Use of astragaloside for preparing a medicine for treating oligoasthenospermia and the medicine

Abstract

The application relates to application of astragaloside B to preparation of a medicine for treating oligoasthenospermia and the medicine, and belongs to the field of traditional Chinese medicine. Through animal experiments, it is shown that the astragaloside B significantly improves the quantity and quality of rat sperms, and no obvious side effects are observed during the treatment. The astragaloside B has the effects of inhibiting oxidation stress, promoting secretion of testosterone, regulating secretion of follicle stimulating hormone and luteinizing hormone, promoting growth and maturation of sperms, effectively improving sperm function, and enhancing the repair capacity of the body to testicular injury. Moreover, no obvious side effects are observed during the treatment of the astragaloside B on oligoasthenospermia.

Description

Technical Field

[0001] This invention belongs to the field of traditional Chinese medicine, and specifically relates to the application of astragaloside A in the preparation of drugs for treating oligoasthenospermia and the drugs themselves. Background Technology

[0002] Oligospermia is a common cause of male infertility, characterized by a lower than normal sperm count, poor sperm motility, or abnormal sperm morphology in the semen. It is a significant contributing factor to male infertility. The causes of oligospermia are varied, potentially including genetic factors, environmental pollution, lifestyle factors, endocrine disorders, and oxidative stress. Current treatments for oligospermia primarily rely on hormone replacement therapy, medication, and assisted reproductive technologies (such as artificial insemination and in-vitro fertilization). Hormone therapy, also known as hormone replacement therapy (e.g., testosterone replacement therapy), is a common treatment for oligospermia. This method improves sperm production by regulating hormone levels and is typically used for oligospermia caused by endocrine disorders. However, hormone therapy often causes side effects such as sexual dysfunction, weight gain, and mood swings, and its effects are often temporary, with symptoms potentially recurring after treatment is discontinued. Antioxidant therapy: Since oxidative stress is one of the main causes of decreased sperm quality, many studies have attempted to use antioxidants (such as vitamin C, vitamin E, selenium, etc.) to improve sperm quality. Although antioxidants can reduce the production of free radicals, their effectiveness varies greatly from person to person, and long-term use may cause some side effects, such as gastrointestinal discomfort. Assisted reproductive technologies are effective, but the treatment process is cumbersome and expensive, and there is a certain failure rate; they cannot fundamentally improve the sperm production process. Therefore, finding a natural drug that can improve sperm count and quality while having fewer side effects has become an important research direction for solving the problem of oligospermia and asthenospermia.

[0003] In the field of traditional Chinese medicine, Astragalus membranaceus, as a traditional medicinal herb, possesses a wide range of pharmacological activities. It has the effects of strengthening the spleen and replenishing the middle jiao, raising yang and lifting prolapse, benefiting wei qi and consolidating the exterior, promoting diuresis, and promoting tissue regeneration. It is mainly used to treat spleen qi deficiency, lung qi deficiency, spontaneous sweating due to qi deficiency, qi and blood deficiency, and sores that are difficult to ulcerate or heal. One of the active ingredients in Astragalus membranaceus, astragaloside IV, is used as a standard for evaluating the quality of Astragalus membranaceus. It has effects such as enhancing the body's immune function, strengthening the heart and lowering blood pressure, lowering blood sugar, promoting diuresis, anti-aging, and anti-fatigue. Currently, there is no research specifically on the treatment of oligospermia and asthenospermia with astragaloside IV, especially no systematic clinical or experimental studies to deeply explore its mechanism and efficacy. Summary of the Invention

[0004] To address the above-mentioned problems, this invention provides the use of astragaloside A in the preparation of a drug for treating oligoasthenospermia and the drug thereof.

[0005] The first object of the present invention is to provide the use of astragaloside A in the preparation of a drug for treating oligoasthenospermia.

[0006] Furthermore, the oligoasthenospermia described herein is caused by a pathogen.

[0007] Furthermore, the pathogenic agent is one or more of di(2-ethylhexyl) phthalate, cyclophosphamide, and tripterygium glycoside.

[0008] The mechanism of action of astragaloside A in treating oligoasthenospermia: Astragaloside A treats oligoasthenospermia by improving the microenvironment of sperm.

[0009] Furthermore, the mechanism of astragaloside A in treating oligoasthenospermia is as follows: astragaloside A treats oligoasthenospermia by inhibiting oxidative stress response, enhancing the activity of antioxidant enzymes (such as superoxide dismutase and catalase), reducing the generation of free radicals, and improving the microenvironment of sperm.

[0010] Furthermore, the mechanism of astragaloside A in treating oligoasthenospermia is as follows: astragaloside A promotes the secretion of testosterone, improves the microenvironment of sperm, and promotes sperm growth and maturation, thereby treating oligoasthenospermia.

[0011] Furthermore, the mechanism of astragaloside A in treating oligoasthenospermia is as follows: astragaloside A improves the sperm microenvironment by regulating the secretion of follicle-stimulating hormone, thereby promoting sperm growth and treating oligoasthenospermia.

[0012] Furthermore, the mechanism of astragaloside A in treating oligoasthenospermia is as follows: astragaloside A regulates the secretion of luteinizing hormone, improves the sperm microenvironment, promotes sperm growth, and treats oligoasthenospermia.

[0013] A second objective of this invention is a pharmaceutical preparation for treating oligoasthenospermia, comprising astragaloside A.

[0014] Furthermore, the pharmaceutical preparations may be oral liquids, injections, granules, tablets, pills, powders, capsules, or drop pills.

[0015] In this invention, astragaloside A can also be made into a gel or patch and applied directly to the skin to treat sperm disorders by means of skin absorption.

[0016] In this invention, astragaloside A can also be formulated into a spray for nasal administration.

[0017] Astragaloside A can be combined with nanocarriers (such as nanoparticles, nanocapsules, and liposomes) to develop nanomedicine dosage forms. Nanotechnology can be used to improve drug bioavailability, duration of action, and targeting. Nanoformulations can ensure more effective drug delivery to the target site and improve therapeutic efficacy by adjusting parameters such as particle size and drug loading.

[0018] The beneficial effects of this invention are:

[0019] This invention relates to the application of astragaloside A in the preparation of drugs for treating oligoasthenospermia and asthenospermia. Animal experiments have shown that astragaloside A significantly improves the quantity and quality of sperm in rats, and no obvious side effects were observed during the treatment. Astragaloside A inhibits oxidative stress, promotes testosterone secretion, regulates the secretion of follicle-stimulating hormone and luteinizing hormone, and also promotes sperm growth and maturation, effectively improving sperm function and enhancing the body's ability to repair testicular damage. Moreover, astragaloside A has no obvious side effects in the treatment of oligoasthenospermia and asthenospermia.

[0020] Other features and advantages of the invention will be set forth in the description which follows, and will be apparent in part from the description, or may be learned by practicing the invention. The objects and other advantages of the invention may be realized and obtained by means of the structures pointed out in the description, claims and drawings. Attached Figure Description

[0021] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the drawings used in the description of the embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0022] Figure 1 A flowchart illustrating the experimental process of SD rats according to an embodiment of the present invention is shown.

[0023] Figure 2 The images show morphological diagrams of rat sperm testes and HE-stained images of testicular and epididymal tissues in each group according to the experimental method of the present invention. Figure 2 Image A shows the morphology of the testes of rats in each group; Figure 2 B shows HE staining of rat testicular tissue, and C shows HE staining of epididymal tissue from each group of rats.

[0024] Figure 3 The diagrams show the number and morphology of rat sperm in each group according to embodiments of the present invention;

[0025] Figure 4 The morphology of rat sperm from the model group according to an embodiment of the present invention is shown;

[0026] Figure 5 The graphs showing the levels of testosterone and follicle-stimulating hormone in the serum of rats in each group according to embodiments of the present invention are illustrated. Figure 5 Figure A shows the serum testosterone levels in each group of rats. Figure 5 Figure B shows the serum follicle-stimulating hormone (FSH) levels in each group of rats.

[0027] Figure 6 The graph shows the luteinizing hormone content in the serum of rats in each group according to an embodiment of the present invention;

[0028] Figure 7 The graphs showing the sperm count and motility of each group of rats according to embodiments of the present invention are shown. Figure 7 Figure A shows the sperm count of rats in each group. Figure 7 Figure B shows the sperm motility of rats in each group;

[0029] Figure 8 The graphs showing the MDA and GSH / GSSG content in the testicular tissue of rats in various groups according to embodiments of the present invention are illustrated. Figure 8 Figure A shows the MDA content in rat testicular tissue. Figure 8 Figure B shows the GSH / GSSG content in rat testicular tissue;

[0030] Figure 9 The graphs showing the contents of GSH-PX and GST in the testicular tissue of rats in various groups according to embodiments of the present invention are shown. Figure 9 Figure A shows the GSH-PX content in rat testicular tissue. Figure 9 Figure B shows the GST content in rat testicular tissue; Detailed Implementation

[0031] To make the objectives, technical solutions, and advantages of the embodiments of the present invention clearer, the technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0032] Animal experiments:

[0033] (1) Laboratory animals

[0034] Male SD rats, weighing 200–220 g, were purchased from Liaoning Changsheng Biotechnology Co., Ltd. (License No.: SCXK(Liaoning)2020-0001). All rats had free access to water and food. The bedding and water in their cages were changed daily to maintain a dry and clean environment.

[0035] (2) Experimental methods

[0036] Sixty SPF-grade male SD rats were divided into six groups. Forty rats were randomly selected and treated for eight consecutive weeks. Ten mice had free access to distilled water. The general condition of the rats was observed daily. Ten rats that did not develop the model served as the normal control group. The 60 rats that successfully developed the model were randomly divided into a normal group, a model group (treated with DEHP via gavage), and each group was treated with the prescribed dosage. The control group received 120 mg / kg body weight of saline intraperitoneally, while the model group and all treatment groups received 60 mg / kg body weight of DEHP via gavage once daily for 28 consecutive days. Starting from the 29th, using a human body weight of 60 kg as the standard, rats in the low, medium, and high dose traditional Chinese medicine treatment groups were intraperitoneally injected with astragaloside IV solution (AS-Ⅳ). The preparation of astragaloside IV solution involves dissolving astragaloside IV in ethanol (generally 100 mg of astragaloside IV is dissolved in 300 μL of ethanol), and then diluting with physiological saline to the corresponding concentration. The doses were 1 mg / kg, 10 mg / kg, and 20 mg / kg, respectively. The control group and model group were given the same volume of purified water by gavage. This treatment was continued for 14 days. During the treatment period, the rats were weighed once a week, and the gavage dose was adjusted according to the new weight. Blood samples were collected from the abdominal aorta between 07:00 and 09:00 on the second day after the treatment. After anesthetizing the rats with chloral hydrate, the testes, epididymis, and other tissues were quickly removed, operated on ice, and adhering blood vessels, ligaments, and fat tissues were removed for later use. The experimental progress of the SD rat experiment (DEHP gavage for four weeks, astragaloside IV intraperitoneal injection for two weeks) is shown in the figure below. Figure 1 As shown.

[0037] Evaluation indicators and effectiveness evaluation:

[0038] (1) Weight monitoring and calculation of testicular and epididymal indices

[0039] After dissecting the rats, the bilateral testes and epididymis were separated and weighed on an electronic balance. The visceral fat coefficient and testicular and epididymal indices of each group of rats were calculated. The effect of astragaloside A on spermatogenesis disorder in the DEHP-induced spermatogenesis model rats is shown in the figure below. Figure 2 As shown in Figure A, from left to right, the images depict the sperm and testes of rats in the control group, model group, low-dose AS-IV group, medium-dose AS-IV group, and high-dose AS-IV group, respectively. Figure 2 As can be seen from A, AS-Ⅳ promotes the growth of rat sperm testes damaged by DEHP, restoring them to their pre-damage morphology.

[0040] Morphological examination of epididymis and testis in oligospermia model rats

[0041] Photographs of the testis and epididymis tissues of rats in both the experimental and control groups were taken, paraffin sections were prepared, and hematoxylin and eosin (HE) staining was performed. The effect of HE staining on the histological changes of the rat testis and epididymis is shown in the figure below. Figure 1 As shown in B (magnification × 200) and C (magnification × 20). Figure 2 Image B is a section of rat testicular tissue. Figure 2 Image C is a section of the rat epididymis;

[0042] from Figure 2 As shown in sections B and C, the testicular tissue structure of the control group rats was intact, with a large number of sperm cells concentrated in the center of the seminiferous tubules. The seminiferous tubules were evenly and densely distributed without atrophy, and the tubular membranes were smooth, intact, and of normal thickness. The Sertoli cells and spermatogenic cells at all levels were arranged regularly and in rich layers, and there were a large number of mature sperm cells in the lumen. In contrast, the number of sperm cells in the epididymal ducts of the model group rats was sparse and diffusely distributed. The seminiferous tubules in the model group showed atrophy, significant shedding of interstitial cells, thickening of the tubular membrane, and a decrease in the number of cell layers. The number of mature sperm cells in the lumen was also low. Furthermore, in the epididymal head of the control group rats, sperm cells filled the entire lumen, while in the epididymal head of the model group rats, sperm cells were distributed on one side of the lumen. Figure 2 Studies B and C confirmed that DEHP can reduce the number of sperm cells in male rats. After intervention with different doses of astragaloside A, varying degrees of improvement were observed in the seminiferous tubule lumen, tubular membrane thickness, cell layer number, and spermatogenic cell number.

[0043] (2) Sperm quality assessment

[0044] The epididymis was cleaned, and the adipose tissue was removed. It was placed in a beaker containing 1 mL of preheated physiological saline at 37°C. Using ophthalmic scissors, 6-8 transverse and longitudinal incisions were made along the tail of the epididymis. The beaker was incubated in a 37°C water bath for 10 minutes to allow sperm to escape from the tail. The sperm suspension was filtered through a 300-mesh filter and maintained at 37°C. A drop of the sperm suspension was placed in the central counting chamber of a Markler sperm counting chamber. The sperm count (n) within a 10×10 square was observed under a 200x optical microscope. The sperm count per milliliter was n×10. 6 Each rat was observed three times, with one column of squares observed each time. The total number of sperm was calculated, and the sperm viability was calculated as follows: Viability / % = (a+b+c) / (a+b+c+d) × 100%. The number of sperm that moved forward was counted, and the sperm motility was calculated as follows: Motility / % = (a+b) / (a+b+c+d) × 100%.

[0045] For details regarding the effects of astragaloside A on sperm count and morphology in rats with spermatogenesis disorders, please refer to [link to relevant documentation]. Figure 3 and Figure 4 , Figure 3The images show the sperm count and morphology of rats in each group. Compared with the control group, the DEHP model group showed a significant decrease in sperm count and the presence of abnormal sperm. However, with the increase of astragaloside A concentration, the sperm count gradually returned to normal. Figure 4 Morphological images of sperm from rats in the DEHP model group. DEHP causes sperm malformation in rats. From left to right, the images show double-tailed sperm, headless sperm, microcephaly, and curled-tailed sperm.

[0046] The results of sperm count and sperm motility are as follows: Figure 7 As shown, where, Figure 7 Figure A shows the sperm count of rats in each group. Figure 7 Figure B shows the sperm motility graphs of rats in each group. Figure 7 Comparison of sperm count and motility among different groups of rats showed that, compared to the model group, the low-, medium-, and high-dose groups of rats injected with astragaloside IV all exhibited improved sperm count and motility. The high-dose group showed the most significant recovery in sperm count and motility, approaching the levels of the control group. This indicates that astragaloside IV helps treat oligospermia in rats affected by DEHP damage and promotes sperm production, increasing sperm count and motility. Data are expressed as mean ± standard deviation (n=3).

[0047] (3) Detection of serum sex hormone levels in rats

[0048] Centrifuged rat serum was collected, and the levels of hormones such as testosterone (T), follicle-stimulating hormone (FSH), and luteinizing hormone (LH) were measured using an ELISA kit according to the kit's instructions. The abbreviated results are as follows: Figure 4-5 As shown. Among them. Figure 5 Figure A shows the serum testosterone levels in rats. Figure 5 Figure B shows the follicle-stimulating hormone (FSH) levels in rat serum. Figure 6 This is a graph showing the luteinizing hormone (LH) content in rat serum.

[0049] from Figure 5 and Figure 6 It can be seen that, compared with the control group, the model group rats had lower serum T content and higher serum FSH and LH content due to DEHP treatment. The serum T content of rats injected with low-dose, medium-dose and high-dose astragaloside was significantly increased. Among them, the serum T content of rats in the high-dose group was close to that of the control group. This indicates that astragaloside can promote the secretion of testosterone in rats after DEHP treatment, improve the microenvironment of sperm in rats after DEHP treatment, promote sperm growth and maturation, and treat oligoasthenospermia.

[0050] In rats injected with low-, medium-, and high-dose astragaloside A, the levels of FSH and LH in the serum gradually decreased. Among them, the levels of FSH and LH in the serum of rats in the high-dose group were close to those in the control group. This also indicates that astragaloside A can regulate the secretion of follicle-stimulating hormone and luteinizing hormone in rats after DEHP treatment, improve the microenvironment of sperm in rats after DEHP treatment, promote sperm growth and maturation, and treat oligoasthenospermia.

[0051] (4) Detection of oxidative stress level in rat testicular tissue

[0052] ELISA was used to detect oxidative stress levels in testicular / epididymal tissues, including MDA (malondialdehyde), GSH (glutathione), T-GSH, GSH / GSSG (GSSG is also known as oxidized glutathione S-transferase), GSH-PX (glutathione peroxidase), and GST (glutathione S-transferase). After collecting rat testes / epididymis, tissue mass (g): extraction liquid volume (mL) was homogenized on ice; centrifuged at 4℃ (8000g for 10 min), and the supernatant was collected and placed on ice for later use. The detection results are as follows: Figure 8-9 As shown.

[0053] in, Figure 8 Figure A shows the MDA content in rat testicular tissue. Figure 8 Figure B shows the GSH / GSSG content in rat testicular tissue. Figure 9 Figure A shows the GSH-PX content in rat testicular tissue. Figure 9 Figure B shows the GST content in rat testicular tissue.

[0054] Figures 5-9 In this study, data are expressed as mean ± standard deviation (n = 3). Significance of differences between the Vcon group (control group) and other groups: *P < 0.05, **P < 0.01, ***P < 0.001. T: Testosterone; FSH: Follicle-stimulating hormone; LH: Luteinizing hormone.

[0055] in, Figure 8 Figure A shows the MDA concentration in the testicular tissue of rats in each group. Figure 8 Figure B shows the GSH / GSSG content in the testicular tissue of rats in each group. Figure 8The results showed that, compared with the control group, the levels of MDA and GSH / GSSG in the testicular tissue of rats in the model group were significantly increased. The levels of MDA and GSH / GSSG in the testicular oxidative stress indicators of rats injected with low-dose, medium-dose and high-dose groups were all reduced. Among them, the oxidative stress indicators of rats in the medium-dose and high-dose groups were close to the levels of rats in the control group, which indicates that astragaloside can reduce the level of oxidative stress and exert a certain antioxidant effect.

[0056] Figure 9 Figure A shows the GSH-PX concentration in the testicular tissue of rats in each group. Figure 9 Figure B shows the GST content in the testicular tissue of rats in each group. Figure 9 The results showed that, compared with the control group, the levels of GSH-PX and GST in the testicular tissue of the model group rats were significantly reduced. The levels of GSH-PX and GST in the testicular tissue of rats injected with low-dose, medium-dose, and high-dose astragaloside were all increased. Among them, the oxidative stress index of rats in the low-, medium-, and high-dose GSH-PX groups was close to the level of the control group rats, while the GST level of rats in the medium- and high-dose astragaloside groups recovered to the level of the control group rats. This indicates that astragaloside can restore the oxidative stress level of DEHP rats and exert a certain antioxidant effect.

[0057] In summary, from Figures 5-9 The results showed that, compared with the control group, the model group had a reduced sperm count and a higher abnormality rate; FSH and LH levels were significantly increased (P < 0.01); and T levels were significantly decreased (P < 0.01), indicating that DEHP could successfully establish a rat oligospermia model, and that high-dose astragaloside IV could indeed significantly increase serum T levels and, to a certain extent, decrease FSH and LH levels in rats. Glutathione depletion and lipid peroxidation are important indicators of oxidative stress. GPX catalyzes the oxidation of GSH to oxidized GSH (GSSG), and GSSG is recycled to reduced GSH via NADPH. Glutathione is synthesized from L-glutamate, L-cysteine, and glycine via γ-glutamylcysteine ​​and glutathione synthase. However, redox reactions are catalyzed by GSH-dependent enzymes, including GSH-PX and glutaredoxins. MDA is an indicator of lipid peroxidation and a marker of oxidative stress-induced DNA damage. These results demonstrate that astragaloside A can exert a certain degree of intervention and therapeutic effect on DEHP-induced oligoasthenospermia in SD rats. Furthermore, it indicates that astragaloside A can regulate oxidative stress and glutathione metabolism-related indicators such as MDA, T-GSH, GSH / GSSG, GSH-PX, and GST in the testes of oligoasthenospermia rats, thereby playing a therapeutic role in oligoasthenospermia.

[0058] Astragaloside A can protect sperm by reducing free radicals and oxidative stress in the body. Oxidative stress is a significant factor leading to decreased sperm quality; excessive free radicals can damage the sperm membrane, reducing sperm motility and function. Astragaloside A improves sperm quality by enhancing the activity of antioxidant enzymes (such as superoxide dismutase and catalase) and reducing free radical production. Astragaloside A can inhibit oxidative stress and improve sperm survival rate and motility.

[0059] Astragaloside A can regulate the endocrine system and promote the secretion of male hormones (such as testosterone), thereby supporting sperm growth and maturation. By regulating the secretion of FSH and LH, astragaloside A helps restore normal reproductive hormone balance and promotes sperm production.

[0060] In summary, astragaloside A significantly improved the quantity and quality of sperm in rats, and no obvious side effects were observed during the treatment. Furthermore, astragaloside A exhibits both antioxidant and hormonal balance-regulating effects in the treatment of oligoasthenospermia, providing a more comprehensive therapeutic effect and significantly improving the clinical symptoms of oligoasthenospermia compared to traditional single-mechanism treatments (such as hormone replacement therapy or simple antioxidant therapy).

[0061] Although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that modifications can still be made to the technical solutions described in the foregoing embodiments, or equivalent substitutions can be made to some of the technical features; and these modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the spirit and scope of the technical solutions of the embodiments of the present invention.

Claims

1. The use of astragaloside IV for preparing a medicament for treating oligoasthenospermia, characterized in that, The oligoasthenospermia is caused by pathogenic substances. The pathogenic substances are one or more of di(2-ethylhexyl) phthalate, cyclophosphamide and tripterygium glycosides.

2. Use according to claim 1, characterized in that, The astragaloside B improves the microenvironment of sperms to treat oligoasthenospermia.

3. Use according to claim 2, characterized in that, The astragaloside B inhibits oxidative stress, reduces the generation of free radicals, improves the microenvironment of sperms to treat oligoasthenospermia.

4. Use according to claim 2, characterized in that, The astragaloside B promotes the secretion of testosterone, improves the microenvironment of sperms, promotes the growth and maturation of sperms to treat oligoasthenospermia.

5. Use according to claim 2, characterized in that, The astragaloside B regulates the secretion of follicle stimulating hormone, improves the microenvironment of sperms, promotes the growth of sperms to treat oligoasthenospermia.

6. Use according to claim 2, characterized in that, The astragaloside B regulates the secretion of luteinizing hormone, improves the microenvironment of sperms, promotes the growth of sperms to treat oligoasthenospermia. ​

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