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

• Journal of Animal Reproduction and Biotechnology
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• v.37 no.2
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• pp.87-95
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• 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
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• v.20 no.1
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• pp.11-22
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• 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.

• Proceedings of the Korean Society of Developmental Biology Conference
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• 2003.10a
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• pp.62-62
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• 2003

Aquaporin (AQP) family protein은 일종의 수분 전달 통로 역할을 하는 단백질로 AQP를 통한 수분의 조절은 삼투압을 통한 물의 이동과 함께 조직내 정상적인 수분의 상성 유지에 필수적이다. 현재까지 11종의 AQP이 신장·뇌·정소·안구 등에서 발현이 확인되었다. AQP9은 물 뿐 아니라 carbamide, polyol, purine, pyrimidine, urea, glycerol 등의 이동에 관여한다. 본 연구에서는 생쥐에서 출생 후 성체에 이르는 동안 정소 내 AQP9의 발현, Leydig cell의 분화에 따른 AQP9의 발현을 조사하였다. 1, 2, 4, 8주령의 정소로부터 semiquantitative RT-PCR 및 real time PCR 법으로 AQP9의 발현을 분석한 결과 1주령에서는 발현되지 않았고 2주령에서는 미량이 발현되기 시작하였고, 4주령에서는 성체의 1/2수준으로 발현량이 급격히 증가하였고 성체에서는 다량으로 발현됨이 확인되었다. Semiquantitative RT-PCR 법과 real time PCR법을 비교할 때 주령별 발현 양상은 유사하였으나 4주령과 성체에서는 두 시험법 사이에 양적인 차이가 있었다. 면역조직화학염색 결과 주로 Leydig cell에서 AQP9의 발현이 확인되었다. 성체의 정소 균질액의 Western blot 상에서 분자량 80, 55, 35 및 23 kDa의 항원이 검출되어 dimer, trimer 형태로 존재할 가능성과 당쇄 결합에 의한 단백질의 변형이 있는 것으로 추정된다. 미성숙 개체의 정소에서는 23 form이 확인되는 반면 성체에서는 35 kDa form이 주로 발현되므로 정소에서 발현되는 AQP9의 경우 Post-translation 수준에서 AQP9의 변형이 수반되는 것으로 사료되며 AQP9의 기능과의 연관성은 추후 연구되어야 할 것이다. Leydig cell은 fetal 및 adult type 2종의 세포가 정소발달 과정에 출현, 사멸, 분화하며 이들은 각기 정소발달, 성숙과 정자형성에 필요한 steroidogenesis에 관여한다. 정소 내 AQP9의 발현은 17beta HSD의 발현 양상과 같게 나타나므로 성적 성숙에 따른 정소 내 AQP9의 발현의 증가는 adult type Leydig cell의 분화와 관련된 것으로 추측된다. 성체의 정소로부터 분리한 Leydig cell-enriched culture에 hCG를 처리한 결과 배양체의 AQP9의 발현이 증가하므로 AQP9은 LH 수용체 하위 신호전달과정을 통해 Leydig cell의 steroidogenesis 또는 생성된 steroids의 분비에 요구되는 수분 및 중성용질의 이동에 관여하는 것으로 사료된다.

• Journal of Environmental Health Sciences
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• v.21 no.1
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• pp.68-73
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• 1995

This study of culturing testicular cell types in vitro has potential to be an invaluable tool for assessing the mechanisms of testicular toxicity, especially those of intragonadal interaction and spermatogenesis. Combined with the Sertoli/germ cell cultures, Leydig cells provide comprehensive and detailed information on the action of testicular toxicants at the level of the testis. Sertoli/germ cell were isolated and incubated well in vitro from 20~30 g rats and Leydig cells from 250~300 g rats. The Sertoli cells isolated from the testis of the SD rats grew into monolayer on about the 2nd~3rd day of culture, an appreciable cell increment being observed between the 4th~5th day. The Leydig cells isolated from the testis of the SD rats grew into a monolayer on about the 3rd-4th day of culture, an appreciable cell increment being observed between the 5th-7th day. These results suggest that Sertoli and Leydig cells can be cultured as a male fertility evaluation method alternative to the in vivo/conventional fertility test method and further study for the physio-chemical determination is needed.

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

• Korean Journal of Veterinary Research
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• v.43 no.4
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• pp.531-539
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• 2003

Changes in the rat testis interstitium from birth to adulthood were studied using Sprague Dawley rats of 1, 7, 14, 21, 28, 40, 60, and 90 days of age to investigate Leydig cell differentiation. In addition, serum testosterone concentrations and luteinizing hormone stimulated (LH; 100 ng/ml) testosterone secretory capacity per testis in vitro were determined via radioimmunoassay. Fetal Leydig cells were present in rat testes from birth to 21 days, and they were only steroidogenic cells in the testis at days 1 and 7. The average volume of a fetal Leydig cell and the absolute volume of fetal Leydig cell per testis were similar at all ages of experimental groups except at day 21 when lower values were observed for both parameters. The number of fetal Leydig cells per testis remained constant from birth through 21 days. Adult Leydig cells were recognized at day 14 and their absolute volume and number per testis increased linearly from 14 to 90 days. The average volume of an adult Leydig cell increased significantly with age and reached maximum size by 60 days of age where the volume was nearly three times bigger than that of at day 14. Total testosterone production per testis in vitro and serum testosterone concentrations were not significantly different at day 1 compared with 7, 14, and 21 days of age. Significant increases were observed at days 40 and 60. Values at days 60 and 90 were not significantly different.

• Development and Reproduction
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• v.9 no.2
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• pp.115-121
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• 2005

The present report aimed at evaluating the effect of bisphenol A(BPA) and diethylstilbestrol(DES) on Leydig or Sertoli cell-lines. To identify the differences in the susceptibility to BPA upon different cell-types, assay of the cell viability was done on TM3(Leydig cells) and TM4(Sertoli cells) cell-lines. The result indicates that Sertoli cells are more sensitive to low dose of BPA than Leydig cells. Also, the BPA- or DES-treated Sertoli cells showed a reduction of phospholipase D(PLD) activity identically. According to the confirmation of the mRNA expression of fas receptor and fas ligand in the BPA-treated cells, fas/fasL system activated by BPA will deliver the apoptosis signal onto Sertoli TM4 cells. However, Fas/FasL system was not activated in the DES-treated cells unlike the BPA-treated cells.

• Journal of Veterinary Clinics
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• v.36 no.2
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• pp.129-131
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• 2019

A 15-year-old intact Yorkshire terrier was presented with anorexia, lethargy, and a pale mucous membrane. A physical examination one year ago revealed right testis mass and subcutaneous petechia. Blood work revealed severe thrombocytopenia and mild anemia, and no abnormalities were found in serum chemistry or ultrasonography. The preoperative serum estrogen concentration was moderately elevated. The enlarged testis was surgically removed. A well-encapsulated mass composed of polyhedral or round with abundant eosinophilic cytoplasm containing fine granular or vacuolation were found in a histological examination of the removed tissue. The nuclei of tumor cells were round, and mitotic figures were low but neoplastic cells showed a mild invasive tendency to adjacent tissues with contained neoplastic cell emboli in one lymphatic lumen. A diagnosis of a malignant Leydig cell tumor was made. The patient recovered from surgery uneventfully, but his condition worsened despite repeated transfusions and supportive therapy, and he was euthanized according to the owner's decision. Leydig cell tumor should be included in estrogen toxicity associated with testicular mass.

• Asian-Australasian Journal of Animal Sciences
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• v.22 no.5
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• pp.618-625
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• 2009

The objective of this work was to study the direct effects of daidzein on steroidogenesis in cultured mouse Leydig cells. Adult mouse Leydig cells were purified by Percoll gradient centrifugation, and the cell purity was determined using a $3{\beta}$-hydroxysteroid dehydrogenase ($3{\beta}$-HSD) staining method. The purified Leydig cells were exposed to different concentrations ($10^{-7}$ M to $10^{-4}$ M) of daidzein for 24 h under basal and human chorionic gonadotropin (hCG)-stimulated conditions. The cell viability and testosterone production were determined, and the related mechanisms of daidzein action were also evaluated using the estrogen receptor antagonist ICI 182,780 and measuring the mRNA levels of steroidogenic acute regulatory protein (StAR), cholesterol side-chain cleavage enzyme (P450scc), and $3{\beta}$-HSD-1 involved in testosterone biosynthesis. The results revealed that daidzein did not influence cell viability. Daidzein increased both basal and hCG-stimulated testosterone production in a dose-dependent manner, and this effect was statistically significant at concentrations of $10^{-5}$ M and $10^{-4}$ M daidzein (p<0.05). ICI 182,780 had no influence on daidzein action. RTPCR results revealed that $10^{-5}$ M and $10^{-4}$ M daidzein did not exert any obvious influence on the mRNA level of P450scc in Leydig cells. However, in the presence of hCG, these concentrations of daidzein significantly increased the StAR and $3{\beta}$-HSD-1 mRNA levels (p<0.05), but in the absence of hCG, only $10^{-5}$ M and $10^{-4}$ M daidzein up-regulated the StAR and $3{\beta}$-HSD-1 mRNA expression (p<0.05), respectively. These results suggest that daidzein has direct effect on Leydig cells. Daidzein-induced increase of testosterone production is probably not mediated by the estrogen receptor but correlates with the increased mRNA levels of StAR and $3{\beta}$-HSD-1.