Novi diagnostični biomarkerji pri anafilaksiji : doktorska disertacija

Vantur Setnikar, Romana (author), Korošec, Peter (mentor), Eržen, Renato (comentor)

Anafilaksija je resna, sistemska akutna alergijska reakcija do katere lahko pride zaradi različnih vzrokov in preko različnih mehanizmov. Določanje serumske triptaze je trenutno najpogosteje uporabljen laboratorijski test, ki ima številne pomanjkljivosti in omejitve. Tekom naše študije smo se osredotočili na potencialne nove diagnostične biomarkerje, ki so bolj kot z mastociti povezani z bazofilci. V našo raziskavo smo skupno vključili 204 anafilaktične bolnike, ki so bili na urgentni oddelek Univerzitetne klinike za pljučne bolezni in alergijo Golnik, pripeljani v časovnem okviru od januarja 2011 do decembra 2018. Bolnikom smo odvzeli parne vzorce: v akutni fazi ob anafilaksiji in v bazalnem stanju (približno 1 mesec po reakciji). Najpogostejši vzrok anafilaktičnih reakcij je bil pik / strup kožekrilca. Največ bolnikov je utrpelo najhujšo, IV stopnjo reakcije po Muellerju. Za primerjavo smo skupno vključili 203 zdrave kontrole. Rezultati so pokazali, da pri anafilaksiji, med izbranimi 11-imi kemokini, pride do signifikantnega povišanja samo pri kemokinskem ligandu z motivom C-C tipa 2 (CCL2), njegove koncentracije in odstotek spremembe glede na njegovo bazalno vrednost, pa so povezane tudi s stopnjo reakcije. CCL2 korelira s triptazo, povišanje CCL2 pa je bilo zabeleženo tudi pri 30 % tistih bolnikov, pri katerih se triptaza ni povišala. Med anafilaksijo pride do signifikantnega znižanja absolutnega števila bazofilcev v krvi ter posledično tudi do znižane relativne genske ekspresije vseh treh bazofilnih markerjev; ?-podenote visokoafinitetnega IgE receptorja Fc?RI (FCER1A), karboksipeptidaze A3 (CPA3) in L-histidin dekarboksilaze (HDC). Signifikantno znižan je tudi kemokinski receptor z motivom C-C tipa 2 (CCR2) na površini bazofilcev in delež CCR2 pozitivnih celic. Ti rezultati potrjujejo hipotezo o potencialni bazofilni migraciji iz krvi na mesta vnetja med anafilaktično reakcijo. In vitro stimulacija polne krvi je pokazala nižje vrednosti receptorja CCR2 na površini bazofilcev in nespremenjene koncentracije CCL2 v plazmi, kar nakazuje, da aktivirani bazofilci bolnikov s predhodno znanimi akutnimi alergijskimi reakcijami, kemokina CCL2 ne proizvajajo de novo in tudi ne izločajo njegovih povišanih koncentracij kot odgovor na ponovno stimulacijo z istim alergenom. Zaključimo lahko, da bazofilno usmerjeni biomarkerji, poleg mastocitne triptaze, predstavljajo nove potencialne (in dodatne) diagnostične biomarkerje anafilaksije.

• Language: Slovenian
• Keywords: anafilaksija, kemokini, CCL2, biomarkerji, triptaza, bazofilci, migracija, kemotaksa, diagnostika, diagnostični testi
• Work type: Doctoral dissertation
• Typology: 2.08 - Doctoral Dissertation
• Organization: BF - Biotechnical Faculty
• Year: 2023
• PID: 20.500.12556/RUL-144896
• COBISS.SI-ID: 146068995
• Publication date in RUL: 22.03.2023

Secondary language

• Language: English
• Title: New diagnostic biomarkers of anaphylaxis : doctoral dissertation

Abstract

Anaphylaxis is a serious, systemic acute allergic reaction that occurs due to various triggers and through a variety of mechanisms. Measurement of serum mast cell tryptase is currently the most commonly used laboratory test, but has many limitations. In our study, we focused on potential new diagnostic biomarkers of anaphylaxis that are more basophil- than mast cell-related. We included a total of 204 anaphylactic patients who were presented to the Emergency Department of University Clinic Golnik in the time frame from January 2011 to December 2018. Paired serum samples were collected: in acute phase during the reaction and later in basal state (approximately 1 month after the reaction). The most common trigger of reactions was Hymenoptera venom and most patients suffered the worst, Mueller's IV grade of reaction. A total of 203 healthy controls were included for comparison. The results showed, that during anaphylaxis, among 11 selected chemokines, there is a significant increase in C-C Motif Chemokine Ligand 2 (CCL2) only, and its concentration and the percentage change from baseline values are both related to the reaction severity. CCL2 strongly correlates with tryptase, and an increase in CCL2 was present also in 30 % of those patients in whom tryptase levels were not elevated. During anaphylaxis, there is a significant decrease in the absolute number of basophils and, consequently, a decrease in relative gene expression of all three basophil markers; the α-subunit of the high-affinity IgE receptor FcεRI (FCER1A), carboxypeptidase A3 (CPA3) and L-histidine decarboxylase (HDC). Additionally, significant decrease is also observed in C-C Motif Chemokine Receptor 2 (CCR2) on the surface of basophils and in the proportion of CCR2 positive cells. These results support the hypothesis of potential basophil migration during an anaphylactic reaction from blood to the site of inflammation. In vitro whole blood stimulations showed lower CCR2 receptors on basophils surface and unchanged concentrations of CCL2 in plasma, suggesting that activated basophils in patients with previously known acute allergic reactions history, do not produce de novo and secrete elevated levels of CCL2 as a response to re-stimulation with the same allergen. We can conclude that basophil-related biomarkers, in addition to mast cell tryptase, represent new potential (and additional) diagnostic biomarkers of anaphylaxis.

• Keywords: anaphylaxis, chemokines, CCL2, biomarkers, tryptase, basophils, migration, chemotaxis, diagnostics, diagnostic tests