A Gly15Arg mutation in the interleukin-10 gene reduces secretion of interleukin-10 in Crohn disease.
BACKGROUND
Genetic susceptibility, probably involving cytokines and their receptors, plays an important role in inflammatory bowel disease (IBD). In this study we examine the potential role of the interleukin-10 (IL-10) gene as a susceptibility gene in IBD.


METHODS
We studied 17 sib-pairs with either Crohn disease (CD) or ulcerative colitis. After microsatellite analysis for allele-sharing, the IL-10 gene of sib-pairs who shared alleles was screened for nucleotide alterations in and around exons and the promoter region. The IL-10 promoter polymorphism at position -1082 was also determined. Function was evaluated by measuring IL-10 secretion by peripheral blood mononuclear cells stimulated with lipopolysaccharide or phorbol ester. The activity of recombinant immature wild-type and mutated IL-10 was tested in a proliferation assay with a human monocytic leukaemia cell line (HL60 cells).


RESULTS
DNA sequencing revealed a G --> A point mutation in exon 1 at base position 43 in one sib-pair, both affected with CD. It was also found in 2 of their healthy siblings, but not in 75 unrelated healthy controls. This mutation results in a glycine to arginine substitution at amino acid position 15 of the leader sequence (Gly15Arg). The in vitro IL-10 secretion by mononuclear cells of the IL-10 Gly15Arg carriers was about 50% of healthy controls, matched for the -1082 polymorphism in the IL-10 promoter region. Incubation of HL60 cells with recombinant mutated IL-10 showed a markedly reduced cell proliferation compared to wild-type IL-10.


CONCLUSIONS
A Gly15Arg mutation in the leader sequence of IL-10 was found in a multiple CD-affected family. This altered leader sequence decreases IL-10 secretion, thereby reducing the anti-inflammatory effect.
van der Linde
K
K
Dept. of Gastroenterology and Hepatology, Erasmus Medical Center Rotterdam, Rotterdam, The Netherlands.
Boor
P P C
PP
Sandkuijl
L A
LA
Meijssen
M A C
MA
Savelkoul
H F J
HF
Wilson
J H P
JH
de Rooij
F W M
FW
D000328:Adult; D001483:Base Sequence / Q000235:genetics; D003424:Crohn Disease / Q000235:genetics* / Q000276:immunology*; D005260:Female; D020022:Genetic Predisposition to Disease / Q000235:genetics; D006801:Humans; D016753:Interleukin-10 / Q000096:biosynthesis* / Q000235:genetics*; D007963:Leukocytes, Mononuclear / Q000276:immunology; D008297:Male; D018895:Microsatellite Repeats / Q000235:genetics; D008875:Middle Aged; D010375:Pedigree; D017354:Point Mutation
D016753:Interleukin-10