Development of an optimal protocol to induce capacitation of boar spermatozoa in vitro

Seung-Ik Jang,Jae-Hwan Jo,Eun-Ju Jung,Woo-Jin Lee,Ju-Mi Hwang,Jeong-Won Bae,Woo-Sung Kwon The Korean Society of Animal Reproduction and Biot 2022 한국동물생명공학회지 Vol.37 No.4

In 1951, Colin Russell Austin and Min Chueh Chang identified "capacitation", a special process involving ejaculated spermatozoa in the female reproductive tract. Capacitation is a phenomenon that occurs in vivo, but almost all knowledge of capacitation has been obtained from in vitro studies. Therefore, numerous trials have been performed to establish in vitro capacitation methods for various studies on reproduction. Although a series of studies have been conducted to develop an optimal protocol for inducing capacitation, most have focused on identifying the appropriate chemical compounds to induce the capacitation of boar spermatozoa in vitro. Therefore, the purpose of this study was to identify the optimal incubation time for inducing capacitation in vitro. Duroc semen was incubated for various periods (60, 90, and 120 min) to induce capacitation. Sperm function (sperm motility, motion kinematic parameters, and capacitation status) was evaluated. The results showed that total sperm motility, rapid sperm motility, progressive sperm motility, curvilinear velocity, and average path velocity significantly decreased in a time-dependent manner. However, the capacitation status did not show any significant changes. Taken together, these results indicate that an incubation time of more than 60 min suppresses sperm motility and motion kinematic parameters. Therefore, we suggest that 60 min may be the best incubation time to induce capacitation without negative effects on sperm motility and motion kinematics in boar spermatozoa in vitro.

Chronic kisspeptin delays puberty and reduces feed intake and body weight in female rats

Sathagopam, Sriravali,Ullewar, Meenal Prabhakar,Harne, Rakhi,Velmurugan, Sathya The Korean Society of Animal Reproduction and Biot 2021 한국동물생명공학회지 Vol.36 No.1

Kisspeptin is a key player in the central control of reproductive axis. Central administration of kisspeptin has been shown to advance puberty in rats. Stimulation of hypothalamic GnRH pulse generating mechanism by kisspeptin has been proposed to be the mechanism behind the onset of puberty. We hypothesized that chronic high doses of kisspeptin administration suppresses the reproductive axis and hence delays the pubertal onset. Hence, we investigated the effect of peripheral administration of chronic high doses of kisspeptin on pubertal onset, feed intake and body weight in female rats. Rats were treated with saline or kisspeptin (100 nmoles per day; intraperitoneal) for 26 days (day 25 to day 50 postnatal) and the day of vaginal opening was marked as day of puberty. Kisspeptin treated rats had delayed pubertal onset and reduced feed intake and body weight. Gonadal GPR54 mRNA was reduced suggesting that chronic high doses of kisspeptin may suppress the reproductive functions possibly by downregulation of GPR54 receptor. However, delay in puberty due to reduction in feed intake and body weight could not be ruled out in this study. Further, our study emphasizes the importance of dosage and duration of kisspeptin administration in the manipulation of reproductive axis. Our study, for the first time, suggests that kisspeptin and its analogues, if proven beneficial, could be used to treat precocious puberty in children. It appears that, though a promising tool for enhancing fertility, kisspeptin acts as a double-edged sword and has to be cautiously used to manipulate reproduction.

Immunohistological expression of cytochrome P450 1A2 (CYP1A2) in the ovarian follicles of prepubertal and pubertal rat

Hwang, Jong-Chan,Park, Byung-Joon,Kim, Hwan-Deuk,Baek, Su-Min,Lee, Seoung-Woo,Jeon, Ryoung-Hoon,Jang, Min,Bae, Seul-Gi,Yun, Sung-Ho,Park, Jin-Kyu,Kwon, Young-Sam,Kim, Seung-Joon,Lee, Won-Jae The Korean Society of Animal Reproduction and Biot 2020 한국동물생명공학회지 Vol.35 No.4

Cytochrome P450 1A2 (CYP1A2) is a member of the cytochrome P450 superfamily enzymes in mammals and plays a major role in metabolizing endogenous hormones in the liver. In recent days, CYP1A2 expression has been found in not only the liver but also other tissues including the pancreas and lung. However, little information is available regarding the expression of CYP1A2 in the ovary, in spite of the facts that the ovarian follicle growth and atresia are tightly associated with controls of endocrine hormonal networks. Therefore, the expression of CYP1A2 in the ovaries of prepubertal and pubertal rats was investigated to assess its expression pattern and puberty-related alteration. It was demonstrated that the expression level of CYP1A2 was significantly (p < 0.01) higher in the pubertal ovaries than prepubertal counterparts. At the ovarian follicle level in both groups, whereas CYP1A2 expression was less detectable in the primordial, primary and secondary follicles, the strongly positive expression of CYP1A2 was localized in the granulosa cell layers in the antral and pre-ovulatory follicles. However, the ratio of CYP1A2-positive ovarian follicle was significantly (p < 0.01) higher in the ovary of pubertal group (73.1 ± 3.1%) than prepubertal one (41.0 ± 10.5%). During the Immunofluorescence, expression of CYP1A2 was mainly localized in Fas-positive follicles, indicating the atretic follicles. In conclusion, these results suggested that CYP1A2 expression was mainly localized at the atretic follicular cells and affected by the onset of puberty. Further study is still necessary but we hypothesize that CYP1A2 expresses in the atretic follicles to metabolize residue of the reproductive hormones. These findings may have important implications for the fields of reproductive biology of animals.

Supplementation of retinoic acid alone in MSC culture medium induced germ cell-like cell differentiation

Kuldeep Kumar,Kinsuk Das,Ajay Kumar,Purnima Singh,Madhusoodan A. P.,Triveni Dutt,Sadhan Bag The Korean Society of Animal Reproduction and Biot 2023 한국동물생명공학회지 Vol.38 No.2

Background: Germ cells undergo towards male or female pathways to produce spermatozoa or oocyte respectively which is essential for sexual reproduction. Mesenchymal stem cells (MSCs) have the potential of trans-differentiation to the multiple cell lineages. Methods: Herein, rat MSCs were isolated from bone marrow and characterized by their morphological features, expression of MSC surface markers, and in vitro differentiation capability. Results: Thereafter, we induced these cells only by retinoic acid supplementation in MSC medium and, could able to show that bone marrow derived MSCs are capable to trans-differentiate into male germ cell-like cells in vitro. We characterized these cells by morphological changes, the expressions of germ cell specific markers by immunophenotyping and molecular biology tools. Further, we quantified these differentiated cells. Conclusions: This study suggests that only Retinoic acid in culture medium could induce bone marrow MSCs to differentiate germ cell-like cells in vitro. This basic method of germ cell generation might be helpful in the prospective applications of this technology.

Hematological characteristics and endocrine profiles of cloned dromedary camels (Camelus dromedaries)

Young-Bum Son,Mohammad Shamim Hossein,Yeon Ik Jeong,Mina Kang,Huijeong Kim,Yura Bae,Kung Ik Hwang,Alex Tinson,Singh Rajesh,Al Shamsi Noura,Woo Suk Hwang The Korean Society of Animal Reproduction and Biot 2024 한국동물생명공학회지 Vol.39 No.1

Background: Somatic cell nuclear transfer (SCNT) is a prominent technology that can preserve superior genetic traits of animals and expand the population in a short time. Hematological characters and endocrine profiles are important elements that demonstrate the stability of the physiological state of cloned animals. To date, several studies regarding cloned camels with superior genes have been conducted. However, detailed hemato-physiological assessments to prove that cloned camels are physiologically normal are limited. In this study, We evaluated the hemato-physiological characteristics of cloned male and female dromedary camels (Camelus dromedaries). Methods: Therefore, we analyzed variations in hematological characteristics and endocrine profiles between cloned and non-cloned age-matched male and female dromedary camels (Camelus dromedaries). Two groups each of male and female cloned and non-cloned camels were monitored to investigate the differences in hemato-physiological characteristics. Results: All the animals were evaluated by performing complete blood count (CBC), serum chemistry, and endocrinological tests. We found no significant difference between the cloned and non-cloned camels. Furthermore, the blood chemistry and endocrine profiles in male and female camels before maturity were similar. Conclusions: These results suggest that cloned and non-cloned camels have similar hematological characteristics and endocrine parameters.

Sperm motility and viability of experimental animals using different cryopreservatives

Won Yong Park,Byoung Boo Seo The Korean Society of Animal Reproduction and Biot 2023 한국동물생명공학회지 Vol.38 No.1

In this study, an experiment was conducted in order to determine what cryopreservatives (CPVs) were more effective in supporting the motility and viability of sperm from experimental animals. The sperm of mice, rats, beagle dogs, and rabbits were frozen using different CPVs, including DMSO, TYB, and Sperm CryoProtec. The results from freezing the sperm of each laboratory animal in Sperm CryoProtec showed a high level of sperm motility and viability in sperm samples from mice, rats, and beagle dogs melted at the end of the first week. For rabbits, a high level of motility was observed in sperm thawed during the first week, whereas a high level of viability was observed in sperm thawed during the second week. The results of analysis of sperm motility and viability using different CPVs according to laboratory animals showed a significantly higher level of sperm motility (26.28%) and viability (36.20%) for mice in Sperm CryoProtec and the lowest levels of motility and viability were observed in DMSO (p < 0.05). Significantly higher levels of motility (27.94%) and viability (37.94%) were observed for rats in Sperm CryoProtec compared with TYB, which showed the lowest levels of motility and viability (p < 0.05). The study findings described above suggest that the selection of appropriate cryopreservatives is required for each experimental animal. This is because there are differences in the levels of sperm motility and viability of experimental animals depending on the CPVs that are typically used for freezing human sperm, including Sperm CryoProtec and TYB.

Reproductive biology of 58 fish species around La Réunion Island (Western Indian Ocean): first sexual maturity and spawning period

Kelig Mahe,Julien Taconet,Blandine Brisset,Claire Gentil,Yoann Aumond,Hugues Evano,Louis Wambergue,Romain Elleboode,Tevamie Rungassamie,David Roos The Korean Society of Animal Reproduction and Biot 2024 한국동물생명공학회지 Vol.39 No.1

Background: The biological information of fish, which include reproduction, is the prerequisite and the basis for the assessment of fisheries. Methods: The aim of this work was to know the reproductive biology with the first sexual maturity (TL₅₀) and the spawning period for 58 mainly fish species in the waters around La Réunion Island (Western Indian Ocean). Twenty families belonging to the Actinopterygii were represented (acanthuridae, berycidae, bramidae, carangidae, cirrhitidae, gempylidae, holocentridae, kyphosidae, labridae, lethrinidae, lutjanidae, malacanthidae, monacanthidae, mullidae, polymixiidae, pomacentridae, scaridae, scorpaenidae, serranidae, sparidae; 56 species; n = 9,751) and two families belonging to the Elasmobranchii (squalidae, centrophoridae; 2 species; n = 781) were sampled. Between 2014 and 2022, 10,532 individuals were sampled covering the maximum months number to follow the reproduction periods of these species. Results: TL₅₀ for the males and the females, respectively, ranged from 103.9 cm (Acanthurus triostegus) to 1,119.3 cm (Thyrsitoides marleyi) and from 111.7 cm (A. triostegus) to 613.1 cm (Centrophorus moluccensis). The reproduction period could be very different between the species from the very tight peak to a large peak covered all months. Conclusions: Most species breed between October and March but it was not the trend for all species around La Réunion Island.

Genetic diversity and relationship of Korean chicken breeds using 12 microsatellite markers

Kim, Yesong,Yun, Ji Hye,Moon, Seon Jeong,Seong, Jiyeon,Kong, Hong Sik The Korean Society of Animal Reproduction and Biot 2021 한국동물생명공학회지 Vol.36 No.3

A number of Korean Chicken breeds were registered in Domestic Animal Diversity Information System (DAD-IS, http://dad.fao.org/) of the Food and Agriculture Organization (FAO). Evaluation of genetic diversity and relationship of local breeds is an important factor towards the identification of unique and valuable genetic resources. Therefore, this study aimed to analysis the genetic diversity and relationship of 22 Korean Chicken breeds using 12 microsatellite (MS) markers. The mean number of alleles for each variety was 5.52, ranging from a 3.75 (Leghorn F; NF) to a 7.0 (Ross). The most diverse breed was the Hanhyup3 (HCC), which had the highest expected heterozygosity (H_Exp) (0.754) and polymorphic information content (PIC) (0.711). The NF was the least diverse population, having the lowest H_Exp (0.467) and PIC (0.413). As a result of the principal coordinates analysis (PCoA) and factorial correspondence analysis (FCA) confirmed that Hy-line Brown (HL) and Lohmann Brown (LO) are very close to each other and that Leghorn and Rhode Island Red (RIR) are clearly distinguished from other groups. Thus, the reliability and power of identification using 12 types of MS markers were improved, and the genetic diversity and probability of individual discrimination were confirmed through statistical analysis. This study is expected to be used as basic data for the identification of Korean chicken breeds, and our results indicated that these multiplex PCR marker sets will have considerable applications in population genetic structure analysis.

Investigation of single nucleotide polymorphism in TSH-β and CaSR associated with body weight in Korean native chickens (Gray Brown)

Oh, Dongyep,Ha, Jae Jung,Yi, Jun Koo,Kim, Dae Hyun,Oh, Seung Min,Kim, Songmi,Han, Kyudong,Park, Yong-Soo The Korean Society of Animal Reproduction and Biot 2021 한국동물생명공학회지 Vol.36 No.3

This study identified single nucleotide polymorphisms (SNPs) that affect the body weight of chickens. Analysis of body weight showed that the Cornish breed had the highest body weight, and the Korean native chicken (Gray Brown) had the lowest body weight. TSH is composed of an α-subunit and a β-subunit, and the TSH-β gene encoding the β-subunit has been reported to be associated with obesity. In chickens, it is located on chromosome 26 and is reported to be associated with growth. The calcium-sensing receptor gene (CaSR) plays a role in the regulation of extracellular calcium homeostasis and is responsible for calcium absorption in the urinary tract, which affects the eggshell quality in poultry. It was shown that TSH-β was strongly correlated with weight in Cornish and Korean native (Gray Brown) chickens, particularly in those with the CC trait. However, CaSR showed no association with body weight in poultry; it was associated with calcium and the eggshell. Thus, selection for TSH-β can be used to produce individuals with more favorable traits in terms of body weight.

Assessment of cryopreserved sperm functions of Korean native brindled cattle (Chikso) from different region research centers of Korea

Ma, Lei,Jung, Dae-Jin,Jung, Eun-Ju,Lee, Woo-Jin,Hwang, Ju-Mi,Bae, Jeong-Won,Kim, Dae-Hyun,Yi, Jun Koo,Lee, Sang Moo,Ha, Jae Jung,Kwon, Woo-Sung The Korean Society of Animal Reproduction and Biot 2021 한국동물생명공학회지 Vol.36 No.2

Sperm cryopreservation is an important method of assisted reproductive techniques and storing genetic resources. It plays a vital role in genetic improvement, livestock industrial preservation of endangered species, and clinical practice. Consequently, the cryopreservation technique is well organized through various studies, especially on Korean native cattle (Hanwoo). However, the cryopreservation technique of Korean native brindled cattle, which is one of the native cattle species in Korea, is not well organized. Therefore, it is necessary to develop a Supplementary Table technique for the cryopreservation of Korean native brindled cattle. For this purpose, it is important to first evaluate the quality of the currently produced cryopreserved sperm of Korean native brindled cattle. In this study, we randomly selected 72 individual Korean native brindled cattle semen samples collected from 8 different region research centers and used them to evaluate sperm functions. We focused on the quality evaluation of cryopreserved Korean native brindled cattle semen following the measurement of motion kinematics, capacitation status, intracellular ATP level, sperm motility, and cell viability. Then, the values of each of the eight groups were derived from various sperm parameters of nine individual samples, including sperm motility, kinematics, cellular motility, and intracellular ATP levels, which were used to compare and evaluate sperm function. Overall, differences in various sperm parameters were observed between most of the research centers. Particularly, the deviations of motility and motion kinematics were high according to the sample. Therefore, we suggest that it is necessary to develop a standard method for the cryopreservation of Korean native brindled cattle semen. We also suggest the need for sperm quality evaluation of the cryopreserved semen of Korean native brindled cattle before using artificial insemination to attain a high fertility rate.