Analiza promotorskih varijanti gena TGFB1 kao faktora rizika i modulatora odgovora na terapiju astme

Abstract

Transformišući faktor rasta beta 1 (TGFB1) je važan inflamatorni modulator i profibrotički medijator u astmi. Promene u regulaciji ekspresije TGFB1 bi mogle biti povezane sa patogenezom astme i sa adekvatnim odgovorom na terapiju. Cilj ovog rada je bio ispitivanje promotorskih varijanti gena TGFB1 kao faktora rizika za nastanak astme i modulatora odgovora na terapiju. Studija je uključila pacijente sa astmom i zdrave ispitanike kod kojih je analiziran 5' regulatorni region gena TGFB1. Detektovane su promotorske varijante -509C>T (rs1800469) i -800G>A (rs1800468) za koje asocijativnom studijom nije pokazana povezanost sa patogenezom astme, ali je uočena potencijalna povezanost između promotorske varijante -509C>T i egzacerbacija bolesti. Uticaj ove promotorske varijante na ekspresiju gena TGFB1 pod dejstvom montelukasta, leka za astmu, je ispitivan ex vivo u ćelijama indukovanog sputuma i in vitro u ćelijama BEAS 2B. Analizom ex vivo je pokazana smanjena ekspresija gena TGFB1, pri čemu je efekat bio izraženiji kod astmatičara sa genotipom -509TT (58,9%) u odnosu na astmatičare sa genotipovima -509CC (49,6%) i -509CT (31,8%). U ćelijama BEAS 2B je ekspresija gena bila smanjena za 27%. Nakon transfekcije ćelija BEAS 2B konstruktima sa promotorskim varijantama -509C ili -509T i tretmana montelukastom, zapažena je povećana aktivnost promotora gena TGFB1. Odgovor varijante -509T je bio intenzivniji i za 139,9% veći u odnosu na odgovor varijante -509C. Ova studija je pokazala da promotorska varijanta -509C>T menja ekspresiju gena TGFB1 u odgovoru na montelukast, ali da bi se njena modulatorna uloga razjasnila, u daljim istraživanjima bi trebalo analizirati i druge genetičke i negenetičke faktore.

Transforming growth factor beta 1 (TGFB1) is an important inflammatory modulator and profibrotic mediator in asthma. The changes in regulation of TGFB1 expression could be associated with asthma pathogenesis and adequate response to therapy. The aim of this study was to investigate TGFB1 gene promoter variants as asthma risk factors and modulators of response to asthma therapy. The 5' regulatory region of the TGFB1 gene was analyzed in asthmatic patients and healthy individuals included in this study. The promoter variants -509C>T (rs1800469) and -800G>A (rs1800468) were detected and the association between these variants and asthma pathogenesis was not found. The association was observed between the presence of variant -509C>T and occurence of exacerbations in patients. The influence of -509C>T variant on TGFB1 gene expression in response to montelukast, used for asthma treatment, was further investigated ex vivo in induced sputum cells and in vitro in BEAS 2B cells. Analysis ex vivo has shown the decrease in the TGFB1 gene expression, with more pronaunced effect in asthmatics with genotype -509TT (58.9%) in comparison with -509CC (49.6%) and -509CT (31.8%) genotypes. In BEAS 2B cells the gene expression was reduced for 27%. After transfection of BEAS 2B cells with constructs carrying promoter variants -509C or -509T and treatment with montelukast the increased activity was observed for both promoter variants. The response of variant -509T was more intensive and 139,3% higher than response of variant -509C. This study showed that promoter variant -509T alters TGFB1 gene expression in response to montelukast, but to clarify its modulatory role other genetic, and nongenetic factors as well, should be taken into consideration in further studies.