Glutathione S-transferase M1, T1, and P1 polymorphisms and periodontitis in a Caucasian population: a case-control study.

BACKGROUND: Glutathione S-transferases (GSTs) play important roles in protecting cells against oxidative stress and toxic chemicals. This study aimed to investigate the distribution of GSTM1, GSTT1, and GSTP1 variants and their roles in periodontitis susceptibility in a Caucasian population. METHODS: We analyzed 406 participants, including 204 healthy controls and 203 periodontitis patients. A multiplex polymerase chain reaction (PCR) approach was used to analyze GSTM1 and GSTT1 loci. GSTP1 variants were detected by PCR-haplotyping method in a subgroup of participants (N = 350). Chi-square or Fisher´s exact tests were used to compare genotypic and allelic differences. The Bonferroni method was applied to correct for multiple comparisons (pcorr). RESULTS: The GSTM1 genotype distribution did not differ significantly between controls and periodontitis patients (p = 0.44). Additionally, the wild/null genotypes of GSTT1, Ile105Val and Ala114Val frequencies of GSTP1 were not significantly different between the two groups after correction for multiple comparisons (p = 0.05, p = 0.55, p = 0.02, pcorr>0.05, respectively). The GSTM1 and GSTP1 Ile105Val gene variants were similarly distributed between non-smokers and smokers in both groups (p = 0.38, p = 0.20, and p = 0.14, p = 0.35, respectively). However, the wild genotype of the GSTT1 and Ala114Ala variant of the GSTP1 genes were present more frequently in non-smoking periodontitis patients than in non-smoking controls (p = 0.03, pcorr>0.05, and p = 0.009, pcorr>0.05, respectively) although their frequencies did not differ between smoking periodontitis patients and smoking controls (p = 0.23, p = 0.68, respectively). CONCLUSIONS: This study in a Czech Caucasian population did not confirm a highly significant association between GST gene variants and susceptibility to periodontitis, as previously reported by Arshad and colleagues in Pakistanis. However, a weak relationship between GSTT1 and GSTP1 rs1138272 polymorphisms and periodontitis in non-smokers was observed.

Gene polymorphisms and serum levels of mannose-binding lectin in Czech patients with recurrent aphthous stomatitis: A case-control study.

BACKGROUND: Recurrent aphthous stomatitis is one of the most prevalent oral mucosal immunological diseases. A recent case-control study in the Egyptian population suggested that single nucleotide polymorphism Gly54Asp (rs1800450) of the mannose-binding lectin 2 gene might affect the mannose-binding lectin serum level and recurrent aphthous stomatitis development. The aim of this study was to determine the distribution of six functional mannose-binding lectin 2 gene polymorphisms and analyse their role in recurrent aphthous stomatitis susceptibility in the Czech population. METHODS: The study included 227 subjects; 137 healthy people and 90 patients with recurrent aphthous stomatitis. Six mannose-binding lectin 2 gene polymorphisms (rs11003125, rs7096206, rs7095891, rs5030737, rs1800450, rs1800451) were analysed by the SNaPshot assay method, mannose-binding lectin serum levels were determined by enzyme-linked immunosorbent assay (ELISA) method in a subgroup of subjects (N = 87). RESULTS: No significant differences in mean of mannose-binding lectin serum levels between healthy controls and patients with recurrent aphthous stomatitis were observed (383 ng/ml ± 249 standard deviation (SD) vs. 316 ng/ml ± 177 SD in remission phase vs. 343 ng/ml ± 254 SD in active phase; p > 0.05), also the allele and genotype frequencies of the studied mannose-binding lectin 2 polymorphisms did not differ significantly between the two groups (p > 0.05, odds ratio (OR): 0.75-1.23). Moreover, the distribution of mannose-binding lectin 2 haplotypes and haplogenotypes was similar in the healthy subjects and patients with recurrent aphthous stomatitis (p > 0.05, OR: 0.75-1.23). CONCLUSIONS: This study did not confirm the previously reported association of the mannose-binding lectin 2 Gly54Asp gene variant and low mannose-binding lectin serum level as the risk factors for susceptibility to recurrent aphthous stomatitis. In addition, no significant relationships between mannose-binding lectin 2 functional haplotypes or haplogenotypes and recurrent aphthous stomatitis were observed.

Association of the angiotensin I converting enzyme (ACE) gene polymorphisms with recurrent aphthous stomatitis in the Czech population: case-control study.

BACKGROUND: Recurrent aphthous stomatitis (RAS) is multifactorial disease with unclear etiopathogenesis. The aim of this study was to determine distribution of the angiotensin I converting enzyme (ACE) gene polymorphisms and their influence on RAS susceptibility in Czech population. METHODS: The study included 230 subjects (143 healthy controls and 87 patients with RAS) with anamnestic, clinical and laboratory data. Five ACE gene polymorphisms (rs4291/rs4305/rs4311/rs4331/rs1799752 = ACE I/D) were determined by TaqMan technique. RESULTS: The allele and genotype distributions of the studied ACE I/D polymorphisms were not significantly different between subjects with/without RAS (Pcorr > 0.05). However, carriers of II genotype were less frequent in the RAS group (OR = 0.48, 95% CI = 0.21-1.12, P = 0.059). Stratified analysis by sex demonstrated lower frequency of II genotype in women (OR = 0.33, 95% CI = 0.09-1.17, P < 0.035, Pcorr > 0.05, respectively) than in men with RAS (P > 0.05). Moreover, the frequency of AGTGD haplotype was significantly increased in RAS patients (OR = 13.74, 95% CI = 1.70-110.79, P = 0.0012, Pcorr < 0.05). In subanalysis, TGD haplotype was significantly more frequent in RAS patients (P < 0.00001) and CGI haplotype was less frequent in RAS patients (P < 0.01), especially in women (P = 0.016, Pcorr > 0.05). CONCLUSIONS: Our study indicates that while the AGTGD and TGD haplotypes are associated with increased risk of RAS development, CGI haplotype might be one of protective factors against RAS susceptibility in Czech population.

Gene variability in matrix metalloproteinases in patients with recurrent aphthous stomatitis.

BACKGROUND: The development of recurrent aphthous stomatitis (RAS), inflammatory disease of oral mucosa, is influenced by both environmental and genetic factors. The aim of this study was to investigate polymorphisms located in seven genes coding different types of matrix metalloproteinases (MMPs)-collagenases (MMP1, MMP8, and MMP13), gelatinases (MMP2 and MMP9), stromelysin (MMP3), and membrane-type metalloproteinase (MMP16) in patients with RAS and healthy controls. METHODS: Totally, 223 subjects were included in this case-control study and their detailed anamnestic, clinical, and laboratory parameters were recorded. Seventy-seven patients with RAS and 146 controls were genotyped for seventeen polymorphisms in the MMPs genes using the real-time polymerase chain reaction (PCR) or PCR with restriction analysis. RESULTS: Allele, genotype, and haplotype frequencies of the studied polymorphisms between RAS patients and controls were similar, except for allele distributions of MMP1 rs1144393, MMP9 rs3918242, and MMP16 rs10429371, which were different between patients with RAS and healthy controls (P = .023, P = .049 and P = .025, all Pcorr  > 0.05, respectively). Moreover, the comparison of genotype frequencies (TT vs CC + CT) of the MMP16 rs10429371 variant showed a marginally significant difference between RAS patients and controls (P = .05, Pcorr  > 0.05, OR = 1.68, 95% CI = 0.95-2.98). CONCLUSIONS: No significant relationship between investigated polymorphisms in seven MMPs genes and RAS development in the Czech population was observed in this study.

Interleukin Gene Variability and Periodontal Bacteria in Patients with Generalized Aggressive Form of Periodontitis.

Host genetic predispositions to dysregulated immune response can influence the development of the aggressive form of periodontitis (AgP) through susceptibility to oral dysbiosis and subsequent host-microbe interaction. This case-control study aimed to perform a multilocus analysis of functional variants in selected interleukin (IL) genes in patients with the generalized form of AgP in a homogenous population. Twelve polymorphisms in IL-1 gene cluster, IL-6 and its receptor, IL-10, IL-17A, and IL-18 were determined in 91 AgP patients and 210 controls. Analysis of seven selected periodontal bacteria in subgingival sulci/pockets was performed with a commercial DNA-microarray kit in a subgroup of 76 individuals. The pilot in vitro study included stimulation of peripheral blood monocytes (PBMC) from 20 individuals with periodontal bacteria and measurement of IL-10 levels using the Luminex method. Only the unctional polymorphism IL­10-1087 A/G (rs1800896) and specific IL-10 haplotypes were associated with the development of the disease (P < 0.05, Pcorr > 0.05). Four bacterial species occurred more frequently in AgP than in controls (P < 0.01, Pcorr < 0.05). Elevated IL-10 levels were found in AgP patients, carriers of IL­10-1087GG genotype, and PBMCs stimulated by periodontal bacteria (P < 0.05, Pcorr > 0.05). We therefore conclude that a combination of genetic predisposition to the altered expression of IL-10 and the presence of specific periodontal bacteria may contribute to Th1/Th2 balance disruption and AgP development.

Chemokine Receptor 2 (CXCR2) Gene Variants and Their Association with Periodontal Bacteria in Patients with Chronic Periodontitis.

Periodontitis, an inflammatory disease caused by subgingival Gram-negative (G-) bacteria, is linked with loss of the connective tissue and destruction of the alveolar bone. In the regulation of inflammatory response, chemokine receptor 2 (CXCR2), a specific receptor for interleukin-8 and neutrophil chemoattractant, plays an important role. The first aim of this study was to investigate the CXCR2 gene variability in chronic periodontitis (CP) patients and healthy nonperiodontitis controls in the Czech population. The second aim was to find a relation between CXCR2 gene variants and the presence of periodontal bacteria. A total of 500 unrelated subjects participated in this case-control study. 329 CP patients and 171 healthy nonperiodontitis controls were analyzed using polymerase chain reaction techniques for three single-nucleotide polymorphisms (SNPs): +785C/T (rs2230054), +1208T/C (rs1126579), and +1440A/G (rs1126580). A DNA microarray detection kit was used for the investigation of the subgingival bacterial colonization, in a subgroup of CP subjects (N = 162). No significant differences in allele, genotype, haplotype, or haplogenotype frequencies of CXCR2 gene variants between patients with CP and healthy controls (P > 0.05) were determined. Nevertheless, Aggregatibacter actinomycetemcomitans was detected more frequently in men positive for the C allele of the CXCR2 +785C/T polymorphism (61.8% vs. 41.1%, P < 0.05; OR = 2.31, 95% CI = 1.03-5.20) and for the T allele of the CXCR2 +1208C/T variant (61.8% vs. 38.9%, P < 0.05; OR = 2.54, 95% CI = 1.13-5.71). In contrast, no statistically significant associations of CXCR2 variants with seven selected periodontal bacteria were found in women. Although none of the investigated SNPs in the CXCR2 gene was associated with CP, the CXCR2 gene variants can be associated with subgingival colonization of G- bacteria in men with CP in the Czech population.

Association of the NOD-like receptor 3 (NLRP3) gene variability with recurrent aphthous stomatitis in the Czech population.

BACKGROUND: Recurrent aphthous stomatitis (RAS) is a multifactorial disease with unclear etiopathogenesis in which disturbance of immunological processes may be involved. The aim of our study was to investigate three single nucleotide polymorphisms (SNPs) rs3806265, rs4612666, rs10754558 in NOD-like receptor 3 (NLRP3), the gene encoding the component of inflammasome, in patients with RAS and healthy controls in the Czech population. METHODS: A total of 207 subjects were included in this case-control study. Sixty-four patients with RAS and 143 healthy controls were genotyped by a method based on polymerase chain reaction using 5' nuclease TaqMan® assays. Detailed anamnestic, clinical, and laboratory data were obtained from all subjects. RESULTS: The allele and genotype frequencies of NLRP3 polymorphisms (rs10754558 and rs3806265) between both groups were similar. However, statistically significant differences in NLRP3 rs4612666 genotypes between the patients with RAS and controls were found; carriers of the TT genotype had a higher risk of developing RAS than subjects with the CT+CC genotypes (OR = 14.69, 95%CI = 1.73-124.72, P = .004, Pcorr < .05). No associations between NLRP3 haplotypes and RAS were observed. CONCLUSIONS: Our study indicates that the NLRP3 rs4612666 polymorphism may be involved in the development of RAS in the Czech population.

Recurrent aphthous stomatitis and gene variability in selected interleukins: a case-control study.

Genetic factors, especially those related to immune system functioning, have been intensively studied to determine their role in the development of recurrent aphthous stomatitis (RAS). The aim of the present study was to analyze gene variability in interleukin (IL)2, IL4 (and its receptor α, IL4Rα), IL10, and IL13, which were selected based on literature review and/or their functional relevance, in Czech patients with RAS and in healthy controls. In total, 252 subjects (178 controls and 74 patients with RAS) were enrolled in this case-control study, and their detailed anamnestic, clinical, and laboratory data were obtained. Nine polymorphisms in the genes encoding interleukins were determined using PCR techniques. There were no significant differences in allele or genotype frequencies of the IL2, IL4, IL4Rα, IL10, and IL13 polymorphisms rs2069762/rs2069763, rs2243250/rs79071878, rs1801275, rs1800896, and rs1800925, respectively, between controls and patients with RAS. The minority alleles rs1800871 and rs1800872, which encode variants of IL10, were associated with a statistically significantly higher risk of RAS, as confirmed by the results of genotype and haplotype analyses. We suggest that variability in the IL10 gene may play an important role in the development of RAS in the Czech population.

Association study of interleukin-1 family, interleukin-6, and its receptor gene polymorphisms in patients with recurrent aphthous stomatitis.

BACKGROUND: Recurrent aphthous stomatitis (RAS) is one of the most common oral chronic ulcerative disease in which proinflammatory cytokines such as interleukin-1 (IL-1) and interleukin-6 (IL-6) are thought to play an important role. The aim of this study was to investigate the possible association between polymorphisms in the IL-1 cytokine family, IL-6 or its receptor and RAS in the Czech population. METHODS: A total of 248 subjects, 184 healthy controls, and 64 patients with RAS were genotyped for IL-1A-889C>T, IL-1B-511C>T, IL-1B+3953C>T, IL-1RN86 bp variable number of tandem repeats (VNTRs) in intron 2, IL-6-597G>A, IL-6-572G>C, IL-6-174G>C, and IL-6R+48992A>C by polymerase chain reaction (PCR) methods. RESULTS: No significant differences between investigated polymorphisms in healthy subjects and patients with RAS were detected (P>.05). In addition, complex analysis also revealed similar IL-1 or IL-6 haplotype frequencies between both groups (P>.05). CONCLUSIONS: In conclusion, IL-1 and IL-6 or its receptor gene variants cannot be used as markers for identification of Czech patients with increased risk of recurrent aphthous stomatitis.

Interleukin-17A Gene Variability in Patients with Type 1 Diabetes Mellitus and Chronic Periodontitis: Its Correlation with IL-17 Levels and the Occurrence of Periodontopathic Bacteria.

Interleukin-17 contributes to the pathogenesis of type 1 diabetes mellitus (T1DM) and chronic periodontitis (CP). We analyzed IL-17A -197A/G and IL-17F +7488C/T polymorphisms in T1DM and CP and determined their associations with IL-17 production and occurrence of periopathogens. Totally 154 controls, 125 T1DM, and 244 CP patients were genotyped using 5' nuclease TaqMan(®) assays. Bacterial colonization was investigated by a DNA-microarray kit. Production of IL-17 after in vitro stimulation of mononuclear cells by mitogens and bacteria was examined by the Luminex system. Although no differences in the allele/genotype frequencies between patients with CP and T1DM + CP were found, the IL-17A -197 A allele increased the risk of T1DM (P < 0.05). Levels of HbA1c were significantly elevated in carriers of the A allele in T1DM patients (P < 0.05). Production of IL-17 by mononuclear cells of CP patients (unstimulated/stimulated by Porphyromonas gingivalis) was associated with IL-17A A allele (P < 0.05). IL-17A polymorphism increased the number of Tannerella forsythia and Treponema denticola in patients with CP and T1DM + CP, respectively (P < 0.05). IL-17A gene variability may influence control of T1DM and the "red complex" bacteria occurrence in patients with CP and T1DM + CP. Our findings demonstrated the functional relevance of the IL-17A polymorphism with higher IL-17 secretion in individuals with A allele.

The effect of IL-4 gene polymorphisms on cytokine production in patients with chronic periodontitis and in healthy controls.

Chronic periodontitis (CP) is an inflammatory disease of the teeth-supporting tissues in which genetic predisposition, dental plaque bacteria, and immune mechanisms all play important roles. The aim of this study was to evaluate the occurrence of IL-4 gene polymorphisms in chronic periodontitis and to investigate the association between polymorphisms and cytokines production after bacterial stimulation. Sixty-two subjects (47 CP patients and 15 healthy controls) with detected two polymorphisms in the IL-4 gene (-590C/T and intron 3 VNTR) were examined. Production of cytokines (IL-1α, IL-1ß, IL-4, IL-5, IL-6, IL-10, IL-17, TNFα, INFγ, and VEGF) was studied after in vitro stimulation of isolated peripheral blood by mitogens (Pokeweed mitogen, Concanavalin A), dental plaque bacteria (Aggregatibacter actinomycetemcomitans, Tannerella forsythia, Porphyromonas gingivalis, and Prevotella intermedia), and Heat Shock Protein (HSP) 70 by the Luminex multiplex cytokine analysis system. The results were correlated with IL-4 genotypes in patients with CP and healthy controls. The mononuclear cells isolated from peripheral blood of CP patients with selected IL-4 polymorphisms significantly altered the production of IFNγ, IL-10, IL-1ß, IL-1α, TNFα, and IL-6 after stimulation by HSP 70 or selected bacteria (from P < 0.001 to P < 0.05). IL-4 gene polymorphisms may influence the function of mononuclear cells to produce not only interleukin-4 but also other cytokines, especially in patients with CP.

Haplotype analysis of interleukin-8 gene polymorphisms in chronic and aggressive periodontitis.

OBJECTIVES: Periodontitis is an inflammatory disease characterized by connective tissue loss and alveolar bone destruction. Interleukin-8 (IL8) is important in the regulation of the immune response. The aim of this study was to analyze four polymorphisms in the IL8 gene in relation to chronic (CP) and aggressive (AgP) periodontitis. METHODS: A total of 492 unrelated subjects were included in this case-control association study. Genomic DNA of 278 patients with CP, 58 patients with AgP, and 156 controls were genotyped, using the 5' nuclease TaqMan assay, for IL8 (rs4073, rs2227307, rs2227306, and rs2227532) gene polymorphisms. Subgingival bacterial colonization was investigated by the DNA-microarray detection kit in a subgroup of subjects (N = 247). RESULTS: Allele and genotype frequencies of all investigated IL8 polymorphisms were not significantly different between the subjects with CP and/or AgP and controls (P > 0.05). Nevertheless, the A(-251)/T(+396)/T(+781) and T(-251)/G(+396)/C(+781) haplotypes were significantly less frequent in patients with CP (2.0% versus 5.1%, P < 0.02, OR = 0.34, 95% CI: 0.15-0.78, resp., 2.0% versus 4.5%, P < 0.05, OR = 0.41, 95% CI: 0.18-0.97) than in controls. CONCLUSIONS: Although none of the investigated SNPs in the IL8 gene was individually associated with periodontitis, some haplotypes can be protective against CP in the Czech population.

Association of Toll-like receptor 9 haplotypes with chronic periodontitis in Czech population.

AIM: Toll-like receptors (TLRs) belong to the pattern recognition receptors family of signal molecules that recognize conserved microbial structures. The aim of this study was to analyse polymorphisms in the TLR genes and their association with chronic periodontitis (CP). MATERIAL AND METHODS: Two polymorphisms (2408G/A, i.e. Arg753Gln and -16934A/T) in TLR-2 and three variants (-1486C/T, -1237C/T and+2848A/G) in the TLR-9 genes were studied in 222 patients with CP and 259 unrelated controls. All polymorphisms were detected using the polymerase chain reaction-restriction fragment length polymorphism methods. Subgingival bacterial colonization was investigated by the VariOr Dento test. RESULTS: No significant differences were found in allele and genotype frequencies of all polymorphisms between patients and controls. Nevertheless, complex analysis revealed differences in TLR9 haplotype frequencies between both groups (p=0.001). Specifically, the haplotype T(-1486)/T(-1237)/A(2848) was significantly more frequent (9.6%versus 2.8%, p<0.000001) and the haplotype T(-1486)/T(-1237)/G (2848) of the TLR9 gene was less frequent (35.9%versus 43.3%, p=0.01) in patients than in controls. However, no significant relationships between periodontal pathogens, TLR polymorphisms and CP were found. CONCLUSIONS: In conclusion, although no significant role of the TLR2 gene in periodontitis was found, our results indicate that TLR9 haplotypes may be associated with susceptibility to CP.

Serotonin transporter gene (SLC6A4) variability in patients with recurrent aphthous stomatitis.

OBJECTIVE: The aim of this study was to investigate five polymorphisms in the SLC6A4 gene in patients with recurrent aphthous stomatitis (RAS) and healthy controls. DESIGN: Totally, 239 subjects were enrolled in this case-control study: 86 patients with RAS and 153 healthy individuals were genotyped for serotonin transporter length polymorphic region (5-HTTLPR) polymorphism, variable number tandem repeat (STin2) and single nucleotide polymorphisms (rs25531, rs3813034, rs1042173) in the SLC6A4 gene by polymerase chain reaction with/without restriction analysis. RESULTS: No significant differences in the allele or genotype frequencies in all studied polymorphisms between RAS patients and healthy controls (P > 0.05) were detected. However, the haplotype analysis detected a higher frequency of LA12 (HTTLPR, rs25531, STin2) haplotype in RAS patients in comparison with healthy controls (P < 0.05, OR = 1.63, 95 % CI = 1.07-2.49). CONCLUSIONS: Our study indicates a possible relationship between SLC6A4 and susceptibility to RAS in the Czech population.

Adipokine gene variability and plasma levels in patients with chronic periodontitis -a case-control study.

Specific variants in genes that encode adipokines and their mRNA and protein expression were previously studied in type 2 diabetes mellitus (T2DM) and obesity, and similar studies have been performed for chronic periodontitis (CP). The aim of this case-control study was to investigate the possible impacts of adiponectin (ADIPOQ), leptin (LEP) and its receptor (LEPR), and resistin (RETN) on the etiopathogenesis of CP. Examinations were performed on 118 non-periodontitis healthy subjects (healthy controls, HC), 205 healthy individuals with CP (H + CP) and 86 type 2 diabetes patients with CP (T2DM + CP). Variants within the ADIPOQ (rs2241766, rs1501299), LEP (rs13228377, rs2167270), LEP receptor (rs1805096), and RETN (rs1862513) genes were determined by qPCR. In addition, the plasma levels of ADIPOQ, LEP, and RETN were analysed by ELISA for 80 individuals. The genotype frequencies of the SNP ADIPOQ +45G/T (rs2241766) differed between the HC and H + CP groups (p=0.03, pcorr>0.05), and carriers of the TT genotype had a lower risk of developing CP compared to carriers of the GG or TG genotypes (p<0.01, pcorr>0.05). However, there were no significant differences in the plasma levels of ADIPOQ, LEP or RETN between the study groups (p > 0.05). Plasma levels of the adipokines were also independent of the gene profiles (p > 0.05). Adipokine plasma levels did not change in patients with H + CP/T2DM + CP compared to HC, but we did identify a specific polymorphism in the ADIPOQ gene that was associated with CP. Although the ADIPOQ +45G/T (rs2241766) gene variant may be a candidate biomarker for CP, further research is required in larger populations with different ethnic backgrounds before any final conclusions can be drawn about the role of this gene in CP.

Interleukin-1 Gene Variability and Plasma Levels in Czech Patients with Chronic Periodontitis and Diabetes Mellitus.

Recent studies have suggested a bidirectional relationship between chronic periodontitis (CP) and diabetes mellitus (DM). Immunoregulatory factors such as cytokines play an important role in etiopathogenesis of both diseases. The aim of this study was to analyze variability in interleukin-1 (IL-1) gene cluster and IL-1ß plasma levels in patients with CP, DM, and a combination of both diseases. A total of 1016 individuals participating in this case-control study-225 healthy controls, 264 patients with CP, 132 with type 1 diabetes (T1DM), and 395 patients with type 2 diabetes (T2DM)-were genotyped using methods based on polymerase chain reaction for IL-1 gene polymorphisms (IL-1A (-889C/T, rs1800587), IL-1B (+3953C/T, rs1143634), and IL-1RN (gene for IL-1 receptor antagonist, IL-1RA, 86 bp tandem repeats in intron 2)). Levels of IL-1ß were measured by Luminex methods in subgroups of controls, CP, T1DM + CP, and T2DM + CP subjects. Although no significant associations were found in the genotype and allele frequencies of IL-1A (-889C/T), significant differences in the allele frequencies of IL-1B (+3953C/T) were observed between controls and CP patients (P < 0.05). In T1DM patients, IL-1RN ∗ S "short" allele and IL-1RN 12 genotype were significantly less frequent than those in controls (P < 0.01). In haplotype analysis, TTL haplotype decreased the risk of CP development (P < 0.01), whereas CCS and CTL haplotypes (P < 0.01 and P < 0.05) were associated with T1DM. Although IL-1ß levels were measured significantly higher in mononuclear cells after stimulation by mitogens, HSP70, or selected periodontal bacteria than in unstimulated cells, IL-1 genotypes did not correlate with circulating IL-1ß levels. In the Czech population, significant associations between the IL-1B polymorphism with CP and the IL-1RN variant with T1DM were found. Haplotype analysis suggests that variability in IL-1 gene cluster may be one of the factors in the CP and T1DM pathogenesis, although single variants of these polymorphisms are not substantial for protein production.

Oral Microbiota Composition and Antimicrobial Antibody Response in Patients with Recurrent Aphthous Stomatitis.

Recurrent aphthous stomatitis (RAS) is the most common disease of the oral mucosa, and it has been recently associated with bacterial and fungal dysbiosis. To study this link further, we investigated microbial shifts during RAS manifestation at an ulcer site, in its surroundings, and at an unaffected site, compared with healed mucosa in RAS patients and healthy controls. We sampled microbes from five distinct sites in the oral cavity. The one site with the most pronounced differences in microbial alpha and beta diversity between RAS patients and healthy controls was the lower labial mucosa. Detailed analysis of this particular oral site revealed strict association of the genus Selenomonas with healed mucosa of RAS patients, whereas the class Clostridia and genera Lachnoanaerobaculum, Cardiobacterium, Leptotrichia, and Fusobacterium were associated with the presence of an active ulcer. Furthermore, active ulcers were dominated by Malassezia, which were negatively correlated with Streptococcus and Haemophilus and positively correlated with Porphyromonas species. In addition, RAS patients showed increased serum levels of IgG against Mogibacterium timidum compared with healthy controls. Our study demonstrates that the composition of bacteria and fungi colonizing healthy oral mucosa is changed in active RAS ulcers, and that this alteration persists to some extent even after the ulcer is healed.

The association of interleukin-4 haplotypes with chronic periodontitis in a Czech population.

BACKGROUND: Cytokine gene polymorphisms are known to influence the susceptibility and disease course of many chronic disorders. Recently, interleukin (IL)-4 gene polymorphisms were associated with aggressive periodontitis. The aim of this study was to test differences in the distribution of the IL-4 alleles, genotypes, and haplotypes between patients with chronic periodontitis (CP) and healthy controls in a Czech population. METHODS: The association study was conducted using an age- and smoking status-matched case-control design in patients with CP (n = 194) and healthy controls (n = 158) using the polymerase chain reaction-restriction fragment length polymorphism methods for the -590C/T, -33C/T, and intron 3 variable number tandem repeat (VNTR) variants of the IL-4 gene. RESULTS: No significant differences between patients and controls were found in allele and genotype frequencies of all three polymorphisms. Nevertheless, complex analysis revealed significant differences in haplotype frequencies between the groups (P = 0.005). The haplotype T(-590)/T(-33)/allele 2 VNTR (70 base pairs)(2) of the IL-4 gene was significantly more frequent in patients with CP than in controls (17.0% versus 11.0%; odds ratio = 1.85; 95% confidence interval: 1.19 to 2.87). CONCLUSION: The three polymorphisms in the IL-4 gene act in a cooperative fashion and suggest that the high-production IL-4 haplotype was associated with an increased risk for CP in the Czech population.

Effects of activated and nonactivated platelet-rich plasma on proliferation of human osteoblasts in vitro.

PURPOSE: The purpose of this study was to evaluate the effect of progressively increasing concentrations of activated and nonactivated platelet-rich plasma (PRP) on proliferation of human osteoblasts in vitro. MATERIALS AND METHODS: Human osteoblasts (hFOB 1.19) obtained from the American Type Culture Collection (ATCC, Manassas, VA) were used in the experiment. PRP was obtained from a 28-year-old healthy male volunteer by means of a Haemonetics gradient density cell separator (Haemonetics, Munich, Germany). Human thrombin was used to activate PRP. Three independent experiments were conducted. Samples containing 10% (0.38x increase in platelet count), 25% (0.95x increase in platelet count), 50% (1.95x increase in platelet count), and 75% (2.86x increase in platelet count) of activated PRP and nonactivated PRP were prepared including controls. After culture periods of 24, 48, and 72 hours osteoblast proliferation was evaluated by counting the number of cells using a Multisizer 3 Coulter Counter (Beckman Coulter, Inc, Fullerton, CA). RESULTS: After 24, 48, and 72 hours of incubation, the number of cells in the control group (without PRP) was higher than that of cells in samples containing activated or nonactivated PRP. Osteoblasts with 10% activated PRP (0.38x increase in platelet count) had the highest viability of all samples containing PRP. CONCLUSIONS: Activated PRP resulted in higher proliferation of osteoblasts compared with nonactivated PRP at concentrations of 10% (0.38x increase in platelet count) and 25% (0.95x increase in platelet count) in culture. This study failed to show significant increases in proliferation of human osteoblasts treated with activated or nonactivated PRP compared with controls in vitro.

Functional polymorphisms in the matrix metalloproteinase-9 gene in relation to severity of chronic periodontitis.

BACKGROUND: Members of the matrix metalloproteinase (MMP) family are implicated in the chronic remodeling in periodontal diseases. Therefore, we performed a case-control study to investigate a plausible association between susceptibility to chronic periodontitis (CP) and the polymorphisms in the MMP-9 (gelatinase B) gene. METHODS: Using polymerase chain reaction with subsequent restriction analyses, MMP-9 -1562C/T and R+279Q variants were determined in 304 subjects (169 patients with CP, 76 with a mild to moderate form and 93 with severe generalized CP, and 135 age- and gender-matched unrelated control subjects). RESULTS: The distribution of the MMP-9 -1562C/T and R+279Q genotypes and alleles did not significantly differ between cases and controls. However, the frequency of the T variant at position -1562 was marginally higher in patients with severe disease compared to those with mild to moderate forms (P <0.05; P(corr) >0.05). In further analysis, the -1562T allele was associated with a severe form of CP in men (odds ratio: 3.87; 95% confidence interval: 1.40 to 10.65; P <0.01 and P(corr) <0.05 for allele; P <0.05 and P(corr) >0.05 for genotype) but not in women. CONCLUSION: These findings suggest that genetic polymorphisms of the MMP-9 gene are not associated with the susceptibility to CP in the Czech population; however, the promoter variant may influence the severity of the disease in men.