Insulin treatment does not affect follicular development but alters granulosa cell gene expression in dairy cows.

The use of strategies to stimulate follicular growth are important, especially for use in timed artificial insemination (TAI) protocols, aiming to increase dairy cow's fertility. The aim of this study was to investigate the effect of insulin on follicular growth, steroid production and expression of genes related to follicular development. For this, cows were submitted to a progesterone (P4) and estradiol (E2) based synchronization protocol. In study 1, eleven primiparous lactating Holstein cows, received a single s.c. application of 0.25 IU/kg human insulin or no treatment (control) on D8 of the protocol. Blood samples were collected, and the dominant follicle diameter was assessed daily via transrectal ultrasonography, from D8 to D12. In study 2, eight multiparous non-pregnant and non-lactating Jersey cows, received a single s.c. application of 0.25 IU/kg human insulin, whereas cows from the control group received a single s.c. injection (1 mL) of saline solution (NaCl 0.9%). Blood samples were collected, and the dominant follicle diameter was assessed daily via transrectal ultrasonography from D6 to D9 of the protocol. Sixteen hours after insulin injection, follicular aspiration was performed. In study 1, insulin treatment decreased systemic glucose levels, but did not affect follicular growth. In study 2, the glucose decrease induced by insulin treatment was accompanied by a tendency of decreased progesterone levels in follicular fluid, along with a decrease in steroidogenic acute regulatory protein (STAR) and insulin like growth factor binding protein 2 (IGFBP2) mRNA abundance in granulosa cells. In conclusion, insulin treatment does not increase follicle growth and estradiol secretion in dairy cows, but decreases IGFBP2 and tends to increase pappalysin (PAPPA) mRNA abundance in granulosa cells, suggesting a positive effect on follicle development.

Evaluation of follicular growth and progesterone production in Bos taurus cows strategically treated with insulin / Avaliação do crescimento folicular e da produção de progesterona em vacas Bos taurus tratadas estrategicamente com insulina

The goal of this study was to evaluate the effects of strategic insulin administration on follicular growth, progesterone production, and pregnancy rate in beef cattle. Two experiments were conducted, in experiment 1, 215 cows crossbreed cows (Bos indicus × Bos taurus) (Control Group, n = 109; Insulin Group, n = 106) were submitted to fixed-time artificial insemination (TAI) with application of 2 mg of estradiol benzoate and a vaginal device containing progesterone on day 0. On day 9, the device was removed and 0.5 mg of estradiol cypionate was administered with 12.5 mg of dinoprost and 0.25 UI kg-1 of insulin for the treated animals. On day 11, artificial insemination (AI) was performed on all animals. On days 9 and 11 according to the protocol, was evaluated follicular growth and estrus manifestation, and 30 days after AI the pregnancy rate. In experiment 2, 85 cows were utilized, including Aberdeen Angus, Hereford, and crossbreeds (Control Group, n = 49 and Insulin Group, n = 36), and were submitted to the protocol for fixed-time embryo transfer (FTET), and progesterone concentrations were evaluated. On day 0, the cows received 2 mg of estradiol benzoate and a vaginal device containing progesterone. On day 5 of the protocol, 0.150 mg of cloprostenol was administered. On day 8, the vaginal device was removed and 20 mg of purified porcine pituitary extract and 0.25 UI kg-1 of insulin was given to the animals in the treatment group, and on day 9, 1 mg of estradiol benzoate was administered. On day 17, an embryo transfer was performed and blood collection for evaluation of progesterone levels in animals submitted to FTET were collected. In experiment 1, the follicular growth rate and the pregnancy rate were similar between groups (P > 0.05) and in experiment 2, the production of P4 was not different between the animals that received the insulin application and the control group...(AU)

O objetivo deste estudo foi avaliar o efeito da aplicação estratégica de insulina sobre o crescimento folicular, produção de progesterona e taxa de prenhez em bovinos de corte. Foram realizados 2 experimentos, no experimento 1, 215 vacas cruzas (Bos indicus x Bos taurus) (Grupo Controle = 109; Grupo Insulina = 106) foram submetidas a um protocolo que consistiu na aplicação de 2 mg de benzoato de estradiol e inserção de um dispositivo intravaginal de liberação lenta de progesterona (P4) sendo este o dia zero do protocolo. No dia 9, foi realizada a remoção do dispositivo de P4, a aplicação de 0,5 mg de cipionato de estradiol, e 12,5 mg de dinoprost. Os animais do grupo insulina receberam ainda a aplicação de 0,25 UI kg-1 de insulina. No dia 11 foi realizada a inseminação artificial de todos os animais. Nos dias 9 e 11 de acordo com o protocolo, foi avaliado crescimento folicular e manifestação de estro, e 30 dias após IA foi avaliado a taxa de prenhez. O experimento 2 foi realizado com 85 vacas da raça Aberdeen Angus, Hereford e cruzas (Grupo Controle = 49; Grupo Insulina = 36) submetidas a um protocolo de Transferência de Embrião em Tempo Fixo (TETF). No dia 0 fez-se aplicação de 2 mg de benzoato de estradiol e a inserção de um dispositivo intravaginal de liberação lenta de progesterona. No 5º dia do protocolo foi aplicado 0,150 mg de d- Cloprostenol. No dia 8 fez-se a remoção do dispositivo de P4 e aplicação de 20 mg de foltropina de pituitária suína e de 0,25 UI kg-1 de insulina nos animais tratados e no dia 9 foi aplicado 1 mg de benzoato de estradiol. No dia 17 procedeu-se com a transferência dos embriões e coleta de sangue para avaliação dos níveis de progesterona nos animais submetidos a TETF. No experimento 1 a taxa de crescimento folicular e a taxa de prenhez foram semelhantes entre os grupos (P > 0,05) e no experimento 2 a produção de P4 não foi diferente entre os animais que receberam a aplicação de insulina e o grupo controle...(AU)

Evaluation of follicular growth and progesterone production in Bos taurus cows strategically treated with insulin / Avaliação do crescimento folicular e da produção de progesterona em vacas Bos taurus tratadas estrategicamente com insulina

The goal of this study was to evaluate the effects of strategic insulin administration on follicular growth, progesterone production, and pregnancy rate in beef cattle. Two experiments were conducted, in experiment 1, 215 cows crossbreed cows (Bos indicus × Bos taurus) (Control Group, n = 109; Insulin Group, n = 106) were submitted to fixed-time artificial insemination (TAI) with application of 2 mg of estradiol benzoate and a vaginal device containing progesterone on day 0. On day 9, the device was removed and 0.5 mg of estradiol cypionate was administered with 12.5 mg of dinoprost and 0.25 UI kg-1 of insulin for the treated animals. On day 11, artificial insemination (AI) was performed on all animals. On days 9 and 11 according to the protocol, was evaluated follicular growth and estrus manifestation, and 30 days after AI the pregnancy rate. In experiment 2, 85 cows were utilized, including Aberdeen Angus, Hereford, and crossbreeds (Control Group, n = 49 and Insulin Group, n = 36), and were submitted to the protocol for fixed-time embryo transfer (FTET), and progesterone concentrations were evaluated. On day 0, the cows received 2 mg of estradiol benzoate and a vaginal device containing progesterone. On day 5 of the protocol, 0.150 mg of cloprostenol was administered. On day 8, the vaginal device was removed and 20 mg of purified porcine pituitary extract and 0.25 UI kg-1 of insulin was given to the animals in the treatment group, and on day 9, 1 mg of estradiol benzoate was administered. On day 17, an embryo transfer was performed and blood collection for evaluation of progesterone levels in animals submitted to FTET were collected. In experiment 1, the follicular growth rate and the pregnancy rate were similar between groups (P > 0.05) and in experiment 2, the production of P4 was not different between the animals that received the insulin application and the control group...

O objetivo deste estudo foi avaliar o efeito da aplicação estratégica de insulina sobre o crescimento folicular, produção de progesterona e taxa de prenhez em bovinos de corte. Foram realizados 2 experimentos, no experimento 1, 215 vacas cruzas (Bos indicus x Bos taurus) (Grupo Controle = 109; Grupo Insulina = 106) foram submetidas a um protocolo que consistiu na aplicação de 2 mg de benzoato de estradiol e inserção de um dispositivo intravaginal de liberação lenta de progesterona (P4) sendo este o dia zero do protocolo. No dia 9, foi realizada a remoção do dispositivo de P4, a aplicação de 0,5 mg de cipionato de estradiol, e 12,5 mg de dinoprost. Os animais do grupo insulina receberam ainda a aplicação de 0,25 UI kg-1 de insulina. No dia 11 foi realizada a inseminação artificial de todos os animais. Nos dias 9 e 11 de acordo com o protocolo, foi avaliado crescimento folicular e manifestação de estro, e 30 dias após IA foi avaliado a taxa de prenhez. O experimento 2 foi realizado com 85 vacas da raça Aberdeen Angus, Hereford e cruzas (Grupo Controle = 49; Grupo Insulina = 36) submetidas a um protocolo de Transferência de Embrião em Tempo Fixo (TETF). No dia 0 fez-se aplicação de 2 mg de benzoato de estradiol e a inserção de um dispositivo intravaginal de liberação lenta de progesterona. No 5º dia do protocolo foi aplicado 0,150 mg de d- Cloprostenol. No dia 8 fez-se a remoção do dispositivo de P4 e aplicação de 20 mg de foltropina de pituitária suína e de 0,25 UI kg-1 de insulina nos animais tratados e no dia 9 foi aplicado 1 mg de benzoato de estradiol. No dia 17 procedeu-se com a transferência dos embriões e coleta de sangue para avaliação dos níveis de progesterona nos animais submetidos a TETF. No experimento 1 a taxa de crescimento folicular e a taxa de prenhez foram semelhantes entre os grupos (P > 0,05) e no experimento 2 a produção de P4 não foi diferente entre os animais que receberam a aplicação de insulina e o grupo controle...

Injectable progesterone in timed artificial insemination programs in beef cows

The aims of this study were I) to compare the follicular diameter, corpus luteum diameter and serum progesterone (P4) concentrations in cows treated with conventional protocol vs. injectable P4 protocol; II) to determine the serum P4 profile in ovariectomized heifers; and III) to compare pregnancy rate between protocols. In experiment I, multiparous cows received a protocol for ovulation synchronization with an intravaginal P4 device (n = 38; device + EB day 0; device removal + PGF2α + eCG + EC day 8) or injectable P4 (n = 38; injection + EB day 0; PGF2α + eCG + EC day 8). In experiment II, ovariectomized heifers (n = 8) were treated with injectable P4 and blood samples were collected to determine the serum P4 profile. In experiment III, multiparous cows were timed AI with two different P4 approaches, intravaginal P4 device (n = 48) or injectable P4 (n = 47). In the first experiment, cows treated with P4 device had higher (P < 0.05) diameter of dominant follicle after ovulation induction (11.6 ± 1.8 vs.10.3 ± 1.8 mm) and ovulation rate (97%, 37/38 vs. 47.3%, 18/38) than cows treated with injectable P4. But, the follicular growth daily was higher (P < 0.05) in cows treated with injectable P4 than intravaginal device (1.3 ± 0.4 vs. 1.0 ± 0.3 mm/day, respectively). In experiment II, the P4 concentration peak occurred within 48 hours (6.54 ng/mL) and decreased after 96 hours (P < 0.05) after P4 injection. In experiment III, cows with P4 device had higher (P < 0.05) pregnancy rate than the injectable P4 group (60.4 vs. 34.0%, respectively). These results demonstrate that although the intravaginal P4 devices showed a higher pregnancy rate, a protocol with injectable P4 represents an easier method and a promising alternative for TAI in cattle.

Injectable progesterone in timed artificial insemination programs in beef cows

The aims of this study were I) to compare the follicular diameter, corpus luteum diameter and serum progesterone (P4) concentrations in cows treated with conventional protocol vs. injectable P4 protocol; II) to determine the serum P4 profile in ovariectomized heifers; and III) to compare pregnancy rate between protocols. In experiment I, multiparous cows received a protocol for ovulation synchronization with an intravaginal P4 device (n = 38; device + EB day 0; device removal + PGF2α + eCG + EC day 8) or injectable P4 (n = 38; injection + EB day 0; PGF2α + eCG + EC day 8). In experiment II, ovariectomized heifers (n = 8) were treated with injectable P4 and blood samples were collected to determine the serum P4 profile. In experiment III, multiparous cows were timed AI with two different P4 approaches, intravaginal P4 device (n = 48) or injectable P4 (n = 47). In the first experiment, cows treated with P4 device had higher (P < 0.05) diameter of dominant follicle after ovulation induction (11.6 ± 1.8 vs.10.3 ± 1.8 mm) and ovulation rate (97%, 37/38 vs. 47.3%, 18/38) than cows treated with injectable P4. But, the follicular growth daily was higher (P < 0.05) in cows treated with injectable P4 than intravaginal device (1.3 ± 0.4 vs. 1.0 ± 0.3 mm/day, respectively). In experiment II, the P4 concentration peak occurred within 48 hours (6.54 ng/mL) and decreased after 96 hours (P < 0.05) after P4 injection. In experiment III, cows with P4 device had higher (P < 0.05) pregnancy rate than the injectable P4 group (60.4 vs. 34.0%, respectively). These results demonstrate that although the intravaginal P4 devices showed a higher pregnancy rate, a protocol with injectable P4 represents an easier method and a promising alternative for TAI in cattle.(AU)

Ovarian follicular dynamics and conception rate in Bos indicus cows with different antral follicle counts subjected to timed artificial insemination.

Two experiments in Nelore cows subjected to timed artificial insemination (TAI) were designed to compare the influence of the antral follicle count (AFC/AFCs) on (1) ovarian follicular dynamics and (2) conception rates. First, multiparous cows with high (≥45 follicles; n=43) or low (≤15 follicles; n=32) AFCs were selected to undergo a TAI protocol to monitor ovarian follicular dynamics. Second, the AFCs of 962 cows also subjected to TAI were determined and classified as groups of high (G-high; ≥45 follicles; n=194), intermediate (G-intermediate; ≥20≤40 follicles, n=397) or low (G-low; ≤15 follicles; n=243) AFCs. In study I, the ovarian measurements (diameter, perimeter and area) were greater (P≤0.05) and there was a greater consistency in number of antral follicles during the period of synchronization for TAI in the high than low group. Effects of the AFC and ultrasonic evaluation interval on the follicular diameter were observed (P≤0.05); however, there was no interaction (P>0.05). Dominant follicles had greater diameters (P≤0.05) in the G-low than in the G-high at D4 (7.3±2.2 vs. 6.2±1.4mm, respectively, P=0.06), D8 (11.2±1.8 vs. 9.5±1.8mm, respectively), D9 (12.3±1.7 vs. 10.6±1.7mm, respectively), and D10 (13.4±1.3 vs. 12.2±1.8mm, respectively), as well as greater estimated diameters of ovulatory follicles (14.4±1.5 vs. 13.4±2.1mm, respectively, P=0.08). In study II, the cows with fewer AFCs had greater (P≤0.05) conception rates (61.7%) than the cows with the intermediate (52.9%) and greater (49.5%) AFCs. Nelore cows with fewer AFCs subjected to synchronization of time of ovulation had a larger follicular diameter and a greater conception rate than the groups with intermediate and a greater AFCs.

Injectable progesterone in timed artificial insemination programs in beef cows.

The aims of this study were I) to compare the follicular diameter, corpus luteum diameter and serum progesterone (P4) concentrations in cows treated with conventional protocol vs. injectable P4 protocol; II) to determine the serum P4 profile in ovariectomized heifers; and III) to compare pregnancy rate between protocols. In experiment I, multiparous cows received a protocol for ovulation synchronization with an intravaginal P4 device (n = 38; device + EB day 0; device removal + PGF2α + eCG + EC day 8) or injectable P4 (n = 38; injection + EB day 0; PGF2α + eCG + EC day 8). In experiment II, ovariectomized heifers (n = 8) were treated with injectable P4 and blood samples were collected to determine the serum P4 profile. In experiment III, multiparous cows were timed AI with two different P4 approaches, intravaginal P4 device (n = 48) or injectable P4 (n = 47). In the first experiment, cows treated with P4 device had higher (P < 0.05) diameter of dominant follicle after ovulation induction (11.6 ± 1.8 vs.10.3 ± 1.8 mm) and ovulation rate (97%, 37/38 vs. 47.3%, 18/38) than cows treated with injectable P4. But, the follicular growth daily was higher (P < 0.05) in cows treated with injectable P4 than intravaginal device (1.3 ± 0.4 vs. 1.0 ± 0.3 mm/day, respectively). In experiment II, the P4 concentration peak occurred within 48 hours (6.54 ng/mL) and decreased after 96 hours (P < 0.05) after P4 injection. In experiment III, cows with P4 device had higher (P < 0.05) pregnancy rate than the injectable P4 group (60.4 vs. 34.0%, respectively). These results demonstrate that although the intravaginal P4 devices showed a higher pregnancy rate, a protocol with injectable P4 represents an easier method and a promising alternative for TAI in cattle.

Influência do diâmetro do maior folículo na taxa de prenhez de vacas Nelore de elite submetidas à IATF / The largest follicle diameter influence on pregnancy rates of elite Nelore cows submitted to TAI

tThe objective was to evaluate the influence of the largest follicle diameter (DMF) in the TAI D10pregnancy rate of Nelore PO cows. 36 females were used, divided into two groups (≤ 10mm and> 10mm) in thefollowing protocol. In the D0 was introduced intravaginal P4 implant (DIB® 1g; Zoetis) and applied 2mL BE(Gonadiol®; Zoetis) IM. In D8 was removed the implant and applied 300 IU (1.5 mL) of eCG (Novormon®;Zoetis) and 2.0 mL of PGF2a (Ciosin®, Zoetis). In the D9 was applied 1mL BE (Gonadiol®; Zoetis) and 24hafter TAI was made. Females were subjected to ultrasound examination by transvaginal the D10 formeasurement of DMF. Pregnancy diagnosis was performed 30 days after TAI. Data were evaluated using SASsoftware (2009) and the pregnancy rate was compared using the chi-square test (P < 0.05). The pregnancy ratefor the group with DF ≤10 mm was 16.6% (6/36) and> 10 mm was 27.7% (10/36) with no statistical difference(P > 0.05).(AU)

Influência do diâmetro do maior folículo na taxa de prenhez de vacas Nelore de elite submetidas à IATF / The largest follicle diameter influence on pregnancy rates of elite Nelore cows submitted to TAI

tThe objective was to evaluate the influence of the largest follicle diameter (DMF) in the TAI D10pregnancy rate of Nelore PO cows. 36 females were used, divided into two groups (≤ 10mm and> 10mm) in thefollowing protocol. In the D0 was introduced intravaginal P4 implant (DIB® 1g; Zoetis) and applied 2mL BE(Gonadiol®; Zoetis) IM. In D8 was removed the implant and applied 300 IU (1.5 mL) of eCG (Novormon®;Zoetis) and 2.0 mL of PGF2a (Ciosin®, Zoetis). In the D9 was applied 1mL BE (Gonadiol®; Zoetis) and 24hafter TAI was made. Females were subjected to ultrasound examination by transvaginal the D10 formeasurement of DMF. Pregnancy diagnosis was performed 30 days after TAI. Data were evaluated using SASsoftware (2009) and the pregnancy rate was compared using the chi-square test (P 10 mm was 27.7% (10/36) with no statistical difference(P > 0.05).

Indução da ovulação com gonadotrofina coriônica humana em éguas Crioulas / Ovulation induction with human chorionic gonadotropin in criollo mares

The effect of age, follicular diameter and month of the breeding season (September to January) on the hCG induction of ovulation was evaluated using 123 Criollo mares. Age varied between two and 24 years and the animals were examined daily by rectal palpation and ultrasonography with a 5 MHz linear transducer. When ovarian follicles reached a diameter of 30 to 35 mm, ovulation was induced with an i.v. injection of 1000 IU (n = 39); 1500 IU (n = 41) or 2000 IU (n = 43) of hCG. The mares were bred the next day and examined daily until ovulation was detected. The percentage of mares ovulating before 24 h of hCG injection was 10.3%, 7.3% and 4.7%; until 48 h after injection 92.3%, 85.3% and 86.0% of the mares treated with 1000, 1500 and 2000 IU of hCG, respectively, ovulated. The month of the breeding season, age of the mares and follicular diameter had no influence on ovulatory response. The three hCG doses used in Criollo mares (P > 0.05) result in the induction of ovulation within 48 h after injection when a pre-ovulatory follicle with a 30 to 35 mm diameter was identified. A single dose of 1000 IU of hCG is efficient to induce ovulation in Criollo mares.

O efeito da idade, diâmetro folicular e mês da estação de monta (setembro a janeiro) na indução da ovulação com hCG foi avaliado em 123 éguas Crioulas. A idade das éguas variou entre dois e 24 anos e os animais foram examinados diariamente por palpação retal e ultrassonografia com transdutor linear de 5 MHz. Quando os folículos ovarianos atingiram diâmetro de 30 a 35 milímetros aplicou-se uma injeção intravenosa com 1000 UI (n = 39); 1500 UI (n = 41) ou 2000 UI (n = 43) de hCG. As éguas foram cobertas no dia seguinte e examinadas diariamente até a detecção da ovulação. O percentual de éguas que ovularam antes de 24 h da injeção de hCG foi de 10,3%, 7,3% e 4,7%, até 48h após a injeção foi de 92,3%, 85,3% e 86,0%, nos grupos com 1000, 1500 e 2000 UI de hCG, respectivamente. O mês da estação de monta, a idade das éguas ou o diâmetro folicular não influenciaram a resposta ovulatória. As três doses de hCG utilizadas em éguas Crioulas (P > 0,05) resultaram na indução da ovulação dentro de 48h após a aplicação, quando foi identificado um folículo pré-ovulatório de 30 a 35 mm de diâmetro. Uma única dose de 1000 UI de hCG é eficiente para induzir a ovulação em éguas Crioulas.

Indução da ovulação com gonadotrofina coriônica humana em éguas Crioulas / Ovulation induction with human chorionic gonadotropin in criollo mares

The effect of age, follicular diameter and month of the breeding season (September to January) on the hCG induction of ovulation was evaluated using 123 Criollo mares. Age varied between two and 24 years and the animals were examined daily by rectal palpation and ultrasonography with a 5 MHz linear transducer. When ovarian follicles reached a diameter of 30 to 35 mm, ovulation was induced with an i.v. injection of 1000 IU (n = 39); 1500 IU (n = 41) or 2000 IU (n = 43) of hCG. The mares were bred the next day and examined daily until ovulation was detected. The percentage of mares ovulating before 24 h of hCG injection was 10.3%, 7.3% and 4.7%; until 48 h after injection 92.3%, 85.3% and 86.0% of the mares treated with 1000, 1500 and 2000 IU of hCG, respectively, ovulated. The month of the breeding season, age of the mares and follicular diameter had no influence on ovulatory response. The three hCG doses used in Criollo mares (P > 0.05) result in the induction of ovulation within 48 h after injection when a pre-ovulatory follicle with a 30 to 35 mm diameter was identified. A single dose of 1000 IU of hCG is efficient to induce ovulation in Criollo mares.(AU)

O efeito da idade, diâmetro folicular e mês da estação de monta (setembro a janeiro) na indução da ovulação com hCG foi avaliado em 123 éguas Crioulas. A idade das éguas variou entre dois e 24 anos e os animais foram examinados diariamente por palpação retal e ultrassonografia com transdutor linear de 5 MHz. Quando os folículos ovarianos atingiram diâmetro de 30 a 35 milímetros aplicou-se uma injeção intravenosa com 1000 UI (n = 39); 1500 UI (n = 41) ou 2000 UI (n = 43) de hCG. As éguas foram cobertas no dia seguinte e examinadas diariamente até a detecção da ovulação. O percentual de éguas que ovularam antes de 24 h da injeção de hCG foi de 10,3%, 7,3% e 4,7%, até 48h após a injeção foi de 92,3%, 85,3% e 86,0%, nos grupos com 1000, 1500 e 2000 UI de hCG, respectivamente. O mês da estação de monta, a idade das éguas ou o diâmetro folicular não influenciaram a resposta ovulatória. As três doses de hCG utilizadas em éguas Crioulas (P > 0,05) resultaram na indução da ovulação dentro de 48h após a aplicação, quando foi identificado um folículo pré-ovulatório de 30 a 35 mm de diâmetro. Uma única dose de 1000 UI de hCG é eficiente para induzir a ovulação em éguas Crioulas.(AU)