Pharmaceutical composition for preventing or relieving sperm motor dysfunction caused by fenvalerate and application thereof

Abstract

The invention relates to the technical field of biological medicine, in particular to a pharmaceutical composition for preventing or relieving sperm motor dysfunction caused by fenvalerate, and the pharmaceutical composition comprises N-acetylcysteine and / or cyclosporin A. Based on the discovery that fenvalerate induces rise of reactive oxygen species (ROS) to cause mitochondrial dysfunction and destroy a cAMP / PKA signaling pathway so as to inhibit sperm motility and induce mitochondrial apoptosis. An in-vitro rat sperm model proves that NAC and CsA can effectively reverse sperm motility reduction, ATP / cAMP level reduction, mitochondrial membrane potential dissipation and cell apoptosis caused by fenvalerate. The invention provides a new prevention and treatment strategy for treating male reproductive function damage caused by exposure of pyrethroid pesticides.

Description

Technical Field

[0001] This invention relates to the field of biomedical technology, specifically to pharmaceutical compositions for preventing or alleviating sperm motility disorders caused by cypermethrin and their applications. Background Technology

[0002] Cypermethrin is a Class II synthetic pyrethroid widely used in agricultural and household pest control. Studies have shown that it induces reproductive toxicity through endocrine disruption mechanisms, including inhibition of testosterone synthesis and disruption of spermatogenesis. Existing research largely focuses on its endocrine-disrupting effects, but its direct toxic mechanism to mature sperm remains unclear. This study found that cypermethrin can directly and rapidly reduce sperm motility, an effect associated with oxidative stress, mitochondrial dysfunction, and disruption of the cAMP / PKA signaling pathway. Currently, there are no effective protective or therapeutic agents specifically targeting the direct reproductive toxicity of this type of pesticide. Summary of the Invention

[0003] The present invention aims to provide a pharmaceutical composition for preventing or alleviating sperm motility dysfunction caused by cypermethrin, wherein the pharmaceutical composition can effectively antagonize the direct toxicity of cypermethrin to sperm.

[0004] Firstly, to achieve the above objectives, the present invention adopts the following technical solution: A pharmaceutical composition for the prevention or relief of sperm motility dysfunction caused by cypermethrin, comprising N-acetylcysteine ​​(NAC) and / or cyclosporine A (CsA). Sperm motility dysfunction is characterized by reduced sperm motility and weakened forward movement.

[0005] In a second aspect, the present invention also provides a medicament comprising the aforementioned medicament composition.

[0006] Furthermore, the concentrations of the drugs used are as follows: N-acetylcysteine ​​(NAC) at 5 μM and cyclosporine A (CsA) at 1 μM.

[0007] Thirdly, the present invention also provides the use of the pharmaceutical composition and the pharmaceutical composition in the preparation of a medicament for preventing or alleviating sperm motility dysfunction caused by cypermethrin.

[0008] Fourthly, the use of at least one of N-acetylcysteine ​​and cyclosporine A in the preparation of a medicament for the prevention or relief of male reproductive dysfunction caused by cypermethrin.

[0009] The beneficial technical effects of the pharmaceutical composition or drug described in this invention include: 1. It can reduce the increase in reactive oxygen species (ROS) levels in sperm caused by cypermethrin exposure.

[0010] 2. Improves the decrease in sperm mitochondrial membrane potential (ΔΨm) caused by cypermethrin exposure.

[0011] 3. It enhances the reduction in sperm ATP and cAMP levels caused by cypermethrin exposure.

[0012] 4. It inhibits the mitochondrial apoptosis pathway activated by cypermethrin exposure and restores the phosphorylation and expression of flagellar motility-related proteins. Inhibition of the mitochondrial apoptosis pathway is manifested by reduced cytochrome C release and decreased levels of cleavage-9 and cleavage-3.

[0013] 5. Improve sperm motility by restoring the phosphorylation level of PKA substrates and / or the protein expression levels of AKAP3 and AKAP4 in sperm flagella.

[0014] 6. NAC and CsA can effectively antagonize the direct toxicity of cypermethrin to sperm and significantly restore sperm motility inhibited by cypermethrin. Attached Figure Description

[0015] Figure 1 Effects of cypermethrin exposure on sperm motility, reactive oxygen species (ROS) levels, and energy metabolism in rats. (A, B) Changes in sperm motility parameters after cypermethrin treatment. (C) Quantitative analysis of ROS levels in sperm of the experimental group. (D, E) ATP and cAMP levels in sperm after cypermethrin exposure. Data are expressed as mean ± standard error (n=10 per group). Statistical significance: P>0.05 (ns, no significant difference), **P<0.01 (compared to control group, or between treatment groups as specified).

[0016] Figure 2 The level of reactive oxygen species (ROS) in sperm cells was detected by flow cytometry using the DCFH-DA probe. When the main peak shifted to the right along the x-axis, the fluorescence intensity increased, indicating an increase in intracellular ROS levels; if the main peak shifted to the left, it indicated a decrease in fluorescence and a decrease in ROS content (indicated by the arrows).

[0017] Figure 3 Effects of cypermethrin exposure on mitochondrial membrane potential (ΔΨM) and GAPDS expression in rat sperm. (A) Quantitative analysis of sperm with low ΔΨM. (B, C) Western blot analysis of GAPDS expression levels in the control and treatment groups. Data are expressed as mean ± standard error (n=10 per group). Statistical significance: P>0.05 (ns, no significant difference), **P<0.01 (compared to the control or designated treatment group).

[0018] Figure 4ΔΨM flow cytometry analysis performed using JC-1 staining. The upper right quadrant Q2-2 shows sperm with high ΔΨM values ​​(red fluorescence), while the lower right quadrant Q2-4 corresponds to low ΔΨM values ​​(green fluorescence, indicated by arrows). The observed decrease in ΔΨM values ​​indicates an alteration in mitochondrial membrane potential (mPTP) after sperm exposure to permethrin.

[0019] Figure 5 Cypermethrin exposure alters the expression of flagellar-related proteins. (A) Immunoblot analysis showed the expression levels of flagellar proteins (AKAP3, AKAP4, DYNLT2A1) and phosphorylated PKR2 (pS99) in each experimental group. (B) Quantitative analysis of protein expression levels. Data are expressed as mean ± standard error (n=10 per group). Statistical significance: P>0.05 (ns, no significant difference), **P<0.01 (compared to the control group or designated treatment group).

[0020] Figure 6 Effects of cypermethrin exposure on phosphorylation levels and spatial distribution of PKA substrates in rat sperm flagella. (A) Immunofluorescence localization of phosphorylated (Ser / Thr) PKA substrates in sperm flagella. Combined images (Alexa Fluor 488 labeled phosphorylated PKA substrates appear green; DAPI nuclear staining appears blue) show differences in phosphorylation patterns in different regions of the flagella: the midpiece and proximal protuberance regions show significant signal intensity, while the distal protuberance and terminal protuberance regions show weaker staining. (B, C) Quantitative analysis of phosphorylated (Ser / Thr) PKA substrate expression in sperm from different treatment groups. Data are expressed as mean ± standard error (n=10 per group). Statistical significance: P>0.05 (ns, no significant difference), **P<0.01 (compared to the control group or designated treatment group).

[0021] Figure 7 Cypermethrin-induced sperm apoptosis in rats. (A) Quantitative analysis of the percentage of apoptotic sperm in different experimental groups. (BE) Immunoblot detection of mitochondrial apoptosis pathway markers showed that mitochondrial exposure to cypermethrin resulted in the release of cytochrome C and activation of executive caspases (cleaved caspase 3 and caspase 9). Data are expressed as mean ± standard error (n=10 per group). Statistical significance: P>0.05 (ns, no significant difference), **P<0.01 (compared to the control group or designated treatment group).

[0022] Figure 8: Flow cytometry analysis of sperm apoptosis was performed using Annexin V-FITC / PI double staining. The upper right quadrant Q2-2 shows late apoptotic cells (PI+ / Annexin V+), and the lower right quadrant Q2-4 shows early apoptotic cells (PI- / Annexin V+). The total apoptosis rate was calculated by summing the proportions of cells in the two quadrants.

[0023] Figure 9 : Schematic diagram of the structure of rat sperm flagella. (A) Optical micrograph showing three different regions of the flagellum: midpiece, principal piece, and end piece. (B) Transmission electron micrograph of a cross-section of the midpiece. The ultrastructure of the flagellum from outside to inside is as follows: plasma membrane (PM), mitochondrial sheath (MS), and nine outer dense fibers (ODFs). In the internal structure, the axoneme contains nine outer microtubule doublets (OMDA), which contain associated dynein arms (DA), radial spokes (RS), and a central pair of microtubules (CP). (C) Transmission electron micrograph of a cross-section of the principal piece, where the mitochondrial sheath is replaced by a fibrous sheath (FS), while other axoneme components remain intact. (D) Transmission electron micrograph of a cross-section of the end piece, with only the plasma membrane and a simplified axoneme structure remaining, while the mitochondrial sheath and fibrous sheath are absent. Specific implementation methods

[0024] The following is a further detailed description through specific implementation methods: I. Materials and methods 1. Chemicals and reagents Fenvalerate (purity 98.8%) was purchased from the National Institute of Metrology of China (Beijing, China). ATP, reactive oxygen species (ROS), Annexin V-FITC apoptosis detection kit, and JC-1 mitochondrial membrane potential probe kit were all purchased from Beyotime Biotechnology Co., Ltd. (Nanjing, China). The commercial ELISA kit for cAMP quantification was purchased from Jimei Biotechnology Co., Ltd. (Wuhan, China). All primary and secondary antibodies are listed in Table 1. Other analytical pure chemicals and solvents were purchased from Sigma-Aldrich Co., Ltd. (Shanghai, China).

[0025] 2. Animals and sperm preparation Sexually mature male Sprague-Dawley rats (8-9 weeks old) used in this study were provided by the Laboratory Animal Center of Zunyi Medical University (license numbers: SCXK (Qian) 2021-0002, SYXK (Qian) 2021-0004). The experimental animals were housed individually in an environment with constant temperature and humidity (24±2°C, 55-65% humidity), using a 12-hour light / 12-hour dark circadian cycle mode, and had free access to food and water. All experimental operations were carried out in accordance with the specifications approved by the Laboratory Animal Welfare and Ethics Committee of Zunyi Medical University, and the ethical approval documents can be provided upon request.

[0026] After CO2 anesthesia, rats were euthanized via cervical dislocation. The epididymis was then removed, adipose tissue was excised, and the junction of the tail and epididymis was clamped. Sperm separation: The tail epididymis was dissected and immersed in 3 ml of preheated M199 medium containing 0.1% bovine serum albumin (BSA). The tissue was longitudinally dissected along the proximal and distal regions. After removing tissue fragments, the sperm suspension was placed in a 37°C incubator and diluted to approximately 2 × 10⁻⁶ with freshly prepared M199 / BSA solution. 6 Sperm concentration per milliliter. Sperm samples from ten rats in each group were mixed and the compound was added, marking the incubation time point as 0 hours.

[0027] 3. Experimental Design The cypermethrin stock solution (40 mM) was prepared with dimethyl sulfoxide (DMSO) and stored at 4°C. Working solutions (1, 10, and 20 mM) were all diluted with DMSO. During treatment, 1 μL of DMSO (solvent control) or cypermethrin working solution was added per mL of M199 medium, resulting in final cypermethrin concentrations of 0, 1, 10, and 20 μM. Preliminary tests confirmed that maintaining a DMSO concentration below 0.1% (volume fraction) did not affect sperm motility.

[0028] Based on preliminary dose-response experiments (see “Results” below), we determined the optimal concentration (20 μM) of cypermethrin and the exposure time (2 hours). Sperm suspensions were then divided into four treatment groups (10 samples per group) and incubated for 2 hours under physiological conditions (37°C, 5% CO2, humid environment): (I) Control group (CG): 0 μM cypermethrin (solvent only); (II) Cypermethrin exposure group (FEG): 20 μM cypermethrin; (III) Cypermethrin + N-acetylcysteine ​​group (Feng): 20 μM cypermethrin + 5 μM NAC; (IV) Cypermethrin + cyclosporine A group (FECG): 20 μM cypermethrin + 1 μM CsA. All suspensions were prepared in M199 medium containing the corresponding compounds.

[0029] 4. Sperm motility analysis Sperm motility was observed using a phase-contrast microscope (200x) and classified according to the World Health Organization (WHO) 2010 standards: Grade A (rapid forward motility), Grade B (slow forward motility), Grade C (no forward motility), and Grade D (no motility). The total percentage of Grade A and Grade B sperm was used as a reference indicator for sperm motility.

[0030] 5. Measurement of reactive oxygen species in sperm cells Intracellular reactive oxygen species (ROS) levels were detected using 2',7'-dichlorodihydrofluorescein diacetate (DCFH-DA; Sigma-Aldrich) according to the manufacturer's instructions. Sperm suspensions were centrifuged (1000×g, 4°C, 1 min), washed twice with ice-cold PBS, and resuspended in 0.5 mL of 15 μM DCFH-DA solution. After incubation at 25°C in the dark for 20 min, intracellular ROS oxidized DCFH-DA to the fluorescent substance DCF. Fluorescence intensity was detected using a NovoCyte flow cytometer (Agilent Technologies, Agilent Biosciences) at excitation wavelengths of 488 nm and emission wavelengths of 525 nm. Data are expressed as mean fluorescence intensity of DCF-positive sperm.

[0031] 6. Quantitative analysis of intracellular ATP and cAMP levels in sperm cells ATP content was detected using a commercial kit from Abcam (UK). The specific procedure was as follows: Sperm pellet was lysed in 100 μL of ice-bath buffer (4°C, 10 min), centrifuged at 10,000 × g for 10 min at 4°C, and 50 μL of the supernatant was mixed with an equal volume of luciferase assay reagent. The luminescence intensity was detected in real time using a Luminoskan chemiluminescence analyzer from Thermo Fisher Scientific. Finally, the ATP content was converted into nanomolar concentration per milligram of protein according to the standard curve.

[0032] cAMP levels were detected using an ELISA kit (R&D Systems, USA). After ultrafiltration, the absorbance was measured at 405 nm using a BioTek microplate reader. The cAMP concentration (pmol / mg protein) was calculated using a standard curve and the inverse relationship between absorbance and concentration.

[0033] 7. Measurement of sperm mitochondrial membrane potential Mitochondrial membrane potential (ΔΨm) was assessed using the JC-1 fluorescent probe. Sperm pellet was resuspended in 0.5 mL of JC-1 working solution (10 μg / mL in PBS) and incubated in the dark at 25°C for 20 min. Dual emission signals were acquired using a BDFACSCanto II flow cytometer (BD Biosciences) with excitation at 488 nm and emission at 530 ± 15 nm (monomers, green) and 585 ± 20 nm (J-aggregates, red). ΔΨm was quantified by the red / green fluorescence intensity ratio, where a decrease indicated mitochondrial depolarization.

[0034] 8. Sperm apoptosis detection Sperm apoptosis was assessed using a double staining method with Annexin V-FITC and propidium iodide (PI). Washed sperm pellets were resuspended in 100 μL PBS (pH 7.4) and incubated with 10 μL Annexin V-FITC (0.01 μg / mL) and 10 μL PI (1 mg / mL) at 25°C in the dark for 20 min. Early apoptotic sperm exhibited phosphatidylserine eversion, binding to Annexin V-FITC (green fluorescence). Late apoptosis and necrosis produced double green / red fluorescence due to membrane rupture allowing PI entry. Flow cytometry analysis was performed. The apoptosis index was defined as the percentage of Annexin V-FITC-positive or Annexin V-FITC-positive / PI-positive sperm in the total population.

[0035] 9. Immunofluorescence localization of phosphorylated serine / threonine (Ser / Thr) PKA substrates in sperm flagella. Sperm motility is regulated by cAMP / PKA-mediated phosphorylation of flagellin proteins, such as dynein. PKA substrate specificity requires the presence of arginine at the -3 position of phosphorylated serine / threonine residues. To achieve spatial resolution of PKA-dependent phosphorylation, researchers performed immunofluorescence experiments. Sperm samples were fixed in 4% paraformaldehyde (5 min), attached to glass slides, permeabilized with 0.5% Triton X-100 membrane, and incubated overnight at 4°C with an antiphosphorylation (serine / threonine) PKA substrate primary antibody. Detection was performed using Alexa Fluor 488-labeled donkey anti-rabbit secondary antibody, and cell nuclei were counterstained with DAPI (5 μg / mL). Image acquisition was performed using a Nikon fluorescence microscope (Tokyo, Japan).

[0036] 10. Western blot analysis Sperm precipitate was lysed and centrifuged (15,000 × g, 15 min), and the supernatant was stored at -20°C. Protein concentration was determined using a BCA kit (Pierce Biotechnology, USA). 50 μg of protein sample was separated by 12% SDS-PAGE electrophoresis and transferred to a PVDF membrane (Merck Millipore, USA). The membrane was blocked with 5% skim milk for 2 hours, washed with TBST, and then incubated overnight at 4°C with primary antibodies against GAPDS, AKAP3, AKAP4, DYNLT1, PKA RIIβ (phosphorylated S99 site), phosphorylated (serine / threonine) PKA substrate, cytochrome c, cleaved caspase-3, and cleaved caspase-9 (see Table 1 for details). After multiple TBST washes, the membrane was incubated at room temperature for 2 hours with rabbit anti-rabbit secondary antibody (labeled with bovine serum albumin, HRP). Protein bands were visualized using an ECL kit (Pierce Biotechnology, USA) and imaged using a ChemiDoc XRS+ imaging system (Bio-Rad Laboratories, USA). Finally, imageJ software (v2.3.0, National Institutes of Health, USA) was used for quantitative analysis of grayscale values, and the data were standardized using an internal control.

[0037] Table 1 Primary and secondary antibodies used for immunofluorescence and immunoblotting analysis.

[0038] 11. Statistical Analysis Data are expressed as mean ± standard error. Statistical analysis was performed using SPSS 17.0 software (SPSS Inc., USA). One-way ANOVA combined with appropriate post-hoc tests was used for inter-group comparisons. A p-value < 0.05 was considered statistically significant, and a p-value < 0.01 was considered highly significant.

[0039] II. Results 1. Determination of Cypermethrin Exposure Parameters and Experimental Grouping Preliminary observation using an optical microscope revealed that, compared to the control group (0 μM), treatment with 20 μM cypermethrin for 2 hours significantly reduced sperm motility in rats (P < 0.01). Figure 1 A). No significant differences were observed in other dose-time combinations (P>0.05). Therefore, a 20 μM concentration and a 2-hour treatment regimen were selected for subsequent experiments.

[0040] To investigate the effects of reactive oxygen species (ROS) and mitochondrial dysfunction on cypermethrin-induced sperm motility disorders, the experiment used the ROS scavenger NAC (5 μM) and the mitochondrial membrane potential (mPTP) inhibitor CsA (1 μM). Sperm samples were divided into four groups: (I) control group (CG, 0.1% dimethyl sulfoxide solvent), (II) cypermethrin exposure group (FEG, 20 μM cypermethrin), (III) cypermethrin combined with NAC group (Feng, 20 μM cypermethrin + 5 μM NAC), and (IV) cypermethrin combined with CsA group (FECG, 20 μM cypermethrin + 1 μM CsA). All samples were incubated under standardized conditions for 2 hours.

[0041] 2. Effects of cypermethrin on sperm motility and the protective effects of NAC and CsA. Compared with the control group, exposure to cypermethrin significantly reduced sperm motility (P<0.01); Figure 1 B). This damage was significantly reduced when NAC or CsA were used in combination (P<0.01 vs. cypermethrin alone). These results indicate that cypermethrin impairs sperm motility through mechanisms of oxidative stress and mitochondrial dysfunction.

[0042] 3. Sperm oxidative stress caused by exposure to cypermethrin Flow cytometry analysis showed that the DCFH-DA fluorescent probe ( Figure 1 C and Figure 2 Analysis revealed that, compared with the control group, the DCF fluorescence intensity in sperm exposed to cypermethrin was significantly increased (P<0.01), indicating a significant increase in oxidative stress levels. Combined treatment with NAC and CsA effectively inhibited the excessive generation of reactive oxygen species (ROS) induced by cypermethrin (P<0.01 compared with the cypermethrin-only group), confirming the antioxidant protective effects of these two substances.

[0043] 4. Effects of cypermethrin on sperm ATP and cAMP levels Given the crucial roles of mitochondrial oxidative phosphorylation and glycolysis in cellular energy metabolism, we performed quantitative analysis of ATP and cAMP levels in sperm exposed to cypermethrin. Figure 1 D and Figure 1 E). Compared with the control group, cypermethrin significantly reduced ATP and cAMP levels (P<0.01). Combined use with NAC or CsA effectively restored these energy metabolites (P<0.01). These results indicate that cypermethrin impairs mitochondrial function and disrupts the cAMP / PKA signaling pathway, which is crucial for sperm motility regulation.

[0044] 5. Effects of cypermethrin on sperm ΔΨm and GAPDS levels We used the JC-1 fluorescent probe to detect sperm ΔΨm levels ( Figure 3 A and Figure 4 The expression level of GAPDS was determined by Western blotting. Figure 3 B and Figure 3 C). Compared with the control group, cypermethrin treatment significantly reduced sperm ΔΨm levels (P<0.01), while the combined use of NAC or CsA effectively restored ΔΨm levels (P<0.01). In contrast, GAPDS expression did not change significantly in any of the treatment groups (P>0.05). These results indicate that cypermethrin impairs sperm function by inhibiting mitochondrial oxidative phosphorylation without affecting glycolysis.

[0045] 6. Cypermethrin interferes with the PKA / AKAP signaling pathway in sperm flagellar regulation. The regulation of sperm flagellar motility relies on a protein kinase A (PKA)-mediated mechanism, which requires phosphorylation of dynein through anchoring via AKAP-mediated binding. Compared to the control group, cypermethrin significantly inhibited PKR2 phosphorylation and AKAP3 / AKAP4 expression. Figure 5 A and Figure 5 B; P<0.01). Combined use of NAC or CsA can restore ATP and cAMP levels (B < P < 0.01). Figure 1 (D and 1E; P<0.01), indicating that these compounds may exert a protective effect by repairing the disruption of flagellar regulatory pathways caused by cypermethrin.

[0046] 7. PKA-mediated phosphorylation that interferes with sperm motility due to cypermethrin. The motility of mammalian sperm is driven by flagella, a structure composed of four parts: the connective segment, the midpiece, the principal segment, and the terminal segment. The core axonal filament exhibits a "9+2" microtubule arrangement—nine peripheral duodenal assemblages (OMDAs) surround the central pair (CP), providing passive structural support through peripheral dense fibers (ODFs). The midpiece contains the mitochondrial sheath (MS) responsible for energy production, while the principal segment is enclosed by a fibrous sheath (FS), limiting the plane of flagellar movement. Figure 9 These structures work synergistically to promote and regulate sperm motility. Adenosine triphosphate (ATP) is the primary energy source for flagellar beating, powering the dynein ATPase in sperm microtubules. As the molecular motor of movement, dynein activity is regulated by the cAMP / PKA signaling pathway.9 Although the mid-mitochondria produce most of the ATP, ATP diffusion is limited due to the greater distance in the distal regions of the flagellum (major and terminal segments). Therefore, these regions mainly rely on glycolysis for energy, with germ cell-specific glyceraldehyde-3-phosphate dehydrogenase-S (GAPDS) playing a crucial role in maintaining local ATP supply.

[0047] Dynein, as the main substrate of pKA in sperm flagella, regulates motility through a phosphorylation-dependent cascade. After the regulatory subunit of pKA is anchored by A-kinase anchored proteins (AKAPs), it phosphorylates the dynein arm, inducing a conformational change, thereby achieving interaction with microtubules and driving flagellar movement. Immunofluorescence assays confirmed significant phosphorylation of the pKA substrate in the flagellar and acrosome regions, with particularly strong signals in the midpiece and main segments. Figure 6 A).

[0048] Cypermethrin specifically impairs the PKA signaling pathway by inhibiting phosphorylation. Although the expression level of Tctex-2A1 protein (DYNLT2A1) remained stable across groups ( Figure 5 A and Figure 5 B; P>0.05), but cypermethrin significantly reduced the phosphorylation level of the regulatory subunit (RII / PKR2), thereby leading to reduced phosphorylation of downstream substrates (B; P>0.05), Figure 5 A, Figure 5 B. Figure 6 B and Figure 6 C; P<0.01). Co-treatment with NAC or CsA effectively restored these phosphorylation defects (P<0.01). These results indicate that benzimidazole esters disrupt sperm motility by specifically inhibiting RII / PKR2 phosphorylation and subsequent PKA substrate activation, without altering the expression levels of core motility components.

[0049] 8. Cypermethrin induces mitochondrial-mediated apoptosis by opening the mitochondrial membrane potential (mPTP) in rat sperm. Cypermethrin exposure significantly increased sperm apoptosis rate ( Figure 7 A and Figure 8 (P<0.01), accompanied by increased cytochrome C release and activation of cleavable Caspase 9 and Caspase 3 ( Figure 7 B~ Figure 7 E; P<0.01). The combined use of NAC or CsA can effectively alleviate these apoptotic effects (P<0.01), confirming that oxidative stress and mitochondrial permeability transition pore (mPTP) opening are key mechanisms for cypermethrin-induced endogenous apoptosis.

[0050] The above descriptions are merely embodiments of the present invention, and common knowledge such as specific technical solutions and / or characteristics are not described in detail here. It should be noted that those skilled in the art can make various modifications and improvements without departing from the technical solutions of the present invention, and these should also be considered within the scope of protection of the present invention. These modifications and improvements will not affect the effectiveness of the implementation of the present invention or the practicality of the patent. The scope of protection claimed in this application should be determined by the content of its claims, and the specific embodiments described in the specification can be used to interpret the content of the claims.

Claims

1. A pharmaceutical composition for preventing or alleviating the sperm motility dysfunction caused by cypermethrin, characterized by comprising: N-acetylcysteine and / or cyclosporin A.

2. A medicament, characterized by: The pharmaceutical composition of claim 1.

3. The medicament according to claim 2, characterized in that: The concentration of the drug used is 5 μM for N-acetylcysteine and 1 μM for cyclosporin A.

4. Use of the pharmaceutical composition of claim 1 or the drug of claim 2 or 3 for the manufacture of a medicament for preventing or alleviating the impairment of male reproductive function caused by cypermethrin.

5. Use of at least one selected from N-acetylcysteine and cyclosporin A for the manufacture of a medicament for preventing or alleviating the impairment of male reproductive function caused by cypermethrin.

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