Phenylethanol glycosides from Cistanche tubulosa improved reproductive dysfunction by regulating testicular steroids through CYP450-3β-HSD pathway
Introduction
Male sexual dysfunction (MSD) is one of the most common health issues, with an estimated prevalence rate of 4.2% worldwide (Hotaling et al., 2015). A variety of factors, including age, smoking, obesity, diabetes and cardiovascular disease et al., can eventually lead to MSD. The pathological changes will severely affect the physical and mental health of individuals, including sexual psychological response, physiological structure, neuroendocrine and vascular function. Evidence demonstrated that reduced level of serum testosterone is the primary cause of MSD and testosterone deficiency dominated the whole process of MSD (Zhu et al., 2016). Improving synthesis of testosterone can alleviate weakened reproductive capacity and improve reproductive function. Therefore, testosterone synthesis is believed to be the most attractive therapeutic target in the treatment of MSD.
The biosynthesis of testosterone takes place in the Leydig cells of testis. Cholesterol in circulation is the major precursor for testosterone synthesis. Steroidogenic acute regulatory protein (StAR) mediates the transport of cholesterol from cytoplasm into mitochondria. Cholesterol is then converted to pregnenolone through cytochrome P450 cholesterol side chain cleavage enzyme (P450scc/CYP11A1). Afterwards, pregnenolone is turned to dehydroepiandrosterone which is eventually converted to testosterone in a series of enzymatic reactions mediated by critical enzymes including 3β-hydroxysteroid dehydrogenase (3β-HSD) (Fig. 1) (Kumar et al., 2008).
Cistanches Herba (CH) is a genus of medicinal plant that has been used in China and other eastern Asian countries since the fifteenth century for the treatment of impotence, seminal emission, infertility, and chronic constipation (Wang et al., 2012). Cistanche tubulosa (Schenk) R. Wight (C. tubulosa) and Cistanche deserticola Y. C. Ma (C. deserticola) have similar chemical compounds and pharmacological activities, and they have been documented in the Chinese Pharmacopoeia (Editorial committee of Chinese pharmacopoeia, 2015). The main medicinal ingredients in CH are phenylethanol glycosides, and the content in C. tubulosa is higher than that of C. deserticola (Song et al., 2016; Liu et al., 2018). C. tubulosa is considered as an alternative to C. deserticola, which is on the verge of extinction due to decades of over-exploitation.
Evidence showed that C. tubulosa could effectively protect against the reproductive system damage of male mice caused by tripterygium glycosides. Moreover, alcohol extract of C. tubulosa could increase the expressions of key enzymes, including CYP11A1 and cytochrome P450 family 17 subfamily A member 1 (CYP17A1), in the synthesis of testosterone in normal male rats (Wang et al., 2016). In addition, the preliminary research of our group indicated that phenylethanol glycosides from C. tubulosa (CPhGs) promoted the release of sex hormones and improved the testicular pathological status of reproductive injury model animals by regulating function of hypothalamus-pituitary-gonadal (HPG) axis (Wang et al., 2018).
Therefore, the aimed of this study was to investigate the effects of CPhGs on the reproductive dysfunction and testosterone levels in the model of hydrocortisone (HCT)-induced reproductive dysfunction of male mice. The regulative effects of CPhGs on key enzymes involved in synthesis of testosterone were explored by in vivo studies.
Section snippets
Preparation of CPhGs
The C. tubulosa was provided and identified by Prof. Peng-fei Tu (Peking University, Beijing, China). The C. tubulosa was collected from the cultivation base of Peking University in Yutian County of Xinjiang Autonomous Region. CPhGs extract was prepared by Tianjin Bei Da Cong Rong Biotechnology Co. Ltd. The dried slices of C. tubulosa (50 kg) were pulverized and filtered through a 20-mesh sieve. Then was extracted twice by water and the extracts were combined. The extracts were subjected to
Quantitative determination of the echinacoside and acteoside contents in CPhGs
The contents of echinacoside and acteoside in CPhGs were quantitated by using HPLC (Fig. 2). The retention times for echinacoside and acteoside were 11.5 min and 15.1 min, respectively at a maximum ultraviolet (UV) absorbance of 330 nm. The standard curves of the echinacoside and acteoside concentrations were y = 13869x - 991.14 (R2 = 1.0000) and y = 4056.9x - 3224.9 (R2 = 0.9998), respectively, where y was the peak area and x was the concentration of echinacoside or acteoside. The echinacoside
Discussion
MSD is one of the common diseases that affect male health and living standard. Decrease in synthesis of testosterone is the major cause of MSD (Helo et al., 2019). Therefore, promoting the production of testosterone and maintaining normal sexual physiology are the key treatment strategies. The synthesis of artificial testosterone provides a feasible approach for the treatment of MSD. However, the application is limited due to multiple side effects (Rao et al., 2015). Traditional Chinese
Conclusions
Our results showed that CPhGs could ameliorate HCT-induced damage of reproductive function and protect mice testis from injury. Further experiments demonstarted that CPhGs regulated synthesis of testosterone via CYP450-3β-HSD pathway. Our study provides experimental evidence that CPhGs extract is an alternative choice for clinical treatment of MSD.
Author contributions
Pengfei TU and Chun LI conceived and designed study; Qixin WANG and Jianteng DONG conducted majority of experiments; Wenji LU, Hao HE and Xiaoqian SUN contributed to hormone analysis and immunofluorescence staining; Qingqing SONG, Ke ZHANG contributed to HPLC analysis of CPhGs; Qixin WANG wrote the manuscript; Yong JIANG and Yong WANG edited the manuscript.
Declaration of competing interest
The authors declare that there is no conflict of interest.
Acknowledgements
This work was supported by the National Key Research and Development Program of China [grant number 2017YFC1702400].
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These two authors contributed equally to this article.