目前针对恶性胶质瘤的治疗手段十分有限,本研究旨在通过揭示天然产物银杏叶提取物(Ginkgo biloba extract,GBE)的抗肿瘤机制,为恶性胶质瘤的治疗提供新的潜在治疗方案。研究发现,125μg/mL GBE处理A172胶质瘤细胞可显著抑制其活力(EC50=...目前针对恶性胶质瘤的治疗手段十分有限,本研究旨在通过揭示天然产物银杏叶提取物(Ginkgo biloba extract,GBE)的抗肿瘤机制,为恶性胶质瘤的治疗提供新的潜在治疗方案。研究发现,125μg/mL GBE处理A172胶质瘤细胞可显著抑制其活力(EC50=128.6μg/mL),TUNEL及瑞氏染色均观察到典型的细胞凋亡形态。机制上,GBE处理导致线粒体膜电位显著下降(JC-1法检测)并引发内质网扩张(ER-Tracker观察)。Western blot分析证实,GBE激活了线粒体自噬与内质网应激,表现为LC3-Ⅱ/Ⅰ比值升高以及葡萄糖调节蛋白78(glucose-regulated protein 78,GRP78)和C/EBP同源蛋白(C/EBP homologous protein,CHOP)表达上调。与此同时,免疫荧光与Western blot结果显示,Notch同源物1(neurogenic locus notch homolog protein 1,Notch1)/转化生长因子-β1(transforming growth factor-beta1,TGF-β1)/Smad家族成员2/3(Smad family member 2/3,Smad2/3)通路被激活,且Notch1胞内结构域(Notch1 intracellular domain,NICD)发生核转位。双荧光素酶报告实验进一步揭示,Smad2/3可直接促进LC3和CHOP的转录。关键的是,使用Notch1抑制剂(DAPT)或TGF-β1抑制剂(SB431542)进行干预,可部分逆转由GBE诱导的细胞器应激并提升细胞存活率。综上所述,GBE通过激活Notch1/TGF-β1/Smad2/3通路,破坏细胞器稳态,进而诱导A172胶质瘤细胞凋亡。本研究的目的在于系统阐明GBE通过“信号通路-细胞器”互作以发挥抗癌作用的全新分子机制,其意义不仅为GBE作为潜在的胶质瘤治疗药物提供了坚实的实验证据,更揭示了Notch1/TGF-β1/Smad2/3通路可作为未来开发抗胶质瘤新药的重要靶点。展开更多
目的在“骨肉不相亲”理论指导下,比较废用性骨质疏松症大鼠股骨、腓肠肌中果蝇母本抗生存因子蛋白2/3(small mother against decapentaplegic 2/3,Smad2/3)、特异性蛋白1(specificity protein 1,SP1)表达在中医不同治法下的影响,分析...目的在“骨肉不相亲”理论指导下,比较废用性骨质疏松症大鼠股骨、腓肠肌中果蝇母本抗生存因子蛋白2/3(small mother against decapentaplegic 2/3,Smad2/3)、特异性蛋白1(specificity protein 1,SP1)表达在中医不同治法下的影响,分析中医药对其的防治机制。方法将60只大鼠随机分为正常组、模型组、补肾组、健脾组、补肾健脾组、西药组6组,并适应性喂养1周,造模组采用尾吊法建立模型,5周后取大鼠的腹主动脉血清、股骨及腓肠肌。通过骨密度检测和对股骨远端横断面处的骨形态进行三维重建观察比较各组大鼠股骨组织结构变化,评估骨质流失情况。采用HE染色和Masson染色观察大鼠腓肠肌组织的形态学变化。采用ELISA法检测各组大鼠血清碱性磷酸酶(alkaline phosphatase,ALP)、抗酒石酸酸性磷酸酶(tartrate resistant acid phosphatase,TRACP)水平和股骨、腓肠肌中Smad2/3、SP1蛋白表达。结果与正常组比较,模型组大鼠股骨骨密度显著降低(P<0.01),骨微结构破坏明显;腓肠肌肌纤维萎缩,排列疏松;血清ALP水平显著降低、TRACP水平显著升高(P<0.01);股骨与腓肠肌中Smad2/3、SP1蛋白表达显著升高(P<0.01)。与模型组比较,各给药组股骨骨密度明显升高(P<0.05),骨微结构修复明显;腓肠肌肌纤维增粗,肌间隙减小;ALP水平明显升高(P<0.05,P<0.01)。与模型组比较,补肾健脾组和西药组TRACP水平降低最显著(P<0.01),其次是补肾组(P<0.05);各给药组股骨与腓肠肌中Smad2/3、SP1蛋白表达显著降低(P<0.01)。结论废用性骨质疏松症的发生、发展可能与骨骼及肌肉中Smad2/3、SP1蛋白表达的升高有关,二者的高表达可能会导致骨-肌失衡。各中医治法对废用性骨质疏松症的防治作用可能与降低骨骼与肌肉中Smad2/3、SP1蛋白表达有关。展开更多
Pathological cardiac hypertrophy is an early and significant cardiac structural characteristic that contributes to the onset and progression of heart failure(HF).Its mainly structural feature is the abnormally enlarge...Pathological cardiac hypertrophy is an early and significant cardiac structural characteristic that contributes to the onset and progression of heart failure(HF).Its mainly structural feature is the abnormally enlarged cardiomyocyte.Effective intervention targets for abnormally enlarged cardiomyocyte remain to be identified.Previous studies have shown that the cellular shape and size can be regulated by the actin related protein 2/3(Arp2/3)complex,which is an actin-binding protein complex involved in the actin nucleation and assembly.However,the roles of the Arp2/3 complex in cardiomyocyte hypertrophy remain unknown.Here our study identifies its novel roles in the occurrence and development of cardiomyocyte hypertrophy.We found that mRNA levels of all subunits from the Arp2/3 complex are significantly upregulated(P<0.05)in the angiotensin Ⅱ(Ang Ⅱ)-induced neonatal rat primary and H9c2 cardiomyocyte hypertrophy.Further studies showed that siRNA-directed ARPC 2 silencing inhibits the reactivation of fetal genes and enlargement of cardiomyocyte area induced by Ang Ⅱ in neonatal rat primary cardiomyocytes(NRCMs)and H9c2 cells(P<0.05).In addition,the upstream activators of the Arp2/3 complex including SH3 protein interacting with Nck,90 kD(SPIN90)and Ras-related C3 botulinum toxin substrate 1(Rac1)/WASp family Verprolin-homologous protein-2(WAVE-2)are upregulated(P<0.05)in Ang Ⅱ-induced neonatal rat primary and H9c2 cardiomyocyte hypertrophy,indicating the excessive activation of the Arp2/3 complex.We further show that CK666,a specific Arp2/3 complex inhibitor,prevents the reactivation of fetal genes and the enlargement of cardiomyocyte area induced by Ang Ⅱ in NRCMs and H9c2 cells(P<0.05).Our results reveal that the Arp2/3 complex plays a crucial role in Ang Ⅱ-induced cardiomyocyte hypertrophy,which is beneficial to further studies about the molecular mechanisms by which the Arp2/3 complex regulates pathological cardiac hypertrophy.展开更多
文摘目前针对恶性胶质瘤的治疗手段十分有限,本研究旨在通过揭示天然产物银杏叶提取物(Ginkgo biloba extract,GBE)的抗肿瘤机制,为恶性胶质瘤的治疗提供新的潜在治疗方案。研究发现,125μg/mL GBE处理A172胶质瘤细胞可显著抑制其活力(EC50=128.6μg/mL),TUNEL及瑞氏染色均观察到典型的细胞凋亡形态。机制上,GBE处理导致线粒体膜电位显著下降(JC-1法检测)并引发内质网扩张(ER-Tracker观察)。Western blot分析证实,GBE激活了线粒体自噬与内质网应激,表现为LC3-Ⅱ/Ⅰ比值升高以及葡萄糖调节蛋白78(glucose-regulated protein 78,GRP78)和C/EBP同源蛋白(C/EBP homologous protein,CHOP)表达上调。与此同时,免疫荧光与Western blot结果显示,Notch同源物1(neurogenic locus notch homolog protein 1,Notch1)/转化生长因子-β1(transforming growth factor-beta1,TGF-β1)/Smad家族成员2/3(Smad family member 2/3,Smad2/3)通路被激活,且Notch1胞内结构域(Notch1 intracellular domain,NICD)发生核转位。双荧光素酶报告实验进一步揭示,Smad2/3可直接促进LC3和CHOP的转录。关键的是,使用Notch1抑制剂(DAPT)或TGF-β1抑制剂(SB431542)进行干预,可部分逆转由GBE诱导的细胞器应激并提升细胞存活率。综上所述,GBE通过激活Notch1/TGF-β1/Smad2/3通路,破坏细胞器稳态,进而诱导A172胶质瘤细胞凋亡。本研究的目的在于系统阐明GBE通过“信号通路-细胞器”互作以发挥抗癌作用的全新分子机制,其意义不仅为GBE作为潜在的胶质瘤治疗药物提供了坚实的实验证据,更揭示了Notch1/TGF-β1/Smad2/3通路可作为未来开发抗胶质瘤新药的重要靶点。
文摘Pathological cardiac hypertrophy is an early and significant cardiac structural characteristic that contributes to the onset and progression of heart failure(HF).Its mainly structural feature is the abnormally enlarged cardiomyocyte.Effective intervention targets for abnormally enlarged cardiomyocyte remain to be identified.Previous studies have shown that the cellular shape and size can be regulated by the actin related protein 2/3(Arp2/3)complex,which is an actin-binding protein complex involved in the actin nucleation and assembly.However,the roles of the Arp2/3 complex in cardiomyocyte hypertrophy remain unknown.Here our study identifies its novel roles in the occurrence and development of cardiomyocyte hypertrophy.We found that mRNA levels of all subunits from the Arp2/3 complex are significantly upregulated(P<0.05)in the angiotensin Ⅱ(Ang Ⅱ)-induced neonatal rat primary and H9c2 cardiomyocyte hypertrophy.Further studies showed that siRNA-directed ARPC 2 silencing inhibits the reactivation of fetal genes and enlargement of cardiomyocyte area induced by Ang Ⅱ in neonatal rat primary cardiomyocytes(NRCMs)and H9c2 cells(P<0.05).In addition,the upstream activators of the Arp2/3 complex including SH3 protein interacting with Nck,90 kD(SPIN90)and Ras-related C3 botulinum toxin substrate 1(Rac1)/WASp family Verprolin-homologous protein-2(WAVE-2)are upregulated(P<0.05)in Ang Ⅱ-induced neonatal rat primary and H9c2 cardiomyocyte hypertrophy,indicating the excessive activation of the Arp2/3 complex.We further show that CK666,a specific Arp2/3 complex inhibitor,prevents the reactivation of fetal genes and the enlargement of cardiomyocyte area induced by Ang Ⅱ in NRCMs and H9c2 cells(P<0.05).Our results reveal that the Arp2/3 complex plays a crucial role in Ang Ⅱ-induced cardiomyocyte hypertrophy,which is beneficial to further studies about the molecular mechanisms by which the Arp2/3 complex regulates pathological cardiac hypertrophy.
