Objectives: To investigate the mechanism through which salidroside inhibits proliferation of gastric cancer (GC) cells focusing on glucose metabolic reprogramming pathways.
Methods: High-throughput sequencing combined with bioinformatics analysis was employed to identify the potential targets of salidroside in human GC MGC-803 cells. Liposome-mediated transfection experiments were carried out to validate the functional and mechanistic roles of these targets. CCK-8 and colony formation assays were used to assess the effects of salidroside on GC cell viability and clonogenic ability. qRT-PCR, Western blotting, and biochemical assay kits were used to analyze the regulatory effects of salidroside on the miR-1343-3p-OGDHL/PDHB enzyme complex-pyruvate metabolic pathway in GC cells.
Results: Bioinformatics analysis suggested that the tumor-suppressive factor miR-1343-3p negatively regulated the key glycolytic enzyme gene oxoglutarate dehydrogenase-like (OGDHL) in GC cells, and OGDHL and pyruvate dehydrogenase E1 subunit beta (PDHB) were both significantly upregulated in GC tissues, which was close by correlated with reduced survival rates of GC patients. In MGC-803 cells, salidroside treatment significantly enhanced the expression level of miR-1343-3p and downregulated OGDHL expression, resulting in disruption of the stability of PDHB, reduced pyruvate oxidative decarboxylation, and consequently decreased production of acetyl-CoA and ATP.
Conclusions: Salidroside inhibits GC cell proliferation possibly by regulating the miR-1343-3p-OGDHL/PDHB enzyme complex-pyruvate metabolic pathway, which provides new insights into its anti-tumor mechanisms and suggests new strategies for targeted therapy for GC.
目的: 探讨红景天苷抗胃肿瘤（GC）机制，强调糖代谢重编程途径。方法: 高通量测序结合生物信息学分析预测红景天苷作用后显著差异表达的miRNA-靶mRNA，mRNA-mRNA相互作用关系，RNA结合蛋白免疫共沉淀实验验证miRNA-mRNA互作，蛋白质免疫共沉淀（Co-IP）实验验证下游靶mRNA-mRNA互作。通过体外及体内实验探究红景天苷与miR-1343-3p对GC的作用机制。体外实验通过CCK-8法确定红景天苷的IC50值并分低、中、高剂量处理细胞，结合克隆实验、RT-qPCR、Western blotting、ELISA及ATP检测评估药物对增殖、基因/蛋白/下游分子表达（miRNA、mRNA、丙酮酸、乙酰CoA）及能量代谢的影响；进一步以脂质体法转染miR-1343-3p模拟物（miR-1343-3p mimic）、抑制剂（miR-1343-3p inhibitor）、靶向OGDHL的siRNA（si-OGDHL）及其阴性对照（NC mimic/NC inhibitor/si-NC）至GC细胞，联合红景天苷处理，通过相同方法验证其协同作用。体内实验通过构建荷瘤裸鼠模型，给予红景天苷和miR-1343-3p激动剂（miR-1343-3p agomir）干预，分析瘤块体积/质量变化及靶基因、蛋白表达与代谢状态。实验分组包括空白对照、单药组及联合组，系统解析红景天苷与miRNA的调控网络。结果: 生物信息学证实，miR-1343-3p与三羧酸（TCA）循环关键酶α-酮戊二酸脱氢酶复合体亚基（OGDHL）呈负相关（P<0.01），且直接相互作用。OGDHL与丙酮酸脱氢酶E1亚基-β（PDHB）在胃癌组织中同步高表达，且二者存在相互作用。体外与体内实验证实，红景天苷以剂量/时间依赖性抑制GC细胞增殖，红景天苷作用GC细胞后，重要抑癌因子miR-1343-3p升高，受其调控的OGDHL的基因表达明显下调，导致与之互作的糖酵解酶蛋白PDHB稳态失衡，丙酮酸氧化脱羧减少，产物乙酰CoA和ATP生成明显减少（P<0.05）。结论: 红景天苷可能通过调控miR-1343-3p-OGDHL/PDHB酶复合体-丙酮酸糖代谢通路抑制GC细胞的增殖，为红景天苷抗肿瘤机制提供了新的见解，为开发基于代谢酶复合体协同调控癌症的精准疗法提供理论依据。.
Keywords: gastric cancer; glucose metabolic reprogramming; miR-1343-3p; oxoglutarate dehydrogenase-like; pyruvate dehydrogenase E1 subunit beta; salidroside.