Abstract:This study aimed at assessing the effect of different concentrations of the growth factor similar to insulin 1 (IGF-1) in the development, survival and ultrastructure of the bovine preantral follicles cultured in situ. Fragments of bovine ovarian cortical tissue were cultured during 1 and 7 days in 1 ml of α-MEM(+) , supplemented with different concentrations of human recombinant IGF-1 (0, 30, 70 and 100 ng/ml), in an incubator at 37°C and 5% of CO2 in 24-well plates with total replacement of the medium every … Show more
“…By chance, IGF‐1 could be efficient with GH in the final preantral stages. However, studies conducted in this laboratory (Jimenez et al, ) with IGF‐1 showed that among the concentrations (30, 70 and 100 ng/ml), 30 ng/ml was more effective for the development of bovine preantral follicles. Another piece of evidence is that IGF‐1 in high concentrations promotes the degeneration of preantral follicles.…”
Section: Discussionmentioning
confidence: 83%
“…After the collection, the ovaries were washed in alcohol 70% and in TCM199, supplemented with 20 mM HEPES, 2,200 mg/L sodium bicarbonate, 25 µg/ml pyruvic acid, 75 µg/ml penicillin and 50 µg/ml streptomycin, named transport TCM199. After the washes, the ovaries were transported in transport TCM199 at 4°C (Jimenez, Araújo, et al, ; Jimenez et al, ). "The ethics committee statement is not required for this research, due we do not have any kind of contact with animals, we worked with slaughterhouse ovary."…”
Section: Methodsmentioning
confidence: 99%
“…The treatments were distributed in the following way: (a) αMEM + during the 14 days (αMEM + ), or αMEM + supplemented with (b) GH during 7 days, followed by 7 days with IGF‐1 (GH → IGF‐1), (c) IGF‐1 during 7 days, followed by 7 days with GH (IGF‐1 → GH) and GH plus IGF‐1 during 14 days of culture (GH + IGF‐1; Figure ). The concentrations used in this experiment were selected based on the best results previously obtained by the research team: 0 ng/ml IGF‐1 (Jimenez et al, ) and 50 ng/ml GH (Jimenez et al, ). The fragments were cultured during 1, 7 and 14 days at 37ºC, air humidified with 5% CO 2 and with total replacement of the medium every 2 days.…”
Section: Methodsmentioning
confidence: 99%
“…Most of the IGF actions that are known are mediated via IGF-1R, and the physiological role of IGF-2R has not yet been clarified (Duarte et al, 2013). In the ovary, IGF-1 has its origins in the granulosa cells and its main role is to stimulate the follicular development in both the preantral (Jimenez et al, 2016a; Magalhães-Padilha, Duarte, et al, 2012;Magalhães-Padilha, Fonseca, et al, 2012) and antral phases (Armstrong, Xia, Gannes, Tekpetey, & Khamsi, 1996). It is also a suppressor of apoptosis (Makarevich & Markkula, 2002), it induces the mitosis of the granulosa cells (Armstrong & Xia, 1993), and it promotes both the oocyte maturation (Lorenzo, Illera, Illera, & Illera, 1994) and the development of bovine embryos (Kaye, 1997).…”
The growth hormone (GH) and growth insulin-like factor-1 (IGF-1) act directly upon the regulation and growth in the different phases of preantral follicles. Thus, it is necessary to define their sequentiality until the in vitro preovulatory development. Therefore, the study aimed to assess the effects of a sequential medium containing GH and/or IGF-1 in the long-duration in vitro culture of preantral ovarian follicles. Ovarian fragments were cultivated: first half (days 1-7), second half (days 7-14) or during 14 culture days. Treatments were identified as: αMEM+; GH → IGF-1; IGF-1 → GH and GH + IGF-1. The culture was designed in 24-well plates, in an incubator at 37°C and 5% CO . The parameters of normality, viability, follicles (primordial/in developing) and follicle diameter were evaluated. In addition, the ultrastructure was confirmed with electron transmission microscopy. The results showed that the culture treated with GH → IGF-1 kept the follicular normality and the viability until the 14th day of culture and increased both in the follicular development until 7th day and in the follicular diameter until 14th day, when compared to the control. The treatments IGF-1 → GH and GH + IGF-1 were not effective in the developing and follicular diameter after 7 days of culture, and also reduced the percentage of viability. It is concluded that the bovine preantral follicles cultured in the sequential medium treated with GH → IGF-1 improved the follicular development until the first half of the culture and kept these parameters with normality, viability and ultrastructure until the second half of the in vitro culture.
“…By chance, IGF‐1 could be efficient with GH in the final preantral stages. However, studies conducted in this laboratory (Jimenez et al, ) with IGF‐1 showed that among the concentrations (30, 70 and 100 ng/ml), 30 ng/ml was more effective for the development of bovine preantral follicles. Another piece of evidence is that IGF‐1 in high concentrations promotes the degeneration of preantral follicles.…”
Section: Discussionmentioning
confidence: 83%
“…After the collection, the ovaries were washed in alcohol 70% and in TCM199, supplemented with 20 mM HEPES, 2,200 mg/L sodium bicarbonate, 25 µg/ml pyruvic acid, 75 µg/ml penicillin and 50 µg/ml streptomycin, named transport TCM199. After the washes, the ovaries were transported in transport TCM199 at 4°C (Jimenez, Araújo, et al, ; Jimenez et al, ). "The ethics committee statement is not required for this research, due we do not have any kind of contact with animals, we worked with slaughterhouse ovary."…”
Section: Methodsmentioning
confidence: 99%
“…The treatments were distributed in the following way: (a) αMEM + during the 14 days (αMEM + ), or αMEM + supplemented with (b) GH during 7 days, followed by 7 days with IGF‐1 (GH → IGF‐1), (c) IGF‐1 during 7 days, followed by 7 days with GH (IGF‐1 → GH) and GH plus IGF‐1 during 14 days of culture (GH + IGF‐1; Figure ). The concentrations used in this experiment were selected based on the best results previously obtained by the research team: 0 ng/ml IGF‐1 (Jimenez et al, ) and 50 ng/ml GH (Jimenez et al, ). The fragments were cultured during 1, 7 and 14 days at 37ºC, air humidified with 5% CO 2 and with total replacement of the medium every 2 days.…”
Section: Methodsmentioning
confidence: 99%
“…Most of the IGF actions that are known are mediated via IGF-1R, and the physiological role of IGF-2R has not yet been clarified (Duarte et al, 2013). In the ovary, IGF-1 has its origins in the granulosa cells and its main role is to stimulate the follicular development in both the preantral (Jimenez et al, 2016a; Magalhães-Padilha, Duarte, et al, 2012;Magalhães-Padilha, Fonseca, et al, 2012) and antral phases (Armstrong, Xia, Gannes, Tekpetey, & Khamsi, 1996). It is also a suppressor of apoptosis (Makarevich & Markkula, 2002), it induces the mitosis of the granulosa cells (Armstrong & Xia, 1993), and it promotes both the oocyte maturation (Lorenzo, Illera, Illera, & Illera, 1994) and the development of bovine embryos (Kaye, 1997).…”
The growth hormone (GH) and growth insulin-like factor-1 (IGF-1) act directly upon the regulation and growth in the different phases of preantral follicles. Thus, it is necessary to define their sequentiality until the in vitro preovulatory development. Therefore, the study aimed to assess the effects of a sequential medium containing GH and/or IGF-1 in the long-duration in vitro culture of preantral ovarian follicles. Ovarian fragments were cultivated: first half (days 1-7), second half (days 7-14) or during 14 culture days. Treatments were identified as: αMEM+; GH → IGF-1; IGF-1 → GH and GH + IGF-1. The culture was designed in 24-well plates, in an incubator at 37°C and 5% CO . The parameters of normality, viability, follicles (primordial/in developing) and follicle diameter were evaluated. In addition, the ultrastructure was confirmed with electron transmission microscopy. The results showed that the culture treated with GH → IGF-1 kept the follicular normality and the viability until the 14th day of culture and increased both in the follicular development until 7th day and in the follicular diameter until 14th day, when compared to the control. The treatments IGF-1 → GH and GH + IGF-1 were not effective in the developing and follicular diameter after 7 days of culture, and also reduced the percentage of viability. It is concluded that the bovine preantral follicles cultured in the sequential medium treated with GH → IGF-1 improved the follicular development until the first half of the culture and kept these parameters with normality, viability and ultrastructure until the second half of the in vitro culture.
“…; Jimenez et al. ,b. 30 follicles were evaluated through repetition (n = 6) for the total of 180 preantral follicles per treatment on day one and on day 7 of in situ culture.…”
The objective of this study was to evaluate different concentrations of growth hormone (GH) on the development of bovine preantral follicles cultured included in the ovarian tissue (in situ) on the rates of morphologically normal, viable, primordial and developing follicles, as well as the oocyte and follicle diameter and ultrastructural analysis. Ovarian fragments collected from cows with no cross-breeds defined were cultured in situ for 1 and 7 days in minimal essential medium (α-MEM+) supplemented with different concentrations of recombinant human GH (0, 10, 25, 50 ng/ml). The ovarian fragments non-cultured (control) and cultured were processed for classic histology, mechanical isolation and electron transmission microscopy (MET). The parameters underwent anova (Tukey's and Dunnett's tests) and chi-square test (χ(2) ). After 7 days of culture, the treatment with 50 ng/ml GH showed no differences with fresh control (p > 0.05) and had greater effectiveness than in the 0, 10 and 25 ng/ml GH concentrations of the morphologically normal follicles. Regarding the primordial follicles, a reduction was observed in the 50 ng/ml GH concentration concomitant with the significant increase in developing follicles, differing from both the fresh control and the other GH concentrations tested. In addition, 50 ng/ml GH showed a larger follicle and oocyte diameter when compared to the other treatments cultured. Similar structures were ultrastructurally observed in the control group, 50 ng/ml GH. Follicles cultured in 10 ng/ml GH showed nuclear invagination, vacuoles and lesioned basal membrane. Hence, it is concluded that 50 ng/ml GH is the most effective concentration for the development of preantral follicles cultured in situ.
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