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Article contents

Establishment of a protocol for the isolation of ovarian preantral follicles derived from collared peccaries (Pecari tajacu)

Published online by Cambridge University Press:  15 August 2019

Lívia Batista Campos
Affiliation:
Laboratory of Animal Germplasm Conservation, Universidade Federal Rural do Semi-Árido (UFERSA), Mossoró, RN, Brazil
Andréia Maria da Silva
Affiliation:
Laboratory of Animal Germplasm Conservation, Universidade Federal Rural do Semi-Árido (UFERSA), Mossoró, RN, Brazil
Samara Sandy Jeronimo Moreira
Affiliation:
Laboratory of Animal Germplasm Conservation, Universidade Federal Rural do Semi-Árido (UFERSA), Mossoró, RN, Brazil
Caio Sergio Santos
Affiliation:
Laboratory of Animal Germplasm Conservation, Universidade Federal Rural do Semi-Árido (UFERSA), Mossoró, RN, Brazil
Carlos Alexandre de Carvalho Apolinário
Affiliation:
Laboratory of Animal Germplasm Conservation, Universidade Federal Rural do Semi-Árido (UFERSA), Mossoró, RN, Brazil
Márcia Viviane Alves Saraiva
Affiliation:
Laboratory of Animal Germplasm Conservation, Universidade Federal Rural do Semi-Árido (UFERSA), Mossoró, RN, Brazil
Moacir Franco de Oliveira
Affiliation:
Laboratory of Animal Germplasm Conservation, Universidade Federal Rural do Semi-Árido (UFERSA), Mossoró, RN, Brazil
Alexandre Rodrigues Silva*
Affiliation:
Laboratory of Animal Germplasm Conservation, Universidade Federal Rural do Semi-Árido (UFERSA), Mossoró, RN, Brazil
*
Address for correspondence: A.R. Silva. Departamento de Ciências Animais, Universidade Federal Rural do Semi-Árido (UFERSA), BR 110, Km 47, Bairro Pres. Costa e Silva. CEP: 59.625-900 Mossoró, RN, Brazil. Fax: +55 84 3317 8360. E-mail: alexrs@ufersa.edu.br

Summary

We compare the efficiency of mechanical or enzymatic methods, and their combination, for the isolation of ovarian preantral follicles (PFs) from collared peccaries. The ovaries from six females were subjected to the different methods investigated here. For the enzymatic method, ovary fragments were exposed to collagenase type IV in TCM-HEPES medium; the mechanical procedure was based on ovarian cortex dissociation by using a scalpel blade. The residual solution obtained after the mechanical isolation was subjected to the enzymatic procedure. The number of isolated PFs was quantified and classified as primordial, primary, or secondary; their viability was assessed using trypan blue dye assay. To confirm the results, PFs derived from the most efficient method were evaluated for integrity using scanning electron microscopy (SEM) and subjected to a 24 h in vitro culture for subsequent evaluation of viability by using fluorescent probes. A higher number of PFs (P < 0.05) was obtained from the enzymatic method (961.7 ± 132.9) in comparison with the mechanical method (434.3 ± 88.9), but no difference was observed between the two methods and their combination (743.2 ± 92.8). The trypan blue assay showed that the enzymatic method (98.7 ± 0.6%) provided the highest percentage of viable follicles (P < 0.05). Furthermore, SEM confirmed the ultrastructural integrity of the surface architecture of peccary PFs isolated by the enzymatic procedure; epifluorescence microscopy was used to confirm their viability (86.0%). In conclusion, we suggest that the enzymatic method investigated here is useful for the isolation of viable ovarian PFs from collared peccaries.

Type
Research Article
Copyright
© Cambridge University Press 2019 

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Establishment of a protocol for the isolation of ovarian preantral follicles derived from collared peccaries (Pecari tajacu)
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