Antimicrobial effect of photodynamic therapy in carious lesions in vivo, using culture and real-time PCR methods.

BACKGROUND: The aim of this study was to evaluate the antimicrobial effect of photodynamic therapy (PDT) in carious lesions in vivo by culture and real-time PCR methods. METHODS: Ten teeth with deep active carious lesions were selected and five portions of carious dentin were removed for each tooth. Two increments were used as control, to represent the superficial and deep dentin, respectively. Methylene blue at 100mg/L was placed in contact with the cavity for 5min, before being irradiated with a halogen light source for 1min. Then, after PDT, other three portions were removed. The samples were processed in laboratory and the number of viable cfu was obtained. The real-time PCR analyses were performed in two increments of carious dentin, removed before and after PDT. The Streptococcus mutans DNA was isolated from carious dentin samples and amplification and detection of DNA were performed with real-time PCR. The cavities were then restored with glass-ionomer cement. RESULTS: Using conventional culture methods, the results demonstrated that viable bacteria were significantly reduced in all of the agar plates following photosensitization. No difference was found between both groups regarding S. mutans DNA quantification by real-time PCR. CONCLUSION: Although PDT may not affect the number of S. mutans DNA copies immediately after the treatment, clear reduction of the number of cfu was found. Despite its promising use for eliminating bacteria in dental caries treatment, further studies are necessary to establish an effective clinical protocol for the PDT.

IL-17 genetic and immunophenotypic evaluation in chronic graft-versus-host disease.

Although interleukin-17 (IL-17) is a recently discovered cytokine associated with several autoimmune diseases, its role in the pathogenesis of chronic graft-versus-host disease (cGVHD) was not established yet. The objective of this study was to investigate the association of IL17A and IL17F genes polymorphisms and IL-17A and IL-17F levels with cGVHD. IL-17A expression was also investigated in CD4(+) T cells of patients with systemic cGVHD. For Part I of the study, fifty-eight allo-HSCT recipients and donors were prospectively studied. Blood samples were obtained to determine IL17A and IL17F genes polymorphisms. Cytokines levels in blood and saliva were assessed by ELISA at days +35 and +100 after HSCT. In Part II, for the immunophenotypic evaluation, eight patients with systemic cGVHD were selected and the expression of IL-17A was evaluated. We found association between recipient AA genotype with systemic cGVHD. No association was observed between IL-17A levels and cGVHD. Lower IL-17A levels in the blood were associated with AA genotype. In flow cytometry analysis, decreased expression of IL-17A was observed in patients with cGVHD after stimulation. In conclusion, IL-17A may have an important role in the development of systemic cGVHD.

Association between IL1B (+3954) polymorphisms and IL-1ß levels in blood and saliva, together with acute graft-versus-host disease.

Graft-versus-host disease (GVHD) is associated with morbidity and mortality in the recipients of allogeneic hematopoietic stem cell transplants (allo-HSCTs). Interleukin-1ß (IL-1ß) is a potent inflammatory mediator involved in different inflammatory conditions. Therefore, we aimed to investigate the association of IL1B gene polymorphism in recipients and donors in cases in which acute GVHD (aGVHD) has been reported and the impact of this gene polymorphism on the level of cytokines in the blood and saliva. Fifty-eight consecutive allo-HSCT recipients and their donors were prospectively studied. Saliva and/or blood samples were obtained from the recipients and donors to identify the IL1B gene polymorphism, and cytokine levels were assessed by ELISA. Samples were collected weekly from 7 days before transplantation (day -7) to 100 days after allo-HSCT (day+100), for a total of 16 weeks or until death. aGVHD occurred in 27 individuals evaluated. A significant association was identified between the IL1B polymorphism in the donor and aGVHD development in the corresponding recipients. However, no significant association was detected between the IL1B polymorphism in recipients and the development of aGVHD. In the recipients who were diagnosed with aGVHD, the level of IL-1ß in the saliva and blood were increased. In the saliva, IL-1ß levels increased progressively from the time before the diagnosis of aGVHD until weeks after the diagnosis, whereas in the blood, IL-1ß peak levels could be observed within the time allotted for diagnosis, followed by a decrease in the levels. In addition, we observed a significant association between the IL1B genotype of the recipient (CC) and high IL-1ß levels in the saliva at week 13. In conclusion, IL-1ß could be considered a useful predictor of aGVHD development.

Evaluation of IL17A expression and of IL17A, IL17F and IL23R gene polymorphisms in Brazilian individuals with periodontitis.

The IL23/Th17 axis plays an important role in the pathogenesis of cell-mediated tissue damage caused either by autoimmunity or immune responses against bacterial infection. Single nucleotide polymorphisms in the IL17A, IL17F and IL23R genes have been associated with several inflammatory diseases. However, these polymorphisms have not yet been studied in periodontitis. The aim of present study was to evaluate the expression of IL17A and occurrence of the IL17A (rs2275913), IL17F (rs763780) and IL23R (rs11209026) gene polymorphisms in different clinical forms or severity of periodontitis in a sample of Brazilian individuals. Peripheral blood was obtained from 30 non-smoker individuals and analyzed by flow cytometry to determine IL-17 expression. Genomic DNA was obtained from oral swabs in 180 individuals and analyzed by Real-time PCR. The study group was composed by individuals without periodontitis (control), with aggressive periodontitis (AP) and with chronic periodontitis (CP). Higher frequency of IL17A+CD4+ T cells was observed in control group. The A+ genotype from IL17A (rs2275913) was associated with lack of disease. No association was found considering the IL17F and IL23R polymorphisms. Our data suggest that IL17A and the presence of IL17A (rs2275913) A allele are associated with the absence of periodontal disease.

Serotonin transporter gene polymorphisms: a case-control study.

Considerable evidence indicates that serotonergic mechanisms, particularly the serotonin transporter, are involved in alcoholism and tobacco use and are influenced by polymorphism of the promoter region of 5HTT (5-HTTLPR). As alcohol and tobacco consumption have been implicated in the pathogenesis of oral cancer, the purpose of this study was to investigate 5-HTTLPR polymorphism in patients with oral squamous cell carcinoma (OSCC) compared with a control group in a sample of Brazilian patients. One hundred and three patients affected by OSCC and 103 volunteers without OSCC were genotyped for 5-HTTLPR. Both groups were matched for age, sex and tobacco use. The chi-squared test was used for statistical analysis (α=0.05). There was no statistically significant difference in 5-HTTLPR genotypes between case and control group (p= 0.408). In conclusion, the present investigation demonstrated that serotonin transporter polymorphisms are not implicated in the OSSC development.

Association between polymorphisms in interleukin-17A and -17F genes and chronic periodontal disease.

OBJECTIVE: Interleukin-17 (IL-17) is a cytokine that induces neutrophil recruitment and the release of inflammatory mediators in several inflammatory conditions; nevertheless, the involvement of IL-17 gene polymorphisms in chronic periodontitis (CP) has not been addressed yet. Our aim was to evaluate the association between periodontal status and the polymorphisms IL-17A G197A and IL-17F C7488T in subjects with CP along with their impact on levels of inflammatory mediators. MATERIAL AND METHODS: Genomic DNA was obtained from 30 CP patients and 30 healthy controls (HCs). IL-17A G197A and IL-17F C7488T polymorphisms were determined using PCR-RFLP. Serum and periodontal tissues were collected and processed for ELISA, myeloperoxidase (MPO), and/or microscopic analysis. RESULTS: The frequencies of genotypes in the CP group were significantly different from those of HC. Odds ratio indicated that increased risks for CP were associated with the -197A allele, not with the -7488T allele. In addition, the -197A allele was correlated with worse clinical parameters, higher MPO activity, and increased expression of inflammatory mediators (IL-17A and IL-8) than the other genotypes. CONCLUSIONS: These results indicate that the IL-17A -197A allele is associated with increased risk for CP, likely because this genotype relates to the enhanced inflammation in periodontal tissues.

Serotonin transporter gene polymorphisms: a case-control study

Considerable evidence indicates that serotonergic mechanisms, particularly the serotonin transporter, are involved in alcoholism and tobacco use and are influenced by polymorphism of the promoter region of 5HTT (5-HTTLPR). As alcohol and tobacco consumption have been implicated in the pathogenesis of oral cancer, the purpose of this study was to investigate 5-HTTLPR polymorphism in patients with oral squamous cell carcinoma (OSCC) compared with a control group in a sample of Brazilian patients. One hundred and three patients affected by OSCC and 103 volunteers without OSCC were genotyped for 5-HTTLPR. Both groups were matched for age, sex and tobacco use. The chi-squared test was used for statistical analysis (α=0.05). There was no statistically significant difference in 5-HTTLPR genotypes between case and control group (p= 0.408). In conclusion, the present investigation demonstrated that serotonin transporter polymorphisms are not implicated in the OSSC development.

Consideráveis evidências indicam que mecanismos serotoninérgicos, particularmente o transportador de serotonina, estão envolvidos no alcoolismo e no uso de fumo e são influenciados pelo polimorfismo da região promotora do 5HTT (5-HTTLPR). Como o consumo de álcool e fumo está implicado na patogênese do câncer, o objetivo deste estudo foi investigar o polimorfismo 5-HTTLPR em pacientes com carcinoma bucal de células escamosas (CBCE) comparado com um grupo controle em uma amostra de pacientes brasileiros. Cento e três pacientes afetados por CBCE e 103 voluntários sem história de CBCE foram genotipados para 5-HTTLPR. Ambos os grupos foram pareados pela idade, gênero e uso de fumo. O teste do qui-quadrado foi usado para análise estatística. Não houve diferença estatística entre os genótipos dos grupos caso e controle (p= 0,408). Concluindo, a presente investigação demonstrou que os polimorfismos do transportador de serotonina não estão implicados no desenvolvimento do CBCE.

Investigation of functional IL-10 gene polymorphism and IL-10 levels in acute graft-versus-host disease.

BACKGROUND: Graft-versus-host disease (GVHD) represents a major complication in allogeneic hematopoietic stem cell transplant (allo-HSCT) recipients. Although studies have been conducted concerning the investigation of cytokine polymorphisms in the development of acute GVHD (aGVHD), the contribution of recipients and donors as regards cytokine levels has not yet been thoroughly assessed. OBJECTIVE: The aim of this study was to investigate the impact of IL-10 polymorphisms on cytokine levels in blood and saliva, in addition to the occurrence and severity of aGVHD. METHODS: Fifty-eight consecutive allo-HSCT recipients and their donors were included in this prospective study. Saliva and/or blood samples were obtained from recipients and donors to determine IL10 polymorphisms. The IL-10 levels in the blood and saliva were also assessed. The samples were collected from seven days before transplant (day -7) to 100 days after allo-HSCT (day +100), once a week or until the death of recipient. RESULTS: No association was found between recipient and donor IL10 polymorphism and IL-10 levels in the saliva with aGVHD. In contrast, IL-10 levels in the blood were associated with the occurrence of aGVHD. The high producer phenotype in the recipient was also associated with high levels of IL-10 in the blood and saliva. CONCLUSION: Although IL10 polymorphisms were not associated with the occurrence and severity of aGVHD, the genetic background of the recipient did in fact influence the production of the cytokine. Furthermore, as IL-10 levels in the blood were associated with the disease development, this parameter may well be a useful predictor of aGVHD development.

Association between oral HSV-1 and survival in allogeneic hematopoietic stem cell transplanted patients.

INTRODUCTION: This study was designed to investigate the effect of oral HSV-1 shedding on the survival of allogeneic hematopoietic stem cell transplanted (allo-HSCT) patients. METHODS: One hundred nineteen allo-HSCT patients were included in the study and divided in three groups: before transplant, 100 days after transplant and 1 year of allo-HSCT. Healthy volunteers matched by age and gender were also selected. Oral swabs were performed and the nested PCR was used to detect HSV-1 presence in the oral mucosa. In statistical analysis, chi-square test was used to test the distribution of HSV1 shedding among the three groups. Time to death after allo-HSCT was displayed by means of the Kaplan-Meier method and the results were compared by the log-rank test. Cox proportional hazards multivariate model was used to evaluate the survival. RESULTS: We observed that HSV-1 shedding was similar at different points after allo-HSCT. However, HSV-1 shedding before allo-HSCT was associated with worst survival rates after allo-HSCT in multivariate analysis. CONCLUSION: Our data demonstrates that HSV-1 shedding in oral mucosa before transplant is associated with worst survival rate of allo-HSCT patients.

Epstein-Barr virus and human herpes virus-8 are not associated with juvenile nasopharyngeal angiofibroma.

BACKGROUND: Nasopharyngeal angiofibroma (also known as juvenile nasopharyngeal angiofibroma) is a rare fibroblastic tumor with a vascular component that occurs in the nasopharynx and posterolateral nasal wall of adolescent boys. The etiology of nasopharyngeal angiofibroma remains elusive. This investigation was undertaken to determine if human herpes simplex virus-8 and Epstein-Barr virus are possible etiologic viruses and to determine if they have any association with the age of the patient and/or the proliferative state of the lesion. MATERIALS AND METHODS: Formalin fixed, routinely processed, and paraffin embedded surgical specimens of 15 angiofibromas were submitted to PCR for EBV and HHV-8, while in situ hybridization was also employed for EBV. Immunohistochemical analysis for ki-67 was performed using MIB immunostaining. RESULTS: None of the tumors were positive for HHV-8. The PCR technique produced a false positive reaction in five cases, with all cases non-reactive with EBV-ISH. The age of the patients did not show correlation with the Ki-67 labeling index. CONCLUSION: Angiofibroma does not appear to be associated with either HHV-8 or EBV, thereby excluding these viruses as potential etiologic agents. The lack of a correlation between the proliferative index and the age of the patient suggests the proposed puberty induced, testosterone-dependent tumor growth may not play a significant role in tumor development.

EBV-1 and HCMV in aggressive periodontitis in Brazilian patients.

The purpose of the present investigation was to compare the presence of Epstein-Barr virus type 1 (EBV-1) and of Human Cytomegalovirus (HCMV) in crevicular fluid samples from deep and shallow periodontal pocket sites of Brazilian patients with aggressive periodontitis. A total of 30 systemically healthy patients with aggressive periodontitis participated in the study. Paper points were inserted into 2 gingivitis sites (<3 mm) and into 2 periodontitis sites (>5 mm) in each patient. PCR assay was used to identify genomic copies of HCMV and EBV-1. Twenty-three patients (77%) were positive for EBV-1, while only 2 patients (6%) were positive for HCMV. The McNemar test revealed a positive association between EBV-1 and periodontal lesions (p=0.043). Thirty-four (57%) out of 60 periodontitis sites were positive for EBV-1, whereas 18 (30%) gingivitis sites were positive (p=0.01). Only two sites (6.7%) were positive for HCMV. No positive association was found between HCMV and periodontitis or gingivitis (p=0.479). The elevated occurrence of EBV-1 DNA in periodontal pockets of patients with aggressive periodontitis supports a possible periodontopathic role of this virus.

Investigation of functional gene polymorphisms: IL-1B, IL-6 and TNFA in benign migratory glossitis in Brazilian individuals.

BACKGROUND: Benign migratory glossitis (BMG) is a very common immunological oral disease of unknown aetiology. METHODS AND SUBJECTS: Fifty-three consecutive subjects affected by BMG and 53 age- and sex-matched control subjects were genotyped for IL-1B, IL-6 and TNFA polymorphisms. Binary logistic regression models were fitted and values of P < 0.05 were considered significant. RESULTS: A significant difference in the distribution of IL-1B genotypes was observed in the group with BMG in univariate analyses (P = 0.01). The multivariate analyses showed that the CT genotype of the IL1-B gene was significantly associated with a high risk to develop BMG (P = 0.02, OR 2.76). The combined presence of IL-1beta high and intermediate producers genotypes was also associated with BMG in multivariate analyses (P = 0.01, OR 3.05). IL-6 and TNFA polymorphisms were not associated with BMG in the univariate and multivariate analyses. CONCLUSION: Our findings demonstrate that the polymorphism +3954 IL-1B is associated with an increased risk of BMG development and suggest a genetic basis for disease development.

Investigation of functional gene polymorphisms IL-1beta, IL-6, IL-10 and TNF-alpha in individuals with recurrent aphthous stomatitis.

UNLABELLED: Recurrent aphthous stomatitis (RAS) is characterized by recurrent episodes of oral ulceration in an otherwise healthy individual. Some reports in the literature indicate that RAS may have immunological, psychological, genetic and microbiological bases. OBJECTIVE: The purpose of the present study was to investigate, using binary logistic regression analyses, a possible association between the functional IL-1beta +3954 (C/T), IL-6 -174 (G/C), IL-10 -1082 (G/A) and TNF-alpha -308 (G/A) genetic polymorphism and RAS in a sample of Brazilian patients, using a multivariate statistical analysis. DESIGN: Sixty-four consecutive subjects affected by minor and major forms of RAS and 64 healthy volunteers were genotyped. To investigate the association between the single nucleotide polymorphisms and risk of RAS, binary logistic regression models were fitted. The associations were expressed by odd ratios (ORs) and adjusted for age and gender, with the corresponding 95% CIs. P-values less than 0.05 were considered significant. RESULTS: A significant increase in the IL-1beta and TNF-alpha heterozygous genotypes were associated with an increased risk of RAS development (OR 2.40 and 3.07, respectively), in the multivariate model. CONCLUSION: Our findings demonstrate that polymorphisms of high IL-1beta and TNF-alpha production were associated with an increased risk of RAS development. Our findings also give additional support to a genetic basis for RAS pathogenesis.

EBV-1 and HCMV in aggressive periodontitis in Brazilian patients

The purpose of the present investigation was to compare the presence of Epstein-Barr virus type 1 (EBV-1) and of Human Cytomegalovirus (HCMV) in crevicular fluid samples from deep and shallow periodontal pocket sites of Brazilian patients with aggressive periodontitis. A total of 30 systemically healthy patients with aggressive periodontitis participated in the study. Paper points were inserted into 2 gingivitis sites (< 3 mm) and into 2 periodontitis sites (> 5 mm) in each patient. PCR assay was used to identify genomic copies of HCMV and EBV-1. Twenty-three patients (77 percent) were positive for EBV-1, while only 2 patients (6 percent) were positive for HCMV. The McNemar test revealed a positive association between EBV-1 and periodontal lesions (p = 0.043). Thirty-four (57 percent) out of 60 periodontitis sites were positive for EBV-1, whereas 18 (30 percent) gingivitis sites were positive (p = 0.01). Only two sites (6.7 percent) were positive for HCMV. No positive association was found between HCMV and periodontitis or gingivitis (p = 0.479). The elevated occurrence of EBV-1 DNA in periodontal pockets of patients with aggressive periodontitis supports a possible periodontopathic role of this virus.

O objetivo do presente estudo foi comparar a presença do vírus Epstein-Barr tipo 1 (EBV-1) e do Citomegalovírus Humano (HCMV) em amostras de fluido crevicular de bolsas periodontais rasas e profundas de pacientes brasileiros com periodontite agressiva. Trinta pacientes sistemicamente saudáveis com periodontite agressiva participaram deste estudo. Cones de papel foram inseridos em 2 sítios de gengivite (< 3 mm) e em 2 sítios de periodontite (> 5 mm) de cada paciente. Reações de PCR foram usadas para identificar cópias de DNA genômico de HCMV e EBV-1. Em 23 pacientes (77 por cento), os testes foram positivos para EBV-1, enquanto apenas 2 pacientes (6 por cento) foram positivos para HCMV. O teste de McNemar apontou associação positiva entre EBV-1 e lesões periodontais (p = 0,043). Trinta e quatro (57 por cento) dos 60 sítios de periodontites foram positivos para o EBV-1, enquanto 18 (30 por cento) dos sítios de gengivites foram positivos (p = 0,01). Apenas 2 sítios (6,7 por cento) foram positivos para o HCMV. Não foi encontrada associação positiva entre HCMV e periodontite ou gengivite (p = 0,479). A alta ocorrência de DNA de EBV-1 em bolsas periodontais de pacientes com periodontite agressiva corrobora a possível função periodontopática deste vírus.

Helicobacter pylori in the oral mucosa of patients submitted to allogeneic haematopoietic stem cell transplantation.

This study was designed to investigate the impact of haematopoietic stem cell transplantation (HSCT) on Helicobacter pylori colonization of the oral mucosa by nested polymerase chain reaction (nested-PCR). Forty six consecutive patients submitted to HSCT and 46 healthy volunteers were included in the study. Oral swabs were taken from the oral mucosa of the patients and control group. The medical records of the patients were reviewed and the following information was retrieved: gender and age of the patient, donor gender, primary disease, stem cell source (bone marrow or blood stem cells), leukocyte, neutrophil and platelet counts, and chronic graft versus host disease (cGVHD) of salivary glands. The results demonstrated an increased frequency of H. pylori in the oral mucosa of HSCT patients compared to controls (rho = 0.002). The presence of H. pylori in the oral mucosa was not related to the severity of cGVHD. The median counts of platelet/mm3, leukocytes/mm3 and neutrophils/mm3 in the group of HSCT patients positive for H. pylori were not statistically different from those of the patients negative for it. In conclusion, the present study shows increased frequency of H. pylori in the oral mucosa of HSCT patients compared to non-transplanted healthy volunteers.

Helicobacter pylori in the oral mucosa of patients submitted to allogeneic haematopoietic stem cell transplantation / Helicobacter pylori na mucosa oral de pacientes submetidos ao transplante de células-tronco hematopoiéticas

This study was designed to investigate the impact of haematopoietic stem cell transplantation (HSCT) on Helicobacter pylori colonization of the oral mucosa by nested polymerase chain reaction (nested-PCR). Forty six consecutive patients submitted to HSCT and 46 healthy volunteers were included in the study. Oral swabs were taken from the oral mucosa of the patients and control group. The medical records of the patients were reviewed and the following information was retrieved: gender and age of the patient, donor gender, primary disease, stem cell source (bone marrow or blood stem cells), leukocyte, neutrophil and platelet counts, and chronic graft versus host disease (cGVHD) of salivary glands. The results demonstrated an increased frequency of H. pylori in the oral mucosa of HSCT patients compared to controls (rho = 0.002). The presence of H. pylori in the oral mucosa was not related to the severity of cGVHD. The median counts of platelet/mm³, leukocytes/mm³ and neutrophils/mm³ in the group of HSCT patients positive for H. pylori were not statistically different from those of the patients negative for it. In conclusion, the present study shows increased frequency of H. pylori in the oral mucosa of HSCT patients compared to non-transplanted healthy volunteers.

O objetivo do estudo é investigar o impacto do transplante de células-tronco hematopoiéticas (TCTH) na colonização da mucosa bucal pela Helicobacter pylori através do "nested-PCR". Quarenta e seis pacientes submetidos ao TCTH e 46 indivíduos saudáveis foram incluídos no estudo. Raspados de mucosa bucal foram realizados nos pacientes do grupo de estudo e grupo controle. Os dados médicos dos pacientes foram revisados e as seguintes informações foram coletadas: gênero e idade do paciente, gênero do doador, doença primária, fonte de células-tronco (medula óssea ou células-tronco sanguíneas), número de leucócitos, neutrófilos e plaquetas, doença do enxerto contra o hospedeiro crônica (DECHc) de glândulas salivares. Os resultados demonstram aumento na freqüência de H. pylori na mucosa bucal de pacientes submetidos ao TCTH comparado com grupo controle (r = 0.002). A presença da H. pylori na mucosa bucal não teve relação com a severidade da DECHc. As medianas de número de plaquetas/mm³, leucócitos/mm³ e neutrófilos/mm³ no grupo de pacientes TCTH positivos para H. pylori não foram estatisticamente diferentes das medianas dos pacientes negativos. Concluindo, o presente estudo mostra um aumento da freqüência da H. pylori na mucosa bucal de pacientes submetidos ao TCTH quando comparada com a de um grupo de voluntários não transplantados saudáveis.

Serotonin transporter gene polymorphism (5-HTTLPR) in patients with recurrent aphthous stomatitis.

BACKGROUND: Considerable evidence indicates that serotonergic mechanisms, particularly the serotonin transporter (5-HTT) may be involved in psychological alterations. Recent findings have demonstrated that depression and stress are influenced by polymorphism of the promoter region of 5-HTT (5-HTTLPR) and that the short allele (S) is associated with reduced transcriptional efficiency resulting in reduced serotonin expression and uptake. As psychological and genetic factors have been implicated in the pathogenesis of recurrent aphthous stomatitis (RAS), the purpose of the present study was to investigate 5-HTTLPR polymorphism in patients with RAS compared with control subjects. METHODS: Sixty-nine consecutive subjects affected by minor and major forms of RAS and 70 healthy volunteers were genotyped at 5-HTTLPR. The chi-square test was used for statistical analysis. RESULTS: A significant increase in the genotype of SS (P = 0.05) and of the allele S (P = 0.04) in the group of RAS were observed. CONCLUSION: Our findings demonstrate that RAS patients have a tendency to show polymorphism associated with anxiety-related traits.