Autoantibodies against desmin and transglutaminase 2 in celiac disease: diagnostic and functional significance

Abstract

Tsöliaakia on päriliku eelsoodumusega krooniline peensoolehaigus, mille vallandajaks on nisus leiduvad gluteeni valgud ning sarnased valgud rukkis ja odras. Haigust esineb kuni 1% Euroopa elanikkonnast ja selle klassikalisteks sümptomiteks on toitainete imendumishäiretest tingitud vaevused. Haiguse patogeneesis on kesksel kohal immuunmehhanismid, mis tekitavad peensoole limaskestas põletiku ja koekahjustuse. Lisaks immuunvastusele gluteeni vastu, tekivad tsöliaakiahaigetel autoantikehad koe enda valkude suhtes. Transglutaminaas 2 (TG2) on tsöliaakia peamine autoantigeen, mille vastased IgA tüüpi antikehad vereseerumis on tundlikuks haigusmarkeriks. Kuna tsöliaakia diagnoosimiseks kasutatakse invasiivset peensoolebiopsia protseduuri, on haigete sõeluuringuks vajalik kõrge diagnostilise võimekusega seroloogiliste testide olemasolu. TG2 on multifunktsionaalne valk, mis osaleb mitmetes bioloogilistes protsessides, seega võib TG2 autoantikehadel olla oluline roll ka tsöliaakia immunopatogeneesis.

Käesolevas töös leidsime, et lisaks senikirjeldatud autoantikehadele leidub enam kui pooltel tsöliaakiahaigetel IgA tüüpi autoantikehi tsütoskeleti valgu desmiini vastu ning autoantikehade tase langeb gluteenivaba dieedi toimel. Desmiinivastased antikehad võivad omada tähendust täiendava koekahjustuse markerina tsöliaakia korral. Järgnevalt töötasime välja uudse testi TG2 autoantikehade määramiseks, kus kasutatakse TG2 seondumist tema loomuliku ligandi fibronektiini külge. Modifitseeritud testi diagnostiline tundlikkus (95%) ja spetsiifilisus (99%) tsöliaakiahaigetel lastel oli parem kui seda võimaldas traditsiooniline meetod. Edasi uurisime, kas töliaakiahaigete autoantikehad mõjutavad TG2 bioloogilist funktsiooni. Leidsime, et autoantikehad pidurdavad TG2 seondumist hepariini ja heparaansulfaadiga ning mõjutavad rakkude kinnitumisvõimet. Kaardistasime ka TG2 hepariinseonduvad alad, kasutades seondumis- ja funktsionaalsetes katsetes sünteetilisi peptiide. Tulemuste alusel püstitasime uudse hüpoteesi, et tsöliaakiahaigete autoantikehad, seondudes rakuvälise TG2-ga, nõrgendavad soole epiteelrakkude kontakte rakuvälise maatriksiga ning võivad seeläbi osaleda tsöliaakia koekahjustuse tekkel.

Celiac disease is a chronic disease of the small intestine, which is triggered in genetically susceptible individuals by gluten proteins of the wheat and similar proteins in rye and barley. It is a widespread condition affecting up to 1% of the western population and displaying various clinical symptoms related to the malabsorption of nutrients. Immune mechanisms are central to the pathogenesis of celiac disease, causing inflammation and tissue damage in the small intestinal mucosa. Besides the immune response against gluten proteins, autoantibodies develop against self-proteins in the patients. Autoantibodies against transglutaminase 2 (TG2) are the hallmark of celiac disease and are widely used as sensitive biomarkers. Because invasive intestinal biopsy procedure is required to diagnose celiac disease, the availability of accurate antibody assays is important for the efficient selection of the patients for biopsy. While TG2 is a multifunctional protein involved in various biological processes, autoantibodies against TG2 could also have a role in the pathogenesis of celiac disease.

In this study we detected IgA class autoantibodies against cytoskeletal protein desmin in blood sera of more than half of celiac disease patients and also observed that autoantibody levels decreased in response to a gluten free diet. Desmin autoantibodies could have a potential as an additional marker for intestinal tissue damage in celiac disease. In the following, we developed a novel test for detection of TG2 autoantibodies by using TG2 binding to its natural ligand fibronectin. The diagnostic sensitivity (95%) and specificity (99%) of the modified test for childhood celiac disease was higher when compared with traditional method. Next we asked, whether autoantibodies could interfere with the biological functions of TG2. We found that autoantibodies from celiac disease patients affected cell adhesion by inhibiting TG2 binding to heparin and heparan sulphate molecules. We also delineated heparin-binding sites of TG2 by using synthetic peptides in binding and functional studies. Based on our results, we draw a hypothesis, that celiac disease autoantibodies weaken the adhesion contacts of intestinal epithelial cells by binding to the extracellular TG2, and therefore, could contribute to the tissue lesion in celiac disease.