Oestrus control in beef cows and heifers using cloprostenol

Se realizaron 3 experimentos con el objetivo de determinar el efecto de tratamientos alternativos para controlar el ciclo estral en un programa de servicio artificial. En el Experimento 1, 80 vaquillonas y 24 vacas secas de razas de carne fueron tratadas con 2 inyecciones de Cloprostenol con un intervalo de 11 días. Luego de la segunda inyección se hizo detección de celo e inseminación artificial durante un período de 30 días, salvo en los días 3 y 4 post.tratamiento donde se inseminó a la totalidad de los animales (I.A. sistemática). Se obtuvo 49% y 25% de preñez en vaquillonas y vacas respectivamente luego de la I.A. sistemática vs 44 y 54% a la primoinseminación en 31 vaquillonas y 11 vacas que sirvieron de testigos. En el período de 30 días se logró en vaquillonas y vacas 72% y 50% en tratadas vs. 45% y 64% en testigos. El experimento II se realizó con vaquillonas en las que se aplicó sólo una inyección de Cloprostenol luego de un período de 5 días en que se detectó celo e inseminó a los animales que lo presentaban; estos animales constituyen el lote 4 (n: 36). La inyección se aplicó a aquellos animales que no manifestaron celo en dicho período.Three trials were carried out to explore alternative ways of controlling the oestrus cycle with Cloprostenol for an Artificial Insemination (A.I.) programma. In the first trial 24 dry cows and 80 heifers were given 2 doses 11 days apart. After the second doses and over the period of 30 days oestrus detection and A.I. on animals in heat was carried out, except at 3rda. and 4th. days of this period when A.I. was applied on the animals (systematic A.I.). Pregnancy rates form systematic A.I. were 49% and 25% for heifers and cows Vs 44% and 54% at primoinsemination on control animals (31 heifers and 11 cows.). Pregnancy rates over the 30 days period were 72 % for heifers and 50% for COW5 VS 45% and 65% in the control group. The second trial used only heifers on which one Cloprostenol injection was applied after a 5 days period in which oestrus detection and A.I. on anima’s in heat were carried out. Thirty six animals in heat were separated from the main lot to form the group 4.Facultad de Ciencias Veterinaria

Oestrus control in beef cows and heifers using cloprostenol

Se realizaron 3 experimentos con el objetivo de determinar el efecto de tratamientos alternativos para controlar el ciclo estral en un programa de servicio artificial. En el Experimento 1, 80 vaquillonas y 24 vacas secas de razas de carne fueron tratadas con 2 inyecciones de Cloprostenol con un intervalo de 11 días. Luego de la segunda inyección se hizo detección de celo e inseminación artificial durante un período de 30 días, salvo en los días 3 y 4 post.tratamiento donde se inseminó a la totalidad de los animales (I.A. sistemática). Se obtuvo 49% y 25% de preñez en vaquillonas y vacas respectivamente luego de la I.A. sistemática vs 44 y 54% a la primoinseminación en 31 vaquillonas y 11 vacas que sirvieron de testigos. En el período de 30 días se logró en vaquillonas y vacas 72% y 50% en tratadas vs. 45% y 64% en testigos. El experimento II se realizó con vaquillonas en las que se aplicó sólo una inyección de Cloprostenol luego de un período de 5 días en que se detectó celo e inseminó a los animales que lo presentaban; estos animales constituyen el lote 4 (n: 36). La inyección se aplicó a aquellos animales que no manifestaron celo en dicho período.Three trials were carried out to explore alternative ways of controlling the oestrus cycle with Cloprostenol for an Artificial Insemination (A.I.) programma. In the first trial 24 dry cows and 80 heifers were given 2 doses 11 days apart. After the second doses and over the period of 30 days oestrus detection and A.I. on animals in heat was carried out, except at 3rda. and 4th. days of this period when A.I. was applied on the animals (systematic A.I.). Pregnancy rates form systematic A.I. were 49% and 25% for heifers and cows Vs 44% and 54% at primoinsemination on control animals (31 heifers and 11 cows.). Pregnancy rates over the 30 days period were 72 % for heifers and 50% for COW5 VS 45% and 65% in the control group. The second trial used only heifers on which one Cloprostenol injection was applied after a 5 days period in which oestrus detection and A.I. on anima’s in heat were carried out. Thirty six animals in heat were separated from the main lot to form the group 4.Facultad de Ciencias Veterinaria

Combined use oíreduced doses oíFSH-P and eCG as superovulatory treatment in cattle

Para evaluarla respuesta ovárica después de administrar dosis reducidas de: FSH-P, eCG o su combinación, se utilizaron 12 vacas y 24 vaquillonas (Angus, condición corporal: 6,9±0,1; x±e.e.; escala 1 a 9). Se distribuyeron aleatoriamente en tres grupos: ½FSH-P: FSH-P, 5 ó 6 mg Armour vía intramuscular (IM) y 10 u 11 mg vía subcutánea (SC) en vaquillonas y vacas, respectivamente, administrados en dosis única y simultánea. ½eCG: 1000 o 1250 UI de eCG, SC, en vaquillonas y vacas, respectivamente. FSH-P + eCG: Vaquillonas: 5 mg de FSH-P, IM, y al mismo tiempo, 1000 UI de eCG y 10 mg de FSH-P, SC. Vacas: Ídem vaquillonas con: 6 mg, 1250 UI y 11 mg, respectivamente. Se inyectó Cloprostenol a las 48 h (500 μg) y 60 h (250 μg) de comenzado el tratamiento superovulatorio [día 10,0±0,1 (x±ee) del ciclo estral]. Se inseminaron a las 60 y 72 h post Cloprostenol con semen congelado/descongelado. Siete días después se recolectaron los ovocitos/embriones. El análisis estadístico se realizó utilizando el SAS. No se observaron efectos de la categoría de animal. La respuesta superovulatoria (p<0,01) fue superior en FSH-P + eCG (100%) comparada con ½FSH-P (6,25%) y ½eCG (25%). El número de CL, embriones totales, transferiblesy congelables fue superior en FSH-P + eCG (p< 0,01); %FSH- P y %eCG no difiriendo entre sí (CL: 7,6±0,8; 1,3±0,2; 1,9±0,4; Embriones totales: 6,3±0,7; 0,06±0,06; 0,7±0,4; transferibles: 4,3±0,5; 0,06±0,06; 0,3±0,2; congelables: 3,0±0,4; 0; 0,06±0,l; x±ee; FSH-P + eCG; ½FSH-P y ½eCG, respectivamente). La presencia de un folículo dominante en crecimiento y la concentración de progesterona afectaron la respuesta superovulatoria. En conclusión, las dosis reducidas utilizadas de FSH-P o de eCG son insuficientes para producir una respuesta ovárica compatible con tratamientos superovulatorios convencionales. La presencia de un folículo dominante en crecimiento y la concentración de progesterona al inicio del tratamiento superovulatorio afectan la respuesta al mismo.To evaluated the ovarian response after administration of reduced doses of FSH-P, eCG or a combination ofboth, 12 cows and 24 heifers (Angus, body condition: 6.9 ±0.1; x±s.d.; scale 1 to 9) were assigned to three treatments: ½FSH-P: FSH-P, 5 or 6 mg Armour intramuscularly (IM) and 10 or 11 mg subcutaneously (SC) in heifers and cows, respectively, administered as a sole and simultaneous dose. ½eCG, 1000 or 1250 UI of eCG were SC administered to heifers and cows, respectively. FSH-P + eCG, heifers: 5 mg FSH-P and, at the same time, 1000 UI eCG + 10 mg FSH-P, SC. The same administration scheme was used for cows with: 6 mg, 1250 UI and 11 mg, respectively. All the animals were injected Cloprostenol at 48 h (500 μg) and 60 h (250 μg) after the beginning superovulatory treatment [day 10.0±0.1 (x±sd) of the estrous cycle]. The animals were inseminated at 60 and 72 h post Cloprostenol with frozen semen. After seven days, oocytes/embryos were collected. Statistical analyses were performed by SAS. No animal category effect was observed. The superovulatory response were greater in the FSH-P + eCG (100%) (p<0.01) than ½FSH-P (6.25%) and ½eCG (25%). The number of CL, total embryos, transferable and freezable from FSH-P + eCG was greater (p<0.01) than ½FSH-P and ½eCG; no difference was observed between the latter (CL: 7.6±0.8; 1.3±0.2; 1.9±0.4; total embryos: 6.3±0.7; 0.06±0.06; 0.7±0.4; transferable: 4.3±0.5; 0.06±0.06; 0.3±0.2; freezable: 3.0±0.4; 0; 0.06 ±0.1; x±sd; FSH-P + eCG; ½FSH-P and ½eCG, respectively). The presence of a growing dominant follicle and progesterone concentration at the beginning of the superovulatory treatment affected negatively treatment response. It is concluded that reduced doses of FSH- P or eCG utilized are insufficient to produce an ovarian response compatible with those of conventional superovulatory treatments. The presence of a growing dominant follicle and the progesterone concentration at the beginning of treatment affect the superovulatory response.Facultad de Ciencias Veterinaria

Morphological quality of in vivo oocytes recovered in diferent follicular waves from FSH-treated cows

Los objetivos del presente trabajo fueron determinar 1) el efecto de un tratamiento con FSH y 2) el período del ciclo estral (metaestro= primera onda folicular o diestro= segunda onda folicular) durante el crecimiento de la onda folicular sobre el número de folículos visualizados y aspirados y el número y la calidad morfológica de complejos cúmulo-ovocitos (COC) recuperados in vivo (ovum pick-up = OPU). Vacas Angus adultas (n= 28) se dividieron en cuatro Grupos. Grupo 1: OPU en día 4 (día 0= ovulación); Grupo 2: FSH en días 0, 1 y 2 (66,6 mg/día, Folltropin®); Grupo 3: OPU en día 13 y Grupo 4: FSH en días 9, 10 y 11 y OPU en día 13. En los Grupos 3 y 4 se realizó aspiración del folículo dominante en el Día 8 para sincronizar la segun- da onda folicular y en cada Grupo se realizaron 30 sesiones totales de OPU. En vacas tratadas con FSH, el número promedio (±EEM) de folículos aspirados fue 11,3 (±0,6) y éste fue diferente (P0,05) en el número de folículos aspira- dos entre ondas foliculares. Los números promedio de COC recuperados y viables fueron mayores (P<0,05) en el grupo FSH que en el grupo control, independientemente de la onda folicular. La tasa de recuperación de COC (COC recuperados/folículos aspirados x 100) en las vacas tratadas con FSH (35,5%) fue menor (P<0,001) que en las vacas no tratadas (70,9%). Se concluye que 1) el tratamiento con FSH permite la recu- peración de un mayor número de ovocitos y ovocitos viables por vaca, comparado con el protocolo para OPU sin el uso de FSH y 2) existe una gran variabilidad en la respuesta de estos parámetros reproductivos de acuer- do al período del ciclo estral (metaestro o diestro) en el cual se desarrolla la onda folicular, debido a la presencia o funcionalidad del CL y a las diferencias fisiológicas de las poblaciones foliculares.The objetives of the present work were determine 1) the effect of FSH treatment and 2) the estrous cycle period during the follicu- lar wave (FW) (metaestrus = first follicular wave or diestrus = second follicular wave) development on the number of visualized and aspirated follicles and number of cumulus-oocyte complexes (COC) recovered (ovum pick-up=OPU) and number of viable COC. Angus adult cows (n=28) were allocated to four treatment Groups. G1: OPU on day 4 (day 0=ovulation), G2: FSH on days 0, 1 and 2 (66.6 mg/day, Folltropin®) and OPU on day 4, G3: OPU on day 13, G4: FSH on days 9, 10 and 11 and OPU on day 13. In the Groups 3 and 4, dominant follicle aspiration was performed on Day 8 to synchronize the second follicular wave. A total of 30 OPU sessions per Group was performed. The mean (±SEM) numbers of aspirated follicles were 11.3 (±0.6) and 2.4 (±0.1) to FSH-treated cows and non-treated cows respectively. The mean number of follicles aspirated was not different (P>0.05) between follicular waves. The mean numbers of recovered and viable COC were greater (P<0.05) in FSH-treated cows than those in non-treated cows regardless of FW. Oocytes recovery rate (recovered COC/aspirated follicles x 100) in FSH-treated cows (35.5%) was lower (P<0.001) than that in non-treated cows (70.9%) regardless the FW. We can conclude that 1) FSH treat- ment increase the number of COC and viable COC in vivo recovered by means of OPU and 2) a great variability in the response of these reproductive parameters was observed, according to the period of the estrous cycle (metaestrus or diestrus) in which the FW is developed, due to the presence or functionality of corpus luteum and the physiological differences of the follicular populations.EEA BalcarceFil: Aller Atucha, Juan Florencio. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce; Argentina.Fil: Callejas, Santiago. Universidad Nacional del Centro de la Provincia de Buenos Aires, Facultad de Ciencias Veterinarias; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina.Fil: Alberio, Ricardo . Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce; Argentina. Universidad Nacional de Mar del Plata, Facultad de Ciencias Agrarias; Argentina

Does seminal plasma improves the fertility of cryopreserved ram semen?

El objetivo de este artículo es proporcionar una revisión actualizada del rol del plasma seminal y de sus componentes proteicos sobre la capacidad fecundante del semen ovino criopreservado. En la especie ovina la inseminación artificial se realiza principalmente por la vía cervical mediante el uso de semen fresco/refrigerado, ya que el empleo de semen congelado/descongelado por la misma vía genera bajos porcentajes de preñez. Con el propósito de mejorar la calidad espermática posdescongelación y alcanzar tasas de preñez similares a las obtenidas con semen fresco se han desarrollado numerosos estudios relacionados al efecto del plasma seminal sobre la calidad y funcionalidad espermática. Los resultados de las investigaciones han demostrado que el plasma seminal y sus proteínas de bajo peso molecular son capaces de mejorar parámetros cualitativos de los espermatozoides criopreservados. Sin embargo, los estudios sobre el efecto del plasma seminal en la funcionalidad espermática siguen siendo contradictorios.The aim of the present article is to provide an updated review about the role of seminal plasma and its protein components on the fertilizing capacity of cryopreserved ram semen. Artificial insemination in sheep is performed cervically using fresh/refrigerated semen since the use of frozen/thawed by the same trial causes low pregnancy rates. With the objective of improving sperm posthawing quality and reach pregnancy rates similar to those obtained with fresh semen, seminal plasma has been the subject of numerous studies. Research results have shown that seminal plasma and their low molecular weight proteins are able to improve qualitative parameters of ram cryopreserved sperm. However, studies on the effect of PS on sperm function remaining contradictory.Fil: Ledesma, Alba. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata; Argentina. Universidad Nacional de Mar del Plata. Facultad de Ciencias Agrarias; ArgentinaFil: Manes, J.. Instituto Nacional de Tecnología Agropecuaria. Centro Regional Buenos Aires Sur. Estación Experimental Agropecuaria Balcarce; ArgentinaFil: Alberio, Ricardo. Instituto Nacional de Tecnología Agropecuaria. Centro Regional Buenos Aires Sur. Estación Experimental Agropecuaria Balcarce; ArgentinaFil: Hozbor, Federico Andrés. Instituto Nacional de Tecnología Agropecuaria. Centro Regional Buenos Aires Sur. Estación Experimental Agropecuaria Balcarce; Argentin

A dose-dependent response to MEK inhibition determines hypoblast fate in bovine embryos

Background: The segregation of the hypoblast and the emergence of the pluripotent epiblast mark the final stages of blastocyst formation in mammalian embryos. In bovine embryos the formation of the hypoblast has been partially studied, and evidence shows that MEK signalling plays a limited role in the segregation of this lineage. Here we explored the role of different signalling pathways during lineage segregation in the bovine embryo using immunofluorescence analysis of NANOG and SOX17 as readouts of epiblast and hypoblast, respectively. Results We show that SOX17 starts to be expressed in 16–32-cell stage embryos, whereas NANOG is first detected from 8-cell stage. SOX17 is first co-expressed with NANOG, but these markers become mutually exclusive by the late blastocyst stage. By assessing the expression kinetics of NANOG/SOX17 we show that inhibition of MEK signalling can eliminate SOX17 expression in bovine blastocysts, without altering NANOG expression. Modulation of WNT, PKC and LIF did not affect NANOG expression in the epiblast when used in combination with the ERK inhibitor. Conclusions This study shows that SOX17 can be used as a reliable early marker of hypoblast in the bovine, and based on its expression profile we show that the hypoblast segregates in day 7 blastocysts. Furthermore, SOX17 expression is abolished using 1 μM of PD0325901, without affecting the NANOG population in the epiblast. Modulation of WNT, PKC and LIF are not sufficient to support enhanced NANOG expression in the epiblast when combined with ERK inhibitor, indicating that additional signalling pathways should be examined to determine their potential roles in epiblast expansion.EEA BalcarceFil: Canizo, Jesica Romina. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce; Argentina. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina.Fil: Ynsaurralde Rivolta, Amada Eugenia. Universidad de Buenos Aires. Facultad de Agronomía; Argentina. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Mercedes; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Vazquez Echegaray, Camila. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Suvá, Mariana. Universidad de Buenos Aires. Facultad de Agronomía; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Alberio, Virgilia. Universidad de Buenos Aires. Facultad de Agronomía; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Aller, Juan Florencio. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce; Argentina.Fil: Guberman, Alejandra S. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina.Fil: Salamone, Daniel.F. Universidad de Buenos Aires. Facultad de Agronomía; Argentina.Fil: Alberio, Ricardo H. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce; Argentina. Universidad Nacional de Mar del Plata. Facultad de Ciencias Agrarias; Argentina.Fil: Alberio, Ramiro. University of Nottingham. School of Biosciences; Reino Unid

Electroejaculation Increases Low Molecular Weight Proteins in Seminal Plasma Modifying Sperm Quality in Corriedale Rams

This study was conducted to evaluate the effect of seminal collection method (artificial vagina or electroejaculation) on the protein composition of seminal plasma and sperm quality parameters in Corriedale rams. To address this question, we assessed the effect of seminal collection method on motility, plasma membrane integrity and functionality, mitochondrial functionality and the decondensation state of nuclear chromatin in sperm cells. Volume, pH, osmolarity, protein concentration, total protein content and protein profile using sodium dodecyl sulphate–polyacrylamide gel electrophoresis (SDS‐PAGE) and 2‐D polyacrylamide electrophoresis of seminal plasma collected with artificial vagina and electroejaculation were also analysed. The main findings from this study were that ejaculates obtained with electroejaculation had (i) a higher number of spermatozoa with intact plasma membrane and functional mitochondria and (ii) a higher proportion of seminal plasma, total protein content and relative abundance of low molecular weight proteins than ejaculates obtained with artificial vagina. Five of these proteins were identified by mass spectrometry: binder of sperm 5 precursor; RSVP14; RSVP22; epididymal secretory protein E1 and clusterin. One protein spot with molecular weight of approximately 31 kDa and isoelectric point of 4.8 was only found in the seminal plasma from electroejaculation.EEA BalcarceFil: Ledesma, Alba. Universidad Nacional de Mar del Plata. Facultad de Ciencias Agrarias; ArgentinaFil: Manes, Jorgelina. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce. Departamento de Producción Animal. Biotecnología de la Reproducción; Argentina.Fil: Cesari, Andreina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; ArgentinaFil: Alberio, Ricardo. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce. Departamento de Producción Animal. Biotecnología de la Reproducción; ArgentinaFil: Hozbor, Federico Andres. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce. Departamento de Producción Animal. Biotecnología de la Reproducción; Argentin

Exogenous human OKSM factors maintain pluripotency gene expression of bovine and porcine iPS-like cells obtained with STEMCCA delivery system

Abstract Objectives The use of induced pluripotent stem (iPS) cells as an alternative to embryonic stem cells to produce transgenic animals requires the development of a biotechnological platform for their generation. In this study, different strategies for the generation of bovine and porcine iPS cells were evaluated. Lentiviral vectors were used to deliver human factors OCT4, SOX2, KLF4 and c-MYC (OKSM) into bovine and porcine embryonic fibroblasts and different culture conditions were evaluated. Results Protocols based on the integrative lentiviral vector STEMCCA produced porcine iPS-like cells more efficiently than in bovine cells. The iPS-like cells generated displayed stem cell features; however, expression of exogenous factors was maintained along at least 12 passages. Since inactivation of the exogenous factors is still a major bottleneck for establishing fully reprogrammed iPS cells, defining culture conditions that support endogenous OKSM expression is critical for the efficient generation of farm animals’ iPS cells

Free cholesterol and cholesterol esters in bovine oocytes: Implications in survival and membrane raft organization after cryopreservation.

Part of the damage caused by cryopreservation of mammalian oocytes occurs at the plasma membrane. The addition of cholesterol to cell membranes as a strategy to make it more tolerant to cryopreservation has been little addressed in oocytes. In order to increase the survival of bovine oocytes after cryopreservation, we proposed not only to increase cholesterol level of oocyte membranes before vitrification but also to remove the added cholesterol after warming, thus recovering its original level. Results from our study showed that modulation of membrane cholesterol by methyl-β-cyclodextrin (MβCD) did not affect the apoptotic status of oocytes and improved viability after vitrification yielding levels of apoptosis closer to those of fresh oocytes. Fluorometric measurements based on an enzyme-coupled reaction that detects both free cholesterol (membrane) and cholesteryl esters (stored in lipid droplets), revealed that oocytes and cumulus cells present different levels of cholesterol depending on the seasonal period. Variations at membrane cholesterol level of oocytes were enough to account for the differences found in total cholesterol. Differences found in total cholesterol of cumulus cells were explained by the differences found in both the content of membrane cholesterol and of cholesterol esters. Cholesterol was incorporated into the oocyte plasma membrane as evidenced by comparative labeling of a fluorescent cholesterol. Oocytes and cumulus cells increased membrane cholesterol after incubation with MβCD/cholesterol and recovered their original level after cholesterol removal, regardless of the season. Finally, we evaluated the effect of vitrification on the putative raft molecule GM1. Cholesterol modulation also preserved membrane organization by maintaining ganglioside level at the plasma membrane. Results suggest a distinctive cholesterol metabolic status of cumulus-oocyte complexes (COCs) among seasons and a dynamic organizational structure of cholesterol homeostasis within the COC. Modulation of membrane cholesterol by MβCD improved survival of bovine oocytes and preserved integrity of GM1-related rafts after vitrification

Combined epidermal growth factor and hyaluronic acid supplementation of in vitro maturation medium and its impact on bovine oocyte proteome and competence

The conditions for in vitro oocyte maturation impact on cytoplasmic and nuclear processes in the oocyte. These events are differentially influenced by the nature of the maturation inducer and the presence of intact cumulus in cumulus–oocyte complexes. Epidermal growth factor is the main growth factor promoting oocyte maturation. Also, hyaluronic acid (HA) produced by cumulus cells is known to be responsible for the correct structural and functional organization of the cumulus during oocyte maturation. Therefore, we evaluated the developmental competence of bovine oocytes matured in vitro in a maturation medium supplemented with both EGF and HA, compared to FSH and fetal bovine serum (FBS). In addition, the impact of IVM conditions on the proteomic profile of metaphase II bovine oocytes was analyzed by two-dimensional electrophoresis. Cumulus–oocyte complexes were matured in two media: (1) 10 ng/mL EGF, 15 μg/mL HA, and 100-μM cysteamine and (2) 0.01 UI/mL rh-FSH and 10% FBS. The percentages of first polar body and embryo production and the kinetics of embryo development and oocyte proteomic profiles were analyzed. Oocytes matured in the presence of EGF-HA showed an increase (6%, P < 0.05) in the percentage of polar body extrusion. The blastocyst rate was 3% (P < 0.05) higher in the FSH-FBS group, but no differences were found in the rate of expanded blastocyst neither in total embryo production between IVM conditions. Cleavage rate of oocytes matured with FSH-FBS was 5% higher (P < 0.05) with respect to EGF-HA–matured oocytes when evaluated 30 hours after fertilization. However, at Day 7, those inseminated oocytes that underwent division at a correct timing showed that although there are still early blastocysts in the FSH-FBS condition, EGF-HA embryos have developed completely into blastocysts. Still, the production rate of those embryos that achieved expansion was similar between both maturation conditions. On the other hand, noncleaved presumptive zygotes at Day 7 developed into the different stages with similar rates (∼4%) independently of the medium condition. Modifications of IVM medium composition markedly affected protein profile of bovine oocytes in a differential manner. The proteomic approach revealed the presence of 68 spots in both treatments, 41 exclusively found in the FSH-FBS group and 64 exclusive for the EGF-HA group. Taken together, these results indicate that combined EGF-HA supplementation of in vitro maturation medium could be used to improve oocyte meiotic competence and ensure a better timing to develop into the blastocyst stage.EEA BalcarceFil: Rios, Glenda Laura. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce. Departamento de Producción Animal. Biotecnología de la Reproducción; ArgentinaFil: Buschiazzo, Jorgelina. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce. Departamento de Producción Animal. Biotecnología de la Reproducción; Argentina. Consejo Nacional de Investigaciones Cientificas y Técnicas. Centro Científico Tecnológico Bahia Blanca. Instituto de Investigaciones Bioquímicas Bahia Blanca; ArgentinaFil: Mucci, Nicolas Crescencio. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce. Departamento de Producción Animal. Biotecnología de la Reproducción; ArgentinaFil: Kaiser, German Gustavo. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce. Departamento de Producción Animal. Biotecnología de la Reproducción; ArgentinaFil: Cesari, Andreina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; ArgentinaFil: Alberio, Ricardo. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce. Departamento de Producción Animal. Biotecnología de la Reproducción; Argentin