Application of curcumin in preparation of medicine for preventing and treating mouse spermatogonium GC1 injury caused by triptolide

Abstract

The curcumin disclosed by the invention has the medicinal activity of preventing and treating mouse spermatogonial cell GC1 injury caused by triptolide, so that the curcumin has potential application value in the aspect of preparing medicines for preventing and treating spermatogonial cell injury caused by a tripterygium wilfordii preparation. Experimental results show that curcumin can be used for protecting spermatogonial cell damage caused by triptolide by regulating oxidative stress and reducing cell apoptosis. Therefore, a new thought and approach can be provided for clinical application of traditional Chinese medicine in male reproductive system injury repair. Furthermore, a new thought and a new approach are provided for clinical adjuvant medication of the triptolide, namely, the tripterygium wilfordii preparation containing the triptolide and the curcumin is developed, so that the anti-cancer drug activity and the like of the triptolide can be exerted, the reproductive injury side effect of the triptolide is inhibited or relieved through the curcumin, and the application prospect is wide. Spermatogonial cell injury induced by triptolide is effectively recovered, and particularly, spermatogonial cell oxidative stress injury and apoptosis caused by triptolide are effectively protected.

Description

Technical Field

[0001] The invention belongs to the field of medical technology, and particularly relates to application of curcumin in preparing a medicine for preventing and treating mouse spermatogonia GC1 damage caused by triptolide. Background Art

[0002] Tripterygium wilfordii is a plant of the genus Tripterygium wilfordii of the Celastraceae family. It is one of the traditional Chinese herbal medicines in my country and has the effects of clearing away heat and detoxifying, relaxing muscles and activating blood circulation, dispelling wind and unblocking meridians, and removing dampness and swelling. Triptolide (TP), also known as triptolide and triptolide alcohol, is an epoxidized diterpene lactone compound isolated from Tripterygium wilfordii. It is one of the main active ingredients of Tripterygium wilfordii in traditional Chinese medicine, and is also the main active ingredient of Tripterygium wilfordii tablets and Tripterygium wilfordii polyglycosides. It has significant anti-cancer, anti-inflammatory, immunosuppressive and other pharmacological activities, and is widely used in the clinic to treat autoimmune diseases such as rheumatoid arthritis and systemic lupus erythematosus. However, the use of TP is prone to adverse reactions, mainly side effects of the reproductive system and blood system, and the adverse reactions of the reproductive system are manifested as menstrual disorders in women, oligospermia, asthenospermia or azoospermia in men, which greatly affects its clinical application. A large number of studies have shown that long-term use of Tripterygium wilfordii preparations can cause toxic accumulation, which can cause puberty gonadal development disorders in male patients, leading to infertility, and testicular atrophy in severe cases. At the same time, TP can induce increased apoptosis of testicular cells in mice, increased reactive oxygen concentrations, and reduced spermatogenic cells, causing varying degrees of damage to the testicles. Therefore, it is crucial to find suitable drugs to improve male reproductive damage caused by TP.

[0003] Curcumin is mainly extracted from the perennial herb turmeric and is the most active low-molecular-weight phenolic substance in turmeric. Turmeric is a traditional and commonly used Chinese medicine with a long history of application in my country. It was first seen in the "Wubi Decoction" in the "Taiping Huimin Hejijufang" of the Song Dynasty. The "Chinese Materia Medica" calls turmeric "pungent, bitter, warm, and enters the spleen and liver meridians; it breaks blood and promotes the flow of qi, and relieves pain. Pharmacological studies have shown that curcumin has multiple functions such as antioxidant, anti-inflammatory, anti-coagulant, anti-tumor and anti-atherosclerotic. Curcumin is considered to be a strong antioxidant. Studies have shown that it can resist oxidative stress by scavenging free radicals, activating antioxidant enzymes such as superoxide dismutase and catalase, and inhibiting lipid peroxidation.

[0004] Studies have found that curcumin has a protective effect on oxidative damage to spermatogenic cells and sperm in male mice, but there are no reports on the protection of curcumin against TP-induced spermatogonia damage. Summary of the invention

[0005] The technical problem to be solved by the present invention is to provide an application of curcumin in the preparation of a medicine for preventing and treating mouse spermatogonia GC1 damage caused by triptolide.

[0006] In order to solve the above technical problems, the present invention adopts the following technical solutions:

[0007] Application of curcumin in the preparation of medicine for preventing and treating spermatogonia damage caused by tripterygium wilfordii preparations.

[0008] Tripterygium wilfordii preparations contain triptolide.

[0009] Spermatogonia damage includes decreased spermatogonia proliferation rate and decreased vitality.

[0010] Curcumin protects spermatogonia from triptolide-induced damage by regulating oxidative stress and reducing cell apoptosis.

[0011] The spermatogonia were mouse spermatogonia GC1.

[0012] A drug for preventing and treating spermatogonia damage caused by tripterygium wilfordii preparations, the drug containing curcumin.

[0013] This medicine is for oral use.

[0014] A tripterygium wilfordii preparation comprising tripterygium wilfordii and curcumin.

[0015] The weight ratio of triptolide to curcumin is 0.2:2 to 0.2:6.

[0016] The weight ratio of triptolide to curcumin is 0.2:4.

[0017] In view of the problem of male reproductive system damage in current tripterygium wilfordii preparations, the inventors have found that curcumin has medicinal activity in preventing and treating mouse spermatogonia GC1 damage caused by tripterygium wilfordii preparations. Therefore, curcumin has potential application value in the preparation of drugs for preventing and treating spermatogonia damage caused by tripterygium wilfordii preparations. The experimental results show that curcumin is non-toxic to mouse spermatogonia within a concentration range below 8μmol / L; 4μmol / L curcumin can significantly improve the decrease in spermatogonia GC1 proliferation rate, oxidative stress level and cell apoptosis caused by tripterygium wilfordii. This shows that curcumin may protect spermatogonia damage caused by tripterygium wilfordii by regulating oxidative stress and reducing cell apoptosis. Based on this, it can provide new ideas and approaches for the clinical application of traditional Chinese medicine in the repair of male reproductive system damage. Furthermore, a new idea and approach is provided for the clinical use of triptolide, that is, the development of a triptolide-turmeric preparation containing both triptolide and curcumin, which can not only exert the anti-cancer and other drug activities of triptolide, but also inhibit or reduce the reproductive damage side effects of triptolide through curcumin, effectively restore triptolide-induced spermatogonia damage, especially effectively protect spermatogonia induced by triptolide from oxidative stress damage and apoptosis. In summary, the present invention has broad application prospects in spermatogonia damage and male reproductive system damage. BRIEF DESCRIPTION OF THE DRAWINGS

[0018] Figure 1 This is a graph showing the toxicity test results of curcumin on spermatogonia in Example 1.

[0019] Figure 2 This is a graph showing the protective effect of different concentrations of curcumin on the decrease in cell proliferation rate caused by triptolide in Example 2.

[0020] Figure 3 The results of the protective effect of curcumin on the oxidative stress process caused by triptolide in Example 3 are shown in Figure a, which are the results of the flow cytometry detection of active oxygen in the control group, TP group and TP+4 μmol / L curcumin group, and b are the statistical results of the active oxygen levels in the control group, TP group and TP+4 μmol / L curcumin group.

[0021] Figure 4 The results of the protective effect of curcumin on the apoptosis caused by triptolide in Example 4 are shown in Figure 4, in which: ac are flow cytometry results of apoptosis in the control group, TP group and TP+4 μmol / L curcumin group, and d is a statistical result of apoptosis levels in the control group, TP group and TP+4 μmol / L curcumin group. DETAILED DESCRIPTION

[0022] Example 1 Detection of cytotoxicity of curcumin on mouse spermatogonia

[0023] Mouse spermatogonia GC1 was selected as a model. Curcumin was dissolved in dimethyl sulfoxide, the mother solution concentration was 1mmol / L, and curcumin solutions with concentrations of 2, 4, 6, 8, 10, 12, 14, 16, 18, and 20μmol / L were prepared by stepwise dilution, and the diluent was DMEM cell culture medium. The control group was treated with an equal volume of 0.1% dimethyl sulfoxide. GC1 cells were seeded in 96-well plates, and the above-mentioned different concentrations of curcumin solution and dimethyl sulfoxide were added when the cells were in the logarithmic growth phase. After 24 hours of treatment, the CCK8 kit was used to detect cell growth.

[0024] The results are as follows Figure 1 As shown, curcumin concentrations below 8 μmol / L did not significantly change the proliferation rate of GC1 cells, that is, it did not affect the proliferation rate of GC1 cells and had no obvious toxicity to mouse spermatogonia.

[0025] Example 2 Protective effect of curcumin on the decrease in spermatogonia proliferation rate in mice induced by triptolide

[0026] GC1 cells (5×10 4 / mL) were inoculated in 96-well culture plates, 100μL / well, and 100μL (curcumin solution) of drug in each well. Curcumin was dissolved in dimethyl sulfoxide, the mother solution concentration was 1mmol / L, and the final concentration was 2, 4, and 6μmol / L by stepwise dilution. It was incubated with 200nmol / L concentration of triptolide solution for 24h, and 5 wells for each concentration. A control group and a model group were set up. The control group was treated with an equal volume of 0.1% dimethyl sulfoxide, and the model group was treated with 200nmol / L concentration of triptolide solution. CCK8 kit was used to detect cell growth after 24h of drug action.

[0027] The results are as follows Figure 2 As shown, curcumin at a concentration of 2-6 μmol / L, especially at a concentration of 4 μmol / L, significantly improved the cell proliferation rate induced by triptolide, indicating that curcumin played a protective role against the decreased viability of mouse spermatogonia caused by triptolide.

[0028] Example 3 Protective effect of curcumin on triptolide-induced oxidative stress in mouse spermatogonia

[0029] GC1 cells (5×10 5 / mL) were inoculated in 6-well culture plates, 1mL / well, 1mL of drug in each well, so that the final concentration was 4μmol / L, and incubated with 200nmmol / L triptolide solution for 24h. A control group and a model group were set up. The control group was treated with an equal volume of 0.1% dimethyl sulfoxide, and the model group was treated with 200nmol / L triptolide solution. After 24h of drug action, the ROS detection kit and flow cytometer were used to detect the cellular oxidative stress level.

[0030] The results are as follows Figure 3 As shown, 4 μmol / L curcumin significantly improved the oxidative stress level of mouse spermatogonia treated with triptolide, indicating that curcumin played a protective role against the oxidative stress of mouse spermatogonia caused by triptolide.

[0031] Example 4 Protective effect of curcumin on triptolide-induced apoptosis of mouse spermatogonia

[0032] GC1 cells (5×10 5 / mL) were inoculated in 6-well culture plates, 1mL / well, 1mL of drug in each well, so that the final concentration was 4μmol / L, and incubated with 200nmmol / L triptolide solution for 24h. A control group and a model group were set up. The control group was treated with an equal volume of 0.1% dimethyl sulfoxide, and the model group was treated with 200nmol / L triptolide solution. After 24h of drug action, the cell apoptosis rate was detected using a cell apoptosis detection kit and a flow cytometer.

[0033] The results are as follows Figure 4 As shown, 4 μmol / L curcumin significantly improved the apoptosis rate of spermatogonia in mice treated with triptolide, which indicates that curcumin can significantly improve the apoptosis of spermatogonia in mice caused by triptolide.

Claims

1. The application of curcumin in the preparation of drugs for preventing and treating spermatogonia damage caused by Tripterygium wilfordii preparations.

2. The use according to claim 1, characterized in that: The tripterygium wilfordii preparation contains tripterygium wilfordii.

3. The use according to claim 1, characterized in that: The spermatogonia damage includes decreased spermatogonia proliferation rate and decreased activity.

4. The use according to claim 1, characterized in that: The curcumin protects spermatogonia from damage caused by triptolide by regulating oxidative stress and reducing cell apoptosis.

5. The use according to claim 1, characterized in that: The spermatogonia are mouse spermatogonia GC1.

6. A drug for preventing and treating spermatogonia damage caused by a Tripterygium wilfordii preparation, characterized in that This medicine contains curcumin.

7. The drug according to claim 6, characterized in that This medicine is for oral use.

8. A tripterygium wilfordii preparation, characterized in that This preparation contains triptolide and curcumin.

9. The tripterygium wilfordii preparation according to claim 7, characterized in that: The weight ratio of triptolide to curcumin is 0.2:2 to 0.2:

6.

10. The tripterygium wilfordii preparation according to claim 7, characterized in that: The weight ratio of triptolide to curcumin is 0.2:4.

Images