2011 (SBTE) 25th Annual Meeting Proceedings - International ...

Acta Scientiae Veterinariae, 2011. 39(Suppl 1): Abstracts - 25th Annual Meeting SBTE-Brazil. August 2011.
A241 SUPPORTIVE BIOTECHNOLOGIES: CRYOPRESERVATION AND CRYOBIOLOGY, IMAGE ANALYSIS AND DIAGNOSIS, MOLECULAR BIOLOGY AND “OMICS”
EFFECT OF ANTIOXIDANT SUBSTANCES IN CRYOPRESER
OPRESERVED SEMEN FROM BUFF
UFFAL
ALOS (
(BUB
UBAL
ALUS BUB
UBALIS
ALIS)
Arnaldo Algaranhar Gonçalves 1 , Sâmia Rubielle Silva de Castro 2 , Alexandre Rossetto Garcia 3 , Alessandra Ximenes Santos 4 , Geanne Rocha Silva 5 & Daniel
Vale Barros 6
1,4,5
PROGRAMA DE PÓS-GRADUAÇÃO EM CIÊNCIA ANIMAL UFPA/EMBRAPA/UFRA, BELEM, PA, BRAZIL. 2 FACULDADES INTEGRADAS DO TAPAJÓS (FIT), SANTARÉM, PA, BRAZIL.
3
LABORATÓRIO DE REPRODUÇÃO ANIMAL, EMBRAPA AMAZÔNIA ORIENTAL, BELÉM, PA, BRAZIL. 6 UNIVERSIDADE FEDERAL RURAL DA AMAZÔNIA (UFRA), BELEM, PA, BRAZIL.
Cryopreservation causes damage to sperm plasma membrane due to the increase in oxygen use by cells, leading to increased
production of free radicals. Therefore, the objective was to evaluate the effect of two antioxidants, alone or in combination on parameters of
cryopreserved semen of buffalo. The work was performed at Embrapa Eastern Amazon, Belém-PA. Five bulls previously selected and with
similar semen quality had semen collected by artificial vagina fortnightly and are used six ejaculates from each bull (n = 30). Each ejaculate was
frozen under four distinct treatments. The control group (CONT) comprised the freezing extender with TES-TRIS, as Vale (2002, Buffalo
Symposium of Americas 1, 156-171). The other groups followed the protocol of the control, but were enriched with 2.5 mM vitamin C (VIT
Group), as recommended by Singh et al. (1996, International Journal Animal Science 11, 131-132) or with 3.5 mM pentoxifylline (Group
PENTOX), according to Marques et al. (2002 Theriogenology 58, 257-260) or with the combination of 2.5 mM vitamin C + 3.5 mM
pentoxifylline (VIT Group/PENTOX). The motility and plasma membrane integrity of sperm after thawing and after thermal resistance test
(TRT) execution, as Rota et al. (1997, Theriogenology 47, 1093-1101). After testing the normality of distribution of variables, means were
compared by ANOVA. If contrasts were significant, the Tukey test and t-Test were applied (P < 0.05). Raw semen presented progressive motility
of 77.7 ± 6.1% and integrity of plasme membrane of 85.4 ± 8.3% before freezing. After thawing progressive motility was 39.5 ± 23.8%
(CONT), 41.9 ± 22.3% (VIT), 44.6 ± 19.9% (PENTOX) and 46.9 ± 19% (VIT/PENTOX) P > 0.05. After the TTR, motility was 15.1 ± 15.3%
(CONT), 25.6 ± 16.3% (VIT), 23.3 ± 14.6% (PENTOX) and 28.6 ± 16, 2% (VIT/PENTOX) with P < 0.05. The integrity of plasma membrane
staining performed with Eosin-Nigrosine observed was 62.2 ± 15.9% (CONT), 63.5 ± 13.3% (VIT), 57.9 ± 15.7% (PENTOX) and 57.6 ±
15.9% (VIT/PENTOX), P > 0.05. After the TRT was 58.0 ± 13.5% (CONT), 58.3 ± 13.3% (VIT), 54.0 ± 14.9% (PENTOX) and 51.8 ± 15.5%
(VIT/PENTOX) P > 0.05. The significant increase in motility parameters after the TTR may be related to antioxidant effects, by decreasing the
action of reactive oxygen species and the injuries on the sperm cells. Thus, the vitamin C, used alone or associated with pentoxifylline, caused
significant shifting on progressive motility after TRT, which is worthy since this test simulates the stress that spermatozoa undergo after semen
takes place into the female reproductive system.
Keywords: acid ascorbic, pentoxifylline, spermatozoa.
A242 SUPPORTIVE BIOTECHNOLOGIES: CRYOPRESERVATION AND CRYOBIOLOGY, IMAGE ANALYSIS AND DIAGNOSIS, MOLECULAR BIOLOGY AND “OMICS”
BULL EFFECT IN THE EMBRYO SURVIV
VIVAL PRODUCED IN VITRO AFTER SUBMISSION TO SLOW FREEZING
M auricio Bar
arr os Fer
ernandes
1 , A nderson Mior
ioranza
2 , V ivian Taís Fer
ernandes Cipr
ipriano
iano 2 , R eginaldo Apar
parecido
V ila 2 , M arli Apar
parecida
Galerani 2 , Claudia Cristina Paz 2 & Raysildo Barbosa Lôbo 2
1
PRO-FIV, SÃO JOSÉ DO RIO PRETO, SP, BRAZIL. 2 FMRP-USP, RIBEIRÃO PRETO, SP, BRAZIL.
N
In the programs of embryo transfer, cryopreservation has fundamental importance on storage, transport, trade and sanitary control
of embryos. One of the bottlenecks in the application of this technology on in vitro-produced embryos (IVP), it is the high sensitivity to freezing.
Thus, the aim of this study was to assess the influence of the bull effect on survival of embryos that were submitted to freezing process. To
evaluate that, 2856 cumulus-oocyte complexes (COCs) recovered from slaughterhouses were matured in TCM199 for 24 h at 38.5°C and 5%
CO2. The COCs were fertilizated in vitro (IVF) using frozen semen from 15 Nelore bulls. For each bull, it was observed embryonic
development at 168 h after IVF, considering viable the embryos in the stages of blastocyst and expanded blastocyst. The selected blastocysts
were growth in the medium containing cryoprotectants, loaded into straws, refrigerated and freezed. Then, the cryoprotectant was removed and
the embryos were washed and growth in another medium. To evaluate the effect of freezing, we analyzed two different groups of blastocysts:
the control group (C), composed by blastocysts maintained under cultivation for 72 h, and the freezing group (FG) composed by blastocysts
that were frozen and subsequently submitted to cultivation for 72 h. The data were analyzed using SAS statistical software using Chi-square
and, when necessary, Fisher’s exact test, considering the significance of 5%. The embryos that passed through the freezing process had a
significantly decreased in their survival rate when compared with the control group. From the bulls submitted to the freezing process, 13 had
a statistically significant (P < 0.05) in hatching blastocysts. Among the bulls with statistically differences, some were more contrasting,
reaching to 83% of embryos in the C group and 0% in the CG group. More interesting, two of the analyzed bulls did not show statistical
differences between the groups (CG and FG), presenting similar amounts of embryos. Our results revealed a possible influence of the bull on
the survival of embryos submitted to freezing process and will be important to commercial applications. Further investigations are needed to
confirm this relationship and to elucidate the components responsible for promoting this change in the survival of frozen embryos.
Keywords: embryos, freezing, in vitro fertilization.
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