Cynomorium songaricum induces spermatogenesis with glial cell-derived neurotrophic factor (GDNF) enhancement in rat testes

doi:10.1016/j.jep.2010.02.020

Journal of Ethnopharmacology

Volume 128, Issue 3, 21 April 2010, Pages 693-696

https://doi.org/10.1016/j.jep.2010.02.020 Get rights and content

Abstract

Aim of the study

Cynomorium songaricum Ruprecht has been used in traditional Korean medicine to treat male infertility, including sexual dysfunction, by improving kidney function. Glial cell-derived neurotrophic factor (GDNF) produced by Sertoli cells induces the proliferation of undifferentiated spermatogonia. We investigated the effects of Cynomorium songaricum on sperm parameters and GDNF expression in rat testes.

Materials and methods

Sperm analysis, RT-PCR, and Western blotting assays were performed after administration of CS to 8-week-old male Wistar rats for 56 consecutive days (1.0 g/kg/day, p.o.), the period of sperm formation in the rat.

Results and conclusions

The CS-treated animals showed significant increases in epididymal sperm count and absolute testes weights compared to the control group. CS also increased the expression of GDNF at both the mRNA and protein levels. These results suggest that CS may improve male fertility by enhancing spermatogenesis and GDNF expression.

Graphical abstract

Cynomorium songaricum (CS) treatment induces sperm count and GDNF expression in rat testes. After 56 days of treatment with CS, the epididymal sperm counts were significantly higher than those in the control group. The testes of rats treated with CS exhibited a significant increase in GDNF mRNA levels compared to the control group, and Western blotting revealed a significant increase in the GDNF-immunoreactive band in CS-treated rats compared to the control group.

Introduction

The incidence rates of male infertility are increasing worldwide, the leading cause of which is low sperm count. It has been reported that sperm counts in Western countries have decreased by 1% annually over the last 50 years (Carlsen et al., 1992). Although there have been many clinical trials aim at improving the number and quality of sperm, they have met with only limited success. Recent studies on rats have indicated that male fertility, including sexual dysfunction, can be improved by medicinal herbs (Yeh et al., 2008).

Glial cell-derived neurotrophic factor (GDNF) is an important growth factor that facilitates communication between Sertoli cells and spermatogonia. GDNF induces the proliferation of undifferentiated spermatogonia under in vivo and in vitro conditions (Meng et al., 2000). In the testis, the GDNF dosage controls the cell fate of undifferentiated spermatogonia (Airaksinen and Saarma, 2002). Gene-targeted mice with one GDNF-null allele show partial depletion of spermatogenic stem cells, whereas those overexpressing GDNF show clusters of undifferentiated spermatogonia. GDNF stimulates DNA synthesis in spermatogonia (Viglietto et al., 2000).

Cynomorium songaricum (CS) is used in traditional Korean medicine as an herbal remedy for kidney and intestinal ailments, as well as impotence (Nickrent et al., 2005) which is a fleshy, monoecious or dioecious, holoparasitic herb that often produces swollen tuberous haustorial root connections to its host plants (Kuijt, 1969). Its chemical constituents include steroids, triterpenes, fructosides, flavonoids, and condensed tannins (Jiang et al., 2001). Several studies have examined the effects of CS extracts on mammalian reproductive cells. In addition, a water extract of CS induced a significant increase in sperm count, improved the percentage of live sperm and their motility, and decreased the number of abnormal sperm (Abdel-Rahman et al., 1999), and there was a marked increase in testicular weight in animals treated with the extracts, which also had a direct spermatogenic influence on the seminiferous tubules of immature rats, presumably by exerting a testosterone-like effect (Abdel-Magied et al., 2001).

However, the relationship between CS and its effects on male reproductive tract malfunction associated with GDNF gene expression and spermatogenesis in vivo have not been elucidated. To investigate the effects of CS on male reproductive system in rats, we performed sperm analysis, including testicular weight, sperm count, and sperm motility. In addition, we assessed GDNF expression at the mRNA and protein levels.

Section snippets

Preparation of CS extract

Cynomorii Herba is the dried fleshy stem of Cynomorium songaricum Ruprecht (Fam. Cynomoiaceae). CS, produced in China, was purchased from Wonkwang Herbal Dug Co. Ltd. (Seoul, Korea). A 300 g sample of dried CS was boiled in 6 L water at 100 °C for 2 h and the suspension was filtered. The filtrate was concentrated under reduced pressure and then lyophilised. The final yield was 26% (dry weight 76.60 g), and the sample was stored at 4 °C. A voucher specimen (SK003) was deposited in our laboratory.

Effects of CS on body and testicular weights

None of the animals died during the study and no noticeable adverse effects were observed in any of the animals after administration of the extract or vehicle. Rats treated with CS showed a slight change in body weight compared to the control group (429.2 ± 34.5 and 420.5 ± 26.9 g, respectively). The absolute weights of the testes of the CS-treated group were slightly but significantly greater than those of the control group (1.64 ± 0.05 and 1.52 ± 0.18 g, respectively; 7.89% increase, P < 0.05).

Effects of CS on sperm counts and sperm motility

After 56

Discussion and conclusion

Infertility is an increasing worldwide concern. The leading cause of male infertility is a low sperm count, and sperm counts in Western countries have decreased by 1% annually over the last 50 years (Carlsen et al., 1992). Although many clinical trials have been performed to improve the number and quality of sperm, they have had limited success. We believe that some medicinal herbs may improve male fertility with relatively few side effects.

CS has been used in traditional Korean medicine to

Acknowledgement

This research was supported by the Kyung Hee University Research Fund in 2008. (KHU-20081565)

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The aging phenomenon often exerts a significant reduction in the reproduction performance of aged animals. The objective of this project was to investigate the effects of dietary Folic acid (FA) supplementation on the reproductive performance of aged broiler breeder roosters. A total of 16 aged ROSS 308 broiler breeder roosters (50-week-old) were randomly divided into two groups. The treatments were basal diet (CON), a basal diet supplemented with 10 mg/kg Folic acid (FAS) for four weeks. At the end of the experiment, semen quality, histopathological studies, serum concentrations of testosterone and relative mRNA and protein expressions of testes were evaluated. The results showed that dietary FA supplementation dramatically improved semen quality of aged roosters, manifested by increasing semen volume, sperm concentration, sperm motility, and sperm membrane functional integrity. Furthermore, seminiferous tubule epithelial height (SEH) and testis scores were increased by dietary supplementation with FA. Dietary FA also remarkably augmented the transcription level of spermatogenesis-related gene (CREM, PCK2, DDX4, and GDNF). No significant differences were observed in serum concentrations of testosterone between FAS and CON groups. We noted significant upregulation Beclin-1 and ATG5 protein expressions, and the ratio of LC3-Ⅱ/Ⅰ, as well as significant downregulation of p-mTOR protein expressions in testicular tissue of aged roosters with FA supplementation. In addition, dietary FA supplementation significantly increased the protein expression of H3K9me2 and reduced the protein expression of H3K27me2. In summary, dietary FA supplementation improved the testicular autophagy through the mTOR-signaling pathway, and altered histone methylation in the testis. Dietary supplementation with FA can ameliorate semen quality and spermatogenesis of aged roosters.
Cynomorium songaricum Rupr demonstrates phytoestrogenic or phytoandrogenic like activities that attenuates benign prostatic hyperplasia via regulating steroid 5-α-reductase
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Cynomorium songaricum Rupr. (CS) belongs to the genus of parasitic perennial flowering plants, mostly used in Chinese traditional medicine for benign prostatic hyperplasia (BPH) treatment. BPH is a chronic disease in men that both androgen and estrogen play a crucial role in promoting its development via their receptors. Previously we have showed that compounds from CS have the phytoestrogenic and/or phytoandrogenic activities that may have the potential suppressive effects on BPH, while the mechanism remains unclear.
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Estradiol/testosterone (1:100) was subcutaneous injected to induce BPH in a castrated rat model, and CS was orally administrated for 45 days. Then the weights of the body and prostate were recorded, the pathogenesis changes of prostate were analyzed by Hematoxylin and eosin (H&E) and immunohistochemical (IHC). The levels of 17β-estradiol (E₂), testosterone, and dihydrotestosterone (DHT) from rats’ serum were measured by enzyme-linked immunosorbent assay (ELISA). In vitro, human benign prostatic epithelial cell BPH-1 was cultured and treated with or without different CS compounds and DHT or E₂. MTT and CCK-8 assays were performed to detect the regulatory effects on cell proliferation. The expressions of PCNA, AR, ERα, ERβ, and steroid 5-α-reductases (SRD5A1 and SRD5A2) were further analyzed by western blotting upon treatment.
Treatment with CS significantly inhibited rat prostate enlargement, improved the pathological feature and reduced the thickness of smooth muscle layer. The up-regulated AR and ERα expressions and down-regulated ERβ in BPH rat prostate were significantly blocked after CS administration. Moreover, the enhanced values of E₂/testosterone and the level of DHT in serum were also strongly inhibited in CS group compared with those in BPH groups. In cellular level, LUT, GA, PA, or Pra significantly inhibited DHT- or E₂- induced BPH-1 cell proliferation and PCNA expressions. Consistently with the data in vivo, compounds from CS interfered the DHT or E₂-regulated AR, ERα and ERβ expressions in BPH-1 cells as well. Importantly, the dramatic increased SRD5A1 and SRD5A2 expressions were observed in BPH rat prostates and DHT or E₂-stimulated BPH-1 cells. However, treatment with CS in rat or with compounds isolated from CS in BPH-1 cells significantly blocked the induction of SRD5A1 and SRD5A2.
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A study has shown that the oligosaccharides from Morinda officinalis were a protective agent for the oxidative damaged sperm DNA.21 Epimedium koreanum Nakai had shown antioxidant activity,22 and Cynomorium songaricum Rupr showed significant increases in epididymal sperm count and absolute testes in rat testes.23 This effect might be a result of their antioxidative properties, which help restore the imbalance of excessive levels of ROS.
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Extracts of Epimedium koreanum Nakai had shown antioxidant activities, which included free radical- and ferric-reducing power and lipid peroxidation inhibition [24], and improved erectile function in aged rats [25]. Cynomorium songaricum Rupr showed significant increases in epididymal sperm counts and absolute testes in rat testes [26]. All of these effects might be a result of their antioxidative properties, which help restore the imbalance in excessive levels of ROS.
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Journal of Ethnopharmacology

Abstract

Aim of the study

Materials and methods

Results and conclusions

Graphical abstract

Introduction

Section snippets

Preparation of CS extract

Effects of CS on body and testicular weights

Effects of CS on sperm counts and sperm motility

Discussion and conclusion

Acknowledgement

References (15)

The effect of aqueous extracts of Cynomorium coccineum and Withania somnifera on testicular development in immature Wistar rats

Journal of Ethnophamacology

Disruption of murine Tenr results in teratospermia and male infertility

Developmental Biology

Ginkgo biloba extract enhances male copulatory behavior and reduces serum prolactin levels in rats

Hormones and Behavior

In vitro estrogenic activities of Chinese medicinal plants traditionally used for the management of menopausal symptoms

Journal of Ethnophamacology

The effect of the aqueous extract of Cynomorium cocconieum on the epydidymal sperm pattern of the rat

Phytotherapy Research

The GDNF family: signalling, biological functions and therapeutic value

Nature Reviews Neuroscience

Chinese Herbal Medicine

Cited by (32)

Dietary folic acid supplementation improves semen quality and spermatogenesis through altering autophagy and histone methylation in the testis of aged broiler breeder roosters

Cynomorium songaricum Rupr demonstrates phytoestrogenic or phytoandrogenic like activities that attenuates benign prostatic hyperplasia via regulating steroid 5-α-reductase

Successful treatment of oligoasthenozoospermia using traditional Korean medicine resulting in spontaneous pregnancy: Two case reports

The effects of traditional Korean medicine in infertile male patients with poor semen quality: A retrospective study

Yang-tonifying traditional Chinese medicinal plants and their potential phytoandrogenic activity

Chemical constituents and pharmacologic actions of cynomorium plants

Ethnopharmacological communicationCynomorium songaricum induces spermatogenesis with glial cell-derived neurotrophic factor (GDNF) enhancement in rat testes

Abstract

Aim of the study

Materials and methods

Results and conclusions

Graphical abstract

Introduction

Section snippets

Preparation of CS extract

Effects of CS on body and testicular weights

Effects of CS on sperm counts and sperm motility

Discussion and conclusion

Acknowledgement

Journal of Ethnophamacology

Developmental Biology

Hormones and Behavior

Journal of Ethnophamacology

The effect of the aqueous extract of Cynomorium cocconieum on the epydidymal sperm pattern of the rat

Phytotherapy Research

The GDNF family: signalling, biological functions and therapeutic value

Nature Reviews Neuroscience

Chinese Herbal Medicine

Ethnopharmacological communication
Cynomorium songaricum induces spermatogenesis with glial cell-derived neurotrophic factor (GDNF) enhancement in rat testes