Efeito da carbamazepina sobre a população de células de Leydig da linhagem adulta, a esteroidogenese e o estabelecimento da puberdade na prole masculina de ratas tratadas na prenhez e lactação
Data
2013
Autores
Andretta, Rhayza Roberta 
Orientadores
Miraglia, Sandra Maria
Tipo
Dissertação de mestrado
Título da Revista
ISSN da Revista
Título de Volume
Resumo
A carbamazepina (CBZ) e um farmaco utilizado no tratamento da epilepsia, da neuralgia do trigemeo e de disturbios afetivos. O uso materno de anticonvulsivantes durante a gravidez, entre eles a CBZ, esta associado ao aumento do risco de malformacoes fetais. Alem disso, esse farmaco pode atuar nas celulas de Leydig, responsaveis pela sintese de testosterona, inibindo a esteroidogenese testicular. Desta forma, processos dependentes dos niveis adequados de testosterona podem ser alterados pela CBZ quando administrada nas fases pre e pos-natal. O objetivo principal deste trabalho foi avaliar os efeitos da CBZ nas proles masculinas oriundas de ratas Wistar, tratadas durante toda a gestacao e a lactacao, investigando a ocorrencia de danos ao processo espermatogenico, bem como as possiveis alteracoes espermaticas, quantitativas da populacao de celulas de Leydig adultas e de estabelecimento da puberdade. As proles masculinas dos grupos Controle (n=24) e CBZ (n=24) foram eutanasiadas aos 41 dias pos-parto (dpp) (peripuberdade, n=12) e aos 63 dpp (puberdade tardia, n=12). As ratas receberam CBZ (20mg/Kg/dia, i.p.), diluida em propilenoglicol (veiculo da droga), por via intraperitoneal, durante todo o periodo de gestacao e lactacao. O grupo Controle recebeu apenas propilenoglicol, seguindo o mesmo protocolo experimental. O ganho de peso corporeo das ratas maes, durante o tratamento, bem como o das suas proles masculinas, a partir do 4º dpp ate o 63º dpp, foi obtido. Os demais parametros analisados incluiram: distancia anogenital (DAG) e indice anogenital (IAG) nos filhotes machos aos 4dpp, tempos de descida testicular (a partir dos 15º dpp) e de separacao prepucial (a partir do 33º dpp). No 41º e 63º dpp foram realizadas analises morfometricas, biometricas, estereologicas e histopatologicas testiculares; os pesos absolutos e relativos do epididimo, da prostata, da vesicula seminal e do testiculo tambem foram obtidos. A imunomarcacao de celulas de Leydig adultas, por meio do anticorpo anti-11 β-HSD- II, e a dosagem dos niveis plasmaticos dos hormonios estradiol, testosterona e luteinizante (LH), utilizando-se o metodo ELISA, tambem foram realizadas, em ambas as idades. Parametros espermaticos de carater quantitativo e qualitativo foram avaliados aos 63 dpp. Os resultados obtidos demonstraram que a CBZ causou reducao do ganho de peso corporeo e aumento significativo do IAG nas proles masculinas, oriundas de maes tratadas com CBZ. Alem disso, houve atraso significante no tempo de descida dos testiculos e de separacao prepucial. O nivel plasmatico de testosterona tambem se mostrou reduzido nos animais de 41 dpp, oriundos de maes tratadas com CBZ, enquanto os niveis dos hormonios estradiol e LH nao apresentaram alteracoes significantes nas duas idades estudadas. A densidade numerica de celulas de Leydig imunomarcadas para a enzima esteroidogenica 11β-HSD-II tambem nao variou significativamente, em ambas as idades. Nos animais do subgrupo CBZ63, a avaliacao espermatica revelou diminuicao no numero de espermatozoides presentes na cauda epididimaria (por orgao e por grama de orgao) bem como aumento da porcentagem de espermatozoides apresentando morfologia anormal. A analise histopatologica dos testiculos mostrou normalidade do epitelio seminifero dos animais do grupo CBZ no 41º dpp; entretanto, a presenca de pequenos vacuolos intra-epiteliais, na regiao basal do tubulo seminifero, foi uma ocorrencia frequente, no subgrupo CBZ63. Os resultados obtidos nesta pesquisa mostram que a CBZ, quando administrada diariamente durante os periodos de gestacao e lactacao, pode interferir no processo de maturacao sexual da prole do sexo masculino, conduzindo tambem a reducao dos niveis de testosterona nos ratos peripuberes alem de causar aumento da frequencia de formas anomalas de espermatozoides bem como reducao da concentracao de espermatozoides na cauda epididimaria, conforme observado na puberdade tardia. Como nao ocorreram alteracoes significativas da DAG, e provavel que a CBZ nao tenha atuado durante a janela de programacao da masculinizacao; por outro lado, levando em conta que foi observada uma reducao do peso corporeo dos filhotes machos recem-nascidos, oriundos de maes tratadas com CBZ e considerando que a CBZ pode interferir com a esteroidogenese, a ocorrencia de um efeito anti-androgenico da CBZ, durante a fase fetal, nao esta ainda totalmente descartada e devera ser investigada mais detalhadamente. E provavel, tambem, que outros mecanismos paracrinos e endocrinos possam estar envolvidos nas alteracoes reprodutivas causadas pela CBZ, tornando ainda mais complexo seu mecanismo de acao sobre a reproducao masculina
Carbamazepine (CBZ) is an anti-epileptic drug that acts on Leydig cells, affecting steroidogenesis and causing foetal malformation. Anticonvulsant administration during pregnancy, including CBZ, can lead to an increase of foetal malformation. In addition, this drug can act on Leydig cells, which are responsible for testosterone synthesis, leading to inhibition of testicular steroidogenesis. Thus, testosterone-dependent events may be affected by CBZ administration in pre-natal and post-natal phases. The aim of this study was to investigate the effects of CBZ on Leydig cells, on spermatogenesis, on the spermatic parameters as well as on the puberty onset of male offspring from mothers submitted to CBZ treatment during pregnancy and lactation. Rat dams were treated with CBZ (20mg/Kg/day, i.p., CBZ groups, n=24) or propileneglycol (Control groups, n=24) during pregnancy and lactation. The anogenital distance (AGD) and the anogenital index (AGI) of the male offspring were obtained at 4 days post-partum (4dpp). Testicular descent and preputial separation were also evaluated at 15dpp and from 33dpp on, respectively. The body weight gain of the mothers (during pregnancy and lactation) and of the male offspring (from 4dpp until the euthanasia day) was daily taken. The male offspring was euthanized at 41dpp (n=12) and at 63dpp (n=12). These ages correspond to peripuberty and late puberty, respectively. The accessory glands were weighed and the testes were collected for histopathological, biometric, morphometric and stereological analyses. The absolute and relative weights of prostate, epididymis, seminal vesicle were also obtained. Levels of testosterone, estradiol and LH were measured using ELISA (Enzyme Linked Immuno Sorbent Assay). Leydig cells were labelled with anti-11HSD antibody and quantified for numerical density investigation. The quantitative and qualitative spermatic parameters were investigated at 63dpp. The results showed that CBZ caused a reduction of body weight gain, an increase of AGI and a delay of testicular descent and of preputial separation. CBZ also caused a decrease of testosterone level at 41dpp and of sperm count and an increase of abnormal sperm at 63dpp. However, LH and estradiol levels and Leydig cell number were not affected. Histopathological alterations of the seminiferous epithelium, such as vacuolation, were observed at 63dpp. These results indicate that CBZ delays puberty onset and affects steroidogenesis and sperm quality. Since a significant alteration of DAG was not observed in CBZ-rats, it seems that this drug did not act during the masculinization programming window; however, taking into account the known interference of CBZ with steroidogenesis and the significant body weight reduction observed in neonates, a possible anti-androgenic effect of CBZ cannot be completely excluded and must be more deeply investigated. It is also possible that endocrine and paracrine mechanisms are involved in the reproductive alterations caused by CBZ, suggesting that the CBZ mechanism of action on reproduction is complex and deserves further studies.
Carbamazepine (CBZ) is an anti-epileptic drug that acts on Leydig cells, affecting steroidogenesis and causing foetal malformation. Anticonvulsant administration during pregnancy, including CBZ, can lead to an increase of foetal malformation. In addition, this drug can act on Leydig cells, which are responsible for testosterone synthesis, leading to inhibition of testicular steroidogenesis. Thus, testosterone-dependent events may be affected by CBZ administration in pre-natal and post-natal phases. The aim of this study was to investigate the effects of CBZ on Leydig cells, on spermatogenesis, on the spermatic parameters as well as on the puberty onset of male offspring from mothers submitted to CBZ treatment during pregnancy and lactation. Rat dams were treated with CBZ (20mg/Kg/day, i.p., CBZ groups, n=24) or propileneglycol (Control groups, n=24) during pregnancy and lactation. The anogenital distance (AGD) and the anogenital index (AGI) of the male offspring were obtained at 4 days post-partum (4dpp). Testicular descent and preputial separation were also evaluated at 15dpp and from 33dpp on, respectively. The body weight gain of the mothers (during pregnancy and lactation) and of the male offspring (from 4dpp until the euthanasia day) was daily taken. The male offspring was euthanized at 41dpp (n=12) and at 63dpp (n=12). These ages correspond to peripuberty and late puberty, respectively. The accessory glands were weighed and the testes were collected for histopathological, biometric, morphometric and stereological analyses. The absolute and relative weights of prostate, epididymis, seminal vesicle were also obtained. Levels of testosterone, estradiol and LH were measured using ELISA (Enzyme Linked Immuno Sorbent Assay). Leydig cells were labelled with anti-11HSD antibody and quantified for numerical density investigation. The quantitative and qualitative spermatic parameters were investigated at 63dpp. The results showed that CBZ caused a reduction of body weight gain, an increase of AGI and a delay of testicular descent and of preputial separation. CBZ also caused a decrease of testosterone level at 41dpp and of sperm count and an increase of abnormal sperm at 63dpp. However, LH and estradiol levels and Leydig cell number were not affected. Histopathological alterations of the seminiferous epithelium, such as vacuolation, were observed at 63dpp. These results indicate that CBZ delays puberty onset and affects steroidogenesis and sperm quality. Since a significant alteration of DAG was not observed in CBZ-rats, it seems that this drug did not act during the masculinization programming window; however, taking into account the known interference of CBZ with steroidogenesis and the significant body weight reduction observed in neonates, a possible anti-androgenic effect of CBZ cannot be completely excluded and must be more deeply investigated. It is also possible that endocrine and paracrine mechanisms are involved in the reproductive alterations caused by CBZ, suggesting that the CBZ mechanism of action on reproduction is complex and deserves further studies.
Descrição
Citação
ANDRETTA, Rhayza Roberta. Efeito da carbamazepina sobre a população de células de Leydig da linhagem adulta, a esteroidogênese e o estabelecimento da puberdade na prole masculina de ratas tratadas na prenhez e lactação. 2013. 91 f. Dissertação (Mestrado em Ciências) – Escola Paulista de Medicina, Universidade Federal de São Paulo. São Paulo, 2013.