Barbara Noack

Identification of a shared genetic susceptibility locus for coronary heart disease and periodontitis.

Recent studies indicate a mutual epidemiological relationship between coronary heart disease (CHD) and periodontitis. Both diseases are associated with similar risk factors and are characterized by a chronic inflammatory process. In a candidate-gene association study, we identify an association of a genetic susceptibility locus shared by both diseases. We confirm the known association of two neighboring linkage disequilibrium regions on human chromosome 9p21.3 with CHD and show the additional strong association of these loci with the risk of aggressive periodontitis. For the lead SNP of the main associated linkage disequilibrium region, rs1333048, the odds ratio of the autosomal-recessive mode of inheritance is 1.99 (95% confidence interval 1.33–2.94; P = 6.9×10−4) for generalized aggressive periodontitis, and 1.72 (1.06–2.76; P = 2.6×10−2) for localized aggressive periodontitis. The two associated linkage disequilibrium regions map to the sequence of the large antisense noncoding RNA ANRIL, which partly overlaps regulatory and coding sequences of CDKN2A/CDKN2B. A closely located diabetes-associated variant was independent of the CHD and periodontitis risk haplotypes. Our study demonstrates that CHD and periodontitis are genetically related by at least one susceptibility locus, which is possibly involved in ANRIL activity and independent of diabetes associated risk variants within this region. Elucidation of the interplay of ANRIL transcript variants and their involvement in increased susceptibility to the interactive diseases CHD and periodontitis promises new insight into the underlying shared pathogenic mechanisms of these complex common diseases.

A 3′ UTR transition within DEFB1 is associated with chronic and aggressive periodontitis

Periodontal diseases are complex inflammatory diseases and affect up to 20% of the worldwide population. An unbalanced reaction of the immune system toward microbial pathogens is considered as the key factor in the development of periodontitis. Defensins have a strong antimicrobial function and are important contributors of the immune system toward maintaining health. Here, we present the first systematic association study of DEFB1. Using a haplotype-tagging single nucleotide polymorphism (SNP) approach, including described promoter SNPs of DEFB1, we investigated the associations of the selected variants in a large population (N=1337 cases and 2887 ethnically matched controls). The 3′ untranslated region SNP, rs1047031, showed the most significant association signal for homozygous carriers of the rare A allele (P=0.002) with an increased genetic risk of 1.3 (95% confidence interval: 1.11–1.57). The association was consistent with the specific periodontitis forms: chronic periodontitis (odds ratio=2.2 (95% confidence interval: 1.16–4.35), P=0.02), and aggressive periodontitis (odds ratio=1.3 (95% confidence interval 1.04–1.68), P=0.02). Sequencing of regulatory and exonic regions of DEFB1 identified no other associated variant, pointing toward rs1047031 as likely being the causative variant. Prediction of microRNA targets identified a potential microRNA-binding site at the position of rs1047031.

Validation of reported genetic risk factors for periodontitis in a large-scale replication study.

AIM Many studies investigated the role of genetic variants in periodontitis, but few were established as risk factors. We aimed to validate the associations of recent candidate genes in aggressive periodontitis (AgP). MATERIAL AND METHODS We analysed 23 genes in 600 German AgP patients and 1441 controls on the Illumina custom genotyping array Immunochip. We tested a suggestive association in a Dutch and German/Austrian AgP case-control sample, and a German chronic periodontitis (CP) case-control sample using Sequenom iPlex assays. We additionally tested the common known risk variant rs1333048 of the gene ANRIL for its association in a Turkish and Italian population. RESULTS None of the analysed genes gave statistical evidence for association. Upon covariate adjustment for smoking and gender, in the pooled German-Austrian AgP sample, IL10 SNP rs6667202 was associated with p = 0.016, OR = 0.77 (95% CI = 0.6-0.95), and in the Dutch AgP sample, adjacent IL10 SNP rs61815643 was associated with p = 0.0009, OR = 2.31 (95% CI = 1.4-3.8). At rs61815643, binding of the transcription factor PPARG was predicted. ANRIL rs1333048 was associated in the Turkish sample (pallelic = 0.026, OR = 1.67 [95% CI = 1.11-2.60]). CONCLUSIONS Previous candidate genes carry no susceptibility factors for AgP. Association of IL-10 rs61815643 with AgP is suggested. ANRIL is associated with periodontitis across different populations.

Genetic evidence for PLASMINOGEN as a shared genetic risk factor of coronary artery disease and periodontitis

Background—Genetic studies demonstrated the presence of risk alleles in the genes ANRIL and CAMTA1/VAMP3 that are shared between coronary artery disease (CAD) and periodontitis. We aimed to identify further shared genetic risk factors to better understand conjoint disease mechanisms. Methods and Results—In-depth genotyping of 46 published CAD risk loci of genome-wide significance in the worldwide largest case–control sample of the severe early-onset phenotype aggressive periodontitis (AgP) with the Illumina Immunochip (600 German AgP cases, 1448 controls) and the Affymetrix 500K array set (283 German AgP cases and 972 controls) highlighted ANRIL as the major risk gene and revealed further associations with AgP for the gene PLASMINOGEN (PLG; rs4252120: P=5.9×10−5; odds ratio, 1.27; 95% confidence interval, 1.3–1.4 [adjusted for smoking and sex]; 818 cases; 5309 controls). Subsequent combined analyses of several genome-wide data sets of CAD and AgP suggested TGFBRAP1 to be associated with AgP (rs2679895: P=0.0016; odds ratio, 1.27 [95% confidence interval, 1.1–1.5]; 703 cases; 2.143 controls) and CAD (P=0.0003; odds ratio, 0.84 [95% confidence interval, 0.8–0.9]; n=4117 cases; 5824 controls). The study further provides evidence that in addition to PLG, the currently known shared susceptibility loci of CAD and periodontitis, ANRIL and CAMTA1/VAMP3, are subjected to transforming growth factor-&bgr; regulation. Conclusions—PLG is the third replicated shared genetic risk factor of atherosclerosis and periodontitis. All known shared risk genes of CAD and periodontitis are members of transforming growth factor-&bgr; signaling.

Replication of the association of chromosomal region 9p21.3 with generalized aggressive periodontitis (gAgP) using an independent case-control cohort

BackgroundThe human chromosomal region 9p21.3 has been shown to be strongly associated with Coronary Heart Disease (CHD) in several Genome-wide Association Studies (GWAS). Recently, this region has also been shown to be associated with Aggressive Periodontitis (AgP), strengthening the hypothesis that the established epidemiological association between periodontitis and CHD is caused by a shared genetic background, in addition to common environmental and behavioural risk factors. However, the size of the analyzed cohorts in this primary analysis was small compared to other association studies on complex diseases. Using our own AgP cohort, we attempted to confirm the described associations for the chromosomal region 9p21.3.MethodsWe analyzed our cohort consisting of patients suffering from the most severe form of AgP, generalized AgP (gAgP) (n = 130) and appropriate periodontally healthy control individuals (n = 339) by genotyping four tagging SNPs (rs2891168, rs1333042, rs1333048 and rs496892), located in the chromosomal region 9p21.3, that have been associated with AgP.ResultsThe results confirmed significant associations between three of the four SNPs and gAgP. The combination of our results with those from the study which described this association for the first time in a meta-analysis of the four tagging SNPs produced clearly lower p-values compared with the results of each individual study. According to these results, the most plausible genetic model for the association of all four tested SNPs with gAgP seems to be the multiplicative one.ConclusionWe positively replicated the finding of an association between the chromosomal region 9p21.3 and gAgP. This result strengthens support for the hypothesis that shared susceptibility genes within this chromosomal locus might be involved in the pathogenesis of both CHD and gAgP.

CDKN2BAS is associated with periodontitis in different European populations and is activated by bacterial infection

Epidemiological studies have indicated a relationship between coronary heart disease (CHD) and periodontitis. Recently, CDKN2BAS was reported as a shared genetic risk factor of CHD and aggressive periodontitis (AgP), but the causative variant has remained unknown. To identify and validate risk variants in different European populations, we first explored 150 kb of the genetic region of CDKN2BAS including the adjacent genes CDKN2A and CDKN2B, covering 51 tagging single nucleotide polymorphisms (tagSNPs) in AgP and chronic periodontitis (CP) in individuals of Dutch origin (n=313). In a second step, we tested the significant SNP associations in an independent AgP and CP population of German origin (n=1264). For the tagSNPs rs1360590, rs3217992, and rs518394, we could validate the associations with AgP before and after adjustment for the covariates smoking, gender and diabetes, with SNP rs3217992 being the most significant (OR 1.48, 95% CI 1.19 to 1.85; p=0.0004). We further showed in vivo gene expression of CDKN2BAS, CDKN2A, CDKN2B, and CDK4 in healthy and inflamed gingival epithelium (GE) and connective tissue (CT), and detected a significantly higher expression of CDKN2BAS in healthy CT compared to GE (p=0.004). After 24 h of stimulation with Porphyromonas gingivalis in Streptococcus gordonii pre-treated gingival fibroblast (HGF) and cultured gingival epithelial cells (GECs), we observed a 25-fold and fourfold increase of CDKN2BAS gene expression in HGFs (p=0.003) and GECs (p=0.004), respectively. Considering the global importance of CDKN2BAS in the disease risk of CHD, this observation supports the theory of inflammatory components in the disease physiology of CHD.

Novel Mutations in the Cathepsin C Gene in Patients with Pre-pubertal Aggressive Periodontitis and Papillon-Lefèvre Syndrome

Aggressive periodontitis (AP) in pre-pubertal children is often associated with genetic disorders like Papillon-Lefèvre syndrome (PLS). PLS is caused by mutations in the cathepsin C (CTSC) gene. We report a novel CTSC mutation (c.566-572del) in an otherwise healthy AP child and two novel compound heterozygous mutations (c.947T>G, c.1268G>C) in a PLS patient. We conclude that at least a subset of pre-pubertal AP is due to CTSC mutations and therefore may be an allelic variant of PLS.

COX-2 Is Associated with Periodontitis in Europeans

COX-2 plays an important role in periodontitis by mediating inflammatory reactions in periodontal tissues, and the COX-2 polymorphisms rs20417 and rs689466 have been reported to be associated with periodontitis in populations of Taiwanese and Chinese ethnicity. To test whether these variants were associated with periodontitis in populations of European ethnicity, we genotyped the single-nucleotide polymorphisms (SNPs) rs689466 and rs6681231, the latter a haplotype tagging SNP (htSNP) for rs20417 (r2>0.95), in our large-analysis population of individuals with aggressive (n = 532) and chronic periodontitis (n = 1052), and 2873 healthy control individuals. The rare G allele of htSNP rs6681231 was associated with aggressive periodontitis prior to and after adjustment for the covariates smoking, diabetes, and gender, with an odds ratio of 1.57 (95% confidence interval 1.18–2.08; p = 0.002). The validation of the association of rs20417 by the htSNP rs6681231 provides evidence for a general genetic risk of COX-2 variants in the pathogenesis of periodontitis.

Functional Cathepsin C mutations cause different Papillon–Lefèvre syndrome phenotypes

AIM The autosomal-recessive Papillon-Lefèvre syndrome (PLS) is characterized by severe aggressive periodontitis, combined with palmoplantar hyperkeratosis, and is caused by mutations in the Cathepsin C (CTSC) gene. This study aimed to identify CTSC mutations in different PLS phenotypes, including atypical forms and isolated pre-pubertal aggressive periodontitis (PAP). MATERIAL AND METHODS Thirteen families with different phenotypes were analysed by direct sequencing of the entire coding region and the regulatory regions of CTSC. The function of novel mutations was tested with enzyme activity measurements. RESULTS In 11 of 13 families, 12 different pathogenic CTSC mutations were found in 10 typical PLS patients, three atypical cases and one PAP patient. Out of four novel mutations, three result in protein truncation and are thus considered to be pathogenic. The homozygous c.854C>T nucleotide exchange (p.P285L) was associated with an almost complete loss of enzyme activity. The observed phenotypic heterogeneity could not be associated with specific genotypes. CONCLUSIONS The phenotypic variability of the PLS associated with an identical genetic background may reflect the influence of additional genetic or environmental factors on disease characteristics. CTSC mutation analyses should be considered for differential diagnosis in all children suffering from severe aggressive periodontitis.

TLR4 and IL-18 gene variants in aggressive periodontitis.

AIM We aimed to assess the association of different genotypes with increased aggressive periodontitis susceptibility by studying functional relevant variants in the pathogen-recognition receptor Toll-like receptor 4 (TLR4) and variants in the promoter region of the pro-inflammatory cytokine interleukin-18 (IL-18). MATERIAL AND METHODS One hundred and eleven patients with aggressive periodontitis and 80 periodontally healthy controls were genotyped for four functional variants in the TLR4 gene (c.896A>G and c.1196C>T) and in the IL-18 promoter (c.-368G>C and c.-838C>A). The genotype and allele frequencies, as well as the frequency of combined genotypes were compared between study groups. RESULTS There were no statistical differences in genotype and allele frequencies within the four variants between the groups. All study subjects were further classified into carriers and non-carriers of at least one variant of both genes. The logistic regression analysis adjusted for gender and smoking showed no association between carrier status of at least one variant of both genes and periodontal status (OR=1.41, 95% CI: 0.43-4.70). CONCLUSIONS Our results reject the hypothesis that functionally relevant IL-18 and TLR4 gene mutations have a major effect on aggressive periodontitis susceptibility alone or in combination.