目的 探究山药Dioscoreae Rhizoma（DR）-黄精Polygonati Rhizoma（PR）药对（DR-PR）改善大鼠少弱精子症（OAS）的药效与作用机制。方法 SD大鼠适应性喂养1周后，随机分为6组：对照组、模型组、DR-PR（1∶1）组(2.5 g·kg^-1DR+2.5 g·kg^-1 PR)、DR-PR（2∶1）组(5.0 g·kg^-1 DR+2.5 g·kg^-1 PR)、DR-PR（1∶2）组(2.5 g·kg^-1 DR+5.0 g·kg^-1 PR)、左卡尼汀(LOS,100 mg·kg^-1)组，除对照组外，其余5组ig给予雷公藤多苷35 mg·kg^-1 7 d诱导OAS模型，从造模开始即给药，于每天9∶00时禁食，15∶00时ig给药干预，共干预21 d。精液常规检测大鼠精子数量和活力；ELISA测定大鼠血清中性激素含量；苏木精-伊红（HE）染色检测DR-PR对OAS大鼠睾丸病理的影响。网络药理学分析DR-PR治疗OAS的主要活性成分及作用机制：分别以山药和黄精为关键词，在中药系统药理数据库和分析平台（TCMSP）中检索主要活性成分，于GeneCards数据库、DrugBank数据库和OMIM数据库中进行OAS靶点搜集；使用VLOOCUP函数将OAS靶点和DR-PR靶点进行交集处理，得到DR-PR治疗OAS的潜在靶点。将潜在靶点导入STRING数据库进行蛋白质-蛋白质相互作用（PPI）分析；将DR-PR治疗OAS的潜在靶点信息导入Metascape数据库进行基因本体（GO）富集和京都基因与基因组百科全书（KEGG）富集分析。TUNEL染色测定各组大鼠睾丸细胞凋亡状况；Western blotting检测各组大鼠睾丸组织热休克蛋白90α型1（HSP90AA1）、半胱氨酸蛋白酶（Caspase）-3、Caspase-9、核因子（NF）-κB、磷酸化NF-κB（p-NF-κB）、p38和磷酸化p38（p-p38）表达。分子对接模拟探索DR-PR关键活性成分和关键靶点的结合能力和作用方式。结果 与模型组相比，DR-PR组大鼠精子浓度、活力、存活率、直线运动速率、曲线运动速率、平均路径速度、运动平直度和运动线性显著恢复（P<0.05、0.01），血清性激素酮和雌二醇含量显著升高（P<0.01），睾丸生精细胞和间质细胞损伤得到缓解（P<0.01）,DR-PR（1∶2）组效果最好。网络药理学结果表明，检索出山药活性成分16种、黄精活性成分12种；DR-PR治疗OAS的关键靶点为MAPK14、NFKB1、TNF、IL6、HSP90AA1、CASP3和CASP9，关键作用途径为IL-17信号通路，关键生物学功能为对脂多糖的反应，关键细胞组分是膜筏和囊泡管腔，关键分子功能为蛋白丝氨酸/苏氨酸激酶活性。与模型组相比，DR-PR组大鼠的睾丸组织细胞凋亡显著降低（P<0.01），睾丸组织HSP90AA1、Caspase-3和Caspase-9（P<0.01）表达下调，NF-κB和p38磷酸化水平降低（P<0.01）,DR-PR（1∶2）组效果最好。分子对接结果表明，DR-PR关键活性成分和关键靶点HSP90AA1、Caspase-3、Caspase-9、NF-κB和p38均可形成自发结合。结论 DR-PR可通过调控HSP90/p38/NF-κB通路减少睾丸组织细胞凋亡治疗OAS。

Objective To investigate the therapeutic efficacy and mechanism of the Dioscoreae Rhizoma（DR） and Polygonati Rhizoma（PR） herb pair（DR-PR） in treating oligoasthenozoospermia（OAS）. Methods One week after SD rats were adaptively fed, they were randomly divided into six groups: the control group, the model group, the DR-PR（1∶1） group(2.5 g·kg^-1 DR + 2.5 g·kg^-1 PR), the DR-PR（2∶1） group(5.0 g·kg^-1 DR + 2.5 g·kg^-1 PR), the DR-PR（1∶2） group(2.5 g·kg^-1 DR + 5.0 g·kg^-1 PR), and the L-carnitine(LOS, 100 mg·kg^-1) group. Except for the control group, the other five groups were ig administered with tripterygium glycosides 35 mg·kg^-1 for 7 d to induce the OAS model. Drug administration started from the beginning of modeling, and the rats were fasted at 9:00 every day and ig administered with the drug at 15:00 for a total of 21 d. Semen routine tests were conducted to detect the sperm count and motility of rats; ELISA was used to determine the content of sex hormones in rat serum; HE staining was used to detect the effect of DR-PR on the testicular pathology of OAS rats. Network pharmacology was used to analyze the main active components and mechanism of action of DR-PR in treating OAS: the keywords "Dioscoreae Rhizoma" and "Polygonati Rhizoma" were used to search for the main active components in the TCMSP database, and the OAS targets were collected from the GeneCards database, DrugBank database, and OMIM database. The VLOOCUP function was used to intersect the OAS targets and DR-PR targets to obtain the potential targets of DR-PR in treating OAS. The potential targets were imported into the STRING database for protein-protein interaction（PPI） analysis; The potential target information of DR-PR in treating OAS was imported into the Metascape database for gene ontology（GO） enrichment and Kyoto Encyclopedia of Genes and Genomes（KEGG） enrichment analysis. TUNEL staining was used to determine the apoptosis status of testicular cells in each group of rats; Western blotting was used to detect the expression of heat shock protein 90α type 1（HSP90 AA1）, Caspase-3, Caspase-9, nuclear factor（NF）-κB, phosphorylated NF-κB（p-NF-κB）, p38, and phosphorylated p38（p-p38） in testicular tissues of each group of rats. Molecular docking simulation was used to explore the binding ability and interaction mode of the key active components and key targets of DR-PR. Results Compared with the model group, the sperm concentration, motility, survival rate, straight-line movement rate, curvilinear movement rate, average path velocity, movement straightness, and movement linearity of rats in the DR-PR groups were significantly restored（P ＜ 0.05, 0.01）, the contents of testosterone and estradiol in rat serum were significantly increased（P ＜ 0.01）, and the damage to spermatogenic cells and interstitial cells in the testis was alleviated（P ＜ 0.01）. The DR-PR（1∶2） group had the best effect. The network pharmacology results showed that 16 active components of Dioscoreae Rhizoma and 12 active components of Polygonati Rhizoma were retrieved; the key targets of DR-PR in treating OAS were MAPK14, NFKB1, TNF, IL6, HSP90 AA1, CASP3, and CASP9, the key action pathway was the IL-17 signaling pathway, the key biological function was the response to lipopolysaccharide, the key cellular component was the membrane raft and the vesicle lumen, and the key molecular function was protein serine/threonine kinase activity. Compared with the model group, the apoptosis of testicular tissue cells in the DR-PR groups was significantly reduced（P ＜ 0.01）, the expression of HSP90 AA1, Caspase-3, and Caspase-9 in testicular tissues was down-regulated（P ＜ 0.01）, and the phosphorylation levels of NF-κB and p38 were decreased（P ＜ 0.01）. The DR-PR（1∶2） group had the best effect. The molecular docking results indicated that the key active components of DR-PR and the key targets HSP90 AA1, Caspase-3, Caspase-9, NF-κB and p38 could all form spontaneous binding. Conclusion DR-PR can reduce testicular tissue cell apoptosis and treat OAS by regulating the HSP90/p38/NF-κB pathway.

R285.5

安徽中医药大学科研基金项目（2024LC052）；江苏卫生健康职业学院“医教协同、中西并举”项目（YIXT-HB202409YJXT-HB202401）；安徽省中医药传承创新科研项目（2024CCCX152）；安徽省高等学校科学研究项目（2024AH050990）