Thymic Stromal Lymphopoietin Concentration Research Articles

Exploring TSLP and IL-33 Serum Levels and Genetic Variants: Unveiling Their Limited Potential as Biomarkers for Mild Asthma in Children.

As the burden of mild asthma is not well understood, the significance of expanding research in the group of patients with mild asthma is emphasized. Thymic stromal lymphopoietin (TSLP) and interleukin 33 (IL-33) are involved in the pathogenesis of atopy and the immune response to inhaled environmental insults, such as allergens, in asthmatic patients. Objectives: The objective of this study was to explore the correlation between specific polymorphisms within the genes encoding TSLP and IL-33, as well as the concentrations of TSLP and IL-33 in the serum, and the occurrence of pediatric mild asthma. Methods: The analysis encompassed 52 pediatric patients diagnosed with mild bronchial asthma, including both atopic and non-atopic cases, and a control group of 26 non-asthmatic children. Recruitment was conducted through a comprehensive questionnaire. Parameters such as allergic sensitization, serum levels of circulating TSLP and IL-33, and the identification of single-nucleotide polymorphisms in TSLP (rs11466750 and rs2289277) and IL-33 (rs992969 and rs1888909) were assessed for all participants. Results: Significantly lower mean serum TSLP concentrations were observed in asthmatic subjects compared to the control group, with atopic asthma patients showing even lower TSLP levels than non-atopic counterparts. No significant differences were found in mean serum IL-33 concentrations between the two groups. Considering the allele model, for both tested SNPs of IL-33, we observed that patients with asthma, atopic asthma, and atopy statistically less frequently possess the risk allele. Conclusions: Our study findings suggest that IL-33 and TSLP do not serve as ideal biomarkers for mild asthma in children. Their effectiveness as biomarkers might be more relevant for assessing disease severity rather than identifying asthma in pediatric patients. Further research focusing on the association between TSLP and IL-33 gene polymorphisms and asthma is expected to significantly advance disease management.

Amygdalin Improves Allergic Asthma via the Thymic Stromal Lymphopoietin–dendritic Cell–OX40 Ligand Axis in a Mouse Model

Asthma, characterized by persistent inflammation and increased sensitivity of the airway, is the most common chronic condition among children. Novel, safe, and reliable treatment strategies are the focus of current research on pediatric asthma. Amygdalin, mainly present in bitter almonds, has anti-inflammatory and immunoregulatory potential, but its effect on asthma remains uninvestigated. Here, the impact of amygdalin on the thymic stromal lymphopoietin (TSLP)–dendritic cell (DC)–OX40L axis was investigated.
 A BALB/c mouse model for allergic asthma was established using the ovalbumin-sensitization method. Amygdalin treatment was administered between days 21 and 27 of the protocol. Cell numbers and hematoxylin and eosin (H&E) staining in bronchoalveolar lavage fluid (BALF) were used to observe the impact of amygdalin on airway inflammation. TSLP, IL-4, IL-5, IL-13, and IFN-γ concentrations were determined via Enzyme-linked immunosorbent assay (ELISA). TSLP, GATA-3, and T-bet proteins were measured using western blotting. Cell-surface receptor expression on DCs (MHC II, CD80, and CD86) was assessed via flow cytometry. OX40L mRNA and protein levels were detected using western blotting and qRT-PCR, respectively.
 Amygdalin treatment attenuated airway inflammation decreased BALF TSLP levels, inhibited DC maturation, restrained TSLP-induced DC surface marker expression (MHCII, CD80, and CD86), and further decreased OX40L levels in activated DCs. This occurred together with decreased Th2 cytokine levels (IL-4, IL-5, and IL-13) and GATA3 expression, whereas Th1 cytokine (IFN-γ) levels and T-bet expression increased.
 Amygdalin thus regulates the Th1/Th2 balance through the TSLP–DC–OX40L axis to participate in inflammation development in the airways, providing a basis for potential allergic asthma treatments.

Thymic stromal lymphopoietin (TSLP) is a substrate for tryptase in patients with mastocytosis

Mastocytosis is a heterogeneous disease associated to uncontrolled proliferation and increased density of mast cells in different organs. This clonal disorder is related to gain-of-function pathogenic variants of the c-kit gene that encodes for KIT (CD117) expressed on mast cell membrane. Thymic stromal lymphopoietin (TSLP) is a pleiotropic cytokine, which plays a key role in allergic disorders and several cancers. TSLP is a survival and activating factor for human mast cells through the engagement of the TSLP receptor. Activated human mast cells release several preformed mediators, including tryptase. Increased mast cell-derived tryptase is a diagnostic biomarker of mastocytosis. In this study, we found that in these patients serum concentrations of TSLP were lower than healthy donors. There was an inverse correlation between TSLP and tryptase concentrations in mastocytosis. Incubation of human recombinant TSLP with sera from patients with mastocytosis, containing increasing concentrations of tryptase, concentration-dependently decreased TSLP immunoreactivity. Similarly, recombinant β-tryptase reduced the immunoreactivity of recombinant TSLP, inducing the formation of a cleavage product of approximately 10 kDa. Collectively, these results indicate that TSLP is a substrate for human mast cell tryptase and highlight a novel loop involving these mediators in mastocytosis.

Peripheral Multiple Cytokine Profiles Identified CD39 as a Novel Biomarker for Diagnosis and Reflecting Disease Severity in Allergic Rhinitis Patients.

Allergic rhinitis (AR) is a common clinical problem, and immune cells and cytokines were proven to be pivotal in its pathogenesis. Our aim is to measure the peripheral concentrations of multiple cytokines in AR patients and identify novel biomarkers for diagnosis and disease severity. Peripheral blood samples were collected from 50 AR patients, including 25 mild AR (MAR) patients and 25 moderate-severe AR patients (MSAR), and 22 healthy controls (HCs), and multiple cytokine profiling was outlined by Luminex assay. Cytokine levels were compared among the three groups, and their correlations with disease severity were evaluated. The candidate cytokines were further verified by enzyme-linked immunosorbent assay (ELISA) in a validation cohort. Multiple cytokine profiling revealed that CD39 and interferon (IFN)-γ levels were reduced, and interleukin (IL)-13, IL-5, IL-33, and thymic stromal lymphopoietin (TSLP) levels were elevated in the AR group than the HC group (P < 0.05). Receiver operating characteristic (ROC) curves presented that serum CD39 and IL-33 exhibited strong diagnostic abilities, and serum CD39 and IL-10 presented capacities in distinguishing disease severity (AUC > 0.8, P < 0.05). Moreover, CD39 concentrations were decreased, and IL-10, IL-5, and TSLP concentrations were enhanced in the MSAR group more than in the MAR group. Correlation analysis results showed that serum CD39, IL-5, and TSLP levels were associated with total nasal symptom score (TNSS) and visual analogue score (VAS) (P < 0.05). Further data in the validation cohort suggested that serum CD39 levels were reduced, and IL-5 and TSLP levels were increased in AR patients, especially in MSAR patients (P < 0.05). ROC results revealed potential values of serum CD39 in diagnosis and disease severity evaluation in AR patients (P < 0.05). This study highlighted that peripheral multiple cytokine profiles were significantly varied in AR patients and associated with disease severity. The results in discover-validation cohorts implied that serum CD39 might serve as a novel biomarker for diagnosing AR and reflecting its disease severity.

The impact of allergen specific immunotherapy with house dust mite allergens on natural course of asthma in children

Objective . To assess the dynamics of bronchial asthma (BA) and allergic rhinitis (AR) caused by house dust mites (HDM) in children during 3 years of allergen specific immunotherapy (ASIT) with HDM allergens and 1 year after treatment cessation. Research methods . 50 patients completed an open, prospective, controlled 5-year study. 25 patients of the main group with BA and sensitization to HDM underwent a year of preliminary observation, 3 years of ASIT with HDM allergens, and 1 year of follow-up: 16 boys (64%), 9 girls (36%), 5 years to 13 years (8.3 [6.7; 11.5]) at inclusion. The control group of 25 patients, who did not receive ASIT, was selected as pairs copies. At inclusion, the concentration of periostin and thymic stromal lymphopoietin (TSLP) in blood serum and nasal material was determined in all patients. The dynamics BA and AR was assessed based on the total index of symptoms and medications for each year of observation. Results . During 3 years of treatment, a significant clinical effect of ASIT was formed, which persisted for 1 year of follow-up after the end of ASIT. At the end of the follow-up year, the overall index of symptoms and medications was 8.77 ± 1.06 points in the ASIT group and 22.01 ± 2.18 points in the control group (p=0.00001). The concentration of periostin before the start of treatment did not affect the effectiveness and features of the ASIT treatment course. A high serum TSLP concentration (&gt;750 pg/ml) is characteristic of the subgroup with a higher incidence of adverse events associated with ASIT; the final effectiveness of ASIT does not depend on the concentration of this marker. Conclusion . ASIT with HDM sublingual drops is the method of first choice in the treatment of children with BA and AR, in cases where the role of HDM allergens in the genesis of the disease has been proven. A successful ASIT has a prolonged effect (at least 1 year of follow-up), which indicates the possibility of changing the natural course of asthma in children. The study of biomarkers does not allow us to predict the effectiveness of ASIT.

Thymic stromal lymphopoietin in bronchial asthma patients of different age groups: correlation with other markers, lung function results and disease control

Objective: to investigate correlation between thymic stromal lymphopoietin and bronchial asthma course and control in patients of different age groups. Materials and methods: one hundred and four patients were included in 1-year long open prospective study. There were three age groups: children (6 –11 y.o., n=38), adolescents (14–17 y.o., n=35) and adults (25 –50 y.o., n=31). we used asthma duration ≥12 months, uncontrolled asthma and acute respiratory infection absence for ≥14 days as inclusion criteria. Clinical history, validated questionnaires, spirometry, common blood count, serum and nasal material to evaluate thymic stromal lymphopoietin were obtained during first visit. Patient were consequently examined twice with 6 months intervals. Statistical analyses included ANOVA (Kruskal-wallis test) and Pearson’s correlation (r). Differences accepted significant with р&lt;0,05. Resuts: prevalence of main risk factors of asthma control lost (poor compliance, obesity, non-atopic phenotype, fixed airway obstruction) was different in age groups. we didn’t find any thymic stromal lymphopoietin in nasal material. Thymic stromal lymphopoietin concentration correlate significantly with duration of uncontrolled asthma in previous 12 months (r=0,74). we have found greater serum thymic stromal lymphopoietin concentration in patients who demonstrated FEV1 below normal at Visit 3. Conclusion. Serum thymic stromal lymphopoietin level can be used as risk factor of asthma future exacerbation and spirometry results decline.

Abstract 1528: Molecular mechanisms of TSLP as a therapy for CRLF2 B-Cell acute lymphoblastic leukemia

Abstract B-cell acute lymphoblastic leukemia (B-ALL) is the most common type of leukemia in children. B-ALL characterized by cytokine receptor-like factor 2 (CRLF2) overexpression (CRLF2 B-ALL) has a survival rate of &amp;lt;30% and is the highest risk sub-group of B-ALL in both adults and children. CRLF2 is a receptor component for the cytokine thymic stromal lymphopoietin (TSLP). TSLP plays a role in the survival and proliferation of B-cell precursors, thus explaining the oncogenic role of increased CRLF2 signaling in CRLF2 B-ALL. To our surprise, we found that high-levels of TSLP eliminated leukemia cells in patient-derived xenograft (PDX) models of CRLF2 B-ALL. CRLF2 and IL-7 receptor-alpha (IL-7Ra) form the heterodimer type-I cytokine receptor for TSLP cytokine. Binding of TSLP to its CRLF2 receptor complex induces JAK-STAT5 and PI3K-AKT pathway signals. TSLP shares the IL-7Ra with Interleukin 7 (IL-7) which has a heterodimer receptor consisting of IL-7Ra and the common gamma chain. High-levels of IL-7 (50 ng/ml) have been shown to induce IL-7Ra internalization and degradation in T-cells. We hypothesize that high-level TSLP induces internalization and degradation of IL-7Ra leading to CRLF2 signal inhibition, death of CRLF2 B-ALL cells and the anti-leukemia effects that we have observed in PDX mice. To test this hypothesis, we treated CRLF2 B-ALL cell lines with different TSLP concentrations to observe the effect of TSLP on its receptor and CRLF2 signaling. Flow cytometry data showed that continuous or a pulse of high-dose TSLP induced a loss of surface IL-7Ra expression for up to 24 hours. Phosphorylation assays showed that cells cultured with high-dose TSLP were unresponsive to subsequent TSLP-induced phosphorylation events (pSTAT5 and pRPS6), indicating CRLF2 signal inhibition. In conclusion, high-dose TSLP induces loss of (IL-7Ra) and inhibition of CRLF2 signaling. These results suggest that TSLP exerts its anti-leukemia effects by shutting down CRLF2-mediated signals possibly via the loss of the IL-7Ra receptor component. Citation Format: Hossam Alkashgari, Cornelia Stoian, Caleb Ruiz-Jimenez, Jacqueline Coats, Carlos A. Casiano, Sinisa Dovat, Kimberly J. Payne. Molecular mechanisms of TSLP as a therapy for CRLF2 B-Cell acute lymphoblastic leukemia [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1528.

Elevated Plasma Thymic Stromal Lymphopoietin After Acute Myocardial Infarction.

BackgroundThymic stromal lymphopoietin (TSLP), a distant paralog of the cytokine IL-7, has been shown to be associated with atherosclerosis. However, the effect of plasma TSLP level after acute myocardial infarction (AMI) remains largely unclear. Thus, we aimed to assess the relationship between the concentration of TSLP at admission and the risk of major adverse cardiovascular events (MACE) in AMI patients.MethodsA total of 175 patients with AMI and 145 unstable angina (UA) controls were recruited in the present study. The clinical characteristics were collected, and MACE was recorded during hospitalization and the follow-up period after discharge.ResultsThe median value (25, 75 percentiles) of TSLP concentrations in the AMI group was higher than that in the UA group [11.18 (8.14–15.22) vs. 8.56 (5.26–11.94) pg/ml, p < 0.001, respectively]. Multivariate linear regression analysis revealed that Troponin-I (standardized β = 0.183, p = 0.004) was an independent factor for TSLP. According to the median of TSLP concentrations, all the AMI patients were divided into the high-level group (TSLP level ≥ 11.18 pg/ml, N = 91) and the low-level group (TSLP <11.18 pg/ml, N = 84). In a receiver operating characteristic curve analysis, the area under the curve for TSLP as a predictor of AMI was 0.674 with a cut-off value of 9.235 pg/ml. After a median follow-up of 14 months, Kaplan-Meier survival analysis showed no significant difference in MACE-free survival between the two groups (p = 0.648). Finally, the multivariate logistic regression analyses demonstrated that TSLP was a negative predictor of MACE in AMI patients (OR:0.778,95% CI:0.733–0.876, p = 0.032).ConclusionsPlasma TSLP levels were elevated in patients with AMI than those in UA. The lower TSLP concentration was associated with MACE after AMI.

Dependence of thymic stromal lymphopoietin serum concentration from bronchial asthma control level and lung function results in patients of different age groups

Introduction. Bronchial asthma (BA) pathogenesis is settled by bronchial wall inflammation, which is the main treatment target.Aim. To study the dependence of thymic stromal lymphopoietin (TSLP) concentration in serum and nasal brush-biopsies from age, spirometry data deterioration and blood eosinophils in patients with uncontrolled BA.Materials and methods. Patients with uncontrolled BA of three age groups: children (6–11 y.o., n = 38), adolescents (14–17 y.o., n = 35) and adults (25–50 y.o., n = 31) were included in 1-year long open prospective study. Clinical history, АСТ/сАСТ questionnaires, lung function test (LFT), common blood count, serum and nasal material to evaluate TSLP were obtained during first visit, if there were no any signs of acute respiratory infection for not less than 14 days. Maintenance therapy was revised based on firstvisit findings. Patient were consequently examined twice with 6 months intervals, examination data, LFT readings, ACT results and eosinophil counts were collected. Statistical analyses included ANOVA (Kruskal-Wallis test) and Pearson’s correlation (r). Differences accepted significant with р &lt; 0,05.Results. Serum TSLP levels did not differ between age groups. TSLP concentration correlate directly and significantly with duration of uncontrolled BA in previous 12 months (r = 0.74). In patients with atopic BA, maximum TSLP level was in dust mites sensitized group (792.6 ± 114.1 pg/ml). We have found greater serum TSLP concentration in patients who demonstrated FEV1 below normal even on optimal controller therapy at Visit 3. Eosinophil count randomly changed inside normal values and did not correlate with asthma control or TSLP level.Conclusion. Prognosis of future asthma exacerbations and LFT decrease is less favourable in patients with high serum TSLP level.

Effect of halometasone cream combined with Simiao pill on the therapeutic effect and expression of serum leukotriene B4 (LTB4) and thymic stromal lymphopoietin (TSLP) in patients with eczema, and the factors influencing its clinical efficacy.

Chronic eczema has the characteristics of a long treatment cycle and repeated attacks, which seriously affects the daily life and work of patients. Topical glucocorticoids are the first-line treatment for chronic eczema. This study aimed to retrospectively analyze the effects of halometasone cream combined with Simiao pill on the efficacy and expression of serum leukotriene B4 (LTB4) and thymic stromal lymphopoietin (TSLP) in patients with eczema, and identify the factors influencing its clinical efficacy. We retrospectively collected the medical records of 195 patients with chronic eczema treated in the dermatology department from January 2020 to May 2021, and divided them into two groups according to medication: 98 cases were treated with halometasone cream (control group) and 97 cases were treated with halometasone ointment combined with Simiao pill (observation group). The severity of eczema, quality of life, clinical efficacy, LTB4 and TSLP levels, and safety were compared between the two groups. Multivariate logistic regression analysis was used to determine the independent factors affecting clinical efficacy. After treatment, the Eczema area and severity index (EASI) and Dermatology Life Quality Index (DLQI) scores in the observation group were markedly lower than those of the control group (P<0.05). The total clinical effective rate of the observation group was 88.8%, which was notably higher than that of the control group 70.1% (P=0.001). The concentrations of serum LTB4 and TSLP in the observation group were markedly lower than those in the control group (P<0.05). Logistic regression analysis showed that the treatment regimen, digestive system symptoms, heavy aching limbs, and damp-heat tongue and pulse were independent factors affecting the curative effect of the patients (P<0.05). Simiao pill combined with halometasone cream can effectively improve chronic eczema and enhance the clinical efficacy of treatment, which may be related to the reduction of serum LTB4 and TSLP levels. The treatment plan, digestive system symptoms, heavy aching limbs, and damp-heat tongue and pulse are the main factors that affecting the clinical curative effect. Thus, clinical intervention programs should be made according to the above factors to improve the quality of life of patients.

Polymorphisms within the TNFSF4 and MAPKAPK2 Loci Influence the Risk of Developing Invasive Aspergillosis: A Two-Stage Case Control Study in the Context of the aspBIOmics Consortium.

Here, we assessed whether 36 single nucleotide polymorphisms (SNPs) within the TNFSF4 and MAPKAPK2 loci influence the risk of developing invasive aspergillosis (IA). We conducted a two-stage case control study including 911 high-risk patients diagnosed with hematological malignancies that were ascertained through the aspBIOmics consortium. The meta-analysis of the discovery and replication populations revealed that carriers of the TNFSF4rs7526628T/T genotype had a significantly increased risk of developing IA (p = 0.00022). We also found that carriers of the TNFSF4rs7526628T allele showed decreased serum levels of TNFSF14 protein (p = 0.0027), and that their macrophages had a decreased fungicidal activity (p = 0.048). In addition, we observed that each copy of the MAPKAPK2rs12137965G allele increased the risk of IA by 60% (p = 0.0017), whereas each copy of the MAPKAPK2rs17013271T allele was estimated to decrease the risk of developing the disease (p = 0.0029). Mechanistically, we found that carriers of the risk MAPKAPK2rs12137965G allele showed increased numbers of CD38+IgM-IgD- plasmablasts in blood (p = 0.00086), whereas those harboring two copies of the allele had decreased serum concentrations of thymic stromal lymphopoietin (p = 0.00097). Finally, we also found that carriers of the protective MAPKAPK2rs17013271T allele had decreased numbers of CD27-IgM-IgD- B cells (p = 0.00087) and significantly lower numbers of CD14+ and CD14+CD16- cells (p = 0.00018 and 0.00023). Altogether, these results suggest a role of the TNFSF4 and MAPKAPK2 genes in determining IA risk.

Periostin and Thymic Stromal Lymphopoietin-Potential Crosstalk in Obstructive Airway Diseases.

Periostin and thymic stromal lymphopoietin (TSLP) are newly described markers of obstructive airway diseases and the mechanism by which both markers participate in immune response remains poorly understood. The aim of our study was to determine periostin and TSLP concentration in serum and induced sputum (IS) in patients with atopic asthma, chronic obstructive pulmonary disease (COPD), and controls, as well as to evaluate the potential link between periostin, TSLP, and Th2 immune response. Serum and IS levels of periostin, TSLP, IL-4, and IL-13 were determined in 12 atopic asthmatics, 16 COPD sufferers, and 10 controls. We noticed a significantly higher IS periostin and TSLP concentration at protein and mRNA level in asthmatics compared to the two other groups; additionally, periostin and TSLP were correlated positively with IS eosinophil count. A strong positive correlation between IS periostin and TSLP protein levels (r = 0.96) as well as mRNA expression level (r = 0.95) was found in patients with asthma. The results of our study show that periostin and TSLP are associated with eosinophilic airway inflammation and seem to be important drivers of atopic asthma but not COPD pathobiology. Very strong correlations between local periostin, TSLP, eosinophils, and IL-4 in asthma point to the link between periostin–TSLP and Th2 response.

Interleukin 25, thymic stromal lymphopoietin and house dust mites in pathogenesis of atopic dermatitis.

Atopic dermatitis (AD) is the most common chronic inflammatory skin disease in the world. It is characterized by recurrent eczematous skin lesions, fluctuating course and chronic pruritus. Increasing evidence suggest that AD is more common in adults than previously thought. The disease is characterized by an impaired skin barrier, aberrant Th2-type cytokine production and intensive pruritus. Epithelial keratinocytes constitute the first physical, chemical and immunological barrier, classified as a part of the innate defense system. These keratinocytes secrete various factors, e.g. alarmins such as thymic stromal lymphopoietin (TSLP) and interleukin 25 (IL-25). Serum levels of substance P (SP) have been reported to be increased in patients with AD and correlated with itch intensity. Several previous studies reported a positive association between AD severity and house dust mites (HDM) sensitization. The aim of the study was to analyze IL-25, TSLP and SP concentrations in blood serum of adult patients with severe AD, depending on the degree of allergy to HDM. There were 31 adult AD patients enrolled into the study and a control group that consisted of 20 healthy subjects. AD was diagnosed on the basis of Hanifin and Rajka criteria. SCORing Atopic Dermatitis (SCORAD) and visual analogue (VAS) scores were used to assess the intensity of pruritus and blood content of specific IgE to HDM, as well as TSLP, IL-25 cytokines and SP was measured. Our study presents the evidence that IL-25 serum concentration is increased in patients with atopic dermatitis and this cytokine plays an important role in pathogenesis of this disease. HDM could stimulate the release of IL-25 which aggravates the disease severity. Our results corroborate previous findings on the role of TSLP in atopic dermatitis.

Eosinophilic gastroenteritis: epidemiology, diagnosis, and treatment.

Although several reviews concerning diagnosis and treatment of eosinophilic esophagitis (EoE) have been presented, there are few in regard to eosinophilic gastroenteritis (EGE). Fortunately, findings related to epidemiology, as well as diagnosis and treatment of this disease have recently been increasing. The rates of incidence of both EoE and EGE have been reported to be increasing. For accurate diagnosis, plasma concentrations of thymic stromal lymphopoietin and IL-33 may be useful as biomarkers, though consensus has not been reached, while increased eosinophil infiltration in gastrointestinal tissue remains a critical factor. Topical glucocorticoid administration, an elimination diet, and molecular target therapy with neutralizing antibodies are potentially effective therapies that have recently been evaluated. As seen with other allergic diseases, EGE seems to be increasing. Several research projects regarding diagnosis and treatment of the disease are currently in progress.

Effects of Honokiol on Activation of Transient Receptor Potential Channel V1 and Secretion of Thymic Stromal Lymphopoietin in HaCaT Keratinocytes

Abstract Objective: This study was performed to investigate the effects of honokiol on the activation of transient receptor potential channel V1 (TRPV1) and the secretion of thymic stromal lymphopoietin (TSLP) in a human benign epidermal keratinocyte line (HaCaT). Methods: HaCaT keratinocytes were cultivated and divided into six groups: capsaicin-induced model control group, capsazepine control group, solvent control group, and three honokiol treatment groups (7.81, 15.63, and 31.25 mg/L of honokiol). The effect of honokiol on calcium (Ca2+) influx was measured by a Ca2+ fluorescence imaging system. The fluorescence intensity (F) of cells was measured. The rate of change in F (ΔF/F 0) was calculated, and the ΔF/F 0–time curve was constructed. HaCaT keratinocytes were stimulated with polyinosinic:polycytidylic acid, recombinant human tumor necrosis factor α, and recombinant human interleukin 4. Different concentrations of honokiol (15.63, 7.81, and 3.91 mg/L) were added to the cells in the respective honokiol groups; 20 mg/L of dexamethasone or 0.5% dimethyl sulfoxide was added to the cells in the positive control group or solvent control group. The TSLP concentration in the HaCaT keratinocytes of each group was detected by enzyme-linked immunosorbent assay. Statistical analysis was performed by one-way analysis of variance and Dunnett t test. Results: The capsazepine-induced Ca2+ fluorescence intensity in HaCaT keratinocytes was significantly inhibited in the 31.25 mg/L honokiol group; ΔF/F 0 at 45 second was 0.76 in the model control group and 0 in the 31.25 mg/L honokiol group. The TSLP level in the 15.63 and 7.81 mg/L honokiol groups was lower than that in the solvent control group (t = 7.382, P = 0.003, and t = 2.766, P = 0.023, respectively), while the TSLP level in the 3.91 mg/L honokiol group was not significantly different from that in the solvent control group (t = 1.872, P = 0.124). Conclusions: Honokiol inhibited the Ca2+ influx induced by capsazepine (TRPV1 agonist) in HaCaT keratinocytes. Honokiol has an inhibitory effect on TSLP secretion in HaCaT keratinocytes.

Epithelial Alarmins in Serum and Exhaled Breath in Patients with Idiopathic Pulmonary Fibrosis: A Prospective One-Year Follow-Up Cohort Study.

Background: Recently, epithelial alarmins have been shown to play important roles in non-allergen driven respiratory diseases like idiopathic pulmonary fibrosis (IPF). Little is known about the expression of the epithelial alarmins in IPF. Methods: This study aimed to prospectively examine interleukin (IL)-25, IL-33, and thymic stromal lymphopoietin (TSLP) levels in the serum and exhaled breath condensate (EBC) in patients with IPF before and after one-year of antifibrotic treatment. A total of 82 volunteers, including 52 patients diagnosed with IPF that qualified for antifibrotic therapy as well as 30 controls, were examined. All study participants underwent baseline peripheral blood and EBC sampling. In 35 out of 52 IPF subjects, a follow-up sampling was performed after 12 months of antifibrotic treatment. Concentrations of alarmins in the serum and EBC were evaluated by means of ELISA. Results: Baseline TSLP concentrations were significantly elevated in patients with IPF compared to controls both in the serum (p < 0.05) and EBC (p < 0.0001). Baseline IL-25 and IL-33 serum and EBC levels did not differ significantly between IPF subjects and controls. Prospective analysis of changes in the epithelial alarmin levels showed significantly decreased IL-25 and TSLP EBC concentrations after 12 months of antifibrotic treatment (p < 0.05), which was observed in the subgroup of IPF patients treated with pirfenidone, but not in those treated with nintedanib. In stable patients with IPF over a study period (absolute forced vital capacity (FVC) % of predicted decline/year ≤ 5%, n = 25), a significant decrease in the EBC levels of both IL-25 and TSLP after 12 months of antifibrotic treatment was noted (p < 0.05), whereas, in progressor IPF patients (absolute FVC % of predicted decline/year > 5%, n = 10), a significant decrease was noted in the IL-25 EBC levels only (p < 0.05). Conclusions: Elevated TSLP levels in patients with IPF and their significant decrease in the lung compartment during antifibrotic therapy in stable patients with IPF, but not in progressors, support its significant contribution to pro-fibrotic type 2 immune responses in IPF. Noted changes in the epithelial alarmins concentration in the lung compartment during pirfenidone therapy may suggest its possible interaction with epithelial alarmins pathways in IPF.

Thymic stromal lymphopoietin in human pancreatic ductal adenocarcinoma: expression and prognostic significance.

Thymic stromal lymphopoietin (TSLP) has emerged as an important, but contradictory, player conditioning tumor growth. In certain contexts, by driving T helper (h) 2 responses via tumor-associated OX40 Ligand (OX40L)+ dendritic cells (DCs), TSLP may play a pro-tumorigenic role. The study elucidates the importance of TSPL in pancreatic ductal adenocarcinoma (PDAC), by analyzing: i) TSLP levels in PDAC cell-line supernatants and plasma from patients with locally-advanced/metastatic PDAC, pre- and post-treatment with different chemotherapeutic protocols, in comparison with healthy donors; ii) TSLP and OX40L expression in PDAC and normal pancreatic tissues, by immunohistochemistry; iii) OX40L expression on ex vivo-generated normal DCs in the presence of tumor-derived TSLP, by flow cytometry; iv) clinical relevance in terms of diagnostic and prognostic value and influence on treatment modality and response.Some PDAC cell lines, such as BxPC-3, expressed both TSLP mRNA and protein. Normal DCs, generated ex vivo in the presence of TSLP-rich-cell supernatants, displayed increased expression of OX40L, reduced by the addition of a neutralizing anti-TSLP polyclonal antibody. OX40L+ cells were detected in pancreatic tumor inflammatory infiltrates. Abnormally elevated TSLP levels were detected in situ in tumor cells and, systemically, in locally-advanced/metastatic PDAC patients. Of the chemotherapeutic protocols applied, gemcitabine plus oxaliplatin (GEMOX) significantly increased circulating TSLP levels. Elevated plasma TSLP concentration was associated with shorter overall survival and increased risk of poor outcome. Plasma TSLP measurement successfully discriminated PDAC patients from healthy controls.These data show that TSLP secreted by pancreatic cancer cells may directly impact PDAC biology and patient outcome.

Innate immune response reflects disease activity in eosinophilic granulomatosis with polyangiitis.

Eosinophilic granulomatosis with polyangiitis (EGPA) is a disease characterized by allergic granulomatosis, necrotizing vasculitis, and peripheral blood eosinophilia. Interleukin (IL)-33, thymic stromal lymphopoietin (TSLP), and type 2 innate lymphoid cells (ILC2) are involved in the innate and type 2 immune responses in EGPA. However, the relationships among these molecules and the mechanisms underlying the development of EGPA remain unknown. We investigated the relationships among peripheral blood eosinophil count, serum IL-33 and TSLP concentration, and peripheral blood ILC2 count in patients with EGPA, chronic eosinophilic pneumonia (CEP), or bronchial asthma (BA). We recruited 86 patients with EGPA in three groups (remission, relapse, and onset), 25 patients with CEP at active or inactive stages of disease, and 11 patients with BA. In patients with EGPA, CEP, or BA, serum IL-33, sST2, and TSLP concentrations were determined using ELISA and peripheral blood ILC2 counts (as Lin-1- CD127+ CRTH2+ cells) were determined using flow cytometry. Peripheral blood eosinophil count or ILC2 count, and serum sST2 or TSLP concentration were higher in patients with EGPA at onset than in those with EGPA at relapse or remission, or in those with BA or CEP. Serum IL-33 concentration was higher in patients with EGPA at relapse than in those with EGPA at onset or remission, or in those with BA or CEP. In a logistic regression model, EGPA disease activity was correlated with serum IL-33 concentration and peripheral blood ILC2 count, but not daily systemic and inhaled corticosteroid dose or immunosuppressant use. Eosinophil count was correlated with peripheral blood ILC2 count and serum TSLP concentration, but not serum IL-33 concentration. Increased peripheral blood ILC2 count and serum IL-33 concentration were associated with disease activity in EGPA. Increases in serum IL-33 concentration may indicate the presence of active vasculitis rather than peripheral or tissue eosinophilia.

Elevated Expression of IL-33 and TSLP in the Airways of Human Asthmatics In Vivo: A Potential Biomarker of Severe Refractory Disease.

The epithelial cytokines IL-33, thymic stromal lymphopoietin (TSLP), and IL-25 have been implicated in asthma pathogenesis because they promote Th2-type cytokine synthesis, but their expression is relatively poorly documented in "real-life" human asthma. Using bronchoalveolar lavage fluid (BALF), we measured airway concentrations of these mediators and compared them with those of Th1- and Th2-type cytokines, airway infiltration of neutrophils and eosinophils, and lung function in a large group of asthmatic patients with a range of disease severity (n = 70) and control subjects (n = 30). The median BALF concentrations of IL-33, TSLP, IL-4, IL-5, IL-13, and IL-12p70, but not IL-25, IL-2, or IFN-γ, were significantly elevated in asthmatics compared with controls (p < 0.05). The concentrations of IL-33 and TSLP, but not IL-25, correlated inversely with the lung function (forced expiratory volume in the first second) of asthmatics (IL-33: r = -0.488, p < 0.0001; TSLP: r = -0.565, p < 0.0001) independently of corticosteroid therapy. When divided according to disease severity and corticosteroid therapy, all subgroups of asthmatics had elevated median numbers of eosinophils in BALF, whereas the patients with more severe disease who were treated with corticosteroids had higher numbers of neutrophils compared with milder asthmatics not so treated and control subjects (p < 0.05). The data implicate TSLP and IL-33 in the pathogenesis of asthma that is characterized by persistent airway inflammation and impaired lung function despite intensive corticosteroid therapy, highlighting them as potential molecular targets.

Evaluation of murine lung epithelial cells (TC-1 JHU-1) line to develop Th2-promoting cytokines IL-25/IL-33/TSLP and genes Tlr2/Tlr4 in response to Aspergillus fumigatus

ObjectiveThe aims of this study were to determine the role of live and heat-killed Aspergillus fumigatus conidia in releasing interleukin (IL)-25, IL-33 and thymic stromal lymphopoietin (TSLP) and to express Toll-like receptor (Tlr)2 and Tlr4 genes. Materials and methodsMurine lung epithelial cells were incubated with live and heat-killed A. fumigatus conidia at 37°C for 6, 24 and 48h. After treatments, ELISA was performed to measure the concentrations of IL-25, IL-33 and TSLP in the supernatants. Quantitative real-time PCR (qPCR) was performed to assess the expression levels of Tlr2 and Tlr4 genes. ResultsThe concentrations of IL-25 and IL-33 significantly increased after exposure to live and heat-killed conidia for various times when compared with untreated control (P<0.05). The secretion of TSLP at different concentrations of heat-killed conidia was significantly higher than both live conidia and untreated control (P<0.05). qRT-PCR results indicated a up-regulation from 1.08 to 3.60-fold for Tlr2 gene expression and 1.20 to 1.80-fold for Tlr4 gene expression exposed to heat-killed conidia. ConclusionA. fumigatus has a potential ability to stimulate murine lung epithelial cells to produce IL-25/IL-33/TSLP, as well as to express Tlr2/Tlr4 genes, indicating an important role of lung epithelial cells in innate immune responses to A. fumigatus interaction.