目的 探讨党参含药血清（Codonopsis Radix containing serum，CRCS）对溃疡性结肠炎（ulcerative colitis，UC）体外模型炎症因子释放及上皮细胞过度凋亡的作用机制。方法 借助生物信息学分析工具预测调控磷酸酶和张力蛋白同源物（phosphatase and tensin homolog，PTEN）上游miRNA，采用qRT-PCR和Western blotting检测5% 2,4,6-三硝基苯磺酸（2,4,6-trinitrobenzenesulfonic acid solution，TNBS）/乙醇复合溶液灌肠诱导的UC大鼠模型结肠组织中PTEN mRNA和蛋白表达水平，以及miR-21、miR-30a-3p、miR-198、miR-152-3p、miR-301a-3p、miR-320、miR-3691-5p表达变化。10 μg/mL脂多糖（lipopolysaccharide，LPS）诱导人正常结肠上皮NCM460细胞构建UC体外模型，给予1%、2%、4%、6%、8% CRCS干预24 h，利用CCK-8法检测细胞活力。分别转染miR-21模拟剂（miRNA-21 mimics、miR-21 mimics）与miR-21抑制剂（miRNA-21 inhibitor、miR-21 inhibitor）至模型细胞，给予6% CRCS完全培养基培养24 h，采用ELISA检测各组白细胞介素-6（interleukin-6，IL-6）、IL-17、IL-1β、IL-8、核因子-κB（nuclear factor-κB，NF-κB）和肿瘤坏死因子-α（tumor necrosis factor-α，TNF-α）水平；qRT-PCR检测各组miR-21表达；Western blotting检测PTEN、B细胞淋巴瘤-2（B-cell lymphoma-2，Bcl-2）、Bcl-2相关X蛋白（Bcl-2 associated X protein，Bax）、剪切型半胱氨酸天冬氨酸蛋白酶-3（cleaved cystein-asparate protease-3，cleaved Caspase-3）和磷脂酰肌醇3-激酶（phosphatidylinositol 3-kinase，PI3K）/蛋白激酶B（protein kinase B，Akt）通路相关蛋白的表达。通过生物信息学预测miR-21与PTEN结合位点，并采用双荧光素酶报告基因检测系统验证CRCS干预后miR-21对PTEN的靶向作用。结果 生物信息学共筛选出7个可能靶向PTEN的miRNA，其中党参干预后miR-21表达水平显著降低（P＜0.01），而miR-30a-3p、miR-198、miR-152-3p、miR-301a-3p、miR-320及miR-3691-5p表达变化均无统计学意义。CCK-8结果表明，2%、4%、6%、8% CRCS均可显著提高细胞活力（P＜0.01）。与模型组比较，CRCS和miR-21 inhibitor可显著上调细胞内PTEN和Bcl-2蛋白表达水平（P＜0.01），并降低miR-21、Bax、cleaved Caspase-3、p-PI3K/PI3K、p-Akt/Akt表达水平（P＜0.01）；与CRCS组比较，miR-21 mimics可显著削弱CRCS对PTEN和Bcl-2表达的上调作用（P＜0.01），以及对miR-21、Bax、cleaved Caspase-3、p-PI3K/PI3K、p-Akt/Akt表达的下调作用（P＜0.05、0.01）。双荧光素酶报告实验结果显示，在转染wt-Luc-PTEN质粒细胞中，CRCS干预后可显著提高荧光素酶活性（P＜0.01），miR-21 mimics可显著降低荧光素酶活性（P＜0.05），联合处理时，miR-21 mimics可部分削弱CRCS对荧光素酶活性升高的作用（P＜0.01）；在转染mut-Luc-PTEN质粒细胞中，CRCS干预及miR-21 mimics均未引起荧光素酶活性显著变化。结论 CRCS可通过抑制促炎因子释放及上皮细胞过度凋亡缓解UC，其机制至少部分依赖于miR-21/PTEN/PI3K/Akt轴。

Objective To investigate the mechanism of Dangshen (Codonopsis Radix) containing serum (CRCS) on inflammatory factors release and epithelial cell excessive apoptosis in vitro model of ulcerative colitis (UC). Methods Bioinformatics analysis tools were used to predict and regulate the upstream miRNAs of phosphatase and tensin homolog (PTEN). qRT-PCR and Western blotting were used to detect the mRNA and protein expression levels of PTEN and the expressions of miR-21, miR-30a-3p, miR-198, miR-152-3p, miR-301a-3p, miR-320 and miR-3691-5p in colon tissues of UC rats model induced by enema 5% 2,4,6-trinitrobenzenesulfonic acid solution (TNBS)/ethanol composite solution. Lipopolysaccharide (LPS, 10 μg/mL) was used to induce human normal colon epithelial NCM460 cells to construct an in vitro model of UC, 1%, 2%, 4%, 6% and 8% CRCS intervention was administered for 24 h. The cell viability was detected by CCK-8 method. Subsequently, miR-21 mimics and miR-21 inhibitor were transfected into model cells, respectively, which were cultured in 6 % CRCS complete medium for 24 h. The levels of inflammatory factors interleukin-6 (IL-6), IL-17, IL-1β, IL-8, nuclear factor-κB (NF-κB) and tumor necrosis factor-α (TNF-α) in each group were detected by ELISA. The expression of miR-21 in each group was detected by qRT-PCR. Western blotting was used to detect the expressions of PTEN, B-cell lymphoma-2 (Bcl-2), Bcl-2 associated X protein (Bax), cleaved cystein-asparate protease-3 (cleaved Caspase-3) and phosphoinositide 3-kinase (PI3K)/protein kinase B (Akt) pathway related proteins. The binding sites of miR-21 and PTEN were predicted by bioinformatics, and the targeting effect of miR-21 on PTEN was verified by dual luciferase reporter gene detection system. Results A total of seven miRNAs that may target PTEN were screened by bioinformatics analysis. Among them, the expression of miR-21 was decreased significantly after CRCS intervention (P < 0.01), while the expressions of miR-30a-3p, miR-198, miR-152-3p, miR-301a-3p, miR-320 and miR-3691-5p had no statistical significance. The results of CCK-8 showed that 2%, 4%, 6% and 8% CRCS could significantly improve cell viability (P < 0.01). Compared with model group, CRCS and miR-21 inhibitor increased the expression levels of PTEN and Bcl-2 proteins in cells (P < 0.01), and decreased the expression levels of miR-21, Bax, cleaved Caspase-3, p-PI3K/PI3K and p-Akt/Akt (P < 0.01). Compared with CRCS group, miR-21 mimics significantly attenuated the up-regulation of PTEN and Bcl-2 expressions by CRCS (P < 0.01) and the down-regulation of miR-21, Bax, cleaved Caspase-3, p-PI3K/PI3K, p-Akt/Akt expressions (P < 0.05, 0.01). The results of dual luciferase reporter assay showed that in the transfected wt-Luc-PTEN plasmocytes, CRCS intervention could significantly increase luciferase activity (P < 0.01), and miR-21 mimics could significantly reduce luciferase activity (P < 0.01). In the combined treatment, miR-21 mimics could partially weaken the effect of CRCS on luciferase activity (P < 0.01). In transfected mut-Luc-PTEN plasmocytes, CRCS intervention and miR-21 mimics did not cause significant changes in luciferase activity. Conclusion CRCS could alleviate UC by inhibiting pro-inflammatory factors release and excessive apoptosis of epithelial cells. The mechanism is at least partially dependent on miR-21/PTEN/PI3K/Akt axis.

R285.5

科技部国家重点研发计划项目（2018YFC1706305）；甘肃省重点研发计划-社会发展类（23YFFA0069）；甘肃省中医药研究中心开放课题（zyzx-2023-10）；西北营养与环境相关疾病中医药防控协同创新中心2025年度开放基金课题（ZYXT-25-04）；兰州市科技计划项目（2024-9-78）；甘肃省优秀研究生“创新之星”项目（2026CXZX-924）