The immunomodulatory effect of extracellular vesicles from L1 larvae of Trichinella spiralis on a respiratory allergy model in BALB/C mice

[S. Glamočlija Jekić]

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Biologija - Imunobiologija / Biology - Immunobiology

Respiratorne alergije predstavljaju rastući zdravstveni problem i posledica su preteranog imunskog odgovora na bezopasne supstance iz spoljne sredine. Istraživanja su pokazala da helminti poput Trichinelle spiralis posredstvom ekskretorno sekretornih produkata larvi (ES L1) mogu da usmere imunski odgovor ka anti-inflamatornom i regulatornom i tako ublaže alergijom indukovanu inflamaciju. Nedavno je pokazano da ES L1 produkti sadrže ekstracelularne vezikule (EV), membranom ograničene pakete koje u ekstracelularni prostor oslobađaju sve vrste ćelija, zbog čega se definišu kao jedan od fundamentalnih mehanizama međućelijske komunikacije. Cilj ove studije je da se ispitaju morfološke karakteristike i imunomodulatorna svojstva EV T. spiralis (TsEV) u in vitro uslovima na humanim ćelijama i u in vivo uslovima na ekspreimentalnom modelu respiratornih alergija. Rezultati glikokarakterizacije TsEV su potvrdili prisustvo specifičnih gliko-epitopa koji mogu da aktiviraju receptore prisutne na dendritskim ćelijama (DC). U in vitro uslovima TsEV su stimulisale diferencijaciju DC tolerogenog fenotipa koji se karakteriše ekspresijom gena uključenih u sazrevanje DC kao i niskom ekspresijom površinskih kostimulatornih molekula a povišenom ekspresijom tolerogenih markera, IDO-1 i ILT3. DC kultivisane u prisustvu TsEV su povećano produkovale IL-10 i TGF-β, regulatornih citokina, dok su u ko-kuluturi sa alogenim T limfocitima stimulisale razvoj Th2, Tr1 i inducibilnih regulatornih FoxP3+ T ćelija. Opisana tolerogena svojstva DC ostala su očuvana čak i nakon izlaganja inflamatornom stimulusu. U cilju ispitivanja terapijskog potencijala TsEV in vivo uslovima sprovedeno je istraživanje na BALB/c miševima sa respiratornom alergijom izazvanom ovalbuminom (OVA). Tretman ovih miševa je podrazumevao intranazalnu primenu TsEV i to u vidu ko-aplikacione primene (davanje TsEV u toku indukcije alergije) i vidu terapijske primene (davanje TsEV nakon indukcije alergije i pre ponovnog susreta sa OVA). Intranazalna primena TsEV značajno je smanjila prisustvo makrofaga, neutrofila i eozinofila i smanjila zastupljenost mukusa u disajnim putevima. Terapijski pristup davanja TsEV doveo je do smanjenja Th2 citokina (IL-4, IL-5, IL-13) i IgE specifičnih za OVA, i uticao na povećanu produkciju IL-10 na lokalnom ali i na sistemskom nivou. Pored toga, zabeležena je ekspanzija regulatornih T ćelija, FoxP3⁺ i Tr1, u plućima i slezini, što je dodatno doprinelo uspostavljanju anti-inflamatornog profila. Ova studija je po prvi put pokazala da TsEV ispoljavaju imunomodulatorni potencijal koji se može iskoristiti za snižavanje inflamacije u respiratornim alergijama te bi se mogle koristiti kao osnova za razvoj inovativnih platformi za tretman respiratornih alergija.

Respiratory allergies represent a growing health concern, stemming from an exaggerated immune response to otherwise harmless environmental substances. Research has shown that helminths such as Trichinella spiralis can direct immune responses toward the anti-inflammatory and regulatory profile through their excretory-secretory (ES L1) products, thus reducing allergic inflammation. Recently, it has been demonstrated that ES L1 products contain extracellular vesicles (EV), membrane-bound packages released by all cell types into the extracellular space, hence considered fundamental mediators of intercellular communication. The aim of this study was to investigate the morphological characteristics and immunomodulatory properties of T. spiralis-derived EV (TsEV) under in vitro conditions using human cells, as well as in vivo using an experimental model of respiratory allergy. Glycan characterization of TsEV confirmed the presence of specific glycoepitopes which are capable of activating receptors expressed on dendritic cells (DCs). In vitro experiments showed that TsEV induced the development of a tolerogenic DC phenotype, as evidenced by the expression of genes involved in DC maturation, low expression of costimulatory surface molecules, and increased expression of tolerogenic markers IDO-1 and ILT3. TsEV-treated DCs exhibited increased production of IL-10 and TGF-β cytokines and promoted the differentiation of Th2 cells, FoxP3⁺ inducible regulatory T cells, and Tr1 cells when cocultured with allogeneic T lymphocytes. Importantly, these tolerogenic properties of DCs were preserved even after exposure to an inflammatory stimulus. To evaluate the therapeutic potential of TsEV in vivo, experiments using BALB/c mice with experimental ovalbumin (OVA)-induced respiratory allergy were conducted. Intranasal TsEV treatment was assesed in an co-application model (TsEV applied during allergy induction) as well as in a therapy model (TsEV applied after allergy induction but prior to allergen re-challenge). Intranasal application of TsEV significantly reduced the presence of macrophages, neutrophils, and eosinophils, as well as mucus accumulation in the airways. The therapeutic approach led to decreased levels of Th2 cytokines (IL-4, IL-5, IL-13) and OVA-specific IgE, alongside increased IL-10 production at both local and systemic levels. Furthermore, an expansion of regulatory T cells, including FoxP3⁺ and Tr1 subsets, was observed in the lungs and spleen, contributing to the establishment of the anti-inflammatory profile. This study is the first to demonstrate that TsEV possess immunomodulatory properties capable of reducing inflammation in respiratory allergies, suggesting their potential to serve as a foundation for the development of innovative therapeutic platforms aimed at treating respiratory allergies.

Trichinella spiralis, ES L1, ekstracelularne vezikule, TsEV, respiratorne alergije, dendritske ćelije, regulatorni T limfociti

Trichinella spiralis, ES L1, extracellular vesicles, TsEV, respiratory allergies, dendritic cells, regulatory T lymphocytes

577.27:616-097(043.3)

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