Genetic analysis of prostatic diseases in canis familiaris
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2014-07-07
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A glândula prostática faz parte do sistema reprodutor masculino e pode ser afectada por várias condições incluindo a hiperplasia prostática benigna (HPB), o cancro da próstata (CaP), a prostatite e quistos. 0 cão (Canis familiaris) é considerado um bom modelo animal para o estudo destas doenças porque, para além das semelhanças morfológicas e funcionais entre as glândulas prostáticas do homem e do cão, estes animais desenvolvem HPB e CaP de forma espontânea. Neste contexto, o estudo da base genética das doenças prostáticas caninas poderá contribuir para o melhor conhecimentos destas patologias e para o desenvolvimento de novas ferramentas de diagnóstico.
0 presente trabalho incluiu uma população de 20 cães machos adultos apresentando diferentes condições prostáticas. O diagnóstico clínico foi realizado através do exame do toque rectal e ultrassonografia e confirmado pela análise citológica do fluido prostático. Foram recolhidas e processadas diferentes amostras biológicas (sangue, urina e fluido da próstata] utilizando protocolos optimizados.
O teste Odelis®CPSE foi analisado em todas as amostras de plasma de forma a testar a sua validade na detecção de doenças prostáticas. Os resultados revelaram que as concentrações plasmáticas de CPSE são mais elevadas em condições patológicas do que em condições normais. No entanto, não foi possível diferenciar a HPB de outras doenças prostáticas.
Após os processos de extração de ADN e ARN, foram implementadas quatro abordagens genéticas diferentes. Devido ao seu papel promissor como ferramenta de diagnóstico, a expressão de gene CPSE foi analisada em cães com diferentes condições prostáticas mas não foi possível associar um perfil de expressão deste gene com uma condição prostática específica. Na segunda abordagem, o gene PCA3 canino foi caracterizado devido ao seu papel central na detecção do CaP no homem. O gene PCA3 canino, localizado no cromossoma 1 do cão, apresentou 79% de identidade com a sequência humana do gene PCA3. O PCA3 não foi expresso nesta população, indicando que a sua expressão génica poderá estar associada apenas com condições neoplásicas. Na terceira abordagem, foram analisadas duas regiões do gene IL-6, permitindo a identificação de duas variações genéticas: uma localizada na região codificante do exão 5 (I5_gl05G>A) e outra na região não codificante (I5_g440G>A). As variações I5g_105G>A e 15_g440G>A não foram correlacionadas com a susceptibilidade para desenvolver uma determinada doença prostática. No entanto, o polimorfismo 15g_105G>A altera o aminoácido codificado e poderá ter efeitos prejudiciais para a proteína IL-6; o polimorfismo I5_g440G>A pode influenciar a susceptibilidade para desenvolver doenças prostáticas, através da regulação dos elementos reguladores do gene IL-6. De forma a sistematizar a informação existente sobre a influência do gene IL-6 e a susceptibilidade para o homem desenvolver CaP, a quarta abordagem incluiu uma revisão sistemática e uma meta-análise com base em estudos de caso-controlo. Em geral, não houve nenhuma associação significativa entre o polimorfismo IL-6 -174G>C e o risco para desenvolver CaP. No entanto, tendo em conta o polimorfismo IL-6 -636G>C, os genótipos CC e GC foram significativamente associados com o risco diminuído para desenvolver CaP numa população asiática.
É essencial complementar este trabalho com novas abordagens, de forma a melhorar a aplicabilidade do teste Odelis®CPSE; obter mais informações sobre genes candidatos e clarificar o papel das variantes genéticas na susceptibilidade para desenvolver doenças da próstata.
The prostate gland is part of the male reproductive system and can be affected by several conditions including benign prostatic hyperplasia (BPH), prostate cancer (PCa), prostatitis and cysts. The domestic dog (Canis familiaris) is considered a good animal model for studying prostatic diseases because, in addition to the morphological and functional similarities between the canine and human prostate glands, these animals spontaneously develop BPH and PCa. In this context, the study of the genetic basis of the canine prostatic diseases might contribute to a better understanding of these diseases and to the development of new diagnostic tools. The present work included a population of 20 adult male dogs presenting different prostatic conditions. The clinical diagnosis was made by rectal palpation and ultrasonography and confirmed by cytological examination of prostatic fluid. Different biological samples (blood, urine and prostatic fluid) were collected and processed using optimized protocols, OdelisOCPSE assay was performed in all plasma samples in order to test its validity in the prostatic diseases detection. The results revealed that CPSE plasma concentrations are higher in pathological conditions than in normal control dogs. Nevertheless, it was not possible to differentiate BPH from the other prostatic diseases. Following DNA and RNA extraction procedures, four different genetic approaches were implemented. Due to its role as a promising diagnostic tool, the CPSE gene expression was analyzed in dogs with different prostatic conditions but it was not possible to associate the expression profile of this gene with a specific condition. In the second approach, canine PCA3 gene was characterized due to its central role in human PCa detection. The assembled canine PCA3 gene was located in dog chromosome 1 and presented 79% of identity with human PCA3 sequence. PCA3 was not expressed in this population, indicating that this gene expression might be associated with neoplasic prostatic condition. In the third approach, two regions of the IL-6 gene were analyzed, allowing the identification of two gene variants: one located in the coding region of exon 5 (I/5_gl05G>A) and another in the untranslated region of exon 5 (I/5_g440G>A). l/5_gl05G>A and I/5_g440G>A gene variants were not correlated with susceptibility to develop a specific prostatic disease. However, the l/5_gl05G>A polymorphism affected the encoded amino acid and may have damage effects for the IL-6 protein; the I/5_g440G>A polymorphism may influence the susceptibility to develop prostatic diseases by affecting the regulatory elements of IL-6 gene. In order to systematically summarize the existing data concerning the IL-6 influence in human PCa susceptibility, the fourth approach included a systematic review and a meta-analysis based upon case-control studies. Overall, there was no significant association between IL-6 -174G>C polymorphism and PCa increased risk. Nevertheless, concerning the IL-6 -636G>C polymorphism, the CC and GC genotypes were statistically significantly associated with PCa decreased risk in an Asian population. It is essential to complement this research with new approaches, in order to improve Odelis®CPSE applicability; accomplish more information about candidate genes and clarify the role of gene variants in prostatic diseases susceptibility.
The prostate gland is part of the male reproductive system and can be affected by several conditions including benign prostatic hyperplasia (BPH), prostate cancer (PCa), prostatitis and cysts. The domestic dog (Canis familiaris) is considered a good animal model for studying prostatic diseases because, in addition to the morphological and functional similarities between the canine and human prostate glands, these animals spontaneously develop BPH and PCa. In this context, the study of the genetic basis of the canine prostatic diseases might contribute to a better understanding of these diseases and to the development of new diagnostic tools. The present work included a population of 20 adult male dogs presenting different prostatic conditions. The clinical diagnosis was made by rectal palpation and ultrasonography and confirmed by cytological examination of prostatic fluid. Different biological samples (blood, urine and prostatic fluid) were collected and processed using optimized protocols, OdelisOCPSE assay was performed in all plasma samples in order to test its validity in the prostatic diseases detection. The results revealed that CPSE plasma concentrations are higher in pathological conditions than in normal control dogs. Nevertheless, it was not possible to differentiate BPH from the other prostatic diseases. Following DNA and RNA extraction procedures, four different genetic approaches were implemented. Due to its role as a promising diagnostic tool, the CPSE gene expression was analyzed in dogs with different prostatic conditions but it was not possible to associate the expression profile of this gene with a specific condition. In the second approach, canine PCA3 gene was characterized due to its central role in human PCa detection. The assembled canine PCA3 gene was located in dog chromosome 1 and presented 79% of identity with human PCA3 sequence. PCA3 was not expressed in this population, indicating that this gene expression might be associated with neoplasic prostatic condition. In the third approach, two regions of the IL-6 gene were analyzed, allowing the identification of two gene variants: one located in the coding region of exon 5 (I/5_gl05G>A) and another in the untranslated region of exon 5 (I/5_g440G>A). l/5_gl05G>A and I/5_g440G>A gene variants were not correlated with susceptibility to develop a specific prostatic disease. However, the l/5_gl05G>A polymorphism affected the encoded amino acid and may have damage effects for the IL-6 protein; the I/5_g440G>A polymorphism may influence the susceptibility to develop prostatic diseases by affecting the regulatory elements of IL-6 gene. In order to systematically summarize the existing data concerning the IL-6 influence in human PCa susceptibility, the fourth approach included a systematic review and a meta-analysis based upon case-control studies. Overall, there was no significant association between IL-6 -174G>C polymorphism and PCa increased risk. Nevertheless, concerning the IL-6 -636G>C polymorphism, the CC and GC genotypes were statistically significantly associated with PCa decreased risk in an Asian population. It is essential to complement this research with new approaches, in order to improve Odelis®CPSE applicability; accomplish more information about candidate genes and clarify the role of gene variants in prostatic diseases susceptibility.
Descrição
Dissertação de Mestrado em Genética Molecular Comparativa e Tecnológica
Palavras-chave
Doenças prostáticas , Expressão gênica , Modelo animal , Genética molecular