• Environmental Analysis Health and Toxicology
• /
• v.19 no.3
• /
• pp.261-269
• /
• 2004

Although the biological functions of metallothioneins (MTs) are still being investigated, they have been suggested to be involved in detoxification of heavy metals, scavenging of free radicals, and protection against alkylating agents. MTs have been reported to be induced in most of animal tissues by heavy metals such as zinc, copper, mercury and cadmium, and the proteins have binding affinities to the metals. However, the presence or induction of MTs was reported not to be clear in leydig cells, which produce testosterone for the maturation of spermatozoa in male testes. In this study, we investigated the inducibility of metallothionein isomers by cadmium in cultured mouse leydig cells. Total RNA was extracted from the near confluent grown leydig cells and RT-PCR was Performed using the Primers which were synthesized on the basis of MT-1, 2, 3 and 4 cDNA from GenBank database. As results, MT-1 and MT-2 mRNA were found to be expressed in cadmium non-treated control cells and MT 1 mRNA expression was dose-dependent when leydig cells were treated with cadmium chloride. But MT-3 which is known to be brain specific and MT-4 which is another isoform of metallothionein, were not expressed. Other genes induced or depressed in cadmium treated leydig cells were also identified by microarray techniques.

• Journal of Animal Reproduction and Biotechnology
• /
• v.37 no.2
• /
• pp.87-95
• /
• 2022

Receptor tyrosine kinase c-Kit, a marker found on interstitial cells of Cajal (ICCs), is expressed in Leydig cells, which are testicular interstitial cells. The expression of other ICC markers has not yet been reported. In this study, we investigated the expression of c-Kit and anoctamin 1 (ANO1), another ICC marker, in mouse testes. In addition, the relationship between c-Kit and ANO1 expression and Leydig cell function was investigated. We observed that c-Kit and ANO1 were predominantly expressed in mouse Leydig cells. The mRNA and protein of c-Kit and ANO1 were expressed in TM3, a mouse Leydig cell line. LH induced an increase in intracellular Ca²⁺ concentration, membrane depolarization, and testosterone secretion, whereas these signals were inhibited in the presence of c-Kit and ANO1 inhibitors. These results show that c-Kit and ANO1 are expressed in Leydig cells and are involved in testosterone secretion. Our findings suggest that Leydig cells may act as ICCs in testosterone secretion.

• Development and Reproduction
• /
• v.26 no.3
• /
• pp.99-105
• /
• 2022

Styrene is the precursor of polystyrene. Human exposure to styrene could occur in occupational and residential settings and via food intake. Styrene is metabolized to styrene-7,8-oxide by cytochrome P450 enzyme. In the present study, we investigated the cytotoxicity mediated by styrene and styrene-7,8-oxide in TM3 testicular Leydig cells in vitro. We first monitored the nuclear fragmentation in Leydig cells after exposure to styrene or styrene-7,8-oxide. Hoechst 33258 cell staining showed that styrene exposure in TM3 Leydig cells did not exhibit nuclear fragmentation at any concentration. In contrast, nuclear fragmentation was seen in styrene-7,8-oxide-exposed cells. These results indicate that cytotoxicity-mediated cell death in Leydig cells is more susceptible to styrene-7,8-oxide than to styrene. Following styrene treatment, procaspase-3 and XIAP protein levels did not show significant changes, and cleaved (active) forms of caspase-3 were not detected. Consistent with the western blot results, the active forms of caspase-3 and XIAP proteins were not prominently altered in the cytoplasm of cells treated with styrene. In contrast to styrene, styrene-7,8-oxide induced cell death in an apoptotic fashion, as seen in caspase-3 activation and increased the expression of XIAP proteins. Taken together, the results obtained in this study demonstrate a fundamental idea that Leydig cells are capable of protecting themselves from cytotoxicity-mediated apoptosis as a result of styrene exposure in vitro. It remains unclear whether the steroid-producing function, i.e., steroidogenesis, of Leydig cells is also unaffected by exposure to styrene. Therefore, further studies are needed to elucidate the endocrine disrupting potential of styrene in Leydig cells.

• Development and Reproduction
• /
• v.27 no.3
• /
• pp.149-157
• /
• 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.

• Development and Reproduction
• /
• v.20 no.1
• /
• pp.11-22
• /
• 2016

The ultrastructures of germ cells and the functions of Leydig cells and Sertoli cells during spermatogenesis in male Kareius bicoloratus (Pleuronectidae) were investigated by electron microscope observation. Each of the well-developed Leydig cells during active maturation division and before spermiation contained an ovoid vesicular nucleus, a number of smooth endoplasmic reticula, well-developed tubular or vesicular mitochondrial cristae, and several lipid droplets in the cytoplasm. It is assumed that Leydig cells are typical steroidogenic cells showing cytological characteristics associated with male steroidogenesis. No cyclic structural changes in the Leydig cells were observed through the year. However, although no clear evidence of steroidogenesis or of any transfer of nutrients from the Sertoli cells to spermatogenic cells was observed, cyclic structural changes in the Sertoli cells were observed over the year. During the period of undischarged germ cell degeneration after spermiation, the Sertoli cells evidenced a lysosomal system associated with phagocytic function in the seminiferous lobules. In this study, the Sertoli cells function in phagocytosis and the resorption of products originating from degenerating spermatids and spermatozoa after spermiation. The spermatozoon lacks an acrosome, as have been shown in all teleost fish spermatozoa. The flagellum or sperm tail of this species evidences the typical 9+2 array of microtubules.

• Journal of Physiology & Pathology in Korean Medicine
• /
• v.29 no.5
• /
• pp.403-408
• /
• 2015

Traditionally, 5 kinds of fruits with "ja(子)" in their name, including Rubus coreanus, Schisandra chinensis, Lycinum chineuse, Torilidis fructus, and Cuscuta seed, collectively called Oja(五子), are long known to enhance stamina. In the present study, we replaced tosaja with gyeolmyeongja(Cassiae semen ) and examined the effects of extracts from these fruits on andropause. This study investigated the antioxidant effect and testosterone synthesis of Oja water extract on Leydig TM3 cells. To investigate whether hydrogen peroxide induces oxidative stress in Leydig cells, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide(MTT) assay, nitric oxide assay, and testosterone assay were performed on mouse Leydig TM3 cells. The results were obtained as follows: Leydig TM3 cells viability was assessed by a modified MTT assay, and the protection effect of Oja water extract against hydrogen peroxide-induced cell oxidative stress were examined by mitochondrial activity. Oja water extract could efficiently protect cytotoxicity induced by H₂O₂. Oja water extract promoted testosterone synthesis. These results suggest that Oja water extract has protective roles and promotes steroidogenesis in Leydig cells through its anti-oxidant action.

• Applied Microscopy
• /
• v.25 no.2
• /
• pp.11-19
• /
• 1995

This research was undertaken to determine the effect of cyclophosphamide(CP) on the Leydig cells and macrophages in the interstitial tissue of the mice(ICR strain). To evaluate how this drug could affect the these cells, during administration(200mg/kg) 1 time to 3 times at intervals of 48hrs. In the Leydig cells of the control and 1 time treated group, a number of microperoxisomes were observed interspersed among the network of smooth endoplasmic reticulum(SER) in cellular regions where the SER predominantes. Microperoxisomes were also founded in close proximity to the cell membrane. The interstitial tissue were exhibited degenerating Leydig cells but macrophages wer containd greatly increased numbers of cytoplasmic inclusion body and secondary lysosomes. In the 1 time treated group. A very small number of Leydig cells were observed, from 2 to 3 time group, but macrophages were more increased than 1 time group in number. CP thus offers a valuable opportunity to study further the interaction between Leydig cells and macrophages in the interstitial tissue. These alteration could be direct mediated by toxic effect of the drug on the interstitial tissue.

• The Korea Journal of Herbology
• /
• v.29 no.6
• /
• pp.111-116
• /
• 2014

Objectives : The purpose of this study was to estimate the antioxidant activity of water extract of Cnidii Fructus (CF) and Torilis Fructus (TF) in Leydig cells. Methods : Free radical scavenging activity of CF and TF against 2,2-diphenyl-1-picrylhydrazyl (DPPH) was determined spectrophotometrically. We investigated the effect of CF and TF in Leydig cells by MTT assay. The protective effects of CF and TF against hydrogen peroxide-induced oxidative stress in Leydig cells. Superoxide dismutase (SOD), and catalase activity assays were performed in Leydig cells. Results : The results showed that CF scavenged DPPH radical in a dose-dependent manner by up to 81.2%, TF scavenged DPPH radical in a dose-dependent manner by up to 63.8%. CF showed cell viability as 121.0, 132.7, 126.6% in 5, 10, $100{\mu}g/ml$ concentrations. TF showed cell viability as 127.5, 111.8% in 5, $100{\mu}g/ml$ concentraions, respectively. The hydrogen peroxide-induced cytotoxicity of Leydig cells were protected to 86.3% by CF at concentration of $10{\mu}g/ml$ and protected to 83.5% by TF at concentration of $100{\mu}g/ml$. Both CF and TF at all concentrations, SOD activity was not significantly changed. Catalase activity was significantly increased at 10, $100{\mu}g/ml$ concentrations of CF, respectively. TF's catalase activity showed no significant difference from that of the control. Conclusions : These results suggest that CF, as an antioxidant, protects Leydig cells in hydrogen peroxide-induced oxidative stress. know that "Kwangjebikeup" played a role in settlement and spreading of foreign knowledge to civilians.

• Journal of Animal Reproduction and Biotechnology
• /
• v.38 no.4
• /
• pp.291-299
• /
• 2023

Background: Leydig cells, crucial for testosterone production, express ion channels like ANO1 that influence hormone secretion. This study investigates the expression and role of the Tandem of P domains in a weak inward rectifying K⁺ channel-related Acid-Sensitive K⁺-1 (TASK-1) channel in these cells, exploring its impact on testicular function and steroidogenesis. Methods: TASK-1 expression in Leydig cells was confirmed using immunostaining, while RT-PCR and Western Blot (WB) validated its expression in the TM3 Leydig cell line. The effect of a TASK-1 channel blocker on cell viability was assessed through live/dead staining and MTT assays. Additionally, the blocker's effect on testosterone secretion was evaluated by measuring testosterone levels. Results: Immunohistochemical analysis revealed a predominant presence of TASK-1, along with c-Kit and ANO-1, in Leydig cells adjacent to seminiferous tubules and also in Sertoli and spermatogenic cells. Expression levels of TASK-1 mRNA and protein were significantly higher in TM3 Leydig cells compared to TM4 Sertoli cells. In addition, blocking TASK-1 in TM3 cells with ML365 induced cell death but did not affect LH-induced testosterone secretion. Conclusions: These findings suggest that TASK-1 in Leydig cells is crucial for their viability and proliferation, highlighting its potential importance in testicular physiology.

• Korean Journal of Animal Reproduction
• /
• v.9 no.1
• /
• pp.56-61
• /
• 1985

This study was conducted in order to obtain the number and size of Leydig cell in the testis in accordance with the growth of Korean native goat. Twenty-one Korean native male goats were examied, and was divided into seven groups by 4 weeks interval from 8 to 32 weeks of age. The results were as follows: 1. The number of Leydig cells in the interstitial tissue were increased in accordance with age. The number of cells were increased twice from 1.3 cells at 8 weeks to 2.6 cells at 12 weeks of age, from 2.6 cells at 12 weeks to 5.5 cells at 24 weeks, respectively. And the cells were slightly increased from 5.5 cells at 16 weeks to 6.7 cells at 32 weeks. 2. The smallest cells were found in goats at 8 weeks of age, being 8.2${\mu}$m, and it showed noncontinual growth among groups of 12, 16 and 20 weeks of age, recording 9.18, 8.82 and 9.05${\mu}$m respectively. Leyding cells in 24, 28 and 32 weeks of age showed matured size being 10.42, 10.81 and 10.67${\mu}$m respectively. 3. Rows or clusters of Leydig cells were scattered in the loose connective tissue around lymphatic simusoids and blood vessels. Nuclei of Leydig cells were found to have 2 types, one type was stained pale and the other were stained dark. As a result of this study, Leydig cells found in the interstitial tissue were considered to grow until 20 weeks of age after birth, and reached to almost full maturated form by 24 weeks of ages.