根皮素通过抑制PDK1-p-PDHA1轴影响谷氨酰胺代谢介导的前列腺癌研究  

Phloretin influences glutaminolysis-mediated prostate cancer by inhibiting PDK1-p-PDHA1 axis

作　　者：赵朋[1] 杨拓[1] 王金铸 蔡科科[1] 刘鹏[1] 念学武 ZHAO Peng;YANG Tuo;WANG Jinzhu;CAI Keke;LIU Peng;NIAN Xuewu(Department of Urology Surgery,Tianjin Nankai hospital,Tianjin 300000,China;Department of Urology Surgery,Tianjin Beichen Hospital,Tianjin 300400,China)

机构地区：[1]天津市南开医院泌尿外科,天津300000 [2]天津市北辰医院泌尿外科,天津300400

出　　处：《中草药》2025年第4期1254-1265,共12页Chinese Traditional and Herbal Drugs

基　　金：天津市科技计划项目(20JCQNJC00550)。

摘　　要：目的探究根皮素通过抑制丙酮酸脱氢酶激酶1(pyruvate dehydrogenase kinase 1,PDK1)-磷酸化丙酮酸脱氢酶E1亚基α1(phosphorylation of pyruvate dehydrogenase E1 subunit alpha 1,p-PDHA1)进而影响谷氨酰胺(glutamine,Gln)代谢介导的前列腺癌的作用机制。方法采用不同剂量根皮素(25、50、100μmol/L)干预人前列腺癌PC-3、DU145、LNCaP细胞与人前列腺RWPE-1细胞,采用细胞计数试剂盒(cell counting kit-8,CCK-8)测定细胞活力。将人前列腺癌PC-3细胞分为对照(二甲基亚砜,dimethyl sulfoxide,DMSO)组、根皮素(100μmol/L)组及顺铂(0.03 mmol/L)组,Transwell法检测细胞侵袭能力、TUNEL法检测细胞凋亡水平。根皮素及Gln单独使用及联合干预PC-3细胞,试剂盒测定Gln消耗水平、谷氨酸与腺嘌呤核苷三磷酸(adenosine triphosphate,ATP)产生水平,Western blotting法测定谷氨酰胺酶1(glutaminase 1,GLS1)蛋白表达水平,同时测定细胞增殖、侵袭、凋亡等细胞生物学行为变化。利用网络药理学及生物信息学分析根皮素、前列腺癌与Gln代谢相关基因的交集。Western blotting法测定各组细胞PDK1蛋白表达水平。将PC-3细胞分为空载体对照(pcDNA3.1)组、PDK1过表达载体(pcDNA3.1-PDK1)组、PDK1敲减载体(KD-PDK1)组及其对照(KD-Control)组、PDHA1过表达载体(pcDNA3.1-PDHA1)组、KD-PDK1+pcDNA3.1-PDHA1组及其对照KD-PDK1+pcDNA3.1组,以及根皮素(100μmol/L)+pcDNA3.1-PDK1组及其对照根皮素+pcDNA3.1组,测定各组细胞增殖、侵袭、凋亡变化、Gln消耗水平、谷氨酸与ATP产生水平及GLS1蛋白表达水平。构建前列腺癌移植瘤小鼠模型,通过根皮素干预治疗,以顺铂作为阳性对照,探究根皮素对体内肿瘤生长的影响。结果根皮素对人前列腺RWPE-1细胞活力无显著影响,但100μmol/L根皮素可显著抑制人前列腺癌PC-3、DU145、LNCaP细胞增殖(P<0.05)。与对照组比较,根皮素组细胞增殖与侵袭能力显著降低(P<0.05)、凋亡水平显著增加(Objective To explore whether phloretin can affect the biological behavior of prostate cancer cells mediated by glutaminolysis by inhibiting pyruvate dehydrogenase kinase 1(PDK1)-phosphorylation of pyruvate dehydrogenase E1 subunit alpha 1(p-PDHA1).Methods Human prostate cancer cell lines PC-3,DU145,LNCaP,and human prostate epithelial cell line RWPE-1 were treated with different concentrations of phloretin(25,50,100μmol/L),and cell proliferation was assessed using the cell counting kit-8(CCK-8).PC-3 cells were divided into control(0.1%DMSO),phloretin(100μmol/L),and cisplatin(DDP,0.03 mmol/L)groups,and cell invasion was detected using the Transwell method while cell apoptosis was detected using the TUNEL method.PC-3 cells were treated with phloretin and/or glutamine(Gln)separately or in combination,and Gln levels,glutamate and adenosine triphosphate(ATP)production levels were measured using kits,while glutaminase 1(GLS1)protein expression was determined by Western blotting in cell proliferation,invasion,and apoptosis were also assessed.Network pharmacology and bioinformatics were used to analyze the intersection of phloretin,prostate cancer,and Gln-related genes.Western blotting was used to measure PDK1 protein expression levels in RWPE-1,PC-3,and PC-3+phloretin groups.Furthermore,The PC-3 cells were divided into control pcDNA3.1 group,pcDNA3.1-PDK1 group,the KD-PDK1 group and its control KD-Control group,pcDNA3.1-PDHA1 group,KD-PDK1+pcDNA3.1-PDHA1 group and its control KD-PDK1+pcDNA3.1 group,and phloretin+pcDNA3.1-PDK1 group and its control phloretin+pcDNA3.1 group,Changes in cell proliferation,invasion,apoptosis,Gln levels,glutamate and ATP production levels,and GLS1 protein expression in each group were measured.A prostate cancer xenograft mouse model was established,and phloretin intervention therapy was administered,with DDP treatment serving as a positive control,to investigate the effect of phloretin on tumor growth in vivo.Results Phloretin had no significant effect on the proliferation of RWPE-1 cells bu

关 键 词：根皮素 前列腺癌 丙酮酸脱氢酶激酶1 丙酮酸脱氢酶E1亚基α1 谷氨酰胺代谢 

分 类 号：R285.5[医药卫生—中药学]