• 한국동물생명공학회지
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• 제36권3호
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• pp.145-153
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• 2021

The role of transient receptor potential vanilloid receptor-1 (TRPV1) has been primarily investigated in pain sensory neurons. Relatively, little research has been performed in testicular cells. TRPV1 is abundantly expressed in Leydig cells of young adult mice. This study was conducted to determine the role of the TRPV1 channel in Leydig cells. TRPV1 modulators and testosterone were treated to the mouse Leydig cell line TM3 cells for 24 h. Capsaicin, a TRPV1 activator, dose-dependently induced cell death, whereas capsazepine, a TRPV1 inhibitor, inhibited capsaicin-induced cell death. Testosterone treatment reduced capsaicin-induced cell death. High concentrations of testosterone decreased TRPV1 mRNA and protein expression levels. However, TRPV1 modulators did not affect testosterone production. These results showed that capsaicin induced cell death of Leydig cells and that testosterone reduced capsaicin-induced cell death. Our findings suggest that testosterone may regulate the survival of Leydig cells in young adult mice by decreasing the expression level of TRPV1.

• 한국발생생물학회지:발생과생식
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• 제21권3호
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• pp.237-247
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• 2017

Mammalian spermatogenesis occurs in a precise and coordinated manner in the seminiferous tubules. One of the attempts to understand the detailed biological process during mammalian spermatogenesis at the molecular level has been to identify the testis specific genes followed by study of the testicular expression pattern of the genes. From the subtracted cDNA library of rat testis prepared using representational difference analysis (RDA) method, a complimentary DNA clone encoding type III member of a DnaJ family protein, DnaJC18, was cloned (GenBank Accession No. DQ158861). The full-length DnaJC18 cDNA has the longest open reading frame of 357 amino acids. Tissue and developmental Northern blot analysis revealed that the DnaJC18 gene was expressed specifically in testis and began to express from postnatal week 4 testis, respectively. In situ hybridization studies showed that DnaJC18 mRNA was expressed only during the maturation stages of late pachytene, round and elongated spermatids of adult rat testis. Western blot analysis with DnaJC18 antibody revealed that 41.2 kDa DnaJC18 protein was detected only in adult testis. Immunohistochemistry study further confirmed that DnaJC18 protein, was expressed in developing germ cells and the result was in concert with the in situ hybridization result. Confocal microscopy with GFP tagged DnaJC18 protein revealed that it was localized in the cytoplasm of cells. Taken together, these results suggested that testis specific DnaJC18, a member of the type III DnaJ protein family, might play a role during germ cell maturation in adult rat testis.

• Toxicological Research
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• 제25권4호
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• pp.203-207
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• 2009

Present study was conducted to investigate potential effects of epichlorohydrin on testicular and epididymal function in male rats. The test chemical was administered to adult male rats by gavage at dose levels of 0, 3.125, 12.5, and 50 mg/kg/day for 7 days. Testicular and epididymal function were assessed by measurement of reproductive organ weight, testicular spermatid count, epididymal sperm count, motility and morphology, and histopathology in rats. At 50 mg/kg, a decrease in the sperm motility and an increase in the incidence of sperm abnormalities were observed. Histopathological examinations revealed an increase in the incidence of histopathological changes including cell debris in the ducts, vacuolization of the epithelial cells, oligospermia, and epithelial disruption in the proximal caput epididymidis. At 12.5 mg/kg, an increase in the incidence of histopathological changes of the epididymidis was found. There were no treatment-related effects at 3.125 mg/kg. These results show that 7-day repeated oral administration of epichlorohydrin to male rats results in adverse effects on sperm motility, sperm morphology, and epididymal histology at $\geq$ 12.5 mg/kg/day.

• Journal of Microbiology and Biotechnology
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• 제16권9호
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• pp.1347-1354
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• 2006

The aims of this study were to establish a simple and effective method for isolating male germline stem cells (GSCs), and to test the possibility of using these cells as a new approach for male infertility treatment. Testes obtained from neonatal and adult mice were manually decapsulated. GSCs were collected from seminiferous tubules by a two-step enzyme digestion method and plated on gelatin-coated dishes. Over 5-7 days of culture, GSCs obtained from neonates and adults gave rise to large multicellular colonies that were subsequently grown for 10 passages. During in vitro proliferation, oct-4 and two immunological markers (Integrin ${\beta}1,\;{\alpha}6$) for GSCs were highly expressed in the cell colonies. During another culture period of 6 weeks to differentiate to later stage germ cells, the expression of oct-4 mRNA decreased in GSCs and Sertoli cells encapsulated with calcium alginate, but the expression of c-kit and testis-specific histone protein 2B(TH2B) mRNA as well as the localization of c-kit protein was increased. Expression of transition protein (TP-l) and localization of peanut agglutinin were not seen until 3 weeks after culturing, and appeared by 6 weeks of culture. The putative spermatids derived from GSCs supported embryonic development up to the blastocyst stage with normal chromosomal ploidy after chemical activation. Thus, GSCs isolated from neonatal and adult mouse testes were able to be maintained and proliferated in our simple culture conditions. These GSCs have the potential to differentiate into haploid germ cells during another long-term culture.

• 한국환경생물학회:학술대회논문집
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• 한국환경생물학회 2003년도 학술대회
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• pp.38-47
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• 2003

Relationships between accumulations of endocrine disruptors and abnormalities in the reproductive organs in the adult male striped field mice, Apodemus agrarius, were reviewed. High levels of phenolic compounds were detected in the mice collected at an agricultural village in Gaduck island and at a place having a sewage problem in Samdong-myeon, Namhae. High levels of organo tin compounds were found in the mice collected at Jiri Mt. a tourist resort. Considerably high levels of phenolic or organo tin compounds were detected in mice with shrunken reproductive organs accounting for 14∼42% of the mice examined in each area, which suggests that the abnormality of reproductive organs may be induced by the endocrine disruptors. This hypothesis is strongly supported by histological observations of shrunken reproductive organs, such as necrosis of testicular germ and epithelial cells. This was found not only in the mice with shrunken reproductive organs but also in the mice with enlarged reproductive organs, both had accumulated high levels of endocrine disruptors in general.

• 한국동물학회지
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• 제33권1호
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• pp.78-93
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• 1990

생쥐가 성숙하는 동안 부정소 상피세포의 분화와 부정소액의 분비 및 흡수와 관련된 미세구조의 변화를 알아보고자, 생후 10, 20, 35 그리고 80일 된 생쥐에서 부정소 상피세포를 전자현미경으로 관찰하였다. 생쥐 부정소의 분화과정은 미세구조의 특징에 따라 생후 20일가지의 미분화기, 생후 35일 전후의 성장 및 분화기, 그리고 성체의 성숙기로 구분되었다. 각 시기는 정소에서 세정관의 강소 형성시기 그리고 정소액과 더불어 정자가 부정소에 유입되는 시기와 밀접한 관계가 있었다. 성체의 부정소 상피세포 중 주세포는 두부 부정소 기부에서 흡수 기능을 갖는 구조였고, 두부의 말부와 체부 그리고 미부에서는 조면소포체와 골지체가 발달되어 단백질 합성과 분비가 왕성한 구조로 관찰되었다. 투명세포는 주로 체부와 미부에 존재하였으며 세포질내에 흡수과립이 많이 존재하였고, 그 속에는 정자에서 분리된 세포질 잔기로 사료되는 막구조물이 관찰되었다. 부정소 부위에 따라 상피세포의 종류와 분포가 달랐고, 동일한 종류이 상피세포라도 부위별로 미세구조가 다르게 관찰되었다.

• 한국발생생물학회지:발생과생식
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• 제27권3호
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• pp.149-157
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• 2023

We investigated the involvement of autophagy with steroidogenesis in testicular Leydig cells. Human chorionic gonadotropin (hCG)-stimulated T production in Leydig cells was not remarkably altered in the presence of an autophagy inhibitor 3-methyladenine (3-MA). Although pretreatment with 3-MA demonstrated a tendency to decrease hCG-induced T production, the differences were significant only at a higher time point of 24 h following hCG. Microtubule associated protein light chain 3 (LC3)-II was detectable in the control cells in all the experiments. The hCG-induced increase in steroidogenic acute regulatory protein (StAR) and cytochrome P450 side chain cleave (P450scc) protein levels were not significantly altered by 3-MA. Leydig cells isolated from immature rat testes 12 h following hCG treatment showed relatively increased levels of LC3-II protein compared to the control group. Furthermore, LC3-II levels shown in these cells reached almost the identical to those from normal adult testes. However, LC3-II protein levels were almost comparable or even slightly lower than the controls at 48 h following hCG. Expression of StAR and P450scc was upregulated at both 12 and 48 h after hCG. We also used MA-10 cells, the mouse Leydig cell line, in this experiment. When dibutyryl cyclic-AMP was treated with MA-10 cells, P4 levels were significantly increased in the cell culture medium. However, P4 levels tended to decrease in the presence of 3-MA, but the difference was not statistically significant. This was consistent with the results of the rat Leydig cell experiments. Together, we believe that although autophagy participates in steroidogenesis and enhances steroidogenic efficacy of Leydig cells, it may not be a decisive cellular process for steroidogenesis, specifically in the mature Leydig cells.

• 한국수정란이식학회:학술대회논문집
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• 한국수정란이식학회 2002년도 국제심포지엄
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• pp.63-65
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• 2002

Identification of spermatogonial stem cell-specific surface molecules is important in understanding the molecular mechanisms underlying the maintenance and differentiation of these cells. We have found that spermatogonia from busulfan treated mice expressed an autoantigen that distinguishes between undifferentiated and differentiated spermatogonia. Four to six weeks after busulfan treatment, germ cells located in the basal compartment of seminiferous epithelium show isotype-specific IgG deposits that form due to autoimmunity. Before busulfan treatment, the level of testicular IgG was very low but IgG levels began to increase after week 4 and peaked at week 6. When cells from the busulfan treated testis were analyzed using laser scanning cytomeoy (LSC), the frequency of cells positive for IgG deposits, 6-integrin, and 1-integrin were 16.5${\pm}$3.8%, 11.8${\pm}$2.6%, and 9.0${\pm}$ 1.4%, respectively. Immunofluorescent staining suggested that most, if not all of the cells with IgG-deposits isolated from a laminin-coated dish, were also positive for a spermatogonial stem cell marker \ulcorner6-integrins as well as for a germ cell-specific marker TRA 98. We determined serum and intratesticular IgG levels and the soundness of seminiferous tubule basement membrane from busulfan treated mice using electron microscopy, in order to study the mechanism responsible for IgG deposits in spermatogonia. We found that the basement membranes of seminiferous tubules from busulfan treated mice were severely impaired when compared to those of normal adult, neonates and w/wv mice. Furthermore, new blood cells were observed in the surface of the damaged basement membrane along the seminiferous tubules. These results suggest that the IgG in spermatogonial stem cells accumulates from circulating blood through the impaired basement membranes induced by busulfan treatment. Taken together, our study suggests that IgG can be used as a new marker for undifferentiated spermatogonia cells.

• 한국동물학회지
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• 제32권3호
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• pp.264-274
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• 1989

본 실험은 생쥐에서 부정소액내의 단백질 성분과 조성에 미치는 정소액과 정자의 영향을 알아보기 위하여, 성적 성숙시기에 따라 정소와 부정소의 조직분화 양상을 관찰하였으며, 강소분화 특징에 따라 체취한 부정소액은 전기영동법으로 단백질을 분석하여 다음과 같은 결과를 얻었다. 정소와 부정소는 생후 10일군에서 미분화 상태였고, 20일군에서 세정관의 강소는 형성되지 않았으나 부정소는 두부에서 미부에까지 강소가 형성되었으며, 35일군에서는 정소내 세정관의 강소가 형성되었고 정세포는 정자로 분화되었고, 부정소의 상피세포는 principal cell과 clear cell로 분화되었으나 부정소로 유입된 정자는 없었다. 80일군에서는 정소와 부정소가 완전히 본화되었고 부정소로 유입된 많은정자가 관찰되었다. 그리고 부정소액의 전기영동상에는 혈청내의 성분과 다른 단백질이 모두 28종이 나타났는데, 그 중 12종은 부정소액에만 존재하는 부정소 특이단백질이었고, 16종은 정소액에도 공통으로 존재하는 단백질이었다. 또한 이 단백질들은 성숙시기에 따라 종류가 다르게 나타났으며, 성체에서 나타난 3종의 단백질은 부정소의 부위에 따라 양적인 변화를 나타냈다. 이상의 결과로 보아 부정소액내 단백질의 성분과 조성은 정자를 포함한 정소액의 유입과 부정소 상피세포의 분비 및 흡수의 조절작용에 의하여 변화되는 것으로 사료된다. 따라서 부정소액내의 TEP와 ESP는 부정소 정자의 성숙에 어떤 중요한 역할을 할 것으로 사료된다.

• 대한수의학회지
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• 제59권2호
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• pp.59-67
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• 2019

We investigated whether ${\beta}$-carotene (${\beta}-CA$) or ellagic acid (EA), originating from various fruits and vegetables, has a preventive effect against male infertility induced by exogenous scrotal hyperthermia. ICR adult mice were intraperitoneally treated with 10 mg/kg of ${\beta}-CA$ or EA daily for 13 days consecutively. During this time, mice were subjected to transient scrotal heat stress in a water bath at $43^{\circ}C$ for 20 min on day 7, and their testes and blood were obtained on day 14 for histopathologic and biochemical analyses. Heat stress induced significant testicular weight reduction, germ cell loss and degeneration, as well as abnormal localization of phospholipid hydroperoxide glutathione peroxidase (PHGPx) and manganese superoxide dismutase (MnSOD) in spermatogenic and Leydig cells. Heat stress also altered the levels of oxidative stress (lipid peroxidation, SOD activity, and PHGPx, MnSOD, and $HIF-1{\alpha}$ mRNAs), apoptosis (Bax, Bcl-xL, caspase 3, $NF-{\kappa}B$, and $TGF-{\beta}1$ mRNAs), and androgen biosynthesis (serological testosterone concentration and $3{\beta}$-hydroxysteroid dehydrogenase mRNA) in testes. These changes were all improved significantly by ${\beta}-CA$ treatment, but only slightly improved by EA treatment. These findings indicate that ${\beta}-CA$, through modulations of oxidative stress, apoptosis, and androgen biosynthesis, is a potent preventive agent against testicular injuries induced by scrotal hyperthermia.