The genetics of canine periodontal disease: a candidate gene approach
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2015-09-09
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A doença periodontal (DP) constitui o grupo de doenças inflamatórias causadas pela placa bacteriana no periodonto, incluindo o espectro de doença desde estádios precoces (gengivite) até estádios avançados (periodontite). É uma doença multifatorial resultante da interação dos mecanismos de defesa do hospedeiro com os microrganismos da placa. A DP tem um enorme impacto na medicina humana e na medicina veterinária, pela sua elevada prevalência e pelas suas implicações locais e sistémicas. O modelo cão tem sido extensivamente usado na investigação das doenças orais, contribuindo significativamente para o conhecimento atual em periodontologia. Os aspetos clínicos mais importantes da DP canina são apresentados neste trabalho e são explorados os diferentes modelos animais, com particular relevância para o papel do cão como o modelo mais útil para estudar a DP e o desenvolvimento de novas medidas profiláticas e terapêuticas.
Nas recentes décadas tem sido consolidada a ideia da influência genética na DP, controlando a gravidade do processo inflamatório e a resposta terapêutica. Vários polimorfismos de nucleótidos simples têm sido identificados como fatores de risco, nomeadamente em genes responsáveis por moléculas envolvidas na imunoregulação e/ou metabolismo, mas muitas dúvidas ainda persistem. Na DP canina este é um assunto completamente inexplorado, mas um campo de investigação muito relevante e promissor considerando a elevada similaridade da doença no cão e no homem, o que permite a partilha de conhecimento entre espécies com benefícios mútuos.
Seguindo uma abordagem de genómica comparativa para selecionar os genes candidatos mais promissores, o principal objetivo deste trabalho foi contribuir para uma melhor caracterização da DP no cão, principalmente da sua base genética. Foram selecionados cinco genes (IL1A, IL1B, IL10, IL6 e LTF) que codificam moléculas com um papel relevante na patogenia da DP (interleucina-1α, interleucina-1β, interleucina-10, interleucina-6 e lactotransferrina, respetivamente) e foram delineados estudos caso-controlo, nos quais foi realizada uma análise molecular de cada gene procurando identificar variações genéticas e avaliar a sua possível associação à DP. Foi colocada a hipótese de que na DP canina, similarmente à DP do homem, uma disfunção ou produção desregulada destas moléculas, resultante de variações genéticas, poderá ser parte da explicação das diferenças de susceptibilidade entre indivíduos.
Foram identificadas vinte e seis variações genéticas, oito nos genes IL1A e IL1B, sete no gene IL10, três no gene IL6 e oito no gene LTF. As variações IL1A/1_g.388A>C e IL1A/1_g.521 T>A apresentaram diferenças estatisticamente significativas entre grupos [OR ajustada (95%CI): 0.15 (0.03-0.76), p=0.022; 5.76 (1.03-32.1), p=0.046, respetivamente], significando que, na população estudada, o alelo IL1A/1_g.388C está associado a um menor risco de DP, enquanto que o alelo IL1A/1_g.521A se associa a um risco acrescido. Relativamente às outras variações genéticas não foram encontradas diferenças estatisticamente significativas, mas as variações IL1A/2_g.515G>T, IL10/2_g.285G>A, IL6/2_g.105G>A, LTF/3_g.411C>T, LTF/3_g.420 G>A e LTF/3_g.482G>A conduzem à alteração do aminoácido codificado, o que pode alterar a estrutura e a função protéicas, como demonstrado por várias ferramentas bioinformáticas.
Previamente à análise molecular do gene IL10, foram delineados dois estudos. Pela inexistência de um consenso claro relativamente à associação de polimorfismos no gene IL10 e a DP no homem, foi realizada uma meta-análise de todos os estudos disponíveis. Foi encontrada associação estatisticamente significativa para os polimorfismos IL10-819(-824)C>T e IL10-592(-597)C>A, em que os alelos IL10-819(-824)T e -592(-597)A conferem um acréscimo relativo do risco para periodontite crónica em Caucasianos. Adicionalmente foi delineado um estudo para avaliar os níveis plasmáticos de interleucina-10 em cães com periodontite, investigando a possível correlação destes níveis com a saúde periodontal, e os níveis plasmáticos medidos foram mais baixos no grupo de doentes comparativamente com o grupo controlo.
Os resultados deste trabalho sugerem que os genes caninos IL1A, IL1B, IL10, IL6 e LTF, tal como acontece nos genes ortólogos na espécie humana, são altamente polimórficos e com algumas variações genéticas que podem ser importantes na susceptibilidade para a DP. Os resultados obtidos para o gene IL1A são particularmente relevantes, mas sendo este o primeiro trabalho nesta temática, estudos futuros são essenciais para os reforçar e clarificar a sua
importância biológica, assim como novos trabalhos com outros genes candidatos. Mas é inegável que avanços nesta área são fundamentais para compreender adequadamente as complexas vias causais da DP e melhorar o seu acompanhamento clínico, particularmente com o desenvolvimento de novas estratégias de avaliação de risco. A abordagem de genes candidatos, suportada em ferramentas de genómica comparativa, é um caminho de investigação promissor para atingir estes objetivos, que podem resultar em enormes benefícios na periodontologia humana e veterinária, principalmente por acrescentarem conhecimento para delinear novas estratégias preventivas e terapêuticas, e em última análise, melhorar a saúde de ambas as espécies.
Periodontal disease (PD) refers to a group of inflammatory diseases caused by bacterial plaque in the periodontium and ranges from an early stage (gingivitis) to an advanced stage (periodontitis). It is a multifactorial disease that results from the interaction of the host defence mechanisms with the plaque microorganisms. PD has an enormous impact on human medicine and veterinary medicine due to its high prevalence as well as its local and systemic implications. Dog model has been extensively used in oral disease research, contributing significantly to the current understanding of periodontology. The most important clinical aspects of canine PD were considered in this work and the various animal models were examined with emphasis on the role of the dog as the most useful model for understanding human PD and to develop new therapeutic and preventive measures. In recent decades, it has been consolidated the idea of the genetics influence in PD by controlling the inflammatory process severity and the therapy responses. Various single nucleotide polymorphisms have been identified as risk factors, mainly in genes responsible for molecules involved in immunoregulation and/or metabolism, but many questions still remain. In canine PD, this is a completely unexplored issue but a highly relevant and promising research field namely because the strong similarity between canine and human disease provides the possibility to share the knowledge attained from one species to another, with mutual benefits. Following a comparative genomics approach to identify the most promising candidate genes, the main goal of this work was to contribute for a better characterization of canine PD, particularly in terms of genetic basis. Five candidate genes (IL1A, IL1B, IL10, IL6 and LTF) encoding molecules with recognized relevant role in the PD pathogenesis (interleukin-1α, interleukin-1β, interleukin-10, interleukin-6 and lactotransferrin, respectively) were selected and case-control studies were delineated, in which a molecular analysis of each gene was performed to identify genetic variations and to evaluate its possible association with PD. It was hypothesized that in canine PD, similar to human PD, a disfunction or a dysregulated production of these molecules resulting from genetic variations can be part of the explanation for the differences in disease susceptibility between individuals. A total of twenty-six genetic variations were identified and analyzed, eight in the IL1A and IL1B genes, seven in the IL10 gene, three in the IL6 gene, and eight in the LTF gene. The IL1A/1_g.388A>C and IL1A/1_g.521T>A variations showed statistically significant differences between groups [adjusted OR (95%CI): 0.15 (0.03-0.76), p=0.022; 5.76 (1.03-32.1), p=0.046, respectively], meaning that, in the studied population, the IL1A/1_g.388C allele is associated with a decreased PD risk, whereas the IL1A/1_g.521A allele is associated with an increased risk. Regarding all the others variations, no statistically significant differences were detected, but the IL1A/2_g.515G>T, IL10/2_g.285G>A, IL6/2_g.105G>A, LTF/3_g.411C>T, LTF/3_g.420G>A and LTF/3_g.482G>A variations resulted in a change of encoded amino acid, which may alter protein structure and function, as demonstrated by different bioinformatics tools. Before the molecular analysis of the IL10 gene, two additional studies were delineated. Considering that no clear consensus has been reached about the association of IL10 polymorphisms and human PD, a meta-analysis of all available studies was performed. It was found statistically significant association of IL10-819(-824)C>T and IL10-592(-597)C>A polymorphisms, with IL10-819(-824)T and -592(-597)A alleles conferring a relative increased risk for chronic periodontits in Caucasians. Additionally, a study to evaluate the levels of interleukin-10 in plasma of dogs with periodontitis was delineated assessing a possible correlation between these levels and periodontal condition, being found lower levels in the periodontitis group comparing with the control group. The outcome from this work suggests that dog IL1A, IL1B, IL10, IL6 and LTF genes, as occurs in the human orthologous genes, are highly polymorphic with genetic variants that may be important in PD susceptibility. The results obtained for the IL1A are particularly relevant, but this is the first work in this issue and further studies are essential to reinforce these findings and to clarify its biological importance; as well as other studies with different candidate genes. But, it is undeniable that advances in this area are fundamental to understand properly the complex causal pathways of PD and to improve the clinical management of PD, particularly with the development of novel strategies of risk assessment. A candidate gene approach supported in comparative genomics tools is a promising path of research to achieve these objectives, which may lead to great benefits in human and veterinary periodontology, adding important knowledge to design new preventive and therapeutic strategies, and ultimately to improve health in both humans and dogs.
Periodontal disease (PD) refers to a group of inflammatory diseases caused by bacterial plaque in the periodontium and ranges from an early stage (gingivitis) to an advanced stage (periodontitis). It is a multifactorial disease that results from the interaction of the host defence mechanisms with the plaque microorganisms. PD has an enormous impact on human medicine and veterinary medicine due to its high prevalence as well as its local and systemic implications. Dog model has been extensively used in oral disease research, contributing significantly to the current understanding of periodontology. The most important clinical aspects of canine PD were considered in this work and the various animal models were examined with emphasis on the role of the dog as the most useful model for understanding human PD and to develop new therapeutic and preventive measures. In recent decades, it has been consolidated the idea of the genetics influence in PD by controlling the inflammatory process severity and the therapy responses. Various single nucleotide polymorphisms have been identified as risk factors, mainly in genes responsible for molecules involved in immunoregulation and/or metabolism, but many questions still remain. In canine PD, this is a completely unexplored issue but a highly relevant and promising research field namely because the strong similarity between canine and human disease provides the possibility to share the knowledge attained from one species to another, with mutual benefits. Following a comparative genomics approach to identify the most promising candidate genes, the main goal of this work was to contribute for a better characterization of canine PD, particularly in terms of genetic basis. Five candidate genes (IL1A, IL1B, IL10, IL6 and LTF) encoding molecules with recognized relevant role in the PD pathogenesis (interleukin-1α, interleukin-1β, interleukin-10, interleukin-6 and lactotransferrin, respectively) were selected and case-control studies were delineated, in which a molecular analysis of each gene was performed to identify genetic variations and to evaluate its possible association with PD. It was hypothesized that in canine PD, similar to human PD, a disfunction or a dysregulated production of these molecules resulting from genetic variations can be part of the explanation for the differences in disease susceptibility between individuals. A total of twenty-six genetic variations were identified and analyzed, eight in the IL1A and IL1B genes, seven in the IL10 gene, three in the IL6 gene, and eight in the LTF gene. The IL1A/1_g.388A>C and IL1A/1_g.521T>A variations showed statistically significant differences between groups [adjusted OR (95%CI): 0.15 (0.03-0.76), p=0.022; 5.76 (1.03-32.1), p=0.046, respectively], meaning that, in the studied population, the IL1A/1_g.388C allele is associated with a decreased PD risk, whereas the IL1A/1_g.521A allele is associated with an increased risk. Regarding all the others variations, no statistically significant differences were detected, but the IL1A/2_g.515G>T, IL10/2_g.285G>A, IL6/2_g.105G>A, LTF/3_g.411C>T, LTF/3_g.420G>A and LTF/3_g.482G>A variations resulted in a change of encoded amino acid, which may alter protein structure and function, as demonstrated by different bioinformatics tools. Before the molecular analysis of the IL10 gene, two additional studies were delineated. Considering that no clear consensus has been reached about the association of IL10 polymorphisms and human PD, a meta-analysis of all available studies was performed. It was found statistically significant association of IL10-819(-824)C>T and IL10-592(-597)C>A polymorphisms, with IL10-819(-824)T and -592(-597)A alleles conferring a relative increased risk for chronic periodontits in Caucasians. Additionally, a study to evaluate the levels of interleukin-10 in plasma of dogs with periodontitis was delineated assessing a possible correlation between these levels and periodontal condition, being found lower levels in the periodontitis group comparing with the control group. The outcome from this work suggests that dog IL1A, IL1B, IL10, IL6 and LTF genes, as occurs in the human orthologous genes, are highly polymorphic with genetic variants that may be important in PD susceptibility. The results obtained for the IL1A are particularly relevant, but this is the first work in this issue and further studies are essential to reinforce these findings and to clarify its biological importance; as well as other studies with different candidate genes. But, it is undeniable that advances in this area are fundamental to understand properly the complex causal pathways of PD and to improve the clinical management of PD, particularly with the development of novel strategies of risk assessment. A candidate gene approach supported in comparative genomics tools is a promising path of research to achieve these objectives, which may lead to great benefits in human and veterinary periodontology, adding important knowledge to design new preventive and therapeutic strategies, and ultimately to improve health in both humans and dogs.
Descrição
Tese de Doutoramento em Ciências Veterinárias
Palavras-chave
Doenças periodontais , Cão , Modelo animal , Variação genética , Polimorfismos , Nucleótido , Genómica clínica