Influência do transplante autólogo de células foliculares na formação e funcionalidade de corpo lúteo decorrente da aspiração folicular em ovinos.
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Data
2019-03-29
Autores
Denadai, Renan
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Universidade Estadual Paulista (Unesp)
Resumo
O objetivo do presente trabalho foi avaliar os corpos lúteos (CL) formados em decorrência da aspiração folicular guiada por laparoscopia (LOPU) na espécie ovina, quanto ao desenvolvimento morfológico, funcionalidade e potencial de desenvolvimento gestacional. Foram realizados dois experimentos, em ambos o estro das ovelhas foi sincronizado utilizando dispositivo intravaginal de acetato de medroxiprogesterona por 14 dias, aplicação de 140 μg cloprostenol no quarto dia e retirada do dispositivo no décimo quarto dia com concomitante administração de gonadotrofina coriônica equina (eCG). No Experimento I, 48 horas após a remoção do dispositivo realizou-se a LOPU dos folículos ovarinos em seis ovelhas (Grupo LOPU – LG), ou a LOPU associada com reposição de células em sete animais (Grupo reposição – RG) em cinco animais a ovulação foi espontânea (Grupo controle – CG). Foi realizada avaliação ultrassonográfica ovariana dos animais diariamente até o momento de uma nova ovulação e coleta de sangue para dosagem de progesterona (P4) plasmática a cada 48 horas até o decimo quinto dia. Os animais do LG (1,7±0,5) e RG (1,4±0,5) formaram mais CLs do que os animais do CG (1,0±0,0). A área lútea individual dos CLs foi menor no LG (0,69±0,30 cm2), em comparação ao RG (0,79±0,25 cm2) e CG (0,85±0,32 cm2). A área lútea total foi semelhante entre o LG (1,2±0,4 cm2) e RG (1,1±0,5 cm2), sendo ambas maiores que a do CG (0,9±0,3 cm2). A concentração de P4 plasmática não diferiu entre os grupos, ficando acima de dois ng/mL na fase estática de desenvolvimento dos CLs. No Experimento II, após 48 horas da retirada do dispositivo intravaginal realizou-se a LOPU de todos os folículos ovarianos visíveis. Os complexos cúmulus oócito (COCs) obtidos foram avaliados por estereomicroscopia, um COC autólogo com expansão das células do cúmulus foi transferido para o interior do oviduto de cada ovelha, ipsilateral ao ovário do maior folículo aspirado e inseminação artificial (IA) guiada por laroscopia no corno uterino do mesmo lado. Após as transferências dos oócitos e IA as fêmeas foram acompanhadas diariamente por ultrassonografia trasnretal e dosagem de P4 plasmática por 18 dias. Ocorreu a formação de 2 CLs em cada animal, estes apresentaram produção de P4 crescente em fase inicial, que se estabilizou em concentrações superiores a 2 ng/mL. Uma das ovelhas apresentou diminuição nas concentrações de P4 e redução da área lútea no final do período das observações. Duas ovelhas se apresentavam gestantes à ultrassonografia realizada 35 dias após a transferência dos oócitos, e os partos normais ocorreram aos 151 e 153 dias, respectivamente. A partir das informações dos dois experimentos conclui-se que a LOPU em momento pré-ovulatório leva a formação de CLs funcionais em ovelhas, que o procedimento leva a formação de CLs acessórios, a transferência autóloga de células foliculares leva ao incremento da área lútea, e os animais apresentam concentrações plasmáticas similares à ovelha com CL decorrente de ovulação espontânea, sendo possível o desenvolvimento de gestação normal.
The aim of the present study was to evaluate the corpus luteum (CL) formed as a result of laparoscopic ovum pick-up (LOPU) in the ovine, regarding the morphological development, functionality and potential of gestational development. Two experiments were carried out, in both of them, the oestrous of the ewes was synchronized using a 14-day intravaginal device of medroxyprogesterone acetate, 140 μg cloprostenol on the fourth day and withdrawal of the device on the fourteenth day with concomitant administration of equine chorionic gonadotrophin (eCG). In Experiment I, 48 hours after the removal of the device, was realized a LOPU of the ovarian follicles in six sheep (LOPU Group - LG), or LOPU associated with cell replacement in seven animals (Replacement Group - RG) was performed in five animals a ovulation was spontaneous (Control Group - CG). Ultrasonographic evaluation of the animals was performed daily until the time of ovulation and blood collection for progesterone plasma concentration (P4) every 48 hours until the 15th day. LG animals (1.7 ± 0.5) and RG (1.4 ± 0.5) formed more CLs than CG animals (1.0 ± 0.0). The individual luteal area of CLs was lower in LG (0.69 ± 0.30 cm2), compared to RG (0.79 ± 0.25 cm2) and CG (0.85 ± 0.32 cm2). The total luteal area was similar between LG (1.2 ± 0.4 cm2) and RG (1.1 ± 0.5 cm2), both larger than CG (0.9 ± 0.3 cm2). Plasma P4 concentration did not differ between groups, being above 2 ng / mL in the static phase of CLs development. In Experiment II, after 48 hours of withdrawal of the intravaginal device, LOPU of all visible ovarian follicles was performed. The oocyte cumulus complexes (COCs) obtained were evaluated by stereomicroscopy, an autologous COC with cumulus cell expansion was transferred into the ovary of each sheep, ipsilateral to the ovary of the largest follicle aspirated and artificial insemination (AI) guided by laparoscopy in the uterine horn on the same side. After oocyte and IA transfer the females were monitored daily by transrectal ultrasonography and plasma P4 dosage for 18 days. The formation of 2 CLs occurred in each animal, which presented increased P4 production in the initial phase, which stabilized at concentrations higher than 2 ng/mL. One of the sheep presented a decrease in P4 concentrations and a reduction of the luteal area at the end of the observation period. Two sheep were pregnant on ultrasonography performed 35 days after oocyte transfer, and normal deliveries occurred at 151 and 153 days, respectively. From the information of the two experiments it was concluded that LOPU in the pre-ovulatory period leads to the formation of functional CLs in sheep, which leads to the formation of accessory CLs, the autologous transfer of follicular cells leads to the increase of the luteal area, and the animals present similar plasma concentrations to the sheep with CL due to spontaneous ovulation, being possible the development of normal gestation.
The aim of the present study was to evaluate the corpus luteum (CL) formed as a result of laparoscopic ovum pick-up (LOPU) in the ovine, regarding the morphological development, functionality and potential of gestational development. Two experiments were carried out, in both of them, the oestrous of the ewes was synchronized using a 14-day intravaginal device of medroxyprogesterone acetate, 140 μg cloprostenol on the fourth day and withdrawal of the device on the fourteenth day with concomitant administration of equine chorionic gonadotrophin (eCG). In Experiment I, 48 hours after the removal of the device, was realized a LOPU of the ovarian follicles in six sheep (LOPU Group - LG), or LOPU associated with cell replacement in seven animals (Replacement Group - RG) was performed in five animals a ovulation was spontaneous (Control Group - CG). Ultrasonographic evaluation of the animals was performed daily until the time of ovulation and blood collection for progesterone plasma concentration (P4) every 48 hours until the 15th day. LG animals (1.7 ± 0.5) and RG (1.4 ± 0.5) formed more CLs than CG animals (1.0 ± 0.0). The individual luteal area of CLs was lower in LG (0.69 ± 0.30 cm2), compared to RG (0.79 ± 0.25 cm2) and CG (0.85 ± 0.32 cm2). The total luteal area was similar between LG (1.2 ± 0.4 cm2) and RG (1.1 ± 0.5 cm2), both larger than CG (0.9 ± 0.3 cm2). Plasma P4 concentration did not differ between groups, being above 2 ng / mL in the static phase of CLs development. In Experiment II, after 48 hours of withdrawal of the intravaginal device, LOPU of all visible ovarian follicles was performed. The oocyte cumulus complexes (COCs) obtained were evaluated by stereomicroscopy, an autologous COC with cumulus cell expansion was transferred into the ovary of each sheep, ipsilateral to the ovary of the largest follicle aspirated and artificial insemination (AI) guided by laparoscopy in the uterine horn on the same side. After oocyte and IA transfer the females were monitored daily by transrectal ultrasonography and plasma P4 dosage for 18 days. The formation of 2 CLs occurred in each animal, which presented increased P4 production in the initial phase, which stabilized at concentrations higher than 2 ng/mL. One of the sheep presented a decrease in P4 concentrations and a reduction of the luteal area at the end of the observation period. Two sheep were pregnant on ultrasonography performed 35 days after oocyte transfer, and normal deliveries occurred at 151 and 153 days, respectively. From the information of the two experiments it was concluded that LOPU in the pre-ovulatory period leads to the formation of functional CLs in sheep, which leads to the formation of accessory CLs, the autologous transfer of follicular cells leads to the increase of the luteal area, and the animals present similar plasma concentrations to the sheep with CL due to spontaneous ovulation, being possible the development of normal gestation.
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Palavras-chave
Corpo Lúteo, Aspiração Folicular, Ovino, Ultrassonografia, Progesterona, Corpus Luteum, Ovine, Laparoscopic ovum pick-up, Ultrasonography, Progesterone