Effect of IGFBP-4 during In Vitro Maturation on Developmental Competence of Bovine Cumulus Oocyte Complexes.

Insulin-like growth factors (IGFs) are essential for oocyte maturation. Their bioavailability is regulated by their respective binding proteins (IGFBPs) and proteases. IGFBP-4 blocks the biological effects of IGFs. High IGFBP-4 expression has been associated with follicle atresia. We hypothesized that IGFBP-4 affects oocyte developmental competence during maturation. Therefore, the aim of this study was to examine the effect of IGFBP-4 on the developmental rate of bovine cumulus-oocyte complexes (COCs) during in vitro embryo production. Abattoir-derived COCs were matured with rbIGFBP-4 (2000, 540, and 54 ng/mL) compared to a control. Cumulus expansion, oocyte maturation, cleavage, blastocyst, and hatching rates were evaluated. Furthermore, blastocyst gene expression of SOCS2, STAT3, SLC2A1, SLCA3, BAX, and POU5F1 transcripts were quantified using RT-qPCR. No statistical differences were detected among the groups for cumulus expansion, maturation, cleavage, blastocyst rates, or all gene transcripts analyzed. However, at day 8 and 9, the number of total hatching and successfully hatched blastocysts was lower in 2000 ng/mL rbIGFBP-4 compared to the control (day 8: total hatching: 17.1 ± 0.21 vs. 31.2 ± 0.11%, p = 0.02 and hatched blastocyst 6.7 ± 0.31 vs. 21.5 ± 0.14%, p = 0.004; day 9 total hatching 36.4 ± 0.18 vs. 57.7 ± 0.10%, p = 0.009 and hatched blastocyst 18.2 ± 0.21 vs. 38.1 ± 0.11%, p = 0.004). We concluded that high concentrations of rbIGFBP-4 might negatively affect the subsequent ability of the embryo to hatch and possibly compromise further elongation.

Melatonin improves rate of monospermic fertilization and early embryo development in a bovine IVF system.

The developmental competence of male and female gametes is frequently reduced under in vitro conditions, mainly due to oxidative stress during handling. The amino-acid derived hormone melatonin has emerged as a potent non-enzymatic antioxidant in many biological systems. The goal of the present study was to evaluate the effects of melatonin on post-thaw sperm quality, fertilizing ability, and embryo development and competence in vitro after in vitro fertilization. Frozen-thawed bovine spermatozoa were incubated either in the presence of 10-11 M melatonin (MT), or its solvent (ethanol; Sham-Control), or plain Tyrode's Albumin Lactate Pyruvate medium (TALP, Control). Computer-Assisted Sperm Analysis (CASA) and flow cytometry data after 30 min, 120 min, and 180 min incubation did not reveal any significant effects of melatonin on average motility parameters, sperm subpopulation structure as determined by hierarchical cluster, or on the percentage of viable, acrosome intact sperm, or viable sperm with active mitochondria. Nevertheless, in vitro matured cumulus-oocyte-complexes fertilized with spermatozoa which had been preincubated with 10-11 M melatonin (MT-Sperm) showed higher (P < 0.01) rates of monospermic fertilization, reduced (P < 0.05) polyspermy and enhanced (P < 0.05) embryo development compared to the Control group. Moreover, the relative abundance of MAPK13 in the in vitro-derived blastocysts was greater (P < 0.05) than observed in the Control group. In conclusion, adding melatonin to the sperm-preparation protocol for bovine IVF improved proper fertilization and enhanced embryonic development and competence in vitro.

Melatonin enhances in vitro developmental competence of cumulus-oocyte complexes collected by ovum pick-up in prepubertal and adult dairy cattle.

Bovine oocytes from prepubertal donors have been used for in vitro embryo production to decrease the generation interval. However, reduced cumulus-oocyte competence, mainly attributed to increased apoptosis, has been observed in oocytes/embryos collected from prepubertal donors. Here, we investigated the effects of the potent antioxidative molecule melatonin on cumulus-oocyte competence and embryo development in prepubertal and adult dairy cattle in vitro. A total of fifteen Holstein Friesian calves, six to ten months old (7.6 ± 1.34 months of age). And fifteen adult cows with one to four calvings (2.3 ± 0.96 calvings) were enrolled as ovum pick up (OPU) donors in this study. Cumulus-oocyte complexes (COCs) were cultured either in the presence or absence of melatonin (0.01 nM). The proportion of cleavage stages, blastocysts, and advanced blastocysts was determined. Embryo quality was assessed via differential staining to determine the total embryonic cells and allocation to the inner cell mass (ICM) and trophectoderm (TE) cells. Melatonin treatment yielded a greater percentage of blastocysts compared to the control group, i.e. oocytes from both adult cows (P = 0.0485; 24.8 ± 3.5% vs. 16.0 ± 3.4%, respectively), and from prepubertal donors (P = 0.0007; Melatonin 23.1 ± 5.1% vs. Control: 11.1 ± 3.5%). Adult cows had significantly (P = 0.0370) greater advanced blastocyst rates than those found in the prepubertal group (13.9%± vs. 7.0±%, respectively). Additionally, the number of ICM, total cells, and the ratios ICM: Total, ICM: TE, respectively, were greater (P < 0.05) after melatonin treatment compared with the control group (39.1 ± 2.8, 98.6 ± 5.7, 0.4 ± 0.01, and 0.7 ± 0.04 vs. 27.3 ± 2.9, 81.2 ± 5.8, 0.34 ± 0.01, and 0.52 ± 0.04, respectively). Blastocysts derived from adult cows had a greater number of TE (P = 0.01) and total embryonic cells (P = 0.0095) compared to the prepubertal donor group (63.5 ± 3.2 and 101.05 ± 4.8 vs. 48.9 ± 4.3 and 78.8 ± 6.5, respectively). Nevertheless, embryonic cell counting in embryos derived from prepubertal COCs equated to that observed from adult donors after melatonin exposure. In conclusion, these results indicate that the presence of melatonin during in vitro maturation improves cumulus-oocyte competence, embryo development, and quality by increasing the allocation of embryonic cells to the ICM compartment and the total number of embryonic cells in both adult and prepubertal bovine donors.

Efecto de los días postparto, predominio racial, número de partos y época del año sobre la respuesta reproductiva de vacas mestizas en anestro tratadas con un progestágeno intravaginal más ecg y pgf2a / Effect of days postpartum, breed, parity and season on reproductive performance of anestrous crossbred cows treated with an intravaginal progestagen plus ecg and pgf2a

Para evaluar el efecto de los días postparto al tratamiento (DPP), predominio racial (PR), número de partos (NP) y época del año (E) sobre la ciclicidad y fertilidad de vacas mestizas en anestro tratadas con esponjas intravaginales (EI) impregnadas con Medroxi-acetato progesterona (MAP) más una inyección de eCG y PGF2a; se condujo un experimento en una finca ubicada en un bosque húmedo-tropical en el municipio Machiques de Perijá-Zulia, Venezuela. Se seleccionaron 72 vacas mestizas Bos taurus × Bos indicus primíparas (47) y multíparas (25) entre 45 y 108 días postparto (DPP) y condición corporal (CC) entre 3 y 3,5. Las vacas recibieron el siguiente esquema hormonal: Día 0: inserción de la EI, inyección de 5 mg de 17b-Estradiol y 50 mg de MAP im; Día 6: 500 UI de eCG y 25 mg de PGF2a im; Día 8: retiro de la EI; Día 9: 1 mg de 17b-Estradiol im. Un grupo fue tratado antes de los 70 DPP (MAP < 70DPP; n = 25) y otro después de los 70 DPP (MAP > 70DPP; n = 47). Se estudió la tasa de sincronización (TS), fertilidad al primer servicio (F), fertilidad global (FG), Intervalo Tratamiento-Concepción (ITC), Intervalo Parto-Concepción (IPC) y periodo de sincronización (PS). Las variables TS, F, y FG se analizaron con PROC LOGISTIC, SAS; mientras que ITC, IPC y PS con PROC GLM, SAS. No se encontró efectos del PR y NP sobre la TS, F, FG, ITC, IPC y PS. Las vacas con PR Bos indicus presentaron mayor F y FG en el grupo MAP < 70DPP comparada con MAP > 70DPP (F: 81,8 vs 43,7%; FG: 84,6 vs 45,5%). La TS fue mayor en las vacas tratadas en la época lluviosa comparada con la seca (96,4 vs 69,8%). La F solo fue mayor en la época lluviosa cuando las vacas fueron tratadas antes de 70DPP vs MAP > 70DPP (87,5 vs 42,1%). El PS fue menor en la época seca (36,9 h) vs la época lluviosa (48,8 h). El IPC fue menor en MAP < 70DPP (95,8 días) comparado con MAP > 70DPP (148,3 días). En conclusión, los días postparto al momento de la aplicación de la EI fue el factor que más ...

An experiment was done to evaluate the effect of postpartum interval to treatment (DPP) breed (PR), number of calving (NP) and season (E) on ciclicity and fertility of anestrous crossbred cows treated with intravaginal sponges (EI) impregnated with progesterone medroxi acetate (MAP) plus im injection of eCG and PGF2a. The study was conducted in a farm located in a tropical rainforest in Machiques de Perijá, Venezuela. Seventy two primiparous (47) and multiparous (25) Bos taurus × Bos indicus crossbred cows, between 45 and 108 days postpartum (DPP) and body condition (CC) between 3.0 and 3.5. Cows were treated as follows: MAP; Day 0, EI insertion + 5 mg 17b-Estradiol and 50 mg MAP im; Day 6, 500 IU eCG and 25 mg PGF2a im; day 8 EI removed; day 9 1 mg 17b-Estradiol im. One group was treated before 70 DPP (MAP < 70DPP; n = 25) and another after 70 DPP (MAP > 70DPP; n = 47). Studied variables were: Synchronization rate (TS), first service conception rate (F), total pregnancy rate (FG), treatment to conception interval (ITC), calving to conception interval (IPC) and synchronization period (PS). Variables TS, F, and FG were analyzed with PROC LOGISTIC, SAS, while ITC, IPC and PS were analyzed with the PROC GLM, option SAS. Variables TS, F, FG, ITC, IPC and PS were not affected by PR and NP. Cows predominantly Bos indicus in MAP < 70DPP had greater (P < 0.05) F and FG than those in MAP > 70DPP (F: 81.8 vs 43.7%; FG: 84.6 vs 45.5%). The TS was greater (P < 0.01) in cows treated during the rainy season than those treated during the dry season (96.4 vs 69.8%). During the rainy season, cows in MAP < 70DPP had greater (P < 0.01) F than did those in MAP > 70DPP (87.5 vs 42.1%, respectively). The PS was shorter (P < 0.01) during the dry season (36.9 h) vs the rainy season (48.8 h). The IPC was shorter (P < 0.01) in MAP < 70DPP (95.8 days) compared to MAP > 70DPP (148.3 days). In conclusion, days to EI insertion after parturition was the most important ...

Uso del protocolo ovsynch en el control del anestro postparto en vacas mestizas de doble propósito / Using of ovsynch protocol in anoestrus control in dual purpose crossbred cows

Para evaluar el efecto del protocolo Ovsynch sobre parámetros de fertilidad en vacas mestizas doble propósito en anetsro y definir el momento optimo para la IA a tiempo fijo (IATF), 37 de 48 vacas acíclicas se asignaron a los siguientes grupos: T1: día 0: GnRH; día 7: PGF2 alfa día 9: GnRH (Ovsynch); + IATF 24 h post-última inyección de GnRH, (n=14); T2: Ovsynch + IATF 16 h post-última inyección de GnRH, (n=11)' C: control, (n=12). Las variables estudiadas fueron: Tasa de Estros Prematuros (EP), Tasa de Concepción (TC), Tasa de Preñez (TP) e Intervalo Tratamiento-Preñez (ITP). Las variables EP, TC y TP fueron medidas con el procedimiento PROC FREQ, del paquete estadístico SAS; mientras que la variable ITP se analizó mediante el procedimiento lineal general PROC GLM del SAS. La tasa de EP fue mayor (P<0,05) en Ovsynch (30,5 por ciento, 11/36) vs C (8,3 por ciento, 1/12). La Tc fue similar para EP (72,7 por ciento, 8/11) y T1 (42,8 por ciento, 6/14); pero T1 presentó una TC mayor (P=0,06) en comparación con T2 (42,8 por ciento, 6/14 vs 9,1 por ciento 1/11). No fueron encontradas diferencias para la TP en T1 vs T2 (50 por ciento, 7/14 vs 45,5 por ciento, 5/11), pero ambos grupos presentaron mayor TP comparados con C (25 por ciento, 3/12) (P<0,05). El ITP fue menor (P<0,05) en T1 (38,9 ± 15,7 días) comparado con T2 y C (62,4 ± 18,8 días y 60,0 ± 35,1 días respectivamente). En conclusión, el tratamiento Ovsynch + IATF 24 h posterior a la última inyección de GnRH resultó en un mayor TC y acortó el ITP en vacas mestizas en anestro. Se demostró que el protocolo Ovsynch representa una alternativa para el control del anestro postparto y mejoramiento de la eficiencia reproductiva de las ganaderías bovinas de doble propósito