Impact of smoking on protein levels of beta-defensins in periodontal disease

Abstract Antimicrobial peptides (AMPs) are important components of the host response against invading pathogens. In addition to their direct antimicrobial activity, they can also participate in the immune system modulation. However, the role of AMPs in the etiopathogenesis of periodontal disease and the risk factors that may influence their expression in the oral cavity are not fully understood. The aim of this study was to determine the impact of smoking on beta-defensin (hBD) 1 and 2 levels analyzing samples from periodontitis patients. Fifty patients with periodontitis, 25 smokers and 25 non-smokers, and 20 periodontally healthy patients were recruited. After periodontal clinical evaluation, gingival crevicular fluid (GCF) samples were collected from healthy sites of patients without periodontal disease and from healthy and diseased sites of patients with periodontitis. Peptides quantification was performed by sandwich ELISA technique. Smokers showed reduced GCF hBD 1 levels and increased hBD 2 levels compared to non-smokers in diseased sites (p <0.05). Higher levels of hBD 1 were observed in healthy sites of patients without periodontal disease than in healthy sites of patients with periodontitis (p<0.0001). Diseased sites of non-smokers presented higher levels of hBD 2 than healthy sites (p <0.05). These results reveal that protein levels of hBDs 1 and 2 can be impaired by cigarette smoking in the presence of periodontal disease.

Resumo Peptídeos antimicrobianos (PAMs) são componentes importantes da resposta do hospedeiro contra patógenos invasores. Além de sua atividade antimicrobiana direta, eles também podem participar da modulação do sistema imunológico. No entanto, o papel dos PAMs na etiopatogenia da doença periodontal e os fatores de risco que podem influenciar a sua expressão na cavidade oral não são totalmente compreendidos. O objetivo deste estudo foi determinar o impacto do tabagismo nos níveis de beta-defensina (hBD) 1 e 2 analisando amostras de pacientes com periodontite. Cinquenta pacientes com periodontite, 25 fumantes e 25 não fumantes e 20 pacientes periodontalmente saudáveis foram recrutados. Após avaliação clínica periodontal, amostras de fluido crevicular gengival (FCG) foram coletadas de sítios saudáveis de pacientes sem doença periodontal e de sítios saudáveis e doentes de pacientes com periodontite. A quantificação dos peptídeos foi realizada pela técnica de ELISA sanduíche. Fumantes apresentaram níveis reduzidos de hBD 1 no FCG e níveis aumentados de hBD 2 em comparação com não fumantes em locais doentes (p <0,05). Níveis mais elevados de hBD 1 foram observados em sítios saudáveis de pacientes sem doença periodontal do que em sítios saudáveis de pacientes com periodontite (p<0,0001). Os sítios doentes de não fumantes apresentaram níveis mais elevados de hBD 2 do que os sítios saudáveis (p<0,05). Esses resultados revelam que os níveis das hBDs 1 e 2 podem ser prejudicados pelo tabagismo na presença de doença periodontal.

Compartmentalized Innate Immune Response of Human Fetal Membranes against Escherichia coli Choriodecidual Infection.

An infectious process into the uterine cavity represents a major endangered condition that compromises the immune privilege of the maternal-fetal unit and increases the risk for preterm birth (PTB) and premature rupture of membranes (PROM). Fetal membranes are active secretors of antimicrobial peptides (AMP), which limit bacterial growth, such as Escherichia coli. Nevertheless, the antibacterial responses displayed by chorioamniotic membranes against a choriodecidual E. coli infection have been briefly studied. The objective of this research was to characterize the profile of synthesis, activity, and spatial distribution of a broad panel of AMPs produced by fetal membranes in response to E. coli choriodecidual infection. Term human chorioamniotic membranes were mounted in a two independent compartment model in which the choriodecidual region was infected with live E. coli (1 × 105 CFU/mL). Amnion and choriodecidual AMP tissue levels and TNF-α and IL-1ß secretion were measured by the enzyme-linked immunosorbent assay. The passage of bacterium through fetal membranes and their effect on structural continuity was followed for 24 h. Our results showed that E. coli infection caused a progressive mechanical disruption of the chorioamniotic membranes and an activated inflammatory environment. After the challenge, the amnion quickly (2-4 h) induced production of human beta defensins (HBD)-1, HBD-2, and LL-37. Afterwards (8-24 h), the amnion significantly produced HBD-1, HBD-2, HNP-1-3, S100A7, sPLA2, and elafin, whereas the choriodecidua induced LL-37 synthesis. Therefore, we noticed a temporal- and tissue-specific pattern regulation of the synthesis of AMPs by infected fetal membranes. However, fetal membranes were not able to contain the collagen degradation or the bacterial growth and migration despite the battery of produced AMPs, which deeply increases the risk for PTB and PROM. The mixture of recombinant HBDs at low concentrations resulted in increased bactericidal activity compared to each HBD alone in vitro, encouraging further research to study AMP combinations that may offer synergy to control drug-resistant infections in the perinatal period.

Gingival Crevicular Fluid Levels of Human Beta-defensin 2 and 3 in Healthy and Diseased Sites of Individuals with and without Periodontitis.

OBJECTIVE: Human beta-defensins (hBDs) play an important role in the susceptibility to periodontitis. This study aimed to evaluate the levels of hBD-2 and hBD-3 in gingival crevicular fluid (GCF) of individuals with and without periodontitis. METHODS: Twenty periodontally healthy individuals (H) and 20 with periodontitis (P) were recruited. GCF samples were collected from healthy sites (Hh; n=20) from H individuals; and from healthy sites (Ph; n=20), sites with gingival inflammation but no attachment loss (Pg; n=20) and sites with inflammation and attachment loss (Pp; n=20) from P individuals. Levels of hBDs (pg/mL) were determined using ELISA. Comparisons between individuals and among sites were performed through hierarchical linear modelling. RESULTS: GCF levels [median(Q3-Q1)] of hBD-2 were: Hh=4.80 (0.00-30.69); Ph=33.29 (28.04-38.25); Pg=27.56 (23.14-35.50); and Pp=26.20 (23.20-42.54); and of hBD-3 were: Hh=0.00 (0.00-0.00); Ph=978.44 (760.48-1268.12); Pg=938.19 (806.75- 1266.38); and Pp=613.63 (325.50-854.68). Periodontitis at the individual level was associated with higher levels of hBD-2 (p=0.023) and hBD-3 (p greater than 0.001). No influence of site phenotype was observed on hBDs levels. CONCLUSION: Individuals with periodontitis presented higher levels of hBD-2 and hBD-3 in the GCF. These levels seemed to be influenced by periodontitis at the individual level but not by periodontal site.

Association between genetic polymorphisms in DEFB1 and microRNA202 with caries in two groups of Brazilian children.

OBJECTIVE: This replication study aimed to evaluate an association between caries experience and polymorphisms in DEFB1 and miRNA202 in two different Brazilian groups. DESIGN: The population consisted in 312 Brazilian children. Genomic DNA for was extracted from buccal cells isolated from saliva. The genotyping analysis of the polymorphisms in DEFB1 and miRNA202 was performed by real-time polymerase chain reactions. The associations between caries experience, genotype and allele distribution was performed, with an alpha of 0.05. RESULTS: A statistical significant difference was observed between allele distribution and the polymorphism rs12355840 in the miRNA202 for permanent dentition in the Manaus group, in which individuals that carry the allele C had almost three times more chance to have caries (p = .021; OR = 2.7, 95% CI = 1.1-6.7). In the Ribeirão Preto group there was a statistical significant difference for the polymorphism rs11362 in the DEFB1 for both dentition in alleles (p = .043) and genotype (p = .020) distributions. The T allele increased in two times the chance to have caries (OR = 2.03; 95% CI = 1.05-4.07). CONCLUSION: In conclusion, the allelic distribution of the polymorphism rs12355840 in miRNA202 was associated with caries experience in the Manaus group. In the Ribeirão Preto group, the allelic and genotypic distributions in the polymorphism rs11362 in DEFB1 were associated with caries experience.

Gingival crevicular fluid levels of human beta-defensin 1 in individuals with and without chronic periodontitis.

BACKGROUND AND OBJECTIVE: Human beta-defensins (hBDs) contribute to innate immunity antimicrobial activity. They are also effective in the adaptive immune response and may play a crucial role in the susceptibility to diseases of the oral cavity. This study aimed to evaluate the levels of hBD-1 in the gingival crevicular fluid of individuals with and without chronic periodontitis. MATERIAL AND METHODS: Twenty periodontally healthy individuals (H) and 20 individuals with chronic periodontitis were recruited. Gingival crevicular fluid samples were collected from: healthy sites (Hh) from periodontally healthy individuals; and healthy sites (Ph), sites with gingivitis (Pg), and sites with periodontitis (Pp) from individuals with periodontitis. The levels of hBD-1 (pg/mL) were measured using enzyme-linked immunosorbent assay. Comparisons of hBD-1 between individuals (H and chronic periodontitis) and among sites (Hh, Ph, Pg, Pp) were performed through hierarchical linear modeling. RESULTS: Gingival crevicular fluid levels of hBD-1 were: Hh = 229.52 ± 138.96 (median 199.26), Ph = 53.88 ± 58.17 (median 35.75), Pg = 57.11 ± 40.18 (median 39.90) and Pp = 55.31 ± 37.28 (median 54.19). No influence of site diagnosis (level 1; health/gingivitis/periodontitis) was observed; however, individual diagnosis (level 2; health/periodontitis) influenced the levels of hBD-1 (P < .001). CONCLUSION: Periodontally healthy individuals showed higher gingival crevicular fluid levels of hBD-1 when compared to individuals with chronic periodontitis. This suggests a potential protective role of hBD-1 in the susceptibility to chronic periodontitis.

Associação entre beta-defensinas e periodontite / Association between beta-defensins and periodontitis: literature review

Objetivo: as beta-defensinas humanas (hBDs) podem ter um papel-chave na susceptibilidade às doenças na cavidade bucal. Além do efeito antimicrobiano direto, as hBDs aumentam a imunidade adaptativa. O objetivo deste estudo foi realizar uma revisão de literatura científica sobre a relação entre beta-defensinas (hBD) e periodontite. Material e métodos: foi realizada uma pesquisa bibliográfica na base de dados PubMed sobre a expressão de hBDs em indivíduos com periodontite. Os termos beta defensins e periodontitis foram utilizados nessa busca. Resultados: foram selecionados, por um revisor, sete artigos para serem incluídos nessa revisão de literatura: dois estudos de intervenção e cinco estudos transversais. Conclusão: o número de estudos sobre a expressão de beta-defensinas em indivíduos com periodontite é reduzido. O conhecimento sobre o papel das beta-defensinas na periodontite pode trazer um maior entendimento de sua etiopatogenia, além de possibilitar novos indicadores de risco e terapias. Estudos adicionais são necessários para a elucidação da relação entre esses peptídeos antimicrobianos e a periodontite.

Objective: human beta-defensins (hBDs) may play a key role in the susceptibility to diseases in the oral cavity. In addition to the direct antimicrobial effect, hBDs enhance adaptive immunity. The objective of this study was to investigate the literature on the relationship between hBD and periodontitis. Material and methods: a literature review was conducted in the PubMed database on the expression of hBDs in subjects with periodontitis. The terms "beta-defensins" and "periodontitis" were used in this search. Results: seven articles were selected being: two intervention studies and fi ve cross-sectional studies. Conclusion: the number of studies on the expression of beta-defensins in individuals with periodontitis is reduced. Knowledge about the role of beta-defensins in periodontitis may lead to a better understanding of their etiopathogenesis, in addition to providing new risk indicators and therapies. Additional studies are needed to elucidate the relationship between these antimicrobial peptides and periodontitis.

An investigation of human beta-defensins and cathelicidin expression in patients with pterygium / Uma investigação da expressão de beta-defensinas humanas e de catelidicina em pacientes com pterígio

ABSTRACT Purpose: To investigate human beta-defensins (HBDs) and cathelicidin LL-37 (LL-37) expressions in patients with pterygium. Methods: In this retrospective consecutive case series, 26 pterygium specimens and 15 normal conjunctival specimens of 15 control subjects were in vestigated. Expressions of HBD-1, HBD-2, HBD-3, and LL-37 were assessed using immuno histochemical staining. A brown color in the cytoplasm and/or nuclei of epithelial cells indicated positive staining for HBDs and LL-37. For each antibody, the intensity of the reaction (negative [-], weak [1+], moderate [2+], or strong [3+]) was determined to describe the immunoreactions. Results: The median age was 52 years in both groups. There were no significant differences in age and sex between the groups (p=0.583, p=0.355, respectively). Of the 26 pterygium specimens, 15 (57.7%) (14 weak, 1 moderate staining) showed HBD-2 expression, which was not observed in any of the control specimens. One (3.8%) pterygium and one (6.7%) control specimen demonstrated weak staining for HBD-3. HBD-2 expression was significantly higher in the pterygium specimens than in the controls (p=0.002). None of the tissue specimens had positive staining for HBD-1 or LL-37 in either group (both; p=1.00). Conclusions: HBD-2 expression was higher in pterygium specimens than in the controls. HBD-2 expression that might be stimulated by inflammatory cytokines may be related to inflammation and fibrovascular proliferation and may play a role in pterygium pathogenesis.

RESUMO Objetivo: Investigar as expressões beta defensinas humanas (HBD) e catelicidina em pacientes com pterígio. Métodos: Nesta série de casos retrospectivos consecutivos, 26 espécimes de pterígio e 15 espécimes conjuntivais normais de 15 indivíduos controle foram investigados. As expressões de HBD-1, HBD-2, HBD-3 e catelicidina (LL-37) foram avaliadas por coloração imuno-histoquímica. Uma cor castanha no citoplasma ou nos núcleos de células epiteliais foi definida como coloração positiva para HBDs e LL-37. Para cada anticorpo foi determinada a intensidade da reação (negativo [-], fraco [1+], moderado [2+] ou forte [3+]) para descrever as imunoreações. Resultados: A idade média foi de 52 anos em ambos os grupos. Não houve diferença significativa entre os grupos em termos de idade e sexo (p=0,583, p=0,355, respectivamente). Das 26 amostras de pterígio, 15 (57,7%) (14 fracas e 1 moderada) demonstraram a expressão de HBD-2 enquanto não foi encontrada em nenhum dos espécimes de controlo. Um dos pterígios (3,8%) e um dos espécimes de controlo (6,7%) demonstraram fraca coloração para HBD-3. A expressão de HBD-2 foi significati vamente maior nos espécimes de pterígio do que nos controles (p=0,002). Nenhum dos espécimes de tecido apresentou coloração positiva para HBD-1 ou LL-37 em ambos os grupos (ambos p=1,00). Conclusão: Encontramos aumento da expressão de HBD-2 em espécimes de pte rígio em relação aos controles. A expressão de HBD-2 que pode ser estimulada por citocinas inflamatórias pode estar relacionada com inflamação e proliferação fibrovascular e pode desempenhar um papel na patogênese do pterígio.