Background: An elevated basal serum tryptase level is associated with severe systemic anaphylaxis, most notably caused by Hymenoptera envenomation. Although clonal mast cell disease is the culprit in some individuals, it does not fully explain this clinical association. Objective: Our aim was to determine the prevalence and associated impact of tryptase genotypes on anaphylaxis in humans. Methods: Cohorts with systemic mastocytosis (SM) and venom as well as idiopathic anaphylaxis from referral centers in Italy, Slovenia, and the United States, underwent tryptase genotyping by droplet digital PCR. Associated anaphylaxis severity (Mueller scale) was subsequently examined. Healthy volunteers and controls with nonatopic disease were recruited and tryptase was genotyped by droplet digital PCR and in silico analysis of genome sequence, respectively. The effects of pooled and recombinant human tryptases, protease activated receptor 2 agonist and antagonist peptides, and a tryptase-neutralizing mAb on human umbilical vein endothelial cell permeability were assayed using a Transwell system. Results: Hereditary ?-tryptasemia (H?T)-a genetic trait caused by increased ?- tryptase-encoding Tryptase-?/ß1 (TPSAB1) copy number resulting in elevated BST level-was common in healthy individuals (5.6% [n = 7 of 125]) and controls with nonatopic disease (5.3% [n = 21 of 398]). H?T was associated with grade IV venom anaphylaxis (relative risk = 2.0; P ( .05) and more prevalent in both idiopathic anaphylaxis (n = 8 of 47; [17%; P = .006]) and SM (n = 10 of 82 [12.2%; P = .03]) relative to the controls. Among patients with SM, concomitant H?T was associated with increased risk for systemic anaphylaxis (relative risk = 9.5; P = .007). In vitro, protease-activated receptor-2- dependent vascular permeability was induced by pooled mature tryptases but not ?- or ß-tryptase homotetramers. Conclusions: Risk for severe anaphylaxis in humans is associated with inherited differences in ?-tryptase-encoding copies at TPSAB1.
COBISS.SI-ID: 27755011
Background. Clonal mast cell disorders and elevated BST of unknown cause(s) are associated with severe Hymenoptera venom-triggered anaphylaxis (HVA). However, some individuals with clonal disease have normal BST ((11.4 ng/mL). Objective. To evaluate whether screening for KIT p.D816V in the blood is a useful clinical tool to risk-stratify patients with venom allergy. Methods. We prospectively recruited 374 patients with Hymenoptera allergy and no overt signs of mastocytosis referred to our center in the years 2018-19. KIT p.D816V was determined in the peripheral blood with qPCR and tryptase genotyping was performed by droplet-digital PCR. Results. 351 patients (93.9%) had normal levels of BST and KIT p.D816V was detected in 8% of patients (28/351), predominantly in patients with the most severe Mueller grade IV anaphylaxis (18.2%[24/132] vs 1.8%[4/88 in grade III; 0/131 in other grades] in lower grades; P(0.001). In grade IV patients with normal BST, KIT p.D816V was associated with more severe symptoms including a significantly higher frequency of loss of consciousness (58.3%[14/24] vs 34.3%[37/108]; P=0.03) and absence of skin symptoms (41.7%[10/24] vs 15.7%[17/108]; P=0.004). Among patients with normal BST, KIT p.D816V (OR [95%CI]: 10.25[3.75-36.14]; P(0.0001) was the major risk factor associated with severe HVA. Hereditary [alpha]-tryptasemia (H[alpha]T), due to increased germline copies of TPSAB1 encoding [alpha]-tryptase was the most common cause (65.2%; 15/23) of elevated BST in patients with HVA and together with KIT p.D816V accounted for 90% (20/23) of BST elevations in HVA patients. Conclusion. These results indicate that routine KIT p.D816V screening identifies clonal disease in high-risk HVA patients regularly missed using BST alone.
COBISS.SI-ID: 56665603
Background: The role of chemokines in anaphylaxis is unclear. Methods: We prospectively recruited 49 patients presenting to the emergency department with an acute episode of anaphylaxis and 28 healthy subjects. We measured serum levels of the chemokines CCL2, CCL5, CCL7, CCL8, CCL11, CCL13, CCL17, CCL21, CCL22, CCL24, and CCL26, tryptase, the absolute number of circulating basophils, monocytes, lymphocytes, and PMNs, and whole blood FCER1A, CPA3 and HDC gene expression at two time points: during the anaphylactic episode and in convalescent samples collected approximately 3 months later. We then investigated the in vitro chemotactic activity of chemokines induced during anaphylaxis for the in vitro migration of the corresponding cells. Results: Only CCL2 chemokine levels were significantly increased in anaphylaxis samples (median 514 pg/ml) compared to convalescent samples (284 pg/ml, P ( 0.0001) and healthy subjects (279 pg/ml, P ( 0.0001); there was no significant difference in any of the other chemokines. There was a significant positive correlation between the rates of increase of serum CCL2 (median [range]: 106.0% [- 44.7% to 557.4%]) and tryptase (133.8% [- 6.6% to 893.4%]; r = 0.68, P ( 0.0001) and between the acute concentration of serum CCL2 and the acute concentration of serum tryptase (r = 0.77, P ( 0.0001). The number of circulating basophils, but not other blood cells, significantly decreased during anaphylaxis (median 5.0 vs. 19.1 cells/µl in convalescent samples; P ( 0.0001); a decrease in whole-blood gene expression of basophil markers (P ? 0.0018) confirmed these changes. Anaphylactic serum enhances the in vitro migration of basophils via CCL2-dependent chemotactic activity; in contrast, no CCL2-dependent chemotactic activity was observed for convalescent samples. Conclusions: Our findings imply an important and specific role for CCL2-mediated chemotactic activity in the pathophysiology of human anaphylaxis.
COBISS.SI-ID: 43048963
Asthma is a severe and chronic disabling disease affecting more than 300 million people world-wide. While in the past few drugs for treatment of asthma were available, new treatment options are currently emerging which appear to be highly effective in certain subgroups of patients. Accordingly there is a need for biomarkers which allow selection of patients for refined and personalized treatment strategies. Recently, serological chip tests based on micro-arrayed allergen molecules and peptides derived from the most common rhinovirus strains have been developed which may discriminate two of the most common forms of asthma, i.e., allergen- and virus-triggered asthma. In this perspective we argue that classification of asthma patients according to these common trigger factors may open new possibilities for personalized management of asthma.
COBISS.SI-ID: 2048639089
Background: T cells play a major role in delayed-type hypersensitivity reactions. Their reactivity can be assessed by measuring the upregulation of the activation marker CD69, followed by proliferation and cytokine production. The aim of our study was to develop a novel, whole blood-based, quantitative, absolute count activation index (AI) analysis of CD69 upregulation on different subsets of T cells in nickel hypersensitive patients and compare it with the previously reported approaches. Methods: Ten patients with nickel allergy and nine healthy controls were included. CD69 expression of CD3+, CD3+CD4+ and CD3+CD8+ T cells in heparinized blood was determined with flow cytometry after incubation with nickel sulfate for 48 h. The absolute cell count of CD69+ cells was determined with microbeads. The production of the cytokines IL-2, IL-5, IL-13, and IFN- [gamma] was determined after nickel sulfate stimulation of PBMNCs for 48 h. Results: We showed that the most sensitive methodology is the absolute AI, which was calculated as the ratio between the absolute count of CD69-positive T cells stimulated with nickel and the absolute count of CD69-positive T cells in non-stimulated blood. This novel quantitative approach was more discriminative than the previously reported approaches in which T cell CD69 percentage AI and cytokine production are measured. Conclusions: Our results demonstrated that measuring the absolute CD69 AI is an accurate new approach to quantify antigen-specific...
COBISS.SI-ID: 2048400753