Innate immunity genes influence the severity of acute appendicitis.

OBJECTIVE: Using acute appendicitis as a model, we tested the hypothesis that polymorphisms in genes involved in host defense can be associated with the severity of local infection-inflammation in humans. SUMMARY BACKGROUND DATA: Innate immunity is the body's front-line system for antimicrobial host defense. Local inflammation is a major innate immune mechanism for containing and destroying microbes, but it may also contribute to tissue injury. METHODS: We studied 134 patients with acute appendicitis treated at an urban hospital. We looked for associations between the severity of appendicitis (uncomplicated vs. perforated or gangrenous), plasma and peritoneal cytokine concentrations, and single nucleotide polymorphisms in genes involved in recognizing bacterial molecules [CD14 (-159 C-->T); TLR4 (896 A-->G)] and in mounting an inflammatory response [IL-6 (-174 G-->C), TNF-alpha (-308 G-->A), IL-1beta (-31 C -->T)]. RESULTS: Ninety-one patients (68%) had uncomplicated appendicitis and 43 (32%) had complicated disease. The SNPs in the CD14, TLR4, IL-1beta, and TNF-alpha genes were not associated with the severity of appendicitis. A strong association was found between C-allele carriage at -174 in the IL-6 gene and decreased risk of complicated disease (adjusted odds ratio = 0.24, 95% CI = 0.07-0.76). Lower plasma and peritoneal fluid IL-6 concentrations in the IL-6 -174 C-carriers than in the GG homozygotes suggest that this polymorphism contributes to decreased IL-6 production in vivo. CONCLUSIONS: Polymorphism in the IL-6 gene was associated with the severity of appendicitis, even after adjustment for duration of symptoms. The risk for developing appendiceal perforation or gangrene may be determined, in part, by variation in the IL-6 gene.

Interleukin-6 promoter haplotypes and interleukin-6 cytokine responses.

Genetic variations contribute to differences in the inflammatory response and are markers for disease risk and outcomes. We studied three single-nucleotide polymorphisms (-597 G-->A, -572 G-->C, and -174 G-->C) in the IL-6 promoter to determine associations with ex vivo LPS-stimulated IL-6 production by leukocytes. We also measured nuclear protein binding to synthetic oligonucleotides representing the -174 polymorphic region, located in proximity to important transcription factor motifs. We determined genotypes at three sites in the IL-6 promoter by pyrosequencing of genomic DNA obtained from 49 healthy control subjects. To determine molecular haplotypes, cloned DNA fragments from heterozygous subjects were sequenced. IL-6 release by whole blood leukocytes was measured after 24 h of ex vivo LPS stimulation. We compared IL-6 concentrations between haplotypes using Kruskall-Wallis and adjusted for covariates by analysis of covariance. Electromobility gel shift assay was carried out by the incubation of nuclear proteins from cultured human mononuclear cells with oligonucleotides representing the alternate -174 alleles. The amount of nuclear protein binding was quantified by densitometry, which was compared using analysis of variance. Genotype and sequence analysis of genomic and cloned DNA characterized three haplotypes. Ex vivo IL-6 production was greatest in individuals who were homozygous for the haplotype containing guanine at -597 and -174. IL-6 production was least for individuals homozygous for the haplotype containing adenine at -597 and cytosine at -174. Nuclear protein bound more avidly to guanine-containing oligonucleotides representing the -174 position than to oligonucleotides containing cytosine at that position. The IL-6 promoter haplotype influences ex vivo IL-6 response to endotoxin. This effect may be due to a functional effect of the -174 G-->C polymorphism.