Serum Cytokine Concentrations Research Articles

Effect of topical drugs on the local concentrations of IL-5 and mometasone furoate in chronic polypous rhinosinusitis

Long-term persistence of inflammation is the main factor of pathogenesis in chronic polypous rhinosinusitis (CRSwNP). Prospectives of current clinical otorhinolaryngology include modification of drugs containing topical glucocorticosteroids in order to increase local bioavailability, reduce the concentration of glucocorticosteroids, and alleviate the local inflammatory response. The aim of our work was to evaluate the effect of urea, magnesium sulfate, and mannitol on the concentrations of mometasone furoate and IL-5 in the polypous tissue of patients with CRSwNP. The study included 146 patients with CRSwNP aged 18 to 62 years. Biopsies of polypous tissue were taken during polypotomy. In the fluid samples obtained by microdialysis of polypous tissues, mometasone concentrations were determined by high-performance liquid chromatography. Concentrations of IL-3, IL-4, IL-5 and IL-10 cytokines in blood serum, and IL-5 in microdialysates of polypous tissue were measured by ELISA technique. The average age of patients with CRSwNP was 42.9±7.2 years, with prevalence of male patients (n = 93, 63.7%) over females (n = 53, 36.3%). Severity parameters of the disease by the SNOT-22 scale were significantly higher in CRSwNP patients than in control group (p ≤ 0.05). The results of biochemical analysis of microdialysates from polypous tissue showed increased content of Na+ ions in the resistant clinical cases of CRSwNP compared with the sensitive and dependent clinical forms (p < 0.05). Curves with the level of mometasone furoate were obtained in all the studied samples. It was found that the addition of magnesium sulfate solution to mometasone increases the Cmax of mometasone to 154 ng/mL in biological fluids after microdialysis. When urea solution is added to mometasone, Cmax increases to 198 ng/mL. In the native nasal polypous tissue dialysate, IL-5 levels were 89±2.01 pg/mL. Two hours after the addition of mometasone furoate, IL-5 concentrations decreased to 61±3.5 pg/mL, upon addition of urea, it changed to to 69±2.98 pg/mL (p ≤ 0.01, compared with the control without the addition of drugs). Analysis of serum cytokine levels showed that IL-5 concentrations in the patients with CRSwNP significantly exceeded (> 3-fold) those in the control group (p ≤ 0.05). There was a trend for increase of IL-3 and IL-4 (2-fold) and to decreased IL-10 levels in blood sera (> 1.5-fold) in patients with CRSwNP compared with controls. These results may be helpful for development of novel clinical approaches, in order to increase bioavailability of topically administered glucocorticosteroids, thus allowing to promote the effectiveness of basic therapy of CRSwNP and to develop therapy for the prevention of resistant forms of polyposis.

Elevated GRO-α and IL-18 in serum and brain implicate the NLRP3 inflammasome in frontotemporal dementia

Neuroinflammation is a hallmark of frontotemporal dementia (FTD), a heterogeneous group of proteinopathies characterized by the progressive degeneration of the frontal and temporal lobes. It is marked by microglial activation and subsequent cytokine release. Although cytokine levels in FTD brain and CSF have been examined, the number of cytokines measured in each study is limited and knowledge on cytokine concentrations in FTD serum is scarce. Here, we assessed 48 cytokines in FTD serum and brain. The aim was to determine common cytokine dysregulation pathways in serum and brain in FTD. Blood samples and brain tissue samples from the superior frontal cortex (SFC) were collected from individuals diagnosed with behavioral variant FTD (bvFTD) and healthy controls, and 48 cytokines were measured using a multiplex immunological assay. The data were evaluated by principal component factor analysis to determine the contribution from different components of the variance in the cohort. Levels of a number of cytokines were altered in serum and SFC in bvFTD compared to controls, with increases in GRO-α and IL-18 in both serum and SFC. These changes could be associated with NLRP3 inflammasome activation or the NFκB pathway, which activates NLRP3. The results suggest the possible importance of the NLRP3 inflammasome in FTD. An improved understanding of the role of inflammasomes in FTD could provide valuable insights into the pathogenesis, diagnosis and treatment of FTD.

ОЦЕНКА ЗНАЧИМОСТИ ВЗАИМОСВЯЗИ МЕЖДУ ПОКАЗАТЕЛЯМИ РЕМОДЕЛИРОВАНИЯ СОСУДОВ, ВОСПАЛИТЕЛЬНОГО ОТВЕТА И ПОЛИМОРФИЗМА ГЕНОВ ДИСФУНКЦИИ ЭНДОТЕЛИЯ У ПАЦИЕНТОВ МУЖСКОГО ПОЛА С ИНФАРКТОМ МИОКАРДА В ЗАВИСИМОСТИ ОТ НАЛИЧИЯ ИЛИ ОТСУТСТВИЯ ГИПЕРТОНИЧЕСКОГО АНАМНЕЗА

Introduction. Endothelial dysfunction, arterial hypertension and surrogate events of myocardial infarction are interrelated phenomena, while the genetic predisposition in these patients is an independent risk factor for the development of cardiovascular pathology. Aim. The aim of the study was to evaluate the significance of the interaction of associations of polymorphic loci of adhesion molecule genes (immunoglobulin VCAM1), MCP-1 chemokine, vascular endothelial growth factor (VEGFA), nitric oxide control enzyme (DDAH1), markers of the inflammatory response, vascular remodeling and endothelial dysfunction in patients with male myocardial infarction, depending on the presence or absence of arterial hypertension in history in the development of myocardial infarction. Material and methods. 667 male patients and 621 practically healthy individuals in the comparison group were examined. Patients underwent determination of general clinical parameters, serum concentrations of monocyte-chemoattractant protein-1 (MCP-1), vascular endothelial growth factor (VEGFA), cytokines (IL-1,6; TNF-α), molecular genetic study, measurement of media intima thickness of the common carotid artery. Selected in the group of patients with myocardial infarction, individuals with or without a history of arterial hypertension. Results and discussion. As a result of a comparative analysis of groups of patients with and without a history of arterial hypertension with a group of healthy individuals, significant differences in the frequencies of genotypes and alleles for the following polymorphic gene loci were revealed: MCP-1, VEGFA, DDAH1, VCAM1. The serum concentration of cytokines (IL-1,6; TNF-α) in patients, regardless of the presence or absence of a history of hypertension, significantly exceeded the level in the comparison group. The VEGFA index in patients with myocardial infarction with arterial hypertension was significantly increased compared to the serum level in patients with myocardial infarction without arterial hypertension. Conclusion. A higher significance of the interaction between indicators of vascular remodeling, inflammation, endothelial dysfunction gene polymorphism in the development of myocardial infarction in patients with a history of hypertension, in contrast to non-hypertensive patients with myocardial infarction, was revealed, which indicates a powerful contribution of arterial hypertension to the etiology and implementation of myocardial infarction.

Peripheral blood immune cell and cytokine profiling of patients receiving corticosteroids for immune checkpoint inhibitor-related adverse events: Steroid dose and duration.

e14694 Background: Immune checkpoint inhibitors (ICI) have dramatically extended survival in numerous malignancies, but many patients treated with ICIs experience severe immune-related adverse events (irAEs) that can affect virtually any organ. Corticosteroids (steroids) tapered over weeks to months are usually effective but may affect ICI anti-tumor activity. Tapers are often imprecise, resulting in over- or undertreatment of some patients. There are no known blood biomarkers to guide steroid taper management. We therefore profiled peripheral blood immune cells and cytokines at key timepoints to identify biomarkers of response to steroids for irAE treatment. Methods: We collected blood samples before ICI, during irAE, at three steroid dose levels (high, intermediate, and low), and post-steroids from patients treated with ICI therapy at Yale Cancer Center. We measured 48 serum cytokines with a multiplexed immunoassay and immunophenotyped 40 markers on peripheral blood mononuclear cells with mass cytometry (CyTOF). We correlated changes in serum cytokine concentration across timepoints with clinical features. We applied a cluster analysis to the CyTOF data to identify immune cell types and their change in frequency across timepoints. Results: We evaluated 12 patients treated with anti-PD-1 + anti-CTLA-4 therapy who experienced at least 1 severe irAE (11 patients with melanoma, 1 patient with renal cell carcinoma). Five patients had colitis, 6 had hepatitis, and 1 had hepatitis and enteritis. Concentrations of TNFα, IFNγ-mediated cytokines (IL-12p40, IL-18, IL-27), and T cell homing chemokines (CXCL9, CXCL10) increased during irAEs and decreased during steroid therapy without significant change across steroid dose levels (p < 0.05). We observed the largest changes in patients who experienced another irAE within 3 months (n = 5). IL-6 was notably higher in patients who required steroid taper escalation for recurrent symptoms before or soon after completing steroid therapy (n = 8, p = 0.02). Compared to other T cell subtypes, the frequency of CD4 T effector memory (Tem) cells increased during irAE and decreased upon completing steroid therapy but did not return to baseline (p < 0.01). Conclusions: In patients treated with anti-CTLA-4 + anti-PD-1 therapy, IL-6 and IFNγ-mediated cytokines were elevated during irAEs and suppressed at multiple steroid dose levels despite symptom recurrence during the steroid taper. Increased CD4 Tem cell frequency during irAEs may reflect circulating pathogenic T cells, and the delayed decrease and incomplete return to baseline could be due to the severity of irAEs represented in this study. Specific cytokines may predict the occurrence of multiple irAEs and the need for a longer steroid taper. Additional longitudinal analyses of steroid effects on irAEs are needed to validate these findings.

Five-year follow-up update of defined-composition CD19 CAR T-cell therapy for relapsed/refractory CLL.

7511 Background: We have described high response rates in patients (pts) with relapsed/refractory (R/R) chronic lymphocytic leukemia (CLL) treated with CD19-targeted CAR T cells. We now report comprehensive analyses of factors associated with duration of response (DOR) with a 68-month median follow-up. Methods: We included 49 pts with R/R CLL and/or Richter’s transformation (n = 10) treated on a phase I/II clinical trial of defined-composition CD19 CAR T-cell therapy (1:1 ratio of CD8+:CD4+). Response rates were investigator-assessed by 2018 iwCLL criteria. Measurable residual disease (MRD) in bone marrow (BM) was evaluated by multiparameter flow cytometry (MFC) and IGH next-generation sequencing (NGS; clonoSEQ assay). CAR T-cell enumeration was assessed by MFC. Serum cytokine concentrations (20 analytes) were measured by Luminex assay. Univariate associations between duration of response (DOR) and patient, disease, CAR T-cell manufacturing, and treatment-related variables were estimated using Cox regression (50+ variables). Association between CAR T-cell in vivo persistence and DOR was modeled with a time-dependent Cox model including blood CAR transgene levels (copies/µg genomic DNA). Results: Median age was 61 years (IQR, 55-67). Forty-six pts (94%) had complex karyotype and/or 17p deletion; 46 (94%) received cyclophosphamide/fludarabine lymphodepletion. In the response-evaluable cohort (n = 47), median follow-up was 68.3 months (IQR, 54.3-81.6). Overall and complete response rates at day+28 were 70% and 17%, respectively, and the median DOR was 22.2 months (95% CI, 10.7-not reached [NR]). MRD-negativity at day+28 by MFC occurred in 33/47 (70%). In pts with MRD-negativity by MFC and available data, MRD negativity by NGS was seen in 18/29 (62%). In NGS MRD-negative responders, the median DOR was 56.8 months (95% CI, 39.5-NR). In all evaluable patients, the 5-year OS and PFS were 35.5% (95% CI, 24.0%-52.5%) and 21.5% (95% CI, 12.0%-38.5%), respectively. In univariate Cox regression, day+28 MRD-negativity by MFC (HR = 0.03, 95% CI, 0.01-0.18, p < 0.001), day+28 MRD-negativity by NGS (HR = 0.21, 95% CI, 0.07-0.61, p = 0.004), peak CD8+ CAR T-cell expansion (HR = 0.49, 95% CI, 0.24-1.00, p = 0.05), CAR T-cell persistence (HR = 0.71, 95% CI, 0.54-0.91, p = 0.01), and lower peak MIP-1β (HR = 3.12, 95% CI, 1.13-8.65, p = 0.03) were associated with longer DOR. Peak MIP-1β remained associated with DOR (HR = 3.83, 95% CI, 1.35-10.9, p = 0.012) in a multivariable model including pre-LD CLL cells in BM, tumor cross-sectional area, peak CD8+ CAR T-cell expansion, and day+28 MRD by MFC. Conclusions: CD19 CAR T-cell therapy achieved durable remissions in R/R CLL pts. Day+28 MRD, CAR T-cell peak expansion, and CAR T-cell persistence were strongly associated with DOR. Lower peak serum levels of MIP-1β, a cytokine known to be produced by CLL cells upon BCR activation, was strongly and independently associated with DOR. Clinical trial information: NCT01865617 .

Features of the blood cytokine profile in gastroesophageal reflux disease in schoolchildren with gastritis and family history of peptic ulcer

Gastroesophageal reflux disease (GERD) is a common acid-dependent disease among the population, including children, with multifactorial genesis. It, like many other acid-dependent diseases (peptic ulcer, etc.) is associated with a family predisposition to the disease. Of interest is the study of the role of cytokines in the regulation of pathology in childhood, depending on the severity of a family history of peptic ulcer disease. Aim: to evaluate the levels of cytokines in the blood serum in case of family history of ulcerative diseases in schoolchildren with gastritis associated with GERD. In the course of a scientific study, 142 children with gastroenterological complaints aged 7-17 years were examined. The diagnosis of GERD was made in the presence of weekly heartburn in accordance with the global consensus on pathology in children. All subjects underwent gastroscopy with taking biopsy material from the gastric mucosa and morphological confirmation of their diagnosis of gastritis in accordance with the Sydney classification. The concentration of cytokines in blood serum (IL-2, IL-4, IL-6, IL-8, IL-10, IL-18, IL-1β, IFNα, TNFα) was obtained by enzyme immunoassay. During statistical processing, the χ2 and Mann–Whitney tests were used. The studies were approved by the ethics committee and informed consents of patients and their parents were obtained prior to the start of the study. The results of the study did not show significant differences in the concentration of cytokines in schoolchildren depending on the presence of GERD. In children with a family burden of peptic ulcer, GERD was detected more often (p = 0.054), which is probably a consequence of their increased acid formation. Changes in the cytokine profile of the blood were noted. During GERD, with aggravation of peptic ulcer, there was an increase in the replication of IL-4 (p = 0.027) and IFNα (p = 0.001). The increase in blood IFNα in children with GERD with family burden is obviously aimed at enhancing immune responses involving the whole body to damage. This is due to its functional role – participation in the immune response. Increased replication of IL-4, obviously, provides an increase in metabolic, immune processes in the body aimed at optimizing the course of proliferative processes in the esophageal mucosa under conditions of increased secretion of hydrochloric acid in the stomach. Thus, when a family history of peptic ulcer is aggravated in schoolchildren with gastritis associated with GERD, a number of links in the cytokine network (IL-4, IFNα) move to the systemic level of regulation.

Immunotoxicity induced by occupational inhalation exposure to waste anesthetic gases: a historical cohort study.

This was a historical cohort study in which 30 male participants with at least one year of work experience in the operating room at the time of the study and 30 unexposed referent subjects were investigated. Exposure levels were quantified by measuring the urinary concentrations of nitrous oxide (N2O), isoflurane, and sevoflurane gases by headspace gas chromatography-mass spectrometry. Serum concentrations of interleukin-4 (IL-4), Th2-type cytokines, and interferon-gamma (IFN-γ) were measured by the ELISA method. Additionally, an automated hematology analyzer was used for the white blood cell count and white blood cell differential test. The data were analyzed using SPSS software for Windows version 21. Mean urinary concentrations of N2O, isoflurane, and sevoflurane were found to be 211.57±75.15, 4.06±0.96, and 19.51±12.96 ppb, respectively. In simplistic statistical data analysis, significant differences were noted between exposed and control groups as far as the mean serum cytokines levels (IFN-γ, IL-4) were concerned. Furthermore, after adjusting for important confounders, statistical analysis showed that the IFN-γ, IL-4, and the ratio of IFN-γ/IL-4 were significantly higher in the exposed group than in the referent subjects. These findings provide corroborative evidence to further substantiate the contention that exposure to anesthetics agents (N2O, isoflurane, and sevoflurane) is associated with subtle, subclinical, prepathological changes in the parameters of immune function. The long-term ramification of these changes requires further investigation.

Levels of chemokines and other inflammatory mediators in patients with mild cognitive impairment undergoing rehabilitation

Alzheimer's disease is the most common neurodegenerative disease in old age. In some cases, it is preceded by mild cognitive impairment (MCI). One of the important components in the pathogenesis of neurodegeneration is chronic neuroinflammation (inflammatory activation of microglia and astrocytes in the brain). Systemic inflammatory response and immune dysregulation may contribute to neuroinflammation. The purpose of this study was to investigate the level of chemokines and other inflammatory mediators in patients with MCI who underwent medical rehabilitation, and to study its associations with the severity of cognitive impairment. The study group included 48 patients with MCI undergoing rehabilitation. Rehabilitation included cognitive therapy, psychotherapy and tasks for unaided performance. Repeated examination was conducted 6 months after the completion of rehabilitation. The control group included 46 healthy volunteers. Multiplex assay was used to determine serum cytokine and chemokine concentrations. Student's t-test was used to assess the significance of differences. Assessment of cognitive functions was performed using international neuropsychological scales. In patients with MCI, we have found an increase in the levels of several cytokines and chemokines (TNFα, CXCL10/IP10, MDC) that regulate systemic inflammation, cellular and humoral mechanisms of adaptive immunity. After the rehabilitation course their levels returned to normal. It was also found that decrease in CCL7 level in the patients before the rehabilitation course is associated with the severity of cognitive impairment. The findings contribute to understanding the role of chemokines in the pathogenesis of MCI, and indicate that their levels can be potential biomarkers of the severity of cognitive impairment. For translation of the findings into clinical practice, their validation in larger studies is needed, as well as assessing the associations between chemokine levels and the severity of cognitive impairment in MCI over long-term follow-up.

Investigation of cytokines in equine serum, conceptuses and endometrium in early pregnancy

Cytokines and their receptor systems are present at the fetomaternal interface of some species and regulate trophoblast biology. The cytokine and chemokine systems are key in the communication between fetal derived trophoblast cells and the maternal tissue. However, evidence for the presence of cytokines and their receptors during early equine pregnancy is lacking. Our objective was to investigate interleukins, chemokine ligands, dendritic cells and TNFα in equine serum, conceptuses and endometrium during early pregnancy. Daily transrectal ultrasound examinations of the mares were performed when in estrus until the day (D) of ovulation (OV). An endometrial biopsy was taken from non-pregnant (NP) mares at: D11 (NP11; n=6) or D13 (NP13; n=7). Mares in the pregnant (P) group were bred by artificial insemination from a single fertile stallion, and transrectal ultrasound was performed to detect and measure conceptuses. On the day of sample collection, conceptuses were collected through a transcervical bivona catheter and uterine lavage using 0.9% NaCl, and this was followed by an endometrial biopsy. Pregnant mares’ conceptuses and endometrial biopsies were retrieved at D11 (P11; n=7), D13 (P13; n=8), or D16 (P16; n=6). A D16 sample was not obtained for NP group. A Bradford assay was performed on extracted endometrium and conceptus tissue to determine total protein concentration,followed by a bead-based equine validated multiplex assay (Luminex Corp. Austin, TX) to quantify concentration of the following equine specific inflammatory markers: IL-4, IL-17, IL-10, IL-1β, CCL2, CCL3, CCL5, CCL11, sCD-14 and TNFα. A restricted maximum likelihood model was performed to compare the protein concentration between groups and days in JMP Pro 16 (Cary, NC) with the mare as a random factor and significance set at p<0.05. Cytokine and chemokine serum concentrations were similar between groups and days of collection. Regarding the endometrium, IL-17 U/mg was higher (p<0.05) in group NP13 [median (interquartile)] [11.4 (8.2, 21.7)] than NP11 [4.1 (2.6, 5.3)]. In pregnant mares, endometrial IL-17 was higher in P13 group [6.3 (3.4, 13.9)] compared to group P16 [2.5 (2.1, 4.8)] (p<0.05). In conceptuses, D11 had significantly higher concentrations of CCL2 [1350.2 (793, 1839.5); p<0.03], CCL3 [1990.9 (998.6, 3083.8); p<0.01], CCL5 [363.2 (149.6, 480.6); p<0.006] and CCL11[3143.7 (1764.2, 5221.9); p<0.01 compared to D16 [66.7 (38.8, 644.8); 67.8 (43, 572.30); 9.1 (6.6, 96.3); 196.2 (150.9, 1436.6)], respectively. Higher chemokine concentrations in the earlier embryos may reflect the recruitment of endometrial leukocytes. Further investigation into the role of cytokines and chemokines in early equine pregnancy is warranted.

Mortality, growth, and egg production do not differ between nontransgenic and transgenic female chickens with ubiquitous expression of the 3D8 single chain variable fragment gene

To date, many transgenic (TG) chicken lines have been developed, but few studies have performed a comparative analysis of their mortality, growth, and egg productivity. Previously, we reported the production of 3D8 scFv TG chickens showing antiviral activity. Here, we performed a biometric characterization of TG offspring female chickens. We selected 40 TG and 40 non-TG offspring female chicks among newly hatched chicks produced via artificial insemination of semen from heterotypic 3D8 scFv males into wild-type female chickens. Serum was collected at 14 wk of age, and serum concentrations of biochemical parameters, cytokines, and sex hormones were analyzed. Mortality and growth were monitored daily from 1 to 34 wk, egg productivity was monitored daily from 20 to 34 wk, and the weekly average values were used for analyses. Some serum parameters and cytokines were significantly different between non-TG and TG offspring female chickens. The levels of phosphorus (PHOS), total protein (TP), albumin (ALB), globulin (GLOB), and alanine aminotransferase (ALT) were significantly higher in non-TG chickens (P < 0.05). The levels of alkaline phosphatase (ALP) and gamma-glutamyltransferase (GGT) were significantly higher in TG chickens (P < 0.05). The levels of insulin growth factor-1 (IGF-1), interferon-gamma (INF-γ), interleukin-4 (IL-4), and IL-8 were significantly lower in TG chickens (P < 0.05). Despite these differences, the mortality rates, body weight, egg production rates, and egg weight were not significantly different in the experimental groups of non-TG and TG offspring female chickens (P > 0.05). In conclusion, ubiquitous expression of the 3D8 scFv gene in TG offspring female chickens does not affect some biometric characteristics, including mortality, growth, and egg productivity.

Association of Peripheral Inflammatory Biomarkers and Growth Factors Levels with Sex, Therapy and Other Clinical Factors in Schizophrenia and Patient Stratification Based on These Data.

Multiple lines of evidence are known to confirm the pro-inflammatory state of some patients with schizophrenia and the involvement of inflammatory mechanisms in the pathogenesis of psychosis. The concentration of peripheral biomarkers is associated with the severity of inflammation and can be used for patient stratification. Here, we analyzed changes in serum concentrations of cytokines (IL-1β, IL-2, IL-4, IL-6, IL-10, IL-21, APRIL, BAFF, PBEF/Visfatin, IFN-α, and TNF-α) and growth/neurotrophic factors (GM-CSF, NRG1-β1, NGF-β, and GDNF) in patients with schizophrenia in an exacerbation phase. IL-1β, IL-2, IL-4, IL-6, BAFF, IFN-α, GM-CSF, NRG1-β1, and GDNF increased but TNF-α and NGF-β decreased in schizophrenia compared to healthy individuals. Subgroup analysis revealed the effect of sex, prevalent symptoms, and type of antipsychotic therapy on biomarker levels. Females, patients with predominantly negative symptoms, and those taking atypical antipsychotics had a more pro-inflammatory phenotype. Using cluster analysis, we classified participants into "high" and "low inflammation" subgroups. However, no differences were found in the clinical data of patients in these subgroups. Nevertheless, more patients (17% to 25.5%) than healthy donors (8.6% to 14.3%) had evidence of a pro-inflammatory condition depending on the clustering approach used. Such patients may benefit from personalized anti-inflammatory therapy.

Serum Concentrations of Thelper2-derived Cytokines in Stevens-Johnson Syndrome and Toxic Epidermal Necrolysis

BACKGROUND: Stevens-Johnson syndrome (SJS) and toxic epidermal necrolysis (TEN) are severe cutaneous adverse drug reactions. Some immunological and genetic factors are believed to be involved in the pathogenesis of SJS/TEN, including T helper 1 and T helper 2 (Th2)-derived cytokines. AIM: This study aims to evaluate the serum levels of Th2-derived cytokines in SJS/TEN, compare to those of erythema multiforme (EM) patients, and the relation between them and the progress of SJS/TEN. METHODS: This was a sectional descriptive study conducted at the National Hospital of Dermatology and Venereology, in Hanoi, Vietnam, from October 2017 to September 2019. 48 SJS/TEN patients, 43 EM patients, and 20 healthy controls (HCs) participated. Serum interleukin (IL)-4, IL-5, and IL-13 levels were measured by using the fluorescence covalent microbead immunosorbent assay (FCMIA) (ProcartaPlex Immunoassay Panels kit, Thermo Fisher Scientific, USA). The Mann-Whitney U test was used to compare the serum IL levels of the two groups. The Wilcoxon tests were used to compare quantitative variables before and after the treatment. Differences were considered to be statistically significant at p &lt; 0.05. RESULTS: 19 SJS patients (39.5%) and 29 TEN patients (60.5%) participated in our study. The mean age was 49.3, range of 19–77 years (47.9% males; 52.1% females). The most common causative drugs were traditional medicine (29.1%), and allopurinol (12.5%). On the day of hospitalization, the serum level of IL-4 in the SJS/TEN group was 3 ± 7.5 pg/mL, statistically significantly higher than that in the HCs group (p &lt; 0.05), but not higher than that in the EM group (p &gt; 0.05); serum levels of IL-5 and IL-13 in the SJS/TEN group were 4.5 ± 9.8 pg/mL and 1.6 ± 0.6 pg/mL, respectively, similar to those in the EM and HCs groups. On the day of re-epithelialization, in SJS/TEN patients, the serum level of IL-5 was 1 ± 2.8 pg/ml, statistically significantly lower than that on the day of hospitalization (3 ± 7.5 pg/mL) with p &lt; 0.05. Regarding serum levels of IL-4 and IL-13, there was no difference between the two- time points. CONCLUSION: The serum concentrations of Th2-derived cytokines (IL-4, IL-5, and IL-13) were not higher in the SJS/TEN group than in the EM group and there was no significant change in the clinical progression of SJS/TEN, except the serum level of IL-5.

Some features of the immune system response in the new coronavirus infection COVID-19

AIM: to study the cytokine link of immunity in patients with the new coronavirus infection COVID-19 and the possibility of using these data to predict the risk of lung tissue damage. MATERIALS AND METHODS: Patients with diagnosed new coronavirus infection COVID-19 were divided into 4 groups depending on the severity of the disease: 1 — asymptomatic patients; 2 — patients with lung tissue damage in the amount of 25%; 3 — patients with lung tissue damage 50%; 4 — patients with lung tissue damage 75% (according to the results computed tomography). Immunological studies were performed upon admission to the clinic, then after 7 days and before discharge. The concentration of cytokines in blood serum was studied by the method of solid-phase enzyme immunoassay using kits manufactured by Vector Best. RESULTS: In all groups, a low content of proinflammatory cytokines was found: tumor necrosis factor and interferon-γ, interleukin-1, interleukin-2. The median level of interleukin-6 in all groups during the entire follow-up period was within the normal range, however, in some patients there was a multiple excess of its concentration. In groups with 50% and 75% lung tissue damage, an increase in the concentration of interleukin-8 was shown. Statistically significant differences were noted only for the level of interleukin-8 between the group with an asymptomatic form of infection and the CT-2 and CT-3 groups. It was found that the concentration of interleukin-1 below 3.58 pg/ml is a significant risk factor for severe disease. In all groups of subjects, low levels of interleukin-4 and interleukin-10 were noted, while there was no significant difference between the groups. Positive correlations were revealed between the degree of lung tissue damage and the level of interleukins 6, 2 and 8. Negative correlations were established between the degree of lung tissue damage and the content of interleukin-1. CONCLUSION: There was no significant activation of the cytokine link of adaptive antiviral immunity in the examined contingent. Serum cytokine levels were within normal values and had no significant intergroup differences. An increase in IL-8 in groups with clinically pronounced signs of infection indicates the activation of indicators characterizing innate immunity.

CAR +and CAR −T cells differentiate into an NK-like subset that is associated with increased inflammatory cytokines following infusion

Abstract Chimeric antigen receptor (CAR) T cells have demonstrable efficacy in treating B-cell malignancies. Factors such as product composition, lymphodepletion and immune reconstitution are known to influence CAR +T cell survival and persistence, however the determinants of CAR +T cells’ fate and differentiation and how the patients’ reconstituting immune system influence therapy outcomes remain poorly understood. We applied single cell multi-omics analysis of RNA, protein and clonal profiles of CAR +T and PBMC in patients receiving donor-derived products from a piggybac CAR19 clinical trial (N=8 patients). We used these data to reconstruct a differentiation trajectory, which explained the observed phenotype and fate of CAR +and CAR −T cells within the first 30 days of post-infusion. Following initial lympho-depletion, endogenous CAR −CD8 +, γδ T cells and CAR+ T cells clonally expanded, and differentiated across heterogenous phenotypes. They transitioned from a dominant resting or proliferating state into precursor of exhausted T cells, and finally into a terminal NK-like phenotype. The subset of terminal NK-like CAR +T cells was associated with increased serum concentrations of proinflammatory cytokines including IL-12 and IL-18. Validation on a published cohort (61 patients) showed that this subset was associated with clinical outcome at 6 months. Finally, we analysed in detail one patient that developed CAR T cell induced lymphoma and discussed the impact of early phase therapy in determining this unexpected outcome. These results demonstrate that CAR- and CAR+ T cells share a differentiation profile which may be determined by non-CAR derived signals and which may influence patients’ immune-recovery and outcome. National Health And Medical Research Funding (Australia) (APP1121643, APP1128416) BD Bioscience

82 Responses to a combined exercise and transport stressor following dietary supplementation of anti-endotoxin IgY in mature horses

Immunoglobulin Y elicits positive effects in domestic livestock with gastrointestinal or inflammatory diseases; however, there are limited studies in horses. To test the hypothesis that dietary anti-endotoxin IgY (IgY) has a positive effect on gastrointestinal health and mitigates systemic inflammation in horses undergoing exercise, 30 stock-type horses (578 ± 59 kg BW; 14 ± 3 yr) were used in a completely randomized design. Horses were stratified by BW, age, and sex and then randomly assigned to treatments: 0 g/d IgY (CON; n = 10), 1 g/d IgY (TRT1; n = 10), or 2 g/d IgY (TRT2; n = 10). Horses were fed every 12 h, with dietary treatments (Camas Inc., Le Center, MN) top-dressed on the morning concentrate for 33 d. Horses were maintained in dry lots with ad libitum access to Coastal bermudagrass hay. Horses were exercised 5 d/wk for 1 h. On d 0 and 30, BW and BCS were recorded, and blood was collected for complete blood cell count (CBC) and serum blood chemistry. On d 32, horses were transported 270 km and underwent a 3.2 km exercise stressor on concrete at a walk with maximum 15 min of trot. Whole blood, serum, and fecal samples collected via rectal palpation were obtained pre-, post-, and 24 h post-trailering. Fecal pH was determined by a portable meter, and serum cytokine (TNFα, IL-4, IL-10, IL-1β, IL-17, IFN-α, IFN-γ) and chemokine (CCL2, 3, 5 and 11) concentrations weremeasured using a multiplex platform. Whole blood DNA was extracted using a commercial DNA extraction kit. For gut permeability analysis, 16s rDNA was analyzed via qPCR and calculated as fold change from d 0. Data were analyzed using PROC MIXED of SAS v9.4. Dietary IgY did not affect BW, BCS, blood chemistry, or CBC (P ≥ 0.17). Fecal pH decreased over time (P ≤ 0.01). There was no treatment × time interaction for 16S rDNA (P = 0.45). Dietary treatment affected circulating 16S rDNA (P < 0.01) where TRT2 (1.89-fold) was greater than CON (0.70-fold), and TRT1 (1.24-fold) was intermediate. A significant time effect (P = 0.001) was observed for 16S rDNA where fold change decreased after the stressor from 1.90-fold for pre-, to 1.02 for post-, and 0.92 for 24 h post- trailering. There tended to be a treatment × time interaction (P = 0.08) for CCL3, as CON was greater at 24 h and TRT2 declined post-trailering, returning to baseline by 24 h. Levels of CCL5 decreased (P < 0.01) and TNFα tended to decline (P = 0.06) through 24 h post trailering in all treatment groups. Other cytokines and chemokines were non-detectible. Despite greater circulating 16s rDNA with higher treatment dosages, there was no effect on systemic inflammatory markers.

Serum interleukin-6 level predicts the prognosis for patients with alcohol-related acute-on-chronic liver failure.

Heavy alcohol consumption is the most common etiology of acute-on-chronic liver failure (ACLF) in Japan. In some patients, ACLF is associated with a fatal outcome in less than 6months. We evaluated the prognosis of patients with alcohol-related ACLF in our cohort and explored the prognostic factors. Forty-six patients with alcoholic liver cirrhosis who fulfilled the Japanese diagnostic criteria for ACLF, including those classified as extended and/or probable, were enrolled in this study. Serum concentrations of inflammatory cytokines (interleukin [IL]-1β, IL-6, IL-8, IL-10, IL-12p70 and TNFα) were measured. We assessed prognosis and identified factors associated with survival. During the median 33-day observation period, 19 patients died, and 3 patients underwent living donor liver transplantation. Cumulative survival rates of patients treated without liver transplantation were 69, 48, 41, and 36% at 1, 3, 6, and 12months, respectively. Eighteen of the 19 deceased patients died within 6months after ACLF diagnosis. Serum concentrations of inflammatory cytokines were significantly elevated, and patients who underwent liver transplantation or who died within 6months after admission had significantly higher serum IL-6 levels than the survival group. Multivariate analysis identified IL-6 > 23.3pg/mL at admission and model for end-stage liver disease (MELD) score ≥ 25 on day 4 of admission as significant independent factors for mortality within 6months. Serum IL-6 level and Day-4 MELD were prognostic factors for alcohol-related ACLF. Early liver transplantation is a potential treatment option for patients whose prognosis is expected to be poor.

Ursolic Acid Alleviates Cancer Cachexia and Prevents Muscle Wasting via Activating SIRT1

Skeletal muscle wasting is the most remarkable phenotypic feature of cancer cachexia that increases the risk of morbidity and mortality. However, there are currently no effective drugs against cancer cachexia. Ursolic acid (UA) is a lipophilic pentacyclic triterpene that has been reported to alleviate muscle atrophy and reduce muscle decomposition in some disease models. This study aimed to explore the role and mechanisms of UA treatment in cancer cachexia. We found that UA attenuated Lewis lung carcinoma (LLC)-conditioned medium-induced C2C12 myotube atrophy and muscle wasting of LLC tumor-bearing mice. Moreover, UA dose-dependently activated SIRT1 and downregulated MuRF1 and Atrogin-1. Molecular docking results revealed a good binding effect on UA and SIRT1 protein. UA rescued vital features wasting without impacting tumor growth, suppressed the elevated spleen weight, and downregulated serum concentrations of inflammatory cytokines in vivo. The above phenomena can be attenuated by Ex-527, an inhibitor of SIRT1. Furthermore, UA remained protective against cancer cachexia in the advanced stage of tumor growth. The results revealed that UA exerts an anti-cachexia effect via activating SIRT1, thereby downregulating the phosphorylation levels of NF-κB and STAT3. UA might be a potential drug against cancer cachexia.

Immunological and prognostic significance of tumour necrosis in colorectal cancer

BackgroundColorectal cancer (CRC) causes the second most cancer deaths worldwide, but the disease course varies according to tumour characteristics and immunological factors. Our objective was to examine the associations of tumour necrosis with tumour characteristics, immune cell infiltrates, serum cytokine concentrations, as well as prognosis in CRC.MethodsThree independent CRC cohorts, including 1413 patients, were analysed. Associations of the areal percentage of tumour necrosis with clinicopathologic parameters, tumour infiltrating immune cells, cytokine concentrations in systemic and mesenteric vein blood, and survival were examined.ResultsHigher tumour necrosis percentage associated with shorter colorectal cancer-specific survival independent of tumour grade, T, N or M-class, mismatch repair status, BRAF status, and other possible confounding factors. In the largest cohort (N = 1100), the HR for high tumour necrosis percentage (≥40% vs. <3%) was 3.22 (95% CI 1.68–6.17, Ptrend < 0.0001). Tumour necrosis percentage positively correlated with peripheral serum levels of CXCL8, a proinflammatory chemokine, and negatively correlated with mesenteric serum levels of CXCL10 and mast cell densities in the invasive margin of the tumour.ConclusionsOur results support the value of tumour necrosis as a prognostic factor in colorectal cancer. CXCL8 may have a role in the systemic effects of tumour necrosis.

Abstract 5544: Association of circulating markers of gut barrier damage and inflammation with colorectal neoplasia stage

Abstract A disturbance in the microbiome facilitated by gut barrier dysfunction and inflammation can influence the progression of colorectal neoplasia. One of the most studied indicators of gut barrier status is lipopolysaccharide (LPS). Increased permeability of the gut barrier leads to extensive translocation of microbes into the lamina propria, which is associated with higher circulating levels of LPS. Additionally, increased serum concentrations of inflammatory cytokines including IFNy, IL6, IL10, and TNFα have been associated with impaired gut integrity and its progressive dysfunction. We investigated the role of gut barrier permeability and inflammation in colorectal cancer (CRC) development by assessing markers of intestinal barrier dysfunction and inflammation in two different patient cohorts; Irish (discovery) and Czech (validation). ELISA assays were used to assess circulating concentrations of lipopolysaccharide binding protein (LBP) and C-reactive protein (CRP) in serum from patients with CRC (n=30), colorectal adenoma (n=40), and negative after colonoscopy/blood donor controls (n=35). Circulating levels of inflammatory cytokines and calprotectin were assessed using an MSD MultiSPOT platform and an MSD R-plex assay (Mesocale Discovery, Rockville, Md, United States), respectively. Differences between pathology groups were assessed using the Kruskal Wallis or Mann-Whitney U test. Receiver operating curve (ROC) analysis was implemented to identify single and combinations of biomarkers capable of discriminating between CRC patients and controls with maximum sensitivity (SE) and specificity (SP) (RStudio, PBC, Boston, MA URL http://www.rstudio.com/). In the discovery cohort, levels of LBP, TNFα, IL6, calprotectin and CRP were significantly higher in CRC patients than in controls after false discovery rate correction (p=0.02, p=0.03, p=0.006, p=0.01, p=0.006, respectively). The area under the ROC curve (AUC) was 0.76 (95% CI: 0.64-0.87, p = 0.003) for LBP, 0.69 (95% CI: 0.55-0.82, p = 0.007) for calprotectin and 0.87(95% CI: 0.74-1.00, p = 0.003) for CRP. The optimal biomarker combination for discriminating between CRC and controls was the combination of the three markers (SP= 0.75, SE = 0.75). In the validation cohort, levels of calprotectin and TNFα, were significantly higher in cancer compared to control (p=0.05, p=0.02, respectively). Furthermore, IFNγ and IL10 were also significantly elevated in cancer compared to control (p= 0.02, p=0.003, respectively). These results support the hypothesis that a dysfunctional gut barrier and attendant inflammation may influence CRC development and progression. Additionally, LBP, CRP and calprotectin may have potential as diagnostic biomarkers for CRC or for risk stratification. Gut bacterial and barrier wall modulation and stabilization has potential for preventing colorectal adenomas and hence colorectal cancer. Citation Format: Flavia Genua, Petr Holý, Pavel Souček, Václav Liška, David J. Hughes. Association of circulating markers of gut barrier damage and inflammation with colorectal neoplasia stage. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5544.

A machine-learning based bio-psycho-social model for the prediction of non-obstructive and obstructive coronary artery disease

BackgroundMechanisms of myocardial ischemia in obstructive and non-obstructive coronary artery disease (CAD), and the interplay between clinical, functional, biological and psycho-social features, are still far to be fully elucidated.ObjectivesTo develop a machine-learning (ML) model for the supervised prediction of obstructive versus non-obstructive CAD.MethodsFrom the EVA study, we analysed adults hospitalized for IHD undergoing conventional coronary angiography (CCA). Non-obstructive CAD was defined by a stenosis < 50% in one or more vessels. Baseline clinical and psycho-socio-cultural characteristics were used for computing a Rockwood and Mitnitski frailty index, and a gender score according to GENESIS-PRAXY methodology. Serum concentration of inflammatory cytokines was measured with a multiplex flow cytometry assay. Through an XGBoost classifier combined with an explainable artificial intelligence tool (SHAP), we identified the most influential features in discriminating obstructive versus non-obstructive CAD.ResultsAmong the overall EVA cohort (n = 509), 311 individuals (mean age 67 ± 11 years, 38% females; 67% obstructive CAD) with complete data were analysed. The ML-based model (83% accuracy and 87% precision) showed that while obstructive CAD was associated with higher frailty index, older age and a cytokine signature characterized by IL-1β, IL-12p70 and IL-33, non-obstructive CAD was associated with a higher gender score (i.e., social characteristics traditionally ascribed to women) and with a cytokine signature characterized by IL-18, IL-8, IL-23.ConclusionsIntegrating clinical, biological, and psycho-social features, we have optimized a sex- and gender-unbiased model that discriminates obstructive and non-obstructive CAD. Further mechanistic studies will shed light on the biological plausibility of these associations.Clinical trial registrationNCT02737982.Graphical abstract