Improvement of Testicular Steroidogenesis Using Flavonoids and Isoflavonoids for Prevention of Late-Onset Male Hypogonadism

Androgen production, being important for male fertility, is mainly accomplished by the Leydig cells from the interstitial compartment of the testis. Testosterone plays a critical role in testis development, normal masculinization, and the maintenance of spermatogenesis. Within seminiferous tubules, appropriate Sertoli cell function is highly dependent on testicular androgen levels and is essential to initiate and maintain spermatogenesis. During aging, testosterone production by the testicular Leydig cells declines from the 30s in humans at a rate of 1% per year. This review outlines the recent findings regarding the use of flavonoids and isoflavonoids to improve testosterone production, contributing to normal spermatogenesis and preventing age-related degenerative diseases associated with testosterone deficiency. With the cumulation of information on the actions of different flavonoids and isoflavonoids on steroidogenesis in Leydig cells, we can now draw conclusions regarding the structure-activity relationship on androgen production. Indeed, flavonoids having a 5,7-dihydroxychromen-4-one backbone tend to increase the expression of the steroidogenic acute regulatory protein (StAR), being critical for the entry of cholesterol into the mitochondria, leading to increased testosterone production from testis Leydig cells. Therefore, flavonoids and isoflavonoids such as chrysin, apigenin, luteolin, quercetin, and daidzein may be effective in delaying the initiation of late-onset hypogonadism associated with aging in males.

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Cyclooxygenase-2 and Prostaglandin F2α in Syrian Hamster Leydig Cells: Inhibitory Role on Luteinizing Hormone/Human Chorionic Gonadotropin-Stimulated Testosterone Production

Endocrinology ◽

10.1210/en.2006-0090 ◽

2006 ◽

Vol 147 (9) ◽

pp. 4476-4485 ◽

Cited By ~ 44

Author(s):

Mónica B. Frungieri ◽

Silvia I. Gonzalez-Calvar ◽

Fernanda Parborell ◽

Martin Albrecht ◽

Artur Mayerhofer ◽

...

Keyword(s):

Chorionic Gonadotropin ◽

Human Chorionic Gonadotropin ◽

Syrian Hamster ◽

Leydig Cells ◽

Regulatory Protein ◽

Hydroxysteroid Dehydrogenase ◽

Steroidogenic Acute Regulatory Protein ◽

Testosterone Production ◽

Cox 2 ◽

Steroidogenic Acute Regulatory

We have previously found that cyclooxygenase-2 (COX-2), a key enzyme in the biosynthesis of prostaglandins (PGs), is present in the testicular interstitial cells of infertile men, whereas it is absent in human testes with no evident morphological changes or abnormalities. To find an animal model for further investigating COX-2 and its role in testicular steroidogenesis, we screened testes from adult species ranging from mice to monkeys. By using immunohistochemical assays, we found COX-2 expression only in Leydig cells of the reproductively active (peripubertal, pubertal, and adult) seasonal breeder Syrian hamster. COX-2 expression in hamster Leydig cells was confirmed by RT-PCR. In contrast, COX-1 expression was not detected in hamster testes. Because COX-2 expression implies PG synthesis, we investigated the effect of various PGs on testosterone production and found that PGF2α stood out because it significantly reduced human chorionic gonadotropin-stimulated testosterone release from isolated hamster Leydig cells in a dose-dependent manner. This mechanism involves a decreased expression of testicular steroidogenic acute regulatory protein and 17β-hydroxysteroid dehydrogenase. Testicular concentration and content of PGF2α in reproductively active hamsters as well as production of PGF2α from isolated hamster Leydig cells were also determined. Moreover, PGF2α receptors were localized in Leydig cells of hamsters and testicular biopsies from patients with Sertoli cell only and germ arrest syndromes. Thus, in this study, we described a COX-2-initiated pathway that via PGF2α production, PGF2α receptors, steroidogenic acute regulatory protein, and 17β-hydroxysteroid dehydrogenase represents a physiological local inhibitory system of human chorionic gonadotropin-stimulated testosterone production in the Syrian hamster testes.

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Effect of Glutathione Depletion on Leydig Cell Steroidogenesis in Young and Old Brown Norway Rats

Endocrinology ◽

10.1210/en.2007-1245 ◽

2008 ◽

Vol 149 (5) ◽

pp. 2612-2619 ◽

Cited By ~ 26

Author(s):

Haolin Chen ◽

Angela S. Pechenino ◽

June Liu ◽

Matthew C. Beattie ◽

Terry R. Brown ◽

...

Keyword(s):

Leydig Cell ◽

Leydig Cells ◽

Regulatory Protein ◽

Glutathione Depletion ◽

In Vivo Studies ◽

Brown Norway ◽

Causative Role ◽

Testosterone Production ◽

Age Related ◽

Norway Rats

Changes in the oxidant/antioxidant environment of aging Leydig cells have been shown to be correlated with the reduced ability of these cells to produce testosterone. With this in mind, we hypothesized that the experimental depletion of glutathione (GSH), an abundant Leydig cell intracellular antioxidant, might result in reduced testosterone production. Incubation of Leydig cells isolated from the testes of adult Brown Norway rats with buthionine sulfoximine (BSO) reduced GSH content by more than 70% and testosterone production by about 40%. The antioxidants vitamin E, N-tert-butyl-α-phenylnitrone and Trolox countered BSO’s effect on steroidogenesis but not on GSH depletion. Together, BSO and glutathione ethyl ester maintained intracellular GSH and also testosterone production, whereas 1,2-dithiole-3-thione, which increases intracellular GSH, increased testosterone production. In vivo studies also were conducted. Young (4 month old) and old (24 month old) rats were injected with BSO twice a day for 7 d, after which Leydig cells were isolated and analyzed in vitro. BSO treatment reduced Leydig cell GSH content by 70% and the ability of the Leydig cells to produce testosterone by more than 50%. As with aging, decreases were seen in LH-stimulated cAMP production, steroidogenic acute regulatory protein, cholesterol side-chain cleavage, 3β-hydroxysteroid dehydrogenase, and 17α-hydroxylase/17,20-lyase. The results of these studies, taken together, are consistent with the hypothesis that alteration in the oxidant/antioxidant environment may play a significant, causative role in the age-related reduced ability of Leydig cells to produce testosterone.

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Regulation of Leydig cells through a steroidogenic acute regulatory protein-independent pathway by a lipophilic factor from macrophages

Journal of Endocrinology ◽

10.1677/joe.0.1580267 ◽

1998 ◽

Vol 158 (2) ◽

pp. 267-275 ◽

Cited By ~ 26

Author(s):

YO Lukyanenko ◽

AM Carpenter ◽

DE Brigham ◽

DM Stocco ◽

JC Hutson

Keyword(s):

Leydig Cells ◽

Regulatory Protein ◽

High Capacity ◽

Steroidogenic Acute Regulatory Protein ◽

Maximal Response ◽

Maximal Dose ◽

Testosterone Production ◽

Highly Active ◽

Chain Cleavage ◽

Steroidogenic Acute Regulatory

The purpose of this investigation was to study the mechanism of action of a macrophage-derived factor that stimulates steroid production by Leydig cells. This factor increased testosterone production within 30 min, and reached a half-maximal response by 6-8 h. At a maximal dose, it stimulated testosterone production 20-fold at 24 h. Its efficacy was consistently higher than that achieved with a maximal dose of human chorionic gonadotropin (hCG). However, Leydig cells treated with a maximal dose of both the macrophage-derived factor and hCG secreted the same amount of testosterone as when given a maximal dose of only the macrophage-derived factor. The macrophage-derived factor did not require new protein synthesis to stimulate testosterone production, nor did it alter the amount of steroidogenic acute regulatory protein (StAR). While the macrophage-derived factor required an active cholesterol side-chain cleavage complex system, it did not alter the capacity of this enzyme complex. Finally, the macrophage-derived factor was unable to stimulate the production of progesterone by isolated mitochondria. In summary, the macrophage-derived factor is a highly active, acute regulator of steroidogenesis that acts through a high capacity StAR-independent pathway.

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Cyclooxygenase-2 Regulation of the Age-Related Decline in Testosterone Biosynthesis

Endocrinology ◽

10.1210/en.2005-0298 ◽

2005 ◽

Vol 146 (10) ◽

pp. 4202-4208 ◽

Cited By ~ 57

Author(s):

XingJia Wang ◽

Chwan-Li Shen ◽

Matthew T. Dyson ◽

Sarah Eimerl ◽

Joseph Orly ◽

...

Keyword(s):

Leydig Cells ◽

Regulatory Protein ◽

Cyclooxygenase 2 ◽

Aged Rats ◽

Inhibitory Effects ◽

Age Related ◽

Testosterone Biosynthesis ◽

Cox2 Inhibitor ◽

Steroidogenic Acute Regulatory ◽

Star Gene

The age-related decline in testosterone biosynthesis in testicular Leydig cells has been well documented, but the mechanisms involved in the decline are not clear. Recent studies have described a cyclooxygenase-2 (COX2)-dependent tonic inhibition of Leydig cell steroidogenesis and expression of the steroidogenic acute regulatory protein (StAR). The present study was conducted to determine whether COX2 protein increases with age in rat Leydig cells and whether COX2 plays a role in the age-related decline in testosterone biosynthesis. Our results indicate that from 3 months of age to 30 months, COX2 protein in aged rat Leydig cells increased by 346% over that of young Leydig cells, StAR protein decreased to 33%, and blood testosterone concentration and testosterone biosynthesis in Leydig cells decreased to 41 and 33%, respectively. Further experiments demonstrated that overexpressing COX2 in MA-10 mouse Leydig cells inhibited StAR gene expression and steroidogenesis and that the inhibitory effects of COX2 could be reversed by blocking COX2 activity. Notably, incubation of aged Leydig cells with the COX2 inhibitor NS398 enhanced their testosterone biosynthesis. Blood testosterone concentrations in aged rats fed the COX2 inhibitor DFU, at doses of 5, 10, 15, and 20 mg/kg body weight per day were increased by 15, 23, 56, and 120%, respectively, over the levels in the rats receiving no DFU. The present study suggests a novel mechanism in male aging involving COX2 and a potential application of the mechanism to delay the age-related decline in testosterone biosynthesis.

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BMP15 Suppresses Progesterone Production by Down-Regulating StAR via ALK3 in Human Granulosa Cells

Molecular Endocrinology ◽

10.1210/me.2013-1233 ◽

2013 ◽

Vol 27 (12) ◽

pp. 2093-2104 ◽

Cited By ~ 47

Author(s):

Hsun-Ming Chang ◽

Jung-Chien Cheng ◽

Christian Klausen ◽

Peter C. K. Leung

Keyword(s):

Granulosa Cells ◽

Critical Role ◽

Regulatory Protein ◽

Granulosa Cell Tumor ◽

Type I ◽

Mrna Levels ◽

Protein Levels ◽

Human Granulosa Cells ◽

Progesterone Production ◽

Steroidogenic Acute Regulatory

In addition to somatic cell-derived growth factors, oocyte-derived growth differentiation factor (GDF)9 and bone morphogenetic protein (BMP)15 play essential roles in female fertility. However, few studies have investigated their effects on human ovarian steroidogenesis, and fewer still have examined their differential effects or underlying molecular determinants. In the present study, we used immortalized human granulosa cells (SVOG) and human granulosa cell tumor cells (KGN) to compare the effects of GDF9 and BMP15 on steroidogenic enzyme expression and investigate potential mechanisms of action. In SVOG cells, neither GDF9 nor BMP15 affects the mRNA levels of P450 side-chain cleavage enzyme or 3β-hydroxysteroid dehydrogenase. However, treatment with BMP15, but not GDF9, significantly decreases steroidogenic acute regulatory protein (StAR) mRNA and protein levels as well as progesterone production. These suppressive effects, along with the induction of Sma and Mad-related protein (SMAD)1/5/8 phosphorylation, are attenuated by cotreatment with 2 different BMP type I receptor inhibitors (dorsomorphin and DMH-1). Furthermore, depletion of activin receptor-like kinase (ALK)3 using small interfering RNA reverses the effects of BMP15 on SMAD1/5/8 phosphorylation and StAR expression. Similarly, knockdown of ALK3 abolishes BMP15-induced SMAD1/5/8 phosphorylation in KGN cells. These results provide evidence that oocyte-derived BMP15 down-regulates StAR expression and decreases progesterone production in human granulosa cells, likely via ALK3-mediated SMAD1/5/8 signaling. Our findings suggest that oocyte may play a critical role in the regulation of progesterone to prevent premature luteinization during the late stage of follicle development.

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Molecular Mechanisms of Insulin-like Growth Factor-I Mediated Regulation of the Steroidogenic Acute Regulatory Protein in Mouse Leydig Cells

Molecular Endocrinology ◽

10.1210/me.2004-0526 ◽

2006 ◽

Vol 20 (2) ◽

pp. 362-378 ◽

Cited By ~ 92

Author(s):

Pulak R. Manna ◽

Syam P. Chandrala ◽

Steven R. King ◽

Youngah Jo ◽

Raymond Counis ◽

...

Keyword(s):

Growth Factor ◽

Leydig Cells ◽

Molecular Mechanisms ◽

Regulatory Protein ◽

Steroidogenic Acute Regulatory Protein ◽

Insulin Like Growth Factor ◽

Factor I ◽

Growth Factor I ◽

Steroidogenic Acute Regulatory

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280. INSL3 is a measure of human Leydig cell functionality both during fetal and adult life

Reproduction Fertility and Development ◽

10.1071/srb08abs280 ◽

2008 ◽

Vol 20 (9) ◽

pp. 80

Author(s):

R. Anand-Ivell ◽

J. Manson ◽

G. Wittert ◽

J. Wohlgemuth ◽

B. Hafen ◽

...

Keyword(s):

Leydig Cell ◽

Leydig Cells ◽

Cell Function ◽

Adult Life ◽

Physiological Role ◽

South Australia ◽

Testicular Descent ◽

Adult Type ◽

Hpg Axis ◽

Age Related

Insulin like factor 3 (INSL3) and testosterone are the two major secretory products of the testis, both produced by the interstitial Leydig cells. The Leydig cells of the testis have two distinct generations, one developing before birth (fetal Leydig cells, FLC) and an adult type (adult Leydig cells, ALC) that become differentiated and functional at puberty. Although these two types of Leydig cells represent distinct populations, rodent studies show that both types produce testosterone and INSL3. Both are presumed to have evolved from a common stem cell pool. We measured INSL3 levels in human amniotic fluids collected at various times of gestation and show for the first time that the human male fetus indeed generates INSL3 at a time appropriate for the first transabdominal phase of testicular descent, which appears to be the primary physiological role for the fetal hormone. INSL3 appears to be independent of androgen production. The adult type Leydig cells (in adult men) secrete INSL3 that can be measured in the peripheral circulation at levels ranging from 0.5 to 2.5 ng/mL. We studied a large randomly recruited cohort of 1183 men from South Australia, comparing serum INSL3 concentrations with age, and a variety of endocrine, cognitive and morphological parameters. INSL3 concentration was observed to decline significantly with age. This however, had no correlation with testosterone or components of the HPG axis. INSL3 is an independent measure of Leydig cell function (quality and number), which appears to be independent of acute control via the HPG axis. Its decline with age reflects a decline in the properties of the Leydig cell population only, and emphasises a gonadal component in the age-related decrease in androgen production. Research supported by ARC Discovery grant DP0773315.

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INSL3: A Marker of Leydig Cell Function and Testis-Bone-Skeletal Muscle Network

Protein and Peptide Letters ◽

10.2174/0929866527666200925105739 ◽

2020 ◽

Vol 27 (12) ◽

pp. 1246-1252

Author(s):

Paolo Facondo ◽

Andrea Delbarba ◽

Filippo Maffezzoni ◽

Carlo Cappelli ◽

Alberto Ferlin

Keyword(s):

Skeletal Muscle ◽

Leydig Cell ◽

Leydig Cells ◽

Cell Function ◽

Cell Damage ◽

Endocrine Function ◽

Testicular Descent ◽

Male Hypogonadism ◽

Adult Men ◽

Leydig Cell Function

This article reviews the role of INSL3 as biomarker of Leydig cell function and its systemic action in testis-bone-skeletal muscle crosstalk in adult men. Insulin-like factor 3 (INSL3) is a peptide hormone secreted constitutively in a differentiation-dependent mode by testicular Leydig cells. Besides the role for the testicular descent, this hormone has endocrine anabolic functions on the bone-skeletal muscle unit. INSL3 levels are low in many conditions of undifferentiated or altered Leydig cell status, however the potential clinical utility of INSL3 measurement is not yet well defined. INSL3 levels are modulated by the long-term cytotropic effect of the hypothalamicpituitary- gonadal axis, unlike testosterone that is acutely sensitive to the stimulus by luteinizing hormone (LH). INSL3 directly depends on the number and differentiation state of Leydig cells and therefore it represents the ideal marker of Leydig cell function. This hormone is more sensitive than testosterone to Leydig cell impairment, and the reduction of INSL3 in adult men can precociously detect an endocrine testicular dysfunction. Low INSL3 levels could cause or contribute to some symptoms and signs of male hypogonadism, above all sarcopenia and osteoporosis. The measurement provided suggested that the measurement of INSL3 levels should be considered in the clinical management of male hypogonadism and in the evaluation of testicular endocrine function. The monitoring of INSL3 levels could allow an early detection of Leydig cell damage, even when testosterone levels are still in the normal range.

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Effects of daidzein on testosterone secretion in cultured immature mouse testis

Asian Biomedicine ◽

10.5372/1905-7415.0802.286 ◽

2014 ◽

Vol 8 (2) ◽

pp. 247-253

Author(s):

Liuping Zhang ◽

Genbao Shao ◽

Yaoqian Pan

Keyword(s):

Direct Action ◽

Regulatory Protein ◽

Mouse Testis ◽

In Vivo Studies ◽

Mrna Levels ◽

Testosterone Secretion ◽

Immature Male ◽

Testosterone Production ◽

Steroidogenic Acute Regulatory

AbstractBackground: Daidzein is a major isoflavone in soybeans. Several in vivo studies have showed that daidzein can affect immature male testosterone production. However, whether daidzein has direct action on immature male testis is unknown.Objective: We investigated the effects of daidzein on testosterone secretion in 3-day-old and 21-day-old mouse Leydig cells with organotypic culture model.Materials and Methods: The testes were exposed to different concentrations (10-7to 10-4M) of daidzein for 72 h with medium changed every 24 h. From 72 to 75 h of culture, 100 ng/ml human chorionic gonadotropin (hCG) was added. The testosterone production was determined, and the related mechanisms of daidzein action were also evaluated by measuring the mRNA levels of steroidogenic acute regulatory protein (StAR), cholesterol side-chain cleavage enzyme (P450scc), and 3β-hydroxysteroid dehydrogenase (3β-HSD-1) involved in testosterone biosynthesis.Results: The results revealed that in the presence of 100 ng/ml hCG, 10-7to 10-5M daidzein had no significant effect on testosterone secretion in cultured 3-day-old mouse testis. But 10-4M daidzein significantly increased testosterone concentration (p < 0.05). Daidzein in range of studied doses had no obvious influence on testosterone production in cultured 21-day-old mouse testis. RT-PCR results showed that 10-4M daidzein had obvious influence on the mRNA levels of StAR, P450scc and 3β-HSD-1 in cultured 3-day-old mouse testis (p < 0.05).Conclusion: These results suggest that daidzein mainly influences neonatal mouse testis function, and the influence is partially related to the upregulation of StAR, P450scc, and 3β-HSD-1 mRNA levels.

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