• Clinical and Experimental Reproductive Medicine
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• v.18 no.2
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• pp.123-131
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• 1991

In order to study the growth and maturation of ovarian follicle, the localization and activity of alkaline phosphatase(ALPase) and adenosine triphosphatase(ATPase) of the granulosa cells and theca layer were examined according to the follicle size, the follicle state and the ovarian cyclic phase in pig. Theca interna of the small follicles was more heavyly localized with reaction product by the activites of ALPase and ATPase than that of the large follicles. It is assumed that, as the follicles proceed to growth and maturation, antrum formation is the result of the follicular fluid accumulation by means of active transport by the activities of ALPase and ATPase in theca interna. The activities of ALPase and ATPase in atretic follicles were higher than those of normal follicles. These results imply that the mechanisms of follicle maturation and atresia are different according to the phase of ovarian cycle.

• Journal of Animal Reproduction and Biotechnology
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• v.37 no.2
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• pp.106-112
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• 2022

Cryopreservation of porcine ovarian tissue by vitrification method is a promising approach to preserve genetic materials for future use. However, information is not enough and technology still remains in a challenge stage in pig. Therefore, the objective of present study was to determine possibility of vitrification method to cryopreserve porcine ovarian tissue and to confirm an occurrence of cryoinjuries. Briefly, cryoinjuries and apoptosis patterns in vitrified-warmed ovarian tissue were examined by histological evaluation and TUNEL assay respectively. In results, a damaged morphology of oocytes was detected among groups and the rate was significantly (p < 0.05) lower in vitrification group (25.8%) than freezing control group (67.7%), while fresh control group (6.6%) showed significantly (p < 0.05) lower than both groups. In addition, cryoinjury that form a wave pattern of tissues around follicles was found in the frozen control group, but not in the fresh control group as well as in the vitrification group. Apoptotic cells in follicle was observed only in freezing control group while no apoptotic cell was found in both fresh control and vitrification. Similarly, apoptotic patterns of tissues not in follicle were comparable between fresh control and vitrification groups while freezing control group showed increased tendency. Conclusively, it was confirmed that vitrification method has a prevention effect against cryoinjury and this method could be an alternative approach for cryopreservation of genetic material in pigs. Further study is needed to examine the viability of oocytes derived from vitrified-warmed ovarian tissue.

• Reproductive and Developmental Biology
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• v.33 no.3
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• pp.183-187
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• 2009

This study analyzed change of proteins in granulosa cells during the porcine follicuar development by proteomics techniques. Granulosa cells of the follicles, of which the diameter is $2{\sim}4\;mm$ and $6{\sim}10\;mm$, were collected from ovary of slaughtered pig that each follicle of diameter $1{\sim}4\;mm$ and $6{\sim}10\;mm$. We extracted glanulosa cell proteins by M-PER Mammalian Protein Extraction Reagent. Proteins were refined by clean-up kit and quantified by Bradford method until total protein was $200{\mu}l$. Immobilized pH gradient(IPG) strip used 18 cm, $3{\sim}10\;NL$. SDS-PAGE used 10% acrylamide gel. After silver staining, Melanie 7 and naked eye test were used for spot analyzation. Increasing proteins in glanulosa cell of $6{\sim}10\;mm$ follicle were 7 spots. This spots were analyzed by MALDI-TOF MS and searched on NCBInr. In results, 7 spots were similar to zinc/ling finger protein 3 precursor (RING finger protein 203), angiomotin, heat shock 60 kDa protein 1 (chaperonin) isoform 1 (HSP60), similar to transducin-like enhancer protein 1 (TLE 1), SH3 and PX domains 2A (SH3PXD2A). Those proteins were related with transfer between cells. Increase of proteins has an effect on follicular development.

• Korean Journal of Animal Reproduction
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• v.16 no.2
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• pp.165-173
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• 1992

In order to study the mechanism of follicle growth and maturation, and also to supplement the criteria identifying the follicle state of normal of atretic, the histochemical investigation on the ovarian follicles according to the ovarian cycle of mouse, rat and pig has been done. The intercellular space of granulosa cells, especailly Call-Exner body, and follicular fluid in the antrum showed positive to PAS, and blue stain by trichrome dye. The resutls suggest that the mucous polysaccharide was synthesized by the granulosa cells, and secreted into the antrum through Call-Exner body so as to be the components of the follicular fluid as the follicles proceeded to growth and maturation. The further the follicles proceeded to atresia the more densely their theca externa were stained blue by follicles proceeded to atresia the more densely their theca externa were stained blue by trichrome dye, and the more densely the granulosa cells were stained red by oil red 0 dye. Therefore, these staining methods can be applied to the criteria identifying the follicle atresia.

• Reproductive and Developmental Biology
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• v.34 no.3
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• pp.217-221
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• 2010

When fully grown oocytes are removed from their follicles, they can resume meiosis and mature spontaneously under in vitro conditions. However, nuclear maturation under in vitro condition is not accompanied by complete cytoplasmic maturation, which is essential for successful fertilization and the initiation of zygotic development. This study analyzed change of proteins in follicular fluids during the porcine follicular development. Follicular fluids were collected from follicles of diameter 1~2 mm, 2~6 mm and 6~10 mm in ovary of slaughtered pigs. Total proteins were extracted from follicular fluids by M-PER Mammalian Protein Extraction Reagent. We confirmed totally 27 same spots, 1 spot from follicle fluid of 2~6 mm follicle and 5 spots from follicle fluid of 6~10 mm in diameter were analyzed by MALDI mass spectrometry and searched on NCBInr. In results, spot No. 28 from 2~6 mm follicle was Ig lambda chain C region, and spot No.32 and 33 from 6~10 mm was Apolipoprotein A-(APOA4). Spot No.29 and 31 were failed to analyze. These results indicate that the porcine oocyte during in vitro maturation depend on specific different expressed proteins may play an important roles in the sequence of molecular events in porcine oocyte maturation and follicular development.

• Molecules and Cells
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• v.38 no.4
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• pp.304-311
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• 2015

Most follicles in the mammalian ovary undergo atresia. Granulosa cell apoptosis is a hallmark of follicle atresia. Our previous study using a microRNA (miRNA) microarray showed that the let-7 microRNA family was differentially expressed during follicular atresia. However, whether the let-7 miRNA family members are related to porcine (Sus scrofa) ovary follicular apoptosis is unclear. In the current study, real-time quantitative polymerase chain reaction showed that the expression levels of let-7 family members in follicles and granulosa cells were similar to our microarray data, in which miRNAs let-7a, let-7b, let-7c, and let-7i were significantly decreased in early atretic and progressively atretic porcine ovary follicles compared with healthy follicles, while let-7g was highly expressed during follicle atresia. Furthermore, flow cytometric analysis and Hoechst33342 staining demonstrated that let-7g increased the apoptotic rate of cultured granulosa cells. In addition, let-7 target genes were predicted and annotated by TargetScan, PicTar, gene ontology and Kyoto encyclopedia of genes and genomes pathways. Our data provide new insight into the association between the let-7 miRNA family in granulosa cell programmed death.

• Reproductive and Developmental Biology
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• v.28 no.1
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• pp.37-43
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• 2004

This study was carried out to investigate PSS (Porcine Stress Syndrome) with the PSE (Pale, Soft, Exudative) in 319 different pigs(Yorkshire 150; Landrace 89 and Duroc 80). The PCR-RFLP method was adapted to detect the ryanodine receptor (RYR 1) gene mutation and to estimate the genotype frequency of the RYR1 gene in breeding pig population. The DNA samples were collected from hair follicles of pigs of Yorkshire, Landrace and Duroc. After DNA amplification by PCR, the PCR products were digested by restriction enzyme, Cfo I. Primary PCR products of ryanodine receptor gene were length of 659 bp in hair follicle and their second PCR products were length of 522 bp in hair follicle. The exon region (522 bp) including point mutation ($C \arrow T; Arg \arrow Cys$) in the porcine ryanodine receptor gene, which is a causal mutation for PSS, was digested with Cfo I restriction enzyme. The RYR1 gene was classifed into three genotypes by agarose gel electrophoresis. The normal homozygous (NN) individuals showed two DNA fragments consisted of 439 and 83 bp. The mutant homozygous (nn) individuals showed only one DNA fragment 522 bp. In addition, all three fragments (522, 439 and 83 bp) were showed in heterozygous (Nn) carrier animals. The normal homozygous (NN), heterozygous (Nn) and mutant homozygous (nn) were 98.00, 2.00 and 0.00% in Yorkshire pigs, 87.64, 11.24 and 1.12% in Landrace, 100.00, 0.00 and 0.00% in Duroc, respectively. The gene frequencies of N and n were 0.990 and 0.010 in Yorkshire pigs, 0.933 and 0.067 in Landrace, 1.000 and 0.000 in Duroc, respectively.

• Journal of Animal Science and Technology
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• v.63 no.1
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• pp.1-15
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• 2021

As a result of intensive breeding, litter size has considerably increased in pig production over the last three decades. This has resulted in an increase in farrowing complications. Prolonged farrowing will shorten the window for suckling colostrum and reduce the chances for high-quality colostrum intake. Studies also agree that increasing litter sizes concomitantly resulted in decreased piglet birth weight and increased within-litter birth weight variations. Birth weight, however, is one of the critical factors affecting the prognosis of colostrum intake, and piglet growth, welfare, and survival. Litters of uneven birth weight distribution will suffer and lead to increased piglet mortality before weaning. The proper management is key to handle the situation. Feeding strategies before farrowing, management routines during parturition (e.g., drying and moving piglets to the udder and cross-fostering) and feeding an energy source to piglets after birth may be beneficial management tools with large litters. Insulin-like growth factor 1 (IGF-1)-driven recovery from energy losses during lactation appears critical for supporting follicle development, the viability of oocytes and embryos, and, eventually, litter uniformity. This paper explores certain management routines for neonatal piglets that can lead to the optimization of their colostrum intake and thereby their survival in large litters. In addition, this paper reviews the evidence concerning nutritional factors, particularly lactation feeding that may reduce the loss of sow body reserves, affecting the growth of the next oocyte generation. In conclusion, decreasing birth weight and compromised immunity are subjects warranting investigation in the search for novel management tools. Furthermore, to increase litter uniformity, more focus should be placed on nutritional factors that affect IGF-1-driven follicle development before ovulation.

• Journal of Animal Reproduction and Biotechnology
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• v.37 no.1
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• pp.34-41
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• 2022

In vivo oocytes grow and mature in ovarian follicles whereas oocytes are matured in vitro in plastic culture dishes with a hard surface. In vivo oocytes show a superior developmental ability to in vitro counterparts, indicating suboptimal environments of in vitro culture. This study aimed to evaluate the influence of an agarose matrix as a culture substrate during in vitro maturation (IVM) on the development of pig oocytes derived from small antral follicles (SAFs). Cumulus-oocyte complexes (COCs) retrieved from SAFs were grown in a plastic culture dish without an agarose matrix and then cultured for maturation in a plastic dish coated without (control) or with a 1% or 2% (w/v) agarose hydrogel. Then, the effect of the soft agarose matrix on oocyte maturation and embryonic development was assessed by analyzing intra-oocyte contents of glutathione (GSH) and reactive oxygen species (ROS), expression of VEGFA, HIF1A, and PFKP genes, and blastocyst formation after parthenogenesis. IVM of pig COCs on a 1% (w/v) agarose matrix showed a significantly higher blastocyst formation, intra-oocyte GSH contents, and transcript abundance of VEGFA. Moreover, a significantly lower intra-oocyte ROS content was detected in oocytes matured on the 1% and 2% (w/v) agarose matrices than in control. Our results demonstrated that IVM of SAFs-derived pig oocytes on a soft agarose matrix enhanced developmental ability by improving the cytoplasmic maturation of oocytes through redox balancing and regulation of gene expression.

• Asian-Australasian Journal of Animal Sciences
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• v.32 no.7
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• pp.956-965
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• 2019

Objective: The main goal of this study was to provide a morphological indicator that could be used to select high-quality oocytes of appropriate meiotic and developmental capabilities in pig. The higher quality of immature oocytes, the higher success rates of in vitro maturation (IVM) and in vitro fertilization (IVF). Thus, prior to the IVM culture, it is important to characterize oocytes morphologically and biochemically in order to assess their quality. Two of the largest indicators of oocyte quality are the presence of cumulus cells and status of chromatin. To investigate the effects of porcine oocyte chromatin configurations on the developmental capacity of blastocysts, we assessed oocyte chromatin status according to follicle size and measured the developmental potency of blastocysts. Methods: To sort by follicle size, we divided the oocytes into three groups (less than 1 mm, 1 to 3 mm, and more than 3 mm in diameter). To assess chromatin configuration, the oocytes were assessed for their stages (surrounded nucleolus [SN] germinal vesicle [GV], non-surrounded nucleolus [NSN] GV, GV breakdown, metaphase I [MI], pro-metaphase II [proMII], and metaphase II [MII]) at different maturation times (22, 44, and 66 h). To assess the development rate, oocytes of each follicle size were subjected to parthenogenetic activation for further development. Finally, GV oocytes were grouped by their chromatin configuration (SN, SN/NSN, and NSN) and their global transcriptional levels were measured. Results: SN GV oocytes were more suitable for IVF than NSN GV oocytes. Moreover, oocytes collected from the larger follicles had a greater distribution of SN GV oocytes and a higher developmental capacity during IVM, reaching MII more quickly and developing more often to blastocysts. Conclusion: Porcine oocytes with high-level meiotic and developmental capacity were identified by analyzing the relationship between follicle size and chromatin configuration. The porcine oocytes from large follicles had a significantly higher SN status in which the transcription level was low and could be better in the degree of meiotic progression and developmental capacity.