Toll-like receptor (TLR) 2 and 4 mutations in periodontal disease

Toll-like receptors (TLR) are signal molecules essential for the cellular response to bacterial cell wall components. Different functional effective polymorphisms for the TLR 4 gene (Asp299Gly; Thr399Ile) and for the TLR 2 gene (Arg677Trp, Arg753Gln) have recently been described that are associated with impaired lipopolysaccharide signal transduction. A total of 122 patients with chronic periodontal disease and 122 healthy unrelated controls were genotyped for the Asp299Gly and Thr399Ile polymorphism of the TLR 4 gene and the Arg677Trp and Arg753Gln mutation of the TLR 2 gene. The mutations were identified with polymerase chain reaction followed by restriction fragment length polymorphism (RFLP) analysis. The prevalence of the Asp299Gly and the Thr399Ile mutant allele was 4.1% (10/244) and 4.5% (11/244) among periodontitis patients. For the healthy controls the prevalence was 3.3% (8/244) for the Asp299Gly (P = 0.810) and 3.7% (9/244) for the Thr399Ile mutant allele (P = 0.819). The Arg753Gln mutant allele was found in 2.9% (7/244) of the periodontitis subjects as compared to 4.1% (10/244) in the control group (P = 0.622). The Arg677Trp mutant allele was not found in any of the study subjects. Unlike in ulcerative colitis there was not observed an association between chronic periodontitis and the various mutations of the TLR 2 and 4 gene.     

Toll-like receptor 4 Asp299Gly and Thr399Ile polymorphisms in gastric cancer of intestinal and diffuse histotypes

In the present study we investigated the potential role of Toll-like receptor 4 (TLR-4) Asp299Gly and Thr399Ile polymorphisms as risk factors in the development of gastric cancer. TLR-4 Asp299Gly and Thr399Ile polymorphisms were investigated in 171 Italian patients with sporadic gastric cancer and in 151 controls. Unconditional regression (odds ratio and 95% confidence intervals) were used to investigate the association of the studied polymorphisms with gastric cancer. TLR-4 Thr399Ile polymorphism is linked with an increased susceptibility to gastric cancer (P = 0.023 and hazard ratio = 3.62). No significant association for TLR-4 Asp299Gly polymorphism was found. In the subgroup of patients with intestinal-type gastric cancer, a significant risk of gastric cancer was associated with TLR-4 Thr399Ile genotype (P = 0.006). Our results demonstrated that TLR-4 Thr399Ile polymorphism is linked with an increased susceptibility to gastric cancer. An increased risk for intestinal gastric cancer in carriers of the TLR4 Thr399Ile allele was observed. Future epidemiological studies should consider the possible interactions between proinflammatory genotypes (such as TLR and interleukin-1R polymorphisms) and other risk factors for cancer such as dietary habits and/or exposure to environmental carcinogens.     

Polymorphisms of toll-like receptor 2 and 4 genes in rheumatoid arthritis and systemic lupus erythematosus

Human toll-like receptors (TLRs) participate in the innate response and signal the activation of adaptive immunity. Therefore, these TLRs may be important in autoimmune diseases such as rheumatoid arthritis (RA) and systemic lupus erythematosus (SLE). We investigated, by using a polymerase chain reaction restriction-fragment length polymorphism method, the possible association between the polymorphisms of TLR2 (Arg677Trp and Arg753Gln) and TLR4 (Asp299Gly and Thr399Ile) genes with the susceptibility or severity of RA and SLE. Our study population consisted of 122 patients with SLE, 224 patients with RA, and a control group of 199 healthy individuals. The TLR2 polymorphisms were very rare in our population; no individual carrying the TLR2-Arg677Trp polymorphism was observed, whereas the TLR2-Arg753Gln polymorphism was present in only 1% of the total population. We found no statistically significant differences in the TLR4-Asp299Gly and the TLR4-Thr399Ile genotype or allele distribution between SLE patients, RA patients, and control individuals. Similarly, no association was found with any of the demographic and clinical parameters tested either in RA or in SLE patients. In conclusion, a case-control study was used to analyze, for the first time, the influence of TLR2 and TLR4 gene polymorphism on the predisposition and clinical characteristics of SLE and RA but provided no evidence for association of TLR2 or TLR4 gene polymorphism with either disease in the population under study.     

Toll-like receptors 2 and 4 gene polymorphisms in a southeastern Chinese population with tuberculosis

Polymorphisms of the toll-like receptor 2 (TLR2) gene (Arg677Trp, Arg753Gln) and the TLR4 gene (Asp299Gly, Thr399Ile) were investigated in 205 tuberculosis (TB) patients and 203 controls. Genetic variations were analysed by DNA sequencing. We revealed that these polymorphisms were rare in the southeastern Chinese population and were not associated with susceptibility to TB.     

Chronic periodontal disease is associated with single-nucleotide polymorphisms of the human TLR-4 gene

Periodontitis is an inflammatory disease affecting the connective tissue surrounding the teeth leading to tooth loss. Pathogens associated with periodontitis interact with Toll-like receptors (TLRs) to induce cytokines causing and aggravating disease. We screened 197 individuals suffering from generalized periodontitis for the presence of Asp299Gly and Thr399Ile of TLR-4 as well as Arg753Gln of TLR-2 in comparison to matched controls. Single-nucleotide polymorphisms (SNPs) of TLR-4 were elevated among patients (odd's ratio 3.650, 95% CI 1.573-8.467, P < or = 0.0001), while no difference was observed for TLR-2. TLR-4 SNPs were correlated with chronic periodontitis (odd's ratio 5.562, 95% CI 2.199-14.04, P < or = 0.0001), but not with aggressive periodontitis. This observation was confirmed employing a group of periodontally healthy probands over 60 years of age. These data demonstrate that genetic variants of TLR-4 may act as risk factors for the development of generalized chronic periodontitis in humans.     

Lack of Toll-like receptor 4 and 2 polymorphisms in Korean patients with bacteremia

Toll-like receptors (TLRs) are pattern-recognition receptors that are important in innate immune responses to bacterial infection. The purpose of this study is to describe the prevalence of TLRs genetic variations in the bacteremic patients in Korea. A total of 154 patients with bacteremia and 179 healthy volunteers were included. The Asp299Gly and Thr399Ile allele of the TLR4 gene and Arg753Gln and Arg677Trp allele of the TLR2 gene were tested by PCR-RFLP. The DNA sequences were determined to confirm the PCR-RFLP results. Contrary to the expectation, no genetic polymorphisms were detected in both groups of this study, suggesting that it is very rare in Korean.     

Microsatelite GT polymorphism in the toll-like receptor 2 is associated with colorectal cancer

Factors underlying genetic predisposition for development of sporadic colorectal cancer are largely unknown. The fact that this cancer is more common in patients suffering from inflammatory bowel disease raises the question of the relationship between chronic inflammation and cancer. Toll-like receptors 2 (TLR2) and 4 (TLR4) are critical in initiating innate immune response and inflammation toward various bacteria commonly found in the intestine. Recent evidence about the association of polymorphisms in these genes with ulcerative colitis and Crohn's disease, as well as other inflammatory conditions, was the basis for our investigation of their role in sporadic colorectal cancer. We assessed genotype and allele frequencies of TLR2 GT microsatelite polymorphism, TLR2 Arg753Gln, TLR4 Asp299Gly and TLR4 Thr399Ile polymorphisms in 89 colorectal cancer patients and 88 age- and sex-matched controls. The frequency of TLR2 GT microsatelite alleles with 20 and 21 GT repeats was decreased (p = 0.0044 and p = 0.001, respectively), while the frequency of the allele with 31 GT repeats was increased (p = 0.0147) in patients. The mutant allele Asp299Gly of TLR4 gene was slightly more frequent in colorectal cancer patients (p = 0.0269). In conclusion, we report an association of microsatelite GT polymorphisms of TLR2 gene and Asp299Gly polymorphism of the TLR4 gene with sporadic colorectal cancer among Croatians.     

Rarity of TLR4 Asp299Gly and Thr399Ile polymorphisms in the Korean population

PURPOSE: Activation of the innate immune system and chronic low-grade inflammation are thought to be involved in the pathogenesis of atherosclerosis and also thought to be associated with type 2 diabetes and its complications. As a receptor for bacterial lipopolysaccharide and heat-shock proteins, Toll-like receptor 4 (TLR4) is one of the central regulators of the immune response. Recent studies have reported an association between TLR4 polymorphisms and diabetes and its complications in Caucasian populations. MATERIALS AND METHODS: In this study, we analyzed the association between TLR4 gene polymorphisms in patients with features of type 2 diabetes and healthy controls in Korea. Two polymorphisms of the TLR4 gene (Asp299Gly and Thr399Ile) were examined in 225 diabetic patients and 153 healthy controls using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and single-strand conformation polymorphism (SSCP). RESULTS: No Asp299Gly or Thr399Ile mutations were detected in any of the 378 subjects. Seven subjects from each group who had slightly different SSCP patterns were selected for sequencing, but we found no TLR4 polymorphisms on Exon3. The Asp299Gly and Thr399Ile TLR4 gene polymorphisms were absent in both groups, which was similar to the results for Japanese and Chinese Han subjects. CONCLUSION: Our data and other Asian data suggest that a racial difference can be found in the frequency of the TLR4 polymorphism.     

A unique display of toll-like receptor 4 Asp299Gly and Thr399Ile polymorphisms in non-Han Chinese Hani population

The toll-like receptor 4 (TLR4) polymorphisms, Asp299Gly and Thr399Ile, were investigated with PCR-RFLP and DNA sequencing methods in 938 and 980 individuals from the Yunnan Hani ethnic minority and the majority Han population, respectively. Six heterozygotes for both Asp299Gly and Thr399Ile were detected in the Hani, a polymorphism frequency of 0.6397%, whereas no variants were found amongst the Han.     

Toll-like receptor (TLR) 4 polymorphisms are associated with a chronic course of sarcoidosis

The aetiology of sarcoidosis, an inflammatory granulomatous multi-system disorder, is unclear. It is thought to be the product of an unknown exogenous antigenic stimulus and an endogenous genetic susceptibility. Toll-like receptors (TLR) are signal molecules essential for the cellular response to bacterial cell wall components. Lipopolysaccharide (LPS), for example, binds to TLR 4. Two different polymorphisms for the TLR4 gene (Asp299Gly and Thr399Ile) have been described recently. This leads to a change in the extracellular matrix function of TLR4 and to impaired LPS signal transduction. We genotyped a total of 141 Caucasian patients with sarcoidosis and 141 healthy unrelated controls for the Asp299Gly and Thr399Ile polymorphisms in the TLR4 gene. The mutations were identified with polymerase chain reaction followed by restriction fragment length polymorphism (RFLP) analysis. Among sarcoidosis patients the prevalence for each Asp299Gly and Thr399Ile mutant allele was 15.6% (22/141). In the control group the prevalence was 5.67% (8/141) (P = 0.07). In the subgroup of patients with acute sarcoidosis there was no difference in the control group (P = 0.93), but there was a highly significant association between patients with a chronic course of sarcoidosis and TLR4 gene polymorphisms (P = 0.01).     

Polymorphisms in Toll-like receptors-2 and -4 are not associated with disease manifestations in acute Q fever

Coxiella burnetii is a macrophage-tropic, Gram-negative organism, which causes acute Q fever infection in humans. This zoonotic infection causes illness ranging from asymptomatic seroconversion to severe and protracted disease featuring hepatitis and pneumonia. Interactions between C. burnetii lipopolysaccharide (LPS) and host Toll-like receptors (TLR)-2 and -4 have been implicated in pathogen recognition, phagocytosis and signaling responses. Nonconservative single nucleotide polymorphisms in the coding regions of TLR-2 (Arg677Trp and Arg753Gln) and TLR-4 (Asp299Gly) have been found to correlate with mycobacterial infections and Gram-negative sepsis respectively. Associations between the TLR-2 and -4 polymorphisms, illness characteristics and immune response parameters were examined in subjects with acute Q fever (n=85) and comparison subjects with viral infections (n=162). No correlation was demonstrated between these polymorphisms and susceptibility to Q fever, illness severity or illness course.     

Single step high-throughput determination of Toll-like receptor 4 polymorphisms

BACKGROUND: Toll-like receptors are central components of host defence in humans, responsible for recognition of pathogen-associated molecular patterns and activation of innate immunity. Toll-like receptor 4 (TLR4) is activated by lipopolysaccharide (LPS) and other microbial components, thereby initiating the expression and release of pro-inflammatory cytokines. The common, frequently co-segregating allelic variants Asp299Gly and Thr399Ile have been related to susceptibility to gram-negative infections and sepsis and may be involved in the development of atherosclerosis. Identification of TLR4 Asp299Gly and Thr399Ile genotypes can be important for examination of genotype/phenotype relationships as well as for individual risk assessment of patients. METHODS: TLR4 Asp299Gly and Thr399Ile genotypes were detected by a single tube polymerase chain reaction (PCR), based on exonuclease degradation of dual labelled allele-specific oligonucleotides. The assay results were compared with conventional restriction fragment length polymorphism (RFLP) analysis. RESULTS: Genotypes of 345 individuals were determined simultaneously in a single PCR assay. Allele frequencies for our population were 6.8% for the TLR4 Asp299Gly polymorphism and 6.4% for the Thr399Ile polymorphism. Validation by RFLP analysis revealed a correct detection of all genotypes. CONCLUSIONS: We have developed a novel method for the detection of the TLR4 Asp299Gly and Thr399Ile mutations, permitting rapid genotyping which should be useful for large-scale population studies as well as applicable for routine clinical testing.     

Toll样受体4多态性与强直性脊柱炎易感性的荟萃分析

目的 用荟萃分析方法评价toll样受体(TLR)4基因多态性与强直性脊柱炎的关系. 方法 检索在2007年2月前在PubMed及中国期刊全文数据库发表的中英文献.用RevMan 4.2对纳入文献进行荟萃分析,评价合并效应量、功效、异质性及发表偏倚. 结果 共纳入4篇有关Asp299Gly多态位点和3篇Thr399Ile多态位点的病例对照研究.未发现异质性及发表偏倚,合并后功效在80%左右.Asp299Gly位点合并效应量OR=0.94(95%CI:0.66～1.35),Thr399Ile位点合并效应量OR=1.08(95%CI:0.70～1.65). 结论 TLR4基因不是强直性脊柱炎的主要易感基因.     

Association between toll-like receptors/CD14 gene polymorphisms and inflammatory bowel disease in Korean population

The innate immune response in patients who develop inflammatory bowel disease (IBD) may be abnormal. However, the exact role of Toll-like receptors (TLRs) / CD14 gene in the pathogenesis of IBD has not been fully elucidated. We aimed to investigate the association between polymorphisms of TLR1, 2, 4, 6, and CD14 gene and susceptibility to IBD in Korean population. A total 144 patients of IBD (99 patients with ulcerative colitis, 45 patients with Crohn's disease) and 178 healthy controls were enrolled. Using a PCR-RFLP, we evaluated mutations of TLR1 (Arg80Thr), TLR2 (Arg753Gln and Arg677Trp), TLR4 (Asp299Gly and Thr399Ile), TLR6 (Ser249Pro) genes and the -159 C/T promoter polymorphism of CD14 gene. No TLR polymorphisms were detected in Korean subjects. T allele and TT genotype frequencies of CD14 gene were significantly higher in IBD patients than in healthy controls. In subgroup analysis, T allelic frequency was higher in pancolitis phenotype of ulcerative colitis. In Korean population, the promoter polymorphism at -159 C/T of the CD14 gene is positively associated with IBD, both ulcerative colitis and Crohn's disease.     

Effects of CD14, TLR2, TLR4, LPB, and IL-6 Gene Polymorphisms on Chlamydia pneumoniae Growth in Human Macrophages In Vitro

Chlamydia pneumoniae is an obligate intracellular gram-negative bacterium, which causes respiratory infections in humans. It can infect various cell types, e.g. vascular endothelial cells, smooth muscle cells and monocyte-derived macrophages in vitro . The susceptibility of macrophages from healthy individuals to C. pneumoniae infection is highly variable. In this study, we evaluated the effects of innate immunity genes CD14 −260 C>T, TLR2 Arg753Gln, TLR4 Asp299Gly, LBP Phe436Leu and IL6 −174 G>C polymorphisms on C. pneumoniae growth in human macrophages in vitro. The growth of C. pneumoniae was highest in CD14 −260 C>T TT genotype cells and the difference to CC and CT genotypes was statistically significant ( P  = 0.032 and 0.022 respectively). The G-allele of the IL6 −174 G>C polymorphism had a positive influence on chlamydial growth; the difference was statistically significant only between CC and GC genotypes ( P  = 0.018). TLR2 Arg 753Gln, TLR4 Asp299Gly, LBP Phe436Leu polymorphisms showed no effect on chlamydial growth.     

Association of Toll-like receptor-4 (Asp299Gly and Thr399Ileu) gene polymorphisms with gastritis and precancerous lesions

A Toll-like receptor-4 (TLR-4) Asp299Gly and Thr399Ileu substitution reduces responsiveness to Helicobacter pylori (H. pylori) lipopolysaccharide. CagA+ strains of H. pylori are known to be associated with gastroduodenal diseases. Therefore we aimed to evaluate association of TLR-4 substitutions and CagA seropositivity with gastritis and precancerous lesions in a northern Indian population. After upper gastrointestinal endoscopy, 130 rapid urease test (RUT)-positive patients with nonulcer dyspepsia (NUD) were included. Patients with NUD were also screened for H. pylori infection using modified Giemsa staining and anti-CagA IgG enzyme-linked immunoabsorbent assay. All patients and 200 asymptomatic control subjects were genotyped for TLR-4 substitutions using polymerase chain reaction-restriction fragment length polymorphism. We observed that frequencies of TLR-4 Asp299Gly variants were comparable between patients and control subjects, and also between positive and negative groups of precancerous lesions in patients. Frequencies of TLR-4 399Ileu allele (8% vs 3%, p = 0.008) and Asp299-Ileu399 haplotype (6.5% vs 3%, p = 0.022) were higher in patients than in control subjects at risk for gastritis (OR = 2.6 and 2.5, respectively). TLR-4 399Ileu allele carriers had higher risk for plasma cell infiltration (p = 0.023, OR = 10.6) that led to atrophy (p = 0.028, OR = 4.2) and intestinal metaplasia (p = 0.009, OR = 4.7). CagA positivity was more frequently associated with lymphoid follicle formation (p = 0.033, OR = 2.53). In conclusion TLR-4 Thr399Ileu substitution may be a risk factor for gastritis and precancerous lesions. CagA positivity may be a risk factor for lymphoid follicle development but not for other precancerous lesions in a northern Indian population.     

Polymorphisms of the lipopolysaccharide-signaling complex in inflammatory bowel disease: association of a mutation in the Toll-like receptor 4 gene with ulcerative colitis

Genes encoding for receptors of the innate immune system are potential candidates for susceptibility to inflammatory bowel disease, e.g., mutations in the cytosolic receptor NOD2/CARD15 were associated with Crohn's disease. Herein, two mutations of the Toll-like receptor (TLR)-4 gene (Asp299Gly and Thr399Ile) resulting in impaired lipopolysaccharide signaling, the -159C/T promotor polymorphism of the CD14 gene, polymorphisms of the lipopolysaccharide binding protein gene and the bactericidal permeability increasing protein gene were evaluated in 102 patients with Crohn's disease, 98 patients with ulcerative colitis and 145 healthy controls. The allele and carrier frequencies for the Thr399Ile mutation in TLR4 gene were significantly increased in ulcerative colitis when compared to the controls (P = 0.014 and P = 0.018, respectively). None of the other five polymorphisms was associated with inflammatory bowel disease. In conclusion, a novel association between a functional polymorphism in TLR4 and ulcerative colitis is reported. This observation underscores the importance of impaired innate immunity in inflammatory bowel disease.     

TLR2 and TLR4 polymorphisms in familial Mediterranean fever

It has been suggested that MEVF mutations offer advantage against infections, including tuberculosis. Bearing in mind the central role of TLR-2 and TLR-4 in the recognition of pathogens, we conducted this study to examine whether the TLR2-R753Q, TLR4-D299G, TLR4-T399I common polymorphisms are associated with susceptibility to familial Mediterranean fever (FMF) or affect the course of the disease. A cohort of 169 FMF patients and 245 healthy bone marrow donors were enrolled in the study. FMF patients appeared with a significantly lower frequency of the TLR4-D299G mutated allele (3.2% vs 6.9%, p = 0.032). No association was observed with the other analyzed polymorphisms. Moreover, we found no association between polymorphisms and the frequency of attacks or the development of amyloidosis. Our results may reinforce the hypothesis that FMF patients display a better defense against pathogens, providing an additional mechanism and suggesting a positive selection advantage in the area of the Mediterranean basin.     

Adiposity and bone mineral density of Chilean elderly women in relation to toll-like receptor 4 gene polymorphisms

BACKGROUND: It has been proposed that the toll-like receptor-4 gene (TLR4) may participate in the development of obesity and osteoporosis, in addition to its well-known role in the immune response. On the other hand, the adipose tissue of obese subjects shows an increased expression of the proinflammatory cytokine, tumour necrosis factor-alpha (TNF-alpha), which is released after lipopolysaccharide recognition by TLR4. AIM: To estimate the allele/genotype frequencies and linkage disequilibrium measures of Asp299Gly and Thr399Ile polymorphisms of the TLR4 gene in the Chilean elderly population, and to screen for their association with variables related to adiposity or bone mineral density. SUBJECTS AND METHODS: The study group included 227 unrelated Chilean elderly women (61-95 years) recruited from a population-based sample. Adiposity and bone mineral density measures were obtained using dual-energy X-ray absorptiometry. RESULTS: The allele frequencies for TNF -308A, TLR4 299Gly and TLR4 -399Ile were 9.3%, 4.6% and 4.4%, respectively, with Asp299Gly and Thr399Ile being in strong linkage disequilibrium (D' = 0.88). Although seriously restricted by the low frequency of the allele variants, no relevant association between genotypes and adiposity-related variables were found. Likewise, no significant association between osteoporosis status (categorized as osteoporosis, osteopenia or normal status) with TLR4 Asp299Gly or TNF -308G>A genotypes was found. CONCLUSION: It is unlikely that TLR4 Asp299Gly, TLR4 Thr399Ile or TNF -308G>A polymorphisms have a major influence on adiposity, bone mineral density or osteoporosis status in Chilean elderly women.