cells. Finally, GLS2 overexpression corrected ROS levels and restored IL-2 production by CD4+ T cells from lupus-prone mice and SLE patients.Our findings suggest that GLS2 has a crucial role in IL-2 production by CD4+ T cells by supporting antioxidant defense, and they offer a new approach to correcting IL-2 production by T cells in SLE.

Objective. To analyze the role of PD-1/PD-L1 signaling pathway in regulating T cell activation and secretion of proinflammatory factors in atrial fibrillation. Methods. Forty-five patients with atrial fibrillation admitted to the cardiology department of our hospital from July 2019 to March 2021 were selected to be included in the atrial fibrillation group, and another 45 healthy volunteers were selected as the control group to compare the changes of T cell CD69 and human leukocyte antigen-DR (HLA-DR) expression in the peripheral blood of the two study groups; compare the changes of programmed death factor-1 on CD4+ and CD8+ lymphocytes in the peripheral blood of the two groups (PD-1) expression changes and PD-L1 and PD-L2 expression changes on peripheral blood myeloid dendritic cells (mDCs) cells; compare the changes of interleukin-2, interleukin-6, interleukin-10, and interleukin-17A (IL-2, IL-6, IL-10, and IL-17), tumor necrosis factor (TNF), and interferon gamma (IFN-γ) concentrations on peripheral blood inflammatory factors in the two groups; and isolate the two groups of peripheral blood mDCs cells; α interferon upregulated PD-L1 expression in the cells and analyzed the effect of PD-L1 expression on the ability of mDCs to stimulate T cells to secrete cytokines.Results. The positive expression rates of CD69 and HLA-DR on peripheral blood CD3+ T lymphocytes were significantly higher in patients in the atrial fibrillation group than in the control group, and the differences were statistically significant ( P < 0.01 ). The positive expression rate of PD-1 on CD4+ lymphocytes was significantly lower in patients in the atrial fibrillation group than in the control group ( P < 0.01 ). There was no statistically significant difference between the two groups in terms of PD-1 positive expression rate on CD8+ lymphocytes ( P > 0.05 ). The positive expression rate of PD-L1 on mDCs cells was significantly lower in patients in the atrial fibrillation group than in the control group ( P < 0.01 ), and there was no statistically significant difference between the two groups in the positive expression rate of PD-L2 on mDCs cells, PD-L1, and PD-L2 on CD4+ and CD8+ T cells ( P > 0.05 ). The concentrations of IL-2, IL-6, IL-10, and IFN-γ in peripheral blood were significantly higher in patients in the atrial fibrillation group than in the control group ( P < 0.05 ), and there was no statistically significant difference in the comparison of IL-17A and TNF concentrations in peripheral blood between the two groups ( P > 0.05 ). In the atrial fibrillation group, the ability of mDCs to stimulate T cells to secrete IL-2 and IFN-γ was significantly higher, and the ability to secrete IL-10 was significantly lower compared with the control group ( P < 0.05 ). After α interferon upregulated PD-L1 expression in cells, the ability of mDCs to stimulate T cells to secrete IL-2, IL-10, and IFN-γ cytokines was reversed in patients in the atrial fibrillation group, and the differences compared with the control group were not statistically significant ( P > 0.05 ). Conclusion. PD-1/PD-L1 signaling pathway may play an immunomodulatory role in the pathogenesis of atrial fibrillation by promoting increased secretion of inflammatory factors through regulating T cell activation.

Systemic lupus erythematosus (SLE) is a heterogeneous autoimmune disorder characterized by disturbed cellular and humoral immune responses. Dysregulations of immune system and immunosuppressive medications predispose SLE patients to infection. This study aims to investigate the alterations and absolute concentrations of lymphocyte subpopulations in SLE patients with different infection and their responses of low-dose IL-2 therapy. A total of 333 patients with SLE without recent infection, 162 patients suffering infection, and age and sex-matched 132 healthy controls (HCs) were recruited. Of them, 54 SLE patients (including 41 non-infected group and 13 infected group) received a 5-day course of low-dose IL-2 administration at a dose of 0.5 million IU per day. Lymphocyte subpopulations were analyzed by flow cytometry. Patients with SLE had lower levels of lymphocyte subpopulations in peripheral blood such as T, B, NK, CD4 + T, CD8+ T, Th1, Th2, Th17, and Treg cells, and the reduction in these cells was more obvious in patients with infection (p p <.01). Low-dose IL-2 effectively expanded T (p p p p p p p

Cancer immunotherapy, which blocks immune checkpoint molecules, is an effective therapeutic strategy for human cancer patients through restoration of tumor-infiltrating (TI) cell function. However, evaluating the efficacy of immune checkpoint inhibitors (ICIs) is difficult because no standard in vitro assay for ICI efficacy evaluation exists. Additionally, blocking a particular immune checkpoint receptor (ICR) is insufficient to restore T cell functionality, because other ICRs still transduce inhibitory signals. Therefore, limiting inhibitory signals transduced via other ICRs is needed to more accurately assess the efficacy of ICIs targeting a particular immune checkpoint. Here, we introduce a newly developed in vitro coculture assay using human peripheral blood mononuclear cells (hPBMCs) and engineered human cancer cell lines. We enriched CD8

Migraine is the second most common neurological disorder. Inflammation plays an important role in the pathology and symptoms of migraine. Although, many studies have analyzed the levels of peripheral cytokines in migraine patients, the conclusions of these studies were not consistent. Meta-analysis for peripheral cytokine levels in migraine is necessary to solve the inconsistency in clinical conclusion.We conducted a systematic search to July 2021, to identify the literatures that measured peripheral cytokine levels in migraine patients and compared them with healthy controls.10 studies were finally included in the meta-analysis: 6 for C-reactive protein (CRP), 2 for interleukin (IL)-1β, 5 for IL-6, 3 for tumor necrosis factor alpha (TNF-α), 1 for IL-2, 2 for IL-10. Compared with healthy controls, we found that the patients with migraine had higher serum levels of CRP (standardized mean difference, SMD = 1.48; P 0.001), IL-1β (SMD = 0.75; P 0.001), IL-6(SMD = 1.18; P0.001) and TNF-α (SMD = 0.69; P = 0.003), while did not have significant difference in serum IL-2(SMD = -0.24; P = 0.25) and IL-10 (SMD = -0.17; P = 0.88).The findings of the meta-analysis provide evidence for higher serum of CRP, IL-1β, IL-6 and TNF-α in migraine patients compared with healthy controls. Our results support that inflammation play a role in the pathophysiology of migraine. However, there was no significant difference in serum IL-2 and IL-10.

Objective The mainstay of treatment for patients with malignant pleural disease is fluid drainage and systemic therapy. A tumor-specific oncolytic virus or T-cell–activating interleukin-2 immunotherapy may provide an opportunity for local control. We previously developed a vaccinia virus–expressing interleukin-2, an oncolytic virus that mediated tumor regression in preclinical peritoneal tumor models with expansion of tumor-infiltrating lymphocytes. We evaluated the antitumor efficacy and immune modulatory effects of vaccinia virus–expressing interleukin-2 in malignant pleural disease. Methods A murine model of malignant pleural disease was established with percutaneous intrapleural deposition of the Lewis lung carcinoma cell line and monitored with bioluminescent imaging. After intrapleural or systemic administration of vaccinia viruses (vaccinia virus yellow fluorescent protein control, vaccinia virus–expressing interleukin-2), systemic anti–programmed cell death-1 antibody, or combination therapy (vaccinia virus–expressing interleukin-2 and anti–programmed cell death-1), tumor mass, immune cell infiltration, T-cell receptor diversity, and survival were assessed. Results Intrapleural vaccinia virus resulted in significant tumor regression compared with phosphate-buffered saline control (P Conclusions Intrapleural vaccinia virus–expressing interleukin-2 reduced tumor burden and enhanced survival in a murine malignant pleural disease model. Increased CD8+ tumor-infiltrating lymphocytes and αβ T-cell receptor diversity are associated with enhanced response. Clinical trials will enable assessment of intrapleural vaccinia virus–expressing interleukin-2 therapy in patients with malignant pleural disease.

Panax notoginseng is the dried root and rhizome of Panax notoginseng, which has the effect of lowering blood lipid, lowering blood pressure and promoting blood circulation to remove blood stasis. At present, the research on Panax notoginseng is mainly focused on its pharmacological action and its compound preparation, but the research on the granule of Panax notoginseng is less. This paper mainly studied the clinical study of compound notoginseng nanoparticles in the treatment of local infection in patients with hydrocephalus after medium craniocerebral injury in neurosurgery. The purpose of this article is to investigate the effects of compound notoginseng nanoparticles on serum TNF-α, IL-2 and IL-6 in rats with craniocerebral injury and to verify the protective effect of compound notoginseng nanoparticles on the body after craniocerebral injury. In this paper, 90 patients admitted to a hospital in this city were divided into a control group, model group and compound notoginseng nanoparticle group. According to the Zealonga method, the neurological function deficit score of experimental rats in each group was evaluated. The levels of TNF-α, IL-2 and IL-6 in the serum of the three groups were observed 1, 3 and 5 days after treatment. RESULTS: Compared with serum TNF-α, IL-2 and IL-6 of the three groups, there were significant differences in the main effects of time and intervention (P0.05). CONCLUSIONS: Compound notoginseng nanoparticles can reduce the contents of TNF-α and IL-6 in serum and increase the expression of IL-2 in rats with craniocerebral injury.

Studies are trying to add immunotherapy to gemcitabine and cisplatin (GP) induction chemotherapy, the standard therapy, in nasopharyngeal carcinoma (NPC) patients with locoregionally advanced disease. However, how the immune system responds to GP remains unknown.We examined the dynamic changes of circulating immune cells and plasma cytokines in NPC patients administered with GP.After GP administration, immunosuppressive myeloid cells, including CD11b+CD14+ monocytes, CD33+ myeloid cells, CD33+CD11+ myeloid cells, total MDSCs (CD33+CD11+HLA-DR-/low), monocytic MDSCs, and granulocytic MDSCs decreased significantly. The regulatory T cells and B cells, two important suppressive lymphocyte subpopulations, also decreased. On the other hand, the levels of CD3+ T cells, total B cells, central memory CD4+ T cells, and pro-inflammatory cytokines (including Interleukin [IL]-1β, IL-6, IL-2, IL-5, and IL-8) increased significantly after GP administration. Besides, GP chemotherapy did not weaken the cytotoxic activity and proliferative capacity of T cells.Our results showed the immune modulation effect of GP induction chemotherapy in locoregionally advanced NPC, providing a solid basis for its combination with immunotherapy.

The clinical course of ulcerative colitis (UC) is variable. There is an unmet clinical need for biomarkers of UC disease behaviour. We aimed to evaluate the association between ex vivo human UC explant conditioned media (explant-CM) secreted protein profiles and UC disease behaviour.UC patients undergoing endoscopy were prospectively recruited. Endoscopic biopsies were collected and explant-CM generated. Association between explant-CM protein secretion profiles and disease progression was evaluated. Disease progression was defined as the requirement for corticosteroid therapy, UC-related hospitalisation, UC-related surgery or the introduction of a new immunomodulatory agent. Association between explant-CM secreted protein profiles and anti-TNF failure status was also evaluated. p values 0.05 were considered significant in analyses.Twenty-four UC patients were included (age [median, range]) 55 [21-72] years; 50% female. Disease progression during follow-up occurred in twelve (50%) patients. Multivariate analysis, including endoscopic remission status, demonstrated reduced IL-2 secretion to be independently associated with UC disease progression, p = 0.01. In univariate analysis, anti-TNF failure status was associated with significantly increased IL-17A/F (p = 0.015) and IL-12 / IL-23p40 (p = 0.044) concentrations. In multivariate analysis, there was a trend towards an association between IL-17A/F and anti-TNF failure status (p = 0.069); FLT-1 was demonstrated to be independently associated with anti-TNF failure status (p = 0.016).Reduced explant-CM secreted IL-2 is associated with UC disease progression. Increased secretion of IL-23 pathway-associated cytokines was observed in anti-TNF failure status consistent with previous reports. Ex vivo human UC explants, generated from endoscopic biopsies, have potential as precision medicine tools in inflammatory bowel disease.

Complete endoscopic mucosal healing is defined as a Mayo endoscopic subscore of 0. Some patients diagnosed with a Mayo endoscopic subscore 0 may present with subsequent clinical relapse. Here, we aimed to demonstrate mucosal cytokine profile as a predictor of clinical relapse in ulcerative colitis patients with a Mayo endoscopic subscore of 0 as a marker of mucosal healing.We conducted prospective observational pilot study to examine the relationship between mucosal cytokine expression and subsequent relapse of UC patients diagnosed with a Mayo endoscopic subscore of 0. We enrolled 55 patients, and expression of cytokines tumor necrosis factor-α, interferon γ, interleukin-1β, interleukin-2, interleukin-4, interleukin-5, interleukin-6, interleukin-7, interleukin-8, interleukin-9, interleukin-10, interleukin-12, interleukin-13, interleukin-15, interleukin-17A, interleukin-17F, interleukin-18, interleukin-21, interleukin-22, interleukin-23, interleukin-27, and interleukin-33 was measured by quantitative real-time PCR using rectal mucosa biopsy materials. Cytokine expression levels were compared between patients who relapsed between March 1, 2016, and March 30, 2020, of the study period and those who remained in remission.Ten cytokines, including interleukin-2, interleukin-4, interleukin-8, interleukin-10, interleukin-12, interleukin-15, interleukin-17A, interleukin-21, interleukin-23, and interleukin-33, were significantly elevated in patients with subsequent relapse compared with those who remained in remission. Interleukin-8 expression was the most useful predictor.In the rectal mucosa of ulcerative colitis patients with Mayo endoscopic subscore 0, levels of several cytokines were elevated in cases of subsequent relapse. Among these, interleukin-8 expression was the most useful for predicting relapse.