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1.
J Therm Biol ; 96: 102842, 2021 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-33627280

RESUMO

Heat stress reduces the reproductive capacity of bulls raised in tropical climate. However, the reestablishment of scrotal thermoregulation and the dynamics of sperm defects emergence after stress are not completely known in buffaloes. Thus, the study aimed to evaluate the effect of short-term heat stress over scrotal thermoregulation and sperm attributes, relating them to spermatogenesis stages. Five buffalo bulls went through scrotal insulation during 48 h (from day 0 to day 2). Semen samples were collected every 7 days (from day -7 to day 49) and analyzed about the progressive motility, viability, and sperm morphology. Heat stress significantly destabilized scrotal thermoregulation (P < 0.001). Scrotal temperature was from 4.2 to 6.3 °C lower than the core body temperature, except on insulation days (P < 0.001), and returned to the basal condition five days after the removal of the stressing stimulus. More significant deleterious effects were observed in sperm morphology than in cell concentration, motility, and viability. The chronology of morphologic defects expression demonstrated tail defects (days 7-14), cytoplasmic droplets (days 14-28), and head defects (day 28), returning to pre-insulation condition 35 days after the thermal challenge. Thus, hyperthermia harmed more intensely spermatozoa in epididymal transit, elongated spermatids, and secondary spermatocytes. It is concluded that water buffalo bulls present a peculiar manifestation of sperm morphology after short-term stress, indicating an important difference related to the bovine species. Therefore, during the andrological evaluation of buffalo bulls, it is necessary to avoid the allometric extrapolation between these species.

2.
Zygote ; : 1-6, 2021 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-33448260

RESUMO

Sex selection through sperm sorting offers advantages in regards selection pressure in high-producing livestock. However, the sex-sorting process results in sperm membrane and DNA damage that ultimately decrease fertility. We hypothesized that given the role of protamines in DNA packaging, protamine deficiency could account, at least partially, for the DNA damage observed following sperm sex sorting. To test this, we compared protamine status between unsexed and sexed spermatozoa from two bulls using the fluorochrome chromomycin A3 (CMA3) and flow cytometry. Then, we assessed embryo development following in vitro fertilization (IVF) using the same sperm treatments. Overall, sperm protamination was not different between sexed and unsexed semen. However, one of the two bulls displayed higher rates of protamine deficiency for both unsexed and sexed semen (P < 0.05). Moreover, unsexed semen from this bull yielded lower blastocyst (P < 0.05) and blastocyst hatching rates than unsexed sperm from the other bull. CMA3-positive staining was negatively correlated with cleavage (R2 85.1, P = 0.003) and blastocyst hatching (R2 87.6, P = 0.006) rates in unsexed semen. In conclusion, while the sex-sorting process had no effect on sperm protamine content, we observed a bull effect for sperm protamination, which correlated to embryo development rates following IVF.

3.
Theriogenology ; 158: 382-390, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-33038824

RESUMO

Consequences of oocyte supplementation with l-carnitine may vary depending on species-specific cellular lipid profile, level of mitochondrial activity, or even on ipid availability in culture medium. This study aimed to evaluate l-carnitine supplementation on competence and gene expression of enzymes related to lipid metabolism in oocytes and cumulus cells from buffalo COCs matured in the presence or absence of fetal bovine serum (FBS). COCs were matured in vitro in FBS (10%) or bovine serum albumin fatty acid-free (BSA-FAF) (0.4%) and with or without supplementation with l-carnitine (3.03 mM). COCs matured in the presence of FBS or BSA-FAF were fertilized and cultured, then supplemented with l-carnitine during in vitro maturation or in vitro embryo culture. Finally, in vivo mature and immature COCs were included for gene expression analysis. COCs matured in culture medium with FBS in the presence of l-carnitine produced a lower blastocyst rate (p ≤ 0.05) compared to controls. In turn, the blastocyst rate from COCs matured with BSA-FAF in the presence of l-carnitine was similar to controls (p > 0.05), and higher than FBS + L-carnitine treated COCs (p ≤ 0.05). Addition of l-carnitine during embryo culture showed no differences in blastocyst production between experimental groups and controls (p > 0.05). In cumulus cells, gene expression of ACACA, SCD and FASN was upregulated in COCs matured in the presence of BSA-FAF + L-carnitine, while all genes in oocytes were significantly expressed upregulated by COCs matured in vivo, and only BSA-FAF + L-carnitine group showed similar expression of the FASN gene. In conclusion, the consequences of l-carnitine supplementation during in vitro maturation of buffalo COCs on oocyte competence vary depending on presence or absence of FBS in culture. With FBS, l-carnitine impairs oocyte competence, while in its absence, gene expression suggests adequate lipid metabolism and increased oocyte competence.

4.
Anim Reprod Sci ; 211: 106220, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31785645

RESUMO

The knowledge about the biological events that regulate lipid metabolism in oocytes and embryos in buffalo is scarce. Lipogenesis, lipolysis, transport and oxidation of fatty acids (FAs) occur in gametes and embryonic cells of all mammalian species, as an intrinsic component of energy metabolism. In oocytes and cumulus cells, degradation of lipids is responsible for the production of ATP that is essential for the metabolic processes that lead to oocyte maturation in in vivo and in vitro culture conditions. Similarly, throughout embryo development, blastomeres have the capacity to use exogenous and/or endogenous lipid reserves to serve as an energy source necessary for early embryonic development. In addition, supplementation of culture media with L-carnitine to promote lipid metabolism during in vitro oocyte maturation and early embryonic development leads to an improved embryo quality. The limited scientific evidence available in buffalo indicates there is relatively greater oocyte lipid content as compared with many other species that undergoes a dynamic distribution during folliculogenesis and follicle maturation and that has a positive effect on oocyte maturation and embryo development when there is L-carnitine supplementation of the media. Advances in the understanding of the biological peculiarities of lipid metabolism, and the consequences of its alteration on the quality of buffalo gametes and embryos, therefore, are necessary to design specific culture media and laboratory procedures as a strategy to increase in vitro-derived embryo production rates.


Assuntos
Búfalos/fisiologia , Técnicas de Cultura Embrionária/veterinária , Técnicas de Maturação in Vitro de Oócitos/veterinária , Metabolismo dos Lipídeos/fisiologia , Oócitos/fisiologia , Animais , Búfalos/embriologia
5.
Anim Reprod ; 16(2): 260-266, 2019 Oct 23.
Artigo em Inglês | MEDLINE | ID: mdl-33224285

RESUMO

Transvaginal follicular aspiration technique together with in vitro embryo production are the biotechnological alternatives currently available to support genetic improvement breeding programs in buffalo species. However, aspects related to animal management, lack of knowledge of the metabolic needs and biochemical peculiarities of gametes and embryos, as well as the reproductive physiology characteristics have hampered progress in the results. Despite the low availability of good quality oocytes collected after OPU in donors as a physiological characteristic of buffalo species, high rates of oocyte maturation, modest embryo cleavage, blastocyst production and pregnancy rates after transvaginal embryo transfer in recipients could be obtained in buffalo in vitro embryo production programs. The results of implementing an in vitro embryo production program in buffaloes in the northern region of Pará state, Brazil, and results published by other groups demonstrate the feasibility of implementing this biotechnology in the routine of breeding programs. Nevertheless, in order to achieve better and consistent results, it is necessary to deepen the knowledge on the peculiarities of reproductive biology in this specie. Selection of donor animals based on ovarian size and ovarian follicular reserve and on the rate of blastocyst production is presented as an effective alternative to increase the efficiency of the in vitro embryo production technique applied to the buffalo species.

6.
Anim Reprod ; 16(2): 249-259, 2019 Oct 24.
Artigo em Inglês | MEDLINE | ID: mdl-33224284

RESUMO

In Vitro Embryo Production (IVP) is widely used to improve the reproductive efficiency of livestock animals, however increasing the embryo development rates and pregnancy outcomes is still a challenge for some species. Thus, the lack of biological knowledge hinders developing specie-specific IVP protocols. Therefore, the contributions of RNA-seq to generate relevant biological knowledge and improve the efficiency of IVP in livestock animals are reviewed herein.

7.
Acta Cir Bras ; 33(5): 431-438, 2018 May.
Artigo em Inglês | MEDLINE | ID: mdl-29924207

RESUMO

PURPOSE: To evaluate the effects of this thymol-rich oil in the proliferation of human adipose tissue-derived stem cells. METHODS: Stem cells were isolated from human adipose tissue by liposuction. After the first passage, cells were cultivated in triplicate for three days in control medium and medium supplemented with three oil samples (1.0 µg/mL, 5.0 µg/mL, and 25.0 µg/mL). Cells were analyzed by the MTT assay at passage 1 (P1), and cell proliferation of control and 1 µg/mL groups was determined with a hemocytometer at P2 and P3. RESULTS: Viability of the essential oil-treated cells was significantly higher than the control group at P1 (p = 0.0008). The treatment with the oil, at a concentration of 1 µg/mL, led to increases of 24.8% at P1 and 43.0% at P3 in the rate of cell proliferation compared with control cells. CONCLUSION: Supplementing culture medium with essential oil of Lippia origanoides increased cell proliferation, especially at later passages.


Assuntos
Antioxidantes/farmacologia , Proliferação de Células/efeitos dos fármacos , Lippia/química , Óleos Voláteis/farmacologia , Óleos Vegetais/farmacologia , Timol/farmacologia , Tecido Adiposo/citologia , Adulto , Meios de Cultura , Humanos , Lipectomia , Óleos Voláteis/química , Óleos Vegetais/química , Células-Tronco/efeitos dos fármacos
8.
Acta cir. bras ; 33(5): 431-438, May 2018. tab, graf
Artigo em Inglês | LILACS | ID: biblio-949342

RESUMO

Abstract Purpose: To evaluate the effects of this thymol-rich oil in the proliferation of human adipose tissue-derived stem cells. Methods: Stem cells were isolated from human adipose tissue by liposuction. After the first passage, cells were cultivated in triplicate for three days in control medium and medium supplemented with three oil samples (1.0 μg/mL, 5.0 μg/mL, and 25.0 μg/mL). Cells were analyzed by the MTT assay at passage 1 (P1), and cell proliferation of control and 1 μg/mL groups was determined with a hemocytometer at P2 and P3. Results: Viability of the essential oil-treated cells was significantly higher than the control group at P1 (p = 0.0008). The treatment with the oil, at a concentration of 1 µg/mL, led to increases of 24.8% at P1 and 43.0% at P3 in the rate of cell proliferation compared with control cells. Conclusion: Supplementing culture medium with essential oil of Lippia origanoides increased cell proliferation, especially at later passages.


Assuntos
Humanos , Adulto , Timol/farmacologia , Óleos Vegetais/farmacologia , Óleos Voláteis/farmacologia , Lippia/química , Proliferação de Células/efeitos dos fármacos , Antioxidantes/farmacologia , Células-Tronco/efeitos dos fármacos , Óleos Vegetais/química , Óleos Voláteis/química , Lipectomia , Tecido Adiposo/citologia , Meios de Cultura
9.
Cell Reprogram ; 19(5): 294-301, 2017 10.
Artigo em Inglês | MEDLINE | ID: mdl-28832180

RESUMO

Somatic cell nuclear transfer (SCNT) success is partially hindered by the low epigenetic reprogramming efficiency of the donor cell. Previous studies suggest cellular heterogeneity among donor nuclei in regard to reprogramming potential, which precludes comparison among different strategies to increase cloning success. In this context, we evaluated the effect of using clonal cell populations (CPs) of bovine adult fibroblasts established by single-cell plating in SCNT. Different CPs were evaluated in regard to proliferation rate, senescence level, and chromosome stability, as well as for POU5F1 (POU class 5 homeobox 1) mRNA expression levels. In total, 9 of 24 CPs (37.5%) were successfully expanded in vitro up to the fourth passage and shown to proliferate following cryopreservation, at which time cell analyses were performed. The use of a CP with low senescence level, normal karyotype, and highest POU5F1 expression levels did not improve embryo development rates or quality following SCNT. As previously suggested, this study supports the notion that levels of POU5F1 expression in the donor nucleus do not impact the SCNT results. Notably, the single-cell seeding approach used herein to isolate CPs may be extended to the evaluation of additional predictor markers of reprogrammability success for SCNT in future experiments.


Assuntos
Embrião de Mamíferos/embriologia , Desenvolvimento Embrionário , Fibroblastos/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Técnicas de Transferência Nuclear , Fator 3 de Transcrição de Octâmero/biossíntese , Animais , Bovinos , Células Cultivadas , Embrião de Mamíferos/citologia , Feminino , Fibroblastos/citologia , Masculino , Fator 3 de Transcrição de Octâmero/genética
10.
Acta cir. bras ; 31(12): 821-825, Dec. 2016. tab, graf
Artigo em Inglês | LILACS | ID: biblio-837663

RESUMO

ABSTRACT PURPOSE: To evaluate the Adipose Stem Cells (ACS) therapy efficacy on the time and quality of wound healing process in rats. METHODS: Nine male Wistar rats were randomly distributed into three groups I) 7 days of healing; II) 14 days of healing; III) 21 days of healing. Four incisions were made on the dorsal surface of each rat and then treated with intralesional ACS, meloxicam, and no treatment and ACS+meloxicam. Macroscopic evaluation was measured by percentage of healing and histopathological by hematoxylin-eosin was performed. RESULTS: All groups have the wound reduced during the three weeks (p<0.001) and after 14 days of healing had greater reduction than others. Wounds treated with ASC had accelerated healing in relation to no treatment and only meloxicam (p<0.001), excepting the ASC+Meloxicam that was similar (p=0.13). There was no difference in histopathological analysis between lesions. CONCLUSION: Adipose stem cell have benefits in reducing time of healing of experimental model of wound in rats, observed 7 days of after application.


Assuntos
Animais , Masculino , Ratos , Células-Tronco/fisiologia , Cicatrização/fisiologia , Ferimentos e Lesões/terapia , Tecido Adiposo/citologia , Transplante de Células-Tronco , Resultado do Tratamento , Ratos Wistar , Modelos Animais de Doenças , Antígenos de Superfície
11.
Rev. biol. trop ; 64(4): 1519-1526, oct.-dic. 2016. ilus
Artigo em Inglês | LILACS-Express | LILACS | ID: biblio-958231

RESUMO

Abstract:The agouti (Dasyprocta sp.) is a hystricomorph rodent found in some regions of the Americas. It is an important cynegetic species, which indicates that the overhunting is a threat to their conservation. Very little is known about this wildlife in relation to what already has been studied in domestic animals. Thus, the knowledge on reproduction of wildlife becomes necessary and essential for the management and conservation of these natural resources. Specifically, studies regarding hormonal monitoring are important as a basic tool for research in modern reproductive biotechnology, and currently, there is no information on the progesterone changes during pregnancy of Dasyprocta sp., compared to other hystricomorphs. The aim of this study was to describe the profile of plasmatic progesterone during pregnancy, and report the restart of ovarian cycle in agouti after parturition. For this purpose, 18 black-rumped agoutis (Dasyprocta prymnolopha) born in captivity were used, and one or more consecutive pregnancies were observed. Copulation was confirmed by the presence of spermatozoa observed in the colpocytological examination. Blood samples were collected two times per week, and concentrations of progesterone, assessed in ten agoutis, were determined by radioimmunoassay. The onset of ovarian activity in six females was observed daily by colpocytological examination, starting on the seventh day postpartum. The gestational period observed in this study was 104.04 days (SD = 1.31) (101-106 days) (Number of cases, N = 26), and the interval between births was 126.03 days (SD = 18.40) (109-184 days). The plasmatic profile of progesterone during pregnancy showed a progressive increase from the 1st to the 5th week. The higher progesterone levels over this period (6.88 ng / mL, SD = 3.01) were detected in the 5th week. This value was similar (One-Way ANOVA, p > 0.05) to that observed in the 4th and 6th weeks, but was statistically different (One-Way ANOVA, p < 0.05) when compared to the other weeks. After the 6th week there was a progressive decrease in plasmatic progesterone levels. The animals showed a postpartum estrus of 12.04 days (SD= 4.29) (7-24 days) (N= 23). It was observed that 80.95 % (N= 19) of copulations during this period were fertile. This work contributed to understanding the dynamic changes in the progesterone levels during the pregnancy in agouti. Nevertheless, more studies are needed for a better appreciation of other endocrine and biological changes, in the mother and feto-placental unit of the agouti. Rev. Biol. Trop. 64 (4): 1519-1526. Epub 2016 December 01.


Resumen:El agouti (Dasyprocta sp.) es un roedor histricomorfo encontrado en algunas regiones de América. Es una especie cinegética importante, lo que indica que la caza excesiva es una amenaza para su conservación. Muy poco es lo que se conoce acerca de la biología de los animales silvestres, en relación al conocimiento acumulado sobre los domésticos. Por lo tanto, el estudio sobre la reproducción de los animales silvestres se hace necesario e imprescindible para el manejo y conservación de la especie como recurso natural. En concreto, los estudios relativos a la monitorización hormonal son una herramienta básica para la investigación en biotecnología reproductiva moderna. No hay información sobre los cambios de progesterona durante la preñez de Dasyprocta sp. en comparación con otros histricomorfos. El objetivo de este estudio fue describir el perfil de la progesterona plasmática durante la preñez, y reportar el reinicio del ciclo ovárico después del parto. Para ello, se utilizaron 18 agoutis de rabo negro (Dasyprocta prymnolopha) nacidos en cautiverio. Fueron estudiadas una o más preñeces consecutivas. La copulación fue confirmada por la presencia de espermatozoides observados en examen colpocitológico. Las muestras de sangre se recogieron dos veces por semana, y las concentraciones de progesterona se determinaron en 10 individuos mediante radioinmunoensayo. Se observó el inicio de la actividad ovárica diaria por examen colpocitológico, comenzando en el día séptimo posparto. El período gestacional observado en este estudio fue de 104.04 días (SD= 1.31, rango entre 101-106 días, N= 26), y el intervalo entre los nacimientos fue 126.03 días (SD= 18.40, rango entre 109-184 días). El perfil plasmático de la progesterona durante el embarazo mostró un aumento progresivo de la 1ª a la 5ª semana. Los mayores niveles de progesterona durante este período (6.88 ng / mL, SD= 3.01) se detectaron en la quinta semana. Este valor fue similar (ANOVA de um factor, p > 0.05) al observado en las semanas cuatro y seis, pero fue estadísticamente diferente (ANOVA de um factor, p<0.05) en comparación con las otras semanas. Después de la sexta semana hubo una disminución progresiva en los niveles plasmáticos de progesterona. Los animales mostraron un estro posparto de 12.04 días (SD= 4.29, rango entre 7-24 días, N= 23). Se observó que el 80.95 % (N= 19) de cópulas durante este período fueron fértiles. Este trabajo contribuyó a la comprensión de los cambios dinámicos en los niveles de progesterona durante la preñez del agouti. Sin embargo, se necesitan más estudios para una mejor comprensión de otros cambios biológicos y endocrinos, y en la unidad materna y feto-placentaria del agouti.

12.
Behav Processes ; 131: 68-73, 2016 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-27553126

RESUMO

The goal of this study was to describe the sexual behavior in female and male collared peccary. Twenty females and twelve males were monitored in familiar and non-familiar units for two 60 days periods. During both phases, we recorded 2747 sexual interactions initiated by 20 different females toward males and 4461 sexual interactions initiated by 12 males toward females. The frequency of sexual interactions initiated per female significantly increased from proestrus to estrus, and they were significantly more frequently courted. Females initiated olfactory inspections 15.42 times more and were mounted 22.6 times more during estrus than during proestrus. Nulliparous and primiparous females copulated only when exposed to non-parental males. After estrus, the frequency of sexual interactions received by females sharply decreased. One mating event was recorded during the first gestation week and 31 mountings were observed after the second week. In conclusion, the behavioral monitoring is a useful procedure for the recognition of estrus. Our results suggest that ovulation may be associated with the end of the estrus, which will support future work in assisted reproduction in this species. To promote good handling practices, females of reproductive age should be removed from their family unit of origin.


Assuntos
Artiodáctilos/fisiologia , Ciclo Estral/fisiologia , Reprodução/fisiologia , Comportamento Sexual Animal/fisiologia , Animais , Copulação/fisiologia , Feminino , Masculino , Gravidez
13.
Rev Biol Trop ; 64(4): 1519-26, 2016 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-29465914

RESUMO

The agouti (Dasyprocta sp.) is a hystricomorph rodent found in some regions of the Americas. It is an important cynegetic species, which indicates that the overhunting is a threat to their conservation. Very little is known about this wildlife in relation to what already has been studied in domestic animals. Thus, the knowledge on reproduction of wildlife becomes necessary and essential for the management and conservation of these natural resources. Specifically, studies regarding hormonal monitoring are important as a basic tool for research in modern reproductive biotechnology, and currently, there is no information on the progesterone changes during pregnancy of Dasyprocta sp., compared to other hystricomorphs. The aim of this study was to describe the profile of plasmatic progesterone during pregnancy, and report the restart of ovarian cycle in agouti after parturition. For this purpose, 18 black-rumped agoutis (Dasyprocta prymnolopha) born in captivity were used, and one or more consecutive pregnancies were observed. Copulation was confirmed by the presence of spermatozoa observed in the colpocytological examination. Blood samples were collected two times per week, and concentrations of progesterone, assessed in ten agoutis, were determined by radioimmunoassay. The onset of ovarian activity in six females was observed daily by colpocytological examination, starting on the seventh day postpartum. The gestational period observed in this study was 104.04 days (SD = 1.31) (101-106 days) (Number of cases, N = 26), and the interval between births was 126.03 days (SD = 18.40) (109-184 days). The plasmatic profile of progesterone during pregnancy showed a progressive increase from the 1st to the 5th week. The higher progesterone levels over this period (6.88 ng / mL, SD = 3.01) were detected in the 5th week. This value was similar (One-Way ANOVA, p > 0.05) to that observed in the 4th and 6th weeks, but was statistically different (One-Way ANOVA, p < 0.05) when compared to the other weeks. After the 6th week there was a progressive decrease in plasmatic progesterone levels. The animals showed a postpartum estrus of 12.04 days (SD= 4.29) (7-24 days) (N= 23). It was observed that 80.95 % (N= 19) of copulations during this period were fertile. This work contributed to understanding the dynamic changes in the progesterone levels during the pregnancy in agouti. Nevertheless, more studies are needed for a better appreciation of other endocrine and biological changes, in the mother and feto-placental unit of the agouti.


Assuntos
Dasyproctidae/sangue , Estro/sangue , Período Pós-Parto/sangue , Progesterona/sangue , Análise de Variância , Animais , Feminino , Idade Gestacional , Gravidez , Valores de Referência , Comportamento Sexual Animal/fisiologia , Fatores de Tempo
14.
Acta Cir Bras ; 31(12): 821-825, 2016 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-28076506

RESUMO

PURPOSE: : To evaluate the Adipose Stem Cells (ACS) therapy efficacy on the time and quality of wound healing process in rats. METHODS: : Nine male Wistar rats were randomly distributed into three groups I) 7 days of healing; II) 14 days of healing; III) 21 days of healing. Four incisions were made on the dorsal surface of each rat and then treated with intralesional ACS, meloxicam, and no treatment and ACS+meloxicam. Macroscopic evaluation was measured by percentage of healing and histopathological by hematoxylin-eosin was performed. RESULTS: : All groups have the wound reduced during the three weeks (p<0.001) and after 14 days of healing had greater reduction than others. Wounds treated with ASC had accelerated healing in relation to no treatment and only meloxicam (p<0.001), excepting the ASC+Meloxicam that was similar (p=0.13). There was no difference in histopathological analysis between lesions. CONCLUSION: : Adipose stem cell have benefits in reducing time of healing of experimental model of wound in rats, observed 7 days of after application.


Assuntos
Tecido Adiposo/citologia , Transplante de Células-Tronco , Células-Tronco/fisiologia , Cicatrização/fisiologia , Ferimentos e Lesões/terapia , Animais , Antígenos de Superfície , Modelos Animais de Doenças , Masculino , Ratos , Ratos Wistar , Resultado do Tratamento
15.
Theriogenology ; 85(2): 323-9, 2016 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-26456184

RESUMO

Glucocorticoids (GCs) are important mediators of key cellular events. Herein, we investigated the effect of adding cortisol to the IVM medium on the acquisition of developmental competency in bovine oocytes. Cortisol (0.01, 0.1, or 1 µg/mL) had no effect on cleavage rates or cell numbers of resulting blastocysts; however, supplementation with 0.1 µg/mL during IVM increased blastocyst rates of in vitro-fertilized bovine oocytes as compared to untreated controls (41 ± 10% vs. 21 ± 1.2%, P < 0.05, respectively). This concentration was chosen to assess changes in the relative expression of potential GC target genes. Oocytes matured in the presence of cortisol and their corresponding cumulus cells did not show changes in expression for genes analyzed as compared to untreated controls. Notably, blastocysts from oocytes matured in cortisol-supplemented medium expressed higher relative levels of glucose transporter 1 (GLUT1), fatty acid synthase (FASN), and heat shock protein 70 (HSP70). This study supports a role for cortisol in the acquisition of bovine oocyte competence. This is evidenced by increased blastocyst development rates and presumably related to elevated embryonic transcripts with roles in glucose and lipid metabolism, as well as the cellular response to stress.


Assuntos
Bovinos , Desenvolvimento Embrionário/efeitos dos fármacos , Hidrocortisona/farmacologia , Técnicas de Maturação in Vitro de Oócitos/métodos , Oócitos/efeitos dos fármacos , Animais , Blastocisto/efeitos dos fármacos , Blastocisto/fisiologia , Meios de Cultura , Células do Cúmulo/química , Técnicas de Cultura Embrionária/veterinária , Ácido Graxo Sintases/genética , Feminino , Fertilização In Vitro , Expressão Gênica , Transportador de Glucose Tipo 1/genética , Proteínas de Choque Térmico HSP72/genética , Hidrocortisona/administração & dosagem , Oócitos/química , Oócitos/fisiologia , RNA Mensageiro/análise
16.
Braz. J. Vet. Res. Anim. Sci. (Online) ; 53(3): 251-259, 2016. ilus, tab
Artigo em Inglês | LILACS, VETINDEX | ID: biblio-875209

RESUMO

A functional understanding of the epididymis makes it possible to increase a species' fertility, since it facilitates the preservation of gametes for use in assisted reproduction techniques. This study analyzed the histological features of the different regions of the epididymis of the Pecari tajacu and the associated pathologies found in the spermatozoa present in this organ. After an orchiectomy surgery to remove the testicles and epididymis obtained from eight adult animals bred in captivity, fixation and histological processing of samples were carried out. The presence of pseudostratified columnar epithelium with stereocilia was observed in all regions of the epididymis (caput, corpus and caudal). Similarities were noted between the average height of the corpus and caudal epithelium, which differed from that found at the caput (1- proximal: region associated with the efferent ducts; 2- distal: region associated with corpus epididymis). It was also observed that the proximal caput presents a smaller average of the tubules diameter compared to other portions of the epididymis. Regarding pathologies of the spermatozoa, thirteen different types of morphological modifications were observed. Underdeveloped spermatozoa and detached heads were most frequently encountered along the epididymis of the collared peccaries. This study is a pioneer in the area and its data will serve as a basis for comparative research on the reproductive biology of artiodactyls, thus contributing to their conservation and reproduction.(AU)


O entendimento funcional do epidídimo possibilita o aumento da fertilidade de uma espécie, pois torna possível a preservação de gametas para o uso em técnicas de reprodução assistida. Este trabalho estudou os aspectos histológicos das diferentes regiões do epidídimo de Pecari tajacu, bem como as patologias associadas encontradas nos espermatozoides presentes nesse órgão. Após procedimento cirúrgico de orquiectomia, para remoção dos testículos e epidídimos, obtidos de oito animais adultos nascidos em cativeiro, realizou-se a fixação e o processamento histológico das amostras. Constatou-se em todas as regiões do epidídimo (cabeça, corpo e cauda) a presença de epitélio pseudoestratificado colunar com estereocílios. Foram observadas semelhanças entre a altura média do epitélio do corpo e da cauda, a qual era diferente da encontrada na cabeça (1- proximal: região associada aos ductos eferentes; 2- distal: região associada ao corpo do epidídimo). Foi verificado que a cabeça proximal apresenta a média de diâmetro dos túbulos menor em relação às demais porções do epidídimo. Em relação às patologias dos espermatozoides, constataram-se treze tipos diferentes de alterações morfológicas. Os espermatozoides subdesenvolvidos e cabeças soltas foram as mais frequentes ao longo do epidídimo de catitus. Os dados deste trabalho são pioneiros, e servirão como base para pesquisas comparativas sobre a biologia reprodutiva dos artiodátilos, contribuindo para a sua conservação e produção.(AU)


Assuntos
Animais , Artiodáctilos , Epididimo/anatomia & histologia , Epididimo/fisiologia , Espermatozoides/patologia , Técnicas Histológicas/veterinária
17.
Mol Reprod Dev ; 81(10): 918-27, 2014 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-25236163

RESUMO

Nitric oxide (NO) is a cell-signaling molecule that regulates a variety of molecular pathways. We investigated the role of NO during preimplantation embryonic development by blocking its production with an inhibitor or supplementing in vitro bovine embryo cultures with its natural precursor, L-arginine, over different periods. Endpoints evaluated included blastocyst rates, development kinetics, and embryo quality. Supplementation with the NO synthase inhibitor N-Nitro-L-arginine-methyl ester (L-NAME) from Days 1 to 8 of culture decreased blastocyst (P < 0.05) and hatching (P < 0.05) rates. When added from Days 1 to 8, 50 mM L-arginine decreased blastocyst rates (P < 0.001); in contrast, when added from Days 5 to 8, 1 mM L-arginine improved embryo hatching rates (P < 0.05) and quality (P < 0.05) as well as increased POU5F1 gene expression (P < 0.05) as compared to the untreated control. Moreover, NO levels in the medium during this culture period positively correlated with the increased embryo hatching rates and quality (P < 0.05). These data suggest exerts its positive effects during the transition from morula to blastocyst stage, and that supplementing the embryo culture medium with L-arginine favors preimplantation development of bovine embryos.


Assuntos
Arginina/farmacologia , Blastocisto/metabolismo , Meios de Cultura , Técnicas de Cultura Embrionária , Desenvolvimento Embrionário/efeitos dos fármacos , Óxido Nítrico/metabolismo , Animais , Blastocisto/citologia , Bovinos , Meios de Cultura/química , Meios de Cultura/farmacologia , Inibidores Enzimáticos/farmacologia , NG-Nitroarginina Metil Éster/farmacologia
18.
Theriogenology ; 81(7): 982-7, 2014 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-24576716

RESUMO

Ovum pick up (OPU) associated with in vitro production (IVP) of embryos has been shown as an important tool in cattle breeding to increase the number of descendants from animals of high genetic value. In herds maintained distant from the laboratory, collecting cumulus-oocyte complexes (COCs) and transporting them to the laboratory may take several hours and decrease COCs viability, representing a challenge for commercial settings. In this study, a prematuration culture to induce temporary meiosis block was evaluated in a commercial scale IVP setting as a strategy to transport bovine OPU-derived COCs from Nelore and Brangus donors. Effects on embryo yield and pregnancy rates were assessed. Viable COCs from each donor were destined to one of the experimental groups (control, blocks 1 and 2). Control group COCs were placed in cryotubes with 1 mL TCM199-HEPES. In block groups (1 and 2), COCs were placed in cryotubes with 300 µL TCM 199 + 12 µM butyrolactone I (block medium). All groups were gassed and kept in a thermos bottle for 4 hours at 36 °C. Next, COCs in the control group were transferred to IVM medium and block 1 group to block medium, and cultured for 22 hours and 15 hours, respectively, at 38.5 °C and 5% CO2 in air. Block 2 COCs were kept in the cryotubes and in the thermos bottle for another 15 hours at 36 °C to simulate long-term transport conditions. After meiosis block in prematuration culture, blocks 1 and 2 COCs were matured in vitro for 22 hours as for the control group. After IVM, COCs in all groups were submitted to IVF and IVC, and blastocyst rates were evaluated on day 7. Embryos were transferred and pregnancy rates evaluated at 60 days of gestation. The mean total number of COCs retrieved by OPU did not differ between Nelore and Brangus donors (16.8 and 17.2, respectively, P > 0.05), but Nelore donors produced more viable COCs than Brangus (10.1 and 7.6, respectively, P < 0.05) and more embryos/cow (3.8 and 2.7, respectively, P < 0.05). Blastocyst rates were similar for control (40.2% and 36.7%), block 1 (37.3% and 34.5%), and block 2 groups (34.7% and 33.6%) for Nelore and Brangus cattle, respectively (P > 0.05). Pregnancy rates did not differ regardless of breed or treatment (36.7%, P > 0.05). In conclusion, temporary meiosis block during prematuration culture did not affect embryo development or pregnancy rates; therefore, this strategy may be used to transport bovine COCs in a commercial IVP setting.


Assuntos
Bovinos/fisiologia , Células do Cúmulo/fisiologia , Meiose/fisiologia , Oócitos/fisiologia , Taxa de Gravidez , Animais , Técnicas de Cultura Embrionária/veterinária , Feminino , Fertilização In Vitro/veterinária , Técnicas de Maturação in Vitro de Oócitos , Gravidez
19.
Indian J Exp Biol ; 52(12): 1159-64, 2014 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-25651608

RESUMO

The objective of this work was to evaluate the effect of using L-arginine during in vitro fertilization (IVF) on in vitro embryonic development using Bos taurus and Bos indicus semen. Effect of different concentrations (0, 1, 10 and 50 mM) of L-arginine, added to the IVF medium, was evaluated on the fertilization rate at 18 h post-fertilization (hpf), NO3(-)/NO2(-) production during IVF by the Griess colorimetric method (30 hpf), cleavage and blastocyst rates (on Day 2 and Day 7 of culture, respectively) and total blastocyst cell number (Day 7 of culture). The results reveal that the addition of 50 mM L-arginine to IVF medium, with either Bos taurus or Bos indicus spermatozoa, decreased the cleavage rate and blastocyst rate compared to the control group. Other concentrations did not affect embryo production. However, 1 mM L-arginine with Bos indicus semen increased the proportion of hatched blastocysts. These results indicate that high L-arginine concentrations may exhibit toxic effects on bovine gametes during in vitro fertilization.


Assuntos
Blastocisto/efeitos dos fármacos , Fertilização In Vitro/efeitos dos fármacos , Espermatozoides/efeitos dos fármacos , Animais , Arginina , Blastocisto/citologia , Blastocisto/metabolismo , Bovinos , Relação Dose-Resposta a Droga , Embrião de Mamíferos/citologia , Embrião de Mamíferos/efeitos dos fármacos , Embrião de Mamíferos/metabolismo , Desenvolvimento Embrionário/efeitos dos fármacos , Feminino , Fertilização/efeitos dos fármacos , Masculino , Microscopia de Fluorescência , Nitratos/metabolismo , Nitritos/metabolismo
20.
Theriogenology ; 81(2): 326-31, 2014 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-24210669

RESUMO

In vitro-produced embryos store high lipid content in cytoplasmic lipid droplets (LD), and reduction or removal of LD has been demonstrated to improve freeze-thaw viability. The Perilipin Adipophilin Tail-interacting Protein of 47 kD (PAT) family of proteins is involved in the formation and regulation of LD in many cell types, but their presence has not been addressed either in cattle oocytes or preimplantation embryos. Therefore, this study aimed to detect the expression of PAT family transcripts (Perilipin-2 [PLIN2] and Perilipin-3 [PLIN3]) in immature and in vitro-matured (IVM) oocytes, and in in vitro-produced embryos at the stages of two to four cells, eight to 16 cells, morulae (MO), and blastocyst (BL). The expression of PLIN3 was downregulated in response to IVM, and PLIN2 was comparatively more expressed than PLIN3 in IVM oocytes (P < 0.001). During the early stages of embryo development, PLIN2 expression reached its peak at the MO stage (P < 0.001) and decreased again at the BL stage. In contrast, PLIN3 was expressed in low levels during the earliest stages of development, slightly upregulated at the MO stage (P < 0.05), and greatly increased its expression at the BL stage (15-fold; P < 0.001). PLIN3 was comparatively more expressed than PLIN2 during embryo culture in most stages analyzed (P < 0.05), except in eight- to 16-cell embryos. These results indicate that PLIN2 might be involved in the maintenance of lipid stocks necessary to support embryo development after fertilization of IVM oocytes. Also, we hypothesize that PLIN3 is the main PAT protein responsible for stabilization of LD formed in consequence of the acute lipid load seen during embryo development. We confirmed the presence of both PLIN2 and PLIN3 proteins in BL at Day 7 using immunocytochemistry: these PAT proteins colocalized with LD stained with BODIPY. PLIN3 seemed to be more ubiquitously spread out in the cytoplasm than PLIN2, consistent with the pattern seen in adipocytes. These findings suggest that both elderly (bigger) and newly formed (smaller) LD, positive for PLIN2 and PLIN3 respectively, coexist in blastocysts. To our knowledge this is the first report showing that transcripts of the PAT family are present in cattle oocytes and embryos.


Assuntos
Bovinos/embriologia , Desenvolvimento Embrionário , Proteínas de Membrana/metabolismo , Oócitos/crescimento & desenvolvimento , Proteínas de Transporte Vesicular/metabolismo , Animais , Fertilização In Vitro/veterinária , Perilipina-2
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