Therapeutic Effects And Mechanisms Of Traditional Chinese Medicine Monomers On Late-Onset Hypogonadism in Aged Male Rats

2. Results

2.1 Effects of the Three TCM Monomers on Body Weight and Reproductive Organ Weights in LOH Model Rats

Treatment with ginsenoside Rg1 significantly increased the body weight, as well as the weights of the seminal vesicles, epididymis, and testes in LOH model rats (P < 0.01), showing the most prominent effect among all groups.

The icariin group also demonstrated significant increases in the weights of the seminal vesicles, epididymis, and testes (P < 0.01), while the verbascoside group showed a moderate but statistically significant increase in seminal vesicle weight (P < 0.05).

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2.2 Effects of the Three TCM Monomers on Testicular Histomorphology in LOH Rats

Hematoxylin-eosin (H&E) staining revealed that all three TCM monomers effectively ameliorated degenerative changes in the seminiferous tubules of LOH model rats. The testicular tissue morphology in the treated groups closely resembled that of healthy controls, with increased numbers of spermatogenic cells and mature sperm.

The histological improvements were comparable to those observed in the testosterone propionate group.

Figure 1 Effects of three Chinese medicine monomers on body weight, seminal vesicle, epididymis and testis weight of LOH model rats

 

Note: LOH is delayed hypogonadism; A is the control group, B is the model group, C is the testosterone propionate group, D is the icariin group, E is the verbascoside group, and F is the ginsenoside group; HE staining ×100 times

 

2.3 Effects of the Three TCM Monomers on Serum Reproductive Hormone Levels

All three TCM monomers significantly reduced serum luteinizing hormone (LH) and follicle-stimulating hormone (FSH) levels while elevating serum testosterone in LOH model rats.

Among them, the icariin group showed the most significant hormonal regulation effects (P < 0.01), and the results were closest to those of the testosterone propionate group.

 

2.4 Effects of the Three TCM Monomers on Androgen Receptor (AR) Expression

All three TCM monomers significantly upregulated both protein and mRNA expression levels of the androgen receptor (AR) in the testicular tissues of LOH rats.

Notably, the icariin group exhibited the most pronounced effect (P < 0.01), with AR expression levels exceeding those observed in the testosterone propionate group.

 

Note: LOH is delayed hypogonadism; A is the control group, B is the model group, C is the testosterone propionate group, D is the icariin group, E is the verbascoside group, and F is the ginsenoside group; HE staining ×100 times

Note: LOH is delayed hypogonadism, AR is androgen receptor, GAPDH
is glyceraldehyde-3-phosphate dehydrogenase, and mRNA is messenger RNA. A is the effect of three Chinese medicine monomers
on the AR protein level in the seminal vesicle, epididymis, and testis of model rats, B is the statistical result of the relative gray value of the Western blot band in each group in Figure A, and C is the effect of different Chinese medicine monomers on the AR messenger RNA level in the seminal vesicle, epididymis, and testis of model rats. * Compared with the control group, P < 0.05; ** Compared with the control group, P < 0.01; # Compared with the model group, P < 0.05; # # Compared with the model group, P < 0.01.

3. Discussion

With the acceleration of population aging in China and globally, late-onset hypogonadism (LOH) has become a growing concern, significantly affecting the physical and mental health and quality of life of middle-aged and elderly males [5].

Studies show that testosterone production and testicular function in aging men decline by 2%–11% annually, with testicular volume decreasing by approximately 13% from age 25 to 80–90, negatively impacting both sperm count and quality [6]. The hypothalamic-pituitary-gonadal (HPG) axis regulates testosterone secretion. When serum testosterone drops, the body compensates via negative feedback, increasing luteinizing hormone (LH) and follicle-stimulating hormone (FSH) levels [7].

 

Mechanisms of the Three Chinese Herbal Monomers

Icariin, the main bioactive component of Epimedium, has been reported to have androgen-like effects, mimicking testosterone in various animal models [8–10].

Verbascoside, a phenylethanoid glycoside extracted from Cistanche deserticola, has been shown to improve testicular atrophy, increase sperm production, and counteract androgen deficiency in mice [11–12].

Ginsenoside Rg1, a major active compound in Panax ginseng, has demonstrated the ability to protect reproductive function, increase sperm motility and density, and alleviate testicular damage in aging models [13–14].

In this study, all three monomers reduced serum LH and FSH levels and elevated serum testosterone, with the icariin group showing the most significant endocrine modulation, closely resembling the effect of testosterone propionate.

 

Histological and Molecular Improvements in LOH Rats

All three TCM compounds effectively alleviated degenerative changes in the seminiferous tubules of LOH rats-restoring testicular structure, increasing the number of spermatogenic cells, and improving spermatogenesis. These results were comparable to exogenous testosterone therapy, but without its known risks.

Androgens play a critical role in male development and reproductive maintenance, primarily through their interaction with the androgen receptor (AR) [15]. AR, a nuclear receptor protein, mediates the transcriptional regulation of androgen-responsive genes [16], and is predominantly expressed in testicular Leydig and Sertoli cells, where it governs spermatogenesis and fertility [17].

Previous studies have shown that exogenous androgens can significantly upregulate AR mRNA expression in testicular interstitial cells [18]. In this study, all three monomers upregulated AR protein and mRNA levels, with icariin demonstrating the most prominent effect, even outperforming testosterone propionate. This may be attributed to its dual action of improving testosterone levels and enhancing AR signaling pathways.

 

Comparative Analysis of the Three Monomers

Ginsenoside Rg1 most significantly increased body weight and reproductive organ weights, likely due to its anabolic and organotropic effects on gonadal tissues.

Icariin showed the strongest regulatory effects on serum reproductive hormones and AR expression, suggesting its potential as a natural testosterone regulator and AR modulator.

Verbascoside offered moderate benefits, particularly in seminal vesicle weight and testicular structure preservation, supporting its traditional usage in yang-tonifying and fertility-enhancing therapies.

All three compounds improved testicular histomorphology and sperm production, providing promising evidence for their potential in safe, plant-based LOH management.

 

Clinical and Translational Implications

This study provides preclinical evidence supporting the use of TCM-derived monomers as potential alternatives or adjuncts to testosterone replacement therapy (TRT). Among the three, icariin demonstrated the most consistent and significant improvements in endocrine regulation and AR expression, laying the foundation for future clinical trials.

In addition, since cyclophosphamide, a widely used chemotherapeutic and immunosuppressive agent, is known for its reproductive toxicity, the observed protective effects of these herbs may also have implications in fertility preservation during chemotherapy.

 

Study Limitations

It is important to note that LOH is a chronic, progressive condition, while the cyclophosphamide-induced model used in this study simulates LOH over a relatively short period via cytotoxic injury. Currently, there is no universally accepted LOH animal model, and thus the results should be interpreted with caution.

 

Conclusion of the Discussion

In summary:

Ginsenoside Rg1 had the greatest impact on body weight and reproductive organ mass in LOH rats.

Icariin most effectively modulated reproductive hormones and upregulated AR expression.

All three TCM monomers improved testicular degeneration and spermatogenic function.

These findings provide valuable theoretical and experimental support for the development of natural, safe, and effective treatments for late-onset hypogonadism in aging males.