GnRH类似物对大鼠睾丸间质细胞MAPK的影响

目的:研究大鼠睾丸间质细胞中GnRH激动剂(GnRHa)和拮抗剂(GnRHant)对促分裂原活化蛋白激酶(MAPK)途径的影响。方法:原代培养大鼠睾丸间质细胞24h后,血清饥饿2h。GnRHa(10-7mol/L)或Gn-RHant(10-6mol/L)处理大鼠睾丸间质细胞不同时间(0、5、15、30、60、90min)后,Western印迹检测磷酸化ERK(p-ERK)和磷酸化p38(p-p38)蛋白水平以0min组为对照。此外,不同终浓度(1、5、10、20μmol/L)的PKC抑制剂GF109203X和GnRHa或GnRHant共同作用后,Western印迹检测p-ERK蛋白水平。结果:GnRHa或GnRHant刺激间质细胞不同时间后,p-p38蛋白水平与对照组比较均无显著性差异(P>0.05);加入不同终浓度的PKC抑制剂GF109203X处理细胞20min后,再用GnRHa刺激细胞10min,发现GF109203X在终浓度为10μmol/L和20μmol/L时p-ERK水平显著下降(P<0.05)。GnRHant刺激间质细胞后,p-ERK水平在15min时升高了约65%(P<0.05);30min时升高了约81%,达到最高水平(P<0.05);60min时开始下降,90min时恢复到基础水平。不同浓度GF109203X处理细胞20min后,再用GnRHant刺激细胞30min,p-ERK水平与对照组比较无显著性差异(P>0.05)。结论:GnRHa可能通过PKC途径来诱导ERKMAPK信号通路的活化,而GnRHant对ERKMAPK信号通路的激活作用并非通过PKC途径来实现的;p38可能不参与间质细胞中GnRH类似物作用的MAPK分子信号通路。     

卵泡抑制蛋白对大鼠睾丸间质细胞分泌睾酮的影响

目的 :探讨卵泡抑制蛋白 ( follistatin,FS)对离体大鼠睾丸间质细胞 ( L eydigcell)分泌睾酮的影响。方法 :用 Percoll梯度分离法分离和培养 Wistar雄鼠 ( 2 2 0～ 2 50 g)睾丸的 L eydig细胞。分别观察 FS( rh FS-2 88)、Ca2 +以及 FS加 Ca2 +在基础状态 (不加h CG)和刺激状态 ( h CG 1 .0 IU/ L )对 Leydig细胞分泌睾酮的影响。结果 :FS抑制基础和刺激状态的睾酮分泌 ,并与剂量相关。 Ca2 +亦有抑制作用 ,但在 1 0 0 mmol/ L时出现逸脱现象。FS加 Ca2 +表现为与剂量相关的抑制作用。结果 :FS呈剂量依赖性抑制睾酮分泌 ,Ca2 +可能是 FS的第二信使 ,单独高浓度 Ca2 +出现抑制逸脱的机制未明。     

Effect of cyclosporine on steroidogenesis in rat Leydig cells

Cyclosporine induces hypoandrogenism in adult male rats. In order to assess whether this effect of CsA may be due to a direct inhibitory effect on Leydig cell function, CsA (0, 50, 500, and 5000 ng/ml) was added to a collagenase-dispersed mixed Leydig cell preparation and incubated with and without hCG (0, 0.1, 0.3, 1.0, 3.0, and 10.0 ng/ml). Testosterone (T) production, mitochondrial cholesterol side chain cleavage (CSCC) and microsomal 17,20-desmolase enzyme activities in Leydig cells were determined after 3 hr of incubation. In the absence of CsA, stimulation of T production was maximal (about 16-fold) with 1.0 ng/ml hCG. With 50 and 500 ng/ml CsA there were no changes in either the hCG-stimulated T levels or the two enzymatic activities. However, 5000 ng/ml CsA significantly (P less than 0.05) reduced the hCG (1 ng/ml)-stimulated T levels, CSCC and 17,20-desmolase activities. The high dosage of CsA (5000 ng/ml) also caused a significant decrease in cell viability (P less than 0.05) during the incubation period. These effects of CsA were not due to cremophor EL, the CsA vehicle. This in vitro data indicate that high dosages of CsA (greater than or equal to 5000 ng/ml) appear to have a cytotoxic effect on rat Leydig cells that results in a decrease in T production. However, lower doses of CsA (less than 500 ng/ml) do not have any direct inhibitory effect on the rat Leydig cells, suggesting that the hypoandrogenic effect of in vivo CsA in rats is not due to any direct effect on the testis.     

Effects of varicocele on testosterone, apoptosis and expression of StAR mRNA in rat Leydig cells

The aim of this study was to explore the effects of varicocele on the morphology and function of Leydig cells in the rat testis. Forty male Sprague-Dawley rats were divided into two groups: the experimental group underwent surgery to create a left varicocele (VC), and the control group underwent a sham operation. Serum testosterone and intratesticular testosterone levels were measured using a radioimmunoassay after 4 and 8 weeks of operation. Leydig cells were studied for apoptosis and expression of steroidogenetic acute regulatory (StAR) protein mRNA levels. Serum testosterone levels declined after 4 and 8 weeks of operation but were not significant (P>0.05). However, the intratesticular testosterone levels after 8 weeks were significantly decreased compared with the control group (P<0.01). The mean apoptosis index of Leydig cells in the experimental group was significantly higher than that in the control group after 4 or 8 weeks (P<0.01). StAR mRNA levels in the Leydig cells of the experimental group were significantly lower compared to those of the control group (P<0.01). Our data show that varicocele did impair Leydig cell function by increasing apoptosis and suppressing the expression of the StAR protein.     

大鼠睾丸Leydig细胞体外增殖研究

目的:探讨通过优化细胞培养体系,实现Leydig细胞的体外增殖培养。方法:联合应用胶原酶消化、不锈钢滤网过滤及差速贴壁法获得3周龄雄性Wistar大鼠睾丸Leydig细胞,贴壁细胞以DMEM/F12培养液及优化培养体系培养,MTT法、细胞计数法检测培养细胞的增殖能力;分别对原代培养2h、4d细胞进行3β-HSD免疫化学染色及流式细胞术分析,检测细胞成分,同时检测培养细胞睾酮在hCG刺激下分泌能力变化。结果:优化培养体系能够明显促进3周龄大鼠睾丸Leydig细胞大量增殖,群体倍增时间为(2.26±0.31)d,传统培养体系培养细胞群体倍增时间为(16.32±2.14)d,两者差异有显著性(P<0.05);原代细胞经流式细胞术鉴定,3β-HSD阳性细胞所占比例分别为(54.3±7.1)%,培养4d后,增殖细胞3β-HSD阳性细胞率为(93.6±4.6)%。增殖细胞均有睾酮生成功能,在hCG刺激下睾酮分泌均明显上升(P<0.05)。结论:优化培养体系能够促进差速贴壁法获得的睾丸Leydig细胞大量增殖。     

Effects of gossypol on rat Sertoli and Leydig cells in primary culture and established cell lines

In order to evaluate the direct effect of gossypol on testicular cells, we used primary cultures of rat Leydig and Sertoli cells. No alteration in Leydig cell survival, morphology, or testosterone production was seen during three days of culture with up to 3 microgram/ml gossypol. With higher concentrations (3 to 7 microgram/ml) of gossypol, there was a reduction in cell survival but no change in androgen secretion. In contrast, there was a marked change in Sertoli cell morphology after five days of gossypol treatment. Large vacuoles and electron dense granules appeared in the cytoplasm, but these effects were reversed within six days of removing gossypol from the medium. There was a significant decrease in androgen binding protein (ABP) secretion by Sertoli cells in the presence of gossypol. We also tested the effect of gossypol on the growth of three established cell lines. Two Sertoli-derived cell lines, TM4 and TR-ST, were more sensitive than a Leydig-derived cell line (TM3). These results suggest that, of the somatic cell types in the testis, the Sertoli cells are most sensitive to gossypol.     

Effects of luteinizing hormone and androgen on the development of rat progenitor Leydig cells in vitro and in vivo

Progenitor Leydig cells are derived from stem cells. The proliferation and differentiation of progenitor Leydig cells significantly contributes to Leydig cell number during puberty. However, the regulation of these processes remains unclear. The objective of the present study was to determine whether luteinizing hormone （LH） or androgen contributes to the proliferation and differentiation of progenitor Leydig cells. Fourteen-day-old male Sprague-Dawley rats were treated for 7 days with NalGlu, which is a gonadotropin- releasing hormone antagonist, to reduce the secretion of LH in the pituitary and thus, androgen in the testis. Rats were co-administered with LH or 7a-methyl-nortestosterone （MENT）, which is an androgen resistant to metabolism by 5a-reductase 1 in progenitor Leydig cells, and the subsequent effects of LH or androgen were measured. 3H-Thymidine was also intravenously injected into rats to study thymidine incorporation in progenitor Leydig cells. Progenitor Leydig cells were examined. NalGlu administration reduced progenitor Leydig cell proliferation by 83%. In addition, LH or MENT treatment restored Leydig cell proliferative capacity to 73% or 50% of control, respectively. The messenger RNA levels of proliferation-related genes were measured using real-time PCR. The expression levels of Igfl, Lifr, Pdgfra, Bcl2, Ccnd3and Pcnawere upregulated by MENT, and those of Pdgfra, Ccnd3and Pcnawere upregulated by LH. Both LH and MENT stimulated the differentiation of progenitor Leydig cells in vitro. We concluded that both LH and MENT were involved in regulating the development of progenitor Leydig cells.     

Intracellular pathways of endocytosed tracers in Leydig cells of the rat

The endocytic activity of Leydig cells was examined by electron microscopy following the injection, into the interstitial space, of tracers used to examine fluid-phase endocytosis, ie, native ferritin and horseradish peroxidase-colloidal gold (horseradish peroxidase-gold), and adsorptive endocytosis, ie, cationic ferritin. At 5 minutes after injection, native ferritin or horseradish peroxidase-gold was present in the interstitial space and free in the lumen of large endocytic vesicles forming at the cell surface. At 15 minutes, these tracers appeared in the matrix of pale multivesicular bodies, while at 30 minutes and 1 hour, the matrix of dense multivesicular bodies became labeled. Beginning at 1 hour, dense membrane-delimited bodies identified cytochemically as lysosomes were labeled. In the case of cationic ferritin, two distinct pathways were taken. In one pathway, cationic ferritin was observed 5 minutes after injection bound to the plasma membrane of Leydig cells and to the membrane of small and large endocytic vesicles. At subsequent time intervals, cationic ferritin appeared consecutively in pale, dense multivesicular bodies and lysosomes. In a second pathway, cationic ferritin was observed at 5, 15, and 30 minutes bound to the membrane of vesicles of intermediate size seen near the cell surface. At 1, 1 1/2 and 2 hours, cationic ferritin-containing intermediate vesicles appeared in increasing number in the Golgi region. However, cationic ferritin was never observed in the Golgi saccules themselves. At later time intervals (3-6 hours), intermediate vesicles labeled with cationic ferritin progressively disappeared from the Golgi region and the cell. Thus in Leydig cells, while fluid-phase tracers reached lysosomes exclusively, cationic ferritin, a tracer of adsorptive endocytosis, not only reached the lysosomes, but was also carried by the intermediate vesicles to the Golgi region of the cell.     

老年大鼠睾丸间质细胞形态及睾酮合成功能变化的研究

目的 探讨衰老对睾丸间质细胞的形态及功能的影响.方法 青年(3月龄)及老年(24月龄)清洁级雄性SD大鼠各10只,麻醉后取血清检测总睾酮浓度,取睾丸组织用HE染色观察睾丸组织形态学变化,并用电镜观察睾丸间质细胞的超微结构改变.通过密度梯度离心分离原代睾丸间质细胞,并用LH刺激睾酮分泌后用Western blot比较青年组和老年组睾丸间质细胞类固醇合成快速调节蛋白(steroidogenic acute regulatory protein,StAR)表达水平的差异,并用ELISA法检测其睾酮分泌的差异.结果 HE染色显示老年大鼠睾丸呈老年退行性改变,电镜下观察到老年大鼠睾丸间质细胞线粒体水肿,线粒体嵴消失.老年大鼠血清睾酮水平显著低于青年组(P＜ 0.05).原代培养的睾丸间质细胞无论LH刺激与否,老年组细胞上清中睾酮浓度及StAR蛋白表达水平均显著低于青年组(LH刺激时P＜0.01,无LH刺激时P＜0.05).结论 衰老造成的睾丸间质细胞线粒体水肿及LH诱导的StAR蛋白表达水平下降与其睾酮合成能力降低密切相关.     

Morphologic response of rat Leydig cells to hemicastration

Hemiorchidectomized rats were followed up to 15 days postsurgery for morphologic evaluation of compensatory testicular response and its correlation to serum testosterone levels. Although gross compensatory testicular hypertrophy (CTH) was not noted, an enlarged interstitium was observed with hypertrophy and hyperplasia of Leydig cells with morphologic changes suggestive of increased cellular activity. These histologic changes were accompanied by compensatory testicular hypersecretion (CTHS) illustrated by the return of the serum testosterone levels to near the intact-control value in the later groups. Ultrastructural studies of the Leydig cells indicated an increase in the amount of smooth endoplasmic reticulum as an underlying mechanism for this response. In view of the previously reported normal serum luteinizing hormone levels after hemicastration, the compensatory hyperactivity/hypersecretion should be considered primarily an intrinsic Leydig cell response, not related to changes in the hypothalamo-hypophyseal axis.     

Leydig cell cooperation in vitro: evidence for communication between adult rat Leydig cells

In short-term incubations (32 C, 3 h) of purified adult rat Leydig cells, increasing the density from 5000 to 50,000 cells/16 mm diameter culture well caused a significant increase in human chorionic gonadotropin (hCG)-stimulated testosterone secretion/cell. Density-dependent stimulation was also observed under basal conditions and in the presence of dibutyryl cyclic adenosine monophosphate, luteinizing hormone-releasing hormone, or 22-hydroxy cholesterol. In contrast, increasing the incubation density of purified Leydig cells by addition of other testicular cells had no effect on basal or hCG-stimulated testosterone secretion. hCG-stimulated testosterone secretion by Leydig cells incubated at low density was also increased by addition of Leydig cell-conditioned medium. This stimulatory activity was removed by charcoal extraction and by ultrafiltration (approximately 30 kDa cut-off). The data indicate that Leydig cells cooperate by secretion of low molecular weight, cell-specific stimulatory factors that support Leydig cell steroidogenesis in vitro, and may also play a role in regulating Leydig cell function in vivo.     

Chronic stress induces ageing-associated degeneration in rat Leydig cells

Several studies have suggested that stress and ageing exert inhibitory effects on rat Leydig cells. In a pattern similar to the normal process of Leydig cell ageing, stress-mediated increases in glucocorticoid levels inhibit steroidogenic enzyme expression that then results in decreased testosterone secretion. We hypothesized that chronic stress accelerates the degenerative changes associated with ageing in Leydig cells. To test this hypothesis, we established a model of chronic stress to evaluate stress-induced morphological and functional alterations in Brown Norway rat Leydig cells; additionally, intracellular lipofuscin levels, reactive oxygen species (ROS) levels and DNA damage were assessed. The results showed that chronic stress accelerated ageing-related changes: ultrastructural alterations associated with ageing, cellular lipofuscin accumulation, increased ROS levels and more extensive DNA damage were observed. Additionally, testosterone levels were decreased. This study sheds new light on the idea that chronic stress contributes to the degenerative changes associated with ageing in rat Leydig cells in vivo.     

Effects of follistatin on testosterone secretion of rat Leydig cell in vitro

<正> Objective:To investigate the effects of follistatin(rhFS-288) on biosynthesis andsecretion of testosterone in rat Leydig cell in vitro.Methods:Leydig cells were isolated from Wistar rat testes by a discontinuous Per-coll gradient procedure.Purified cells were incubated in 24-well plate(10~5 cell/ml/well)and maintained for 24 h in a CO_2 incubator,rhFS-288 and Ca~(2+) were added to the wellsindependently or jointly in both baseline (without hCG) and stimulation condition (1.0IU/ml of hCG) to observe the change of testosterone concentration in the media.Results:rhFS-288 showed a dose-dependent inhibiting effect on testosterone releasein baseline and stimulating condition.Ca~(2+) presented inhibitory effect either.Whereas,escape phenomenon emerged while Ca~(2+) concentration reached to 100 mmol/L.A com-bination of rhFS-288 with Ca~(2+) displayed a dose-dependent inhibition on testosterone se-cretion.Conclusion:rhFS-288 inhibits testosterone secretion in a dose-dependent manner.Calcium is thought to be the second messenger of FS action.The mechanism of escapephenomenon during high dose of Ca~(2+) along is unknown.     

Demonstration of direct effect of serotonin on rat Leydig cells.

Nachweis eines direkten Einflusses von Serotonin auf die Testosteron-Synthese der Leydig-Zellen bei der Ratte Im Tierexperiment an hypophysektomierten Ratten wurde der Einfluß von Serotonin (30 mg/kg über 4 Tage, beginnend 24 Std. nach der Hypophysektomie bei gleichzeitiger Gabe von 400 IE HCG und 1 IE FSH tägl.) auf die Testosteron-Synthese der Leydig-Zellen untersucht. Es wurde beobachtet, daß Plasma- und Testis-Testosteron unter Serotonin deutlich abfielen, während die Konzentration von Pregnenolon im Hoden signifikant höher lag als in der Kontrollgruppe. Die Testis-HCG-Rezeptor-Bindungskapazität war in beiden Gruppen gleich.     

Effects of bromocriptine on pituitary-testicular function in the rat: possible inhibition of in vitro production of androgen by Leydig cells

The effect of bromocriptine (BR) on pituitary-testicular function has been investigated in vivo and in vitro in adult male rats. Testosterone production in vitro by collagenase- dispersed Leydig cells from 84-day-old rats was evaluated in the presence and absence of hCG and/or different doses of BR. In the presence of 1.5 X 10^-5 M BR, both basal and hCG-stimulated testosterone production were decreased whereas at lower doses BR was ineffective. In vivo 60-day-old rats were injected sc with BR (150 μg/rat or 750 μg/rat twice daily) or vehicle for 24 days. This treatment reduced the plasma level and pituitary content of prolactin, slightly increased the plasma levels of LH and FSH but did not affect pituitary gonadotrophin content. Irrespective of the dose of BR injected, plasma levels of androgen did not change, but with the large dose of BR a decrease in testicular content of testosterone (P = 0.05) was observed. In the same animals the number of LH/hCG receptors was significantly reduced, and the sensitivity of the isolated Leydig cells to hCG stimulation in vitro was reduced; however, both the basal secretion and the maximum testosterone response to hCG were unaffected. These results show impairment of pituitary-testicular function in BR-treated rats, either as a result of BR-induced hypoprolactinaemia or as a consequence of direct effects of BR on the Leydig cells.     

A morphological study of the testis in patients with idiopathic male infertility--quantification and ultrastructure of Leydig cells

Testicular biopsy was performed on 51 patients with idiopathic male infertility and 13 normal fertile adults. The biopsied specimens were examined by light and electron microscopy. A quantitative evaluation of Leydig cell number was made by determining the mean number of Leydig cells per one cross section of seminiferous tubule in the entire histologic section of each specimen, which was defined as "Leydig cell index". In both oligospermic and azoospermic groups, the Leydig cell index was significantly elevated as compared with in that the normal group, which indicated the presence of Leydig cell hyperplasia in the infertile testis. In addition, this index significantly correlated with serum luteinizing hormone and follicle stimulating hormone levels but not with serum testosterone level. Leydig cells were classified into three types based upon their morphological characteristics. Type I Leydig cells were functionally active and mature ones, with a smooth-surfaced round or oval nucleus, had abundant smooth endoplasmic reticulum (SER), characteristic mitochondria and relatively few lysosomes. Type II Leydig cells were functionally less active, and contained an irregularly shaped nucleus, less abundant SER, mitochondria with undeveloped cristae and relatively few lysosomes. Type III Leydig cells included immature and regressive cells, which were considered to have almost no hormone secreting function. The immature Leydig cells were spindle-shaped and had few cell organelles. The regressive cells had poorly developed SER and many secondary lysosomes in the cytoplasm. In the normal group, type I Leydig cells are the most numerous but in the infertile groups type II Leydig cells are the most numerous. However, in the latter, there is no significant difference in relative number of each Leydig cell type among the groups classified according to the number of sperm or Leydig cell. In summary, Leydig cell hyperplasia observed in the testis of the infertile patients was supposed to be induced by the elevation of the serum LH. Despite of the significant increase in Leydig cell number, the serum testosterone was not elevated, and this was considered to be due to the fact that the hyperplastic cells are mainly composed of functionally less active type II Leydig cells. In addition, especially in the severe hyperplastic group, not only type II but also type I Leydig cells increased in number. However, in this group, the serum testosterone level was not elevated. Therefore, the function of type I Leydig cells was suggested to be impaired.     

Evidence for the presence of androgen receptors in purified rat Leydig cells

Leydig cells, purified on two sequential Percoll gradients to purities of 89 +/- 1%, were used to study the binding of 17-beta-hydroxy-17 alpha-methyl-estra-4,9,11-triene-3-one(3H-R1881). The accumulation of 3H-R1881 in the nuclear fraction of these cells was time- and temperature-dependent. Specific binding was saturable with an apparent Ka of 0.14 nM-1 and a single class of binding sites at a concentration of 721 fmol/mg DNA. A fraction of the bound radioactivity in the nuclear pellet could be extracted with 0.4 M KCl, and a portion of this extracted steroid was associated with macromolecular species. The nuclear accumulation was androgen-specific. These data are consistent with the presence of androgen receptors in rat Leydig cells.     

Green tea polyphenols inhibit testosterone production in rat Leydig cells

This study investigated the acute effects of green tea extract （GTE） and its polyphenol constituents, （-）-epigallocatechin-3-gallate （EGCG） and （-）-epicatechin （EC）, on basal and stimulated testosterone production by rat Leydig cells in vitro. Leydig cells purified in a Percoll gradient were incubated for 3 h with GTE, EGCG or EC and the testosterone precursor androstenedione, in the presence or absence of either protein kinase A （PKA） or protein kinase C （PKC） activators. The reversibility of the effect was studied by pretreating cells for 15 min with GTE or EGCG, allowing them to recover for 1 h and challenging them for 2 h with human chorionic gonadotropin （hCG）, luteinizing hormone releasing hormone （LHRH）, 22（R）-hydroxycholesterol or androstenedione. GTE and EGCG, but not EC, inhibited both basal and kinase-stimulated testosterone production. Under the pretreatment conditions, the inhibitory effect of the higher concentration of GTE/EGCG on hCG/LHRH-stimulated or 22（R）- hydroxycholesterol-induced testosterone production was maintained, whereas androstenedione-supported testosterone production returned to control levels. At the lower concentration of GTE/EGCG, the inhibitory effect of these polyphenols on 22（R）-hydroxycholesterol-supported testosterone production was reversed. The inhibitory effects of GTE may be explained by the action of its principal component, EGCG, and the presence of a gallate group in its structure seems important for its high efficacy in inhibiting testosterone production. The mechanisms underlying the effects of GTE and EGCG involve the inhibition of the PKA/PKC signalling pathways, as well as the inhibition of P450 side-chain cleavage enzyme and 17β-hydroxysteroid dehydrogenase function.     

Partial characterization of alcohol dehydrogenase activity in purified rat Leydig cells

Ethanol metabolism to acetaldehyde by NAD+-dependent alcohol dehydrogenase (ADH) activity reduces, in part, androgen secretion by rat Leydig cells. ADH in Leydig cells is proposed to decrease the NAD+/NADH ratio and thereby inhibit NAD+-dependent delta 5-3 beta hydroxysteroid dehydrogenase-isomerase activity and increase NADH-dependent 5 alpha-androstane-3 beta-hydroxysteroid dehydrogenase activity. Although the reciprocal changes in these steroidogenic enzyme activities by ethanol are attributed to ADH activity, there is very little information about this enzyme in purified Leydig cells. The present studies examined specific characteristics of this enzyme in metrizamide-gradient purified Leydig cells. ADH activity was linear with respect to protein concentration and incubation time. The activity was concentrated in the soluble fraction, and the most effective cofactor was NAD+. The apparent Km for ethanol was 0.50 mM, and the Vmax was 53 nmol NADH/10 min/mg protein. When Leydig cell cytosol was incubated with a fixed ethanol concentration (50 mM) and increasing NAD+ and the data were plotted according to Lineweaver-Burk, a biphasic curve was observed with apparent Km's of 0.032 and 0.17 mM. The optimum pH for the enzyme was 8.2, and the enzyme was inhibited in a dose-dependent manner by 4-methylpyrazole. These studies further characterize ADH activity in purified Leydig cells and demonstrate that this enzyme exhibits many characteristics similar to the more widely studied liver enzyme(s).(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of Basella alba and Hibiscus macranthus extracts on testosterone production of adult rat and bull Leydig cells

Aim： To determine the androgenic effects of Basella alba and Hibiscus macranthus extracts in the rat and the bull, and to develop a novel in vitro test system using Leydig cells from bull testes. Methods： The effect of methanol extracts from both plants on testosterone production in isolated Leydig cells from the rat and the bull was analyzed using ^125I-radioimmunoassay （^125I-RIA）. Rat Leydig cells were obtained by common methods, whereas a novel technique was used to purify Leydig cells from bull testes. Results： Bull testes from the slaughter house were a cheap source of pure Leydig cells. In culture, these cells produced testosterone for 5-6 days, which can be stimulated by human chorionic gonadotrophin （hCG）. Basella alba extracts significantly enhanced testosterone production in bull and rat Leydig cells in a concentration-dependent manner. Hibiscus macranthus showed no androgenic effect but was shown to inhibit testosterone production at higher concentrations. Conclusion： Leydig cells purified from bull testes can be used as an alternative tool in experimental animal research. Certain fractions of Basella alba extract demonstrated androgenic potential whereas Hibiscus macranthus extracts did not.