RT Journal Article
SR Electronic
T1 Widely used commercial ELISA for human Zonulin reacts with Complement C3 rather than preHaptoglobin2
JF bioRxiv
FD Cold Spring Harbor Laboratory
SP 157578
DO 10.1101/157578
A1 Lucas Scheffler
A1 Alyce Crane
A1 Henrike Heyne
A1 Anke Tönjes
A1 Dorit Schleinitz
A1 Christian H. Ihling
A1 Michael Stumvoll
A1 Peter Kovacs
A1 John T. Heiker
YR 2017
UL http://biorxiv.org/content/early/2017/06/30/157578.abstract
AB BACKGROUND There is increasing evidence for the role of impaired intestinal permeability in obesity and associated metabolic diseases. Zonulin is an established serum marker for intestinal permeability and supposedly identical to pre-haptoglobin2. Here, we aimed to investigate the relationship between intestinal permeability represented by circulating zonulin and metabolic traits related to obesity.METHODS Serum zonulin was measured by using a widely used commercially ELISA kit in 376 subjects from the metabolically well-characterized cohort of Sorbs from Germany. In addition, haptoglobin genotype was determined in DNA samples from all study subjects.RESULTS The genotype frequencies for haptoglobin genotypes HP1/1, HP1/2 and HP2/2 were 15.8%, 47.6% and 36.6% respectively. Since zonulin concentrations did not correspond to the haptoglobin genotypes we investigated the specificity of the zonulin ELISA assay using immunoprecipitation experiments, mass spectrometry and Western blot analysis. Thus we demonstrated by mass spectrometry that the kit is mainly capturing complement factor C3 and derived fragments. Circulating complement factor C3 was significantly increased in patients with diabetes and obesity and correlated strongly with markers of the lipid and glucose metabolism.CONCLUSIONS Our study supports the role of the complement C3 in the pathophysiology of obesity and necessitates critical revision of published data on zonulin as a marker for altered intestinal permeability from studies that have used the ELISA kit investigated here.