Background:The purpose of this research is to predict the mechanisms of the experienced prescription Jiangtang decoction for treating diabetic nephropathy based on network pharmacology,the predicted targets and pathwa...Background:The purpose of this research is to predict the mechanisms of the experienced prescription Jiangtang decoction for treating diabetic nephropathy based on network pharmacology,the predicted targets and pathways were validated by celluar experiments.Methods:The active ingredients of the experienced prescription Jiangtang decoction and their putative targets were collected from TCMSP Database and SwissTargetPrediction platform.Diabetic nephropathy-related targets were excavated from GeneCards and DisGeNET database.Then,the interactions of potential targets of the experienced prescription Jiangtang decoction with well-known diabetic nephropathy targets were used to construct the protein-protein interaction network by STRING database and Cytoscape,and screened the core targets via topological analysis.Gene Ontology and Kyoto Encyclopedia of Genes and Genomes(KEGG)enrichment analysis were performed by Metascape platform.Finally,we conducted in vitro experiments to verify this prediction of the network pharmacology.A diabetic nephropathy model was established by treating mesangial cells with D-ribose,in which the therapeutic effects of the experienced prescription Jiangtang decoction were evaluated.CCK-8 and LDH assay were used to test cell viability and cell toxicity,cell apoptosis was evaluated by Hoechst 33258 staining,AO/EB staining,levels of ROS were detected by fluorescent probe,the expression levels of MAPK signaling pathway-associated proteins and apoptosis-related proteins Bax were measured by western blotting assay.Results:A total of 74 active ingredients contained and 871 potential identified targets were retrieved from databases.Simultaneously,803 diabetic nephropathy-associated targets were also obtained,180 overlapped targets were considered as potential therapeutic targets of the experienced prescription Jiangtang decoction against diabetic nephropathy.By constructing a protein-protein interaction network and topological analysis,57 core targets were screened.Gene Ontology analysis highlighted 1655 Gene Ontology terms main including apoptotic signaling pathway,regulation of reactive oxygen species metabolic process and positive regulation of cell migration.KEGG enrichment analysis revealed that core targets were enriched mainly in MAPK,AGE-RAGE,TNF,PI3K-Akt signaling pathways.Cellular experiments further demonstrated D-ribose decreased mesangial cells viability,increased LDH release and the ROS level,induced apoptosis and activated the p38/JNK MAPK signal pathways,while the experienced prescription Jiangtang decoction could be useful in attenuation of D-ribose-induced damage.Conclusion:Network pharmacology intuitively shows the multi-component,multi-target and multi-pathway therapeutic effects of the experienced prescription Jiangtang decoction on diabetic nephropathy.By in vitro experiment,it revealed that the experienced prescription Jiangtang decoction has potential therapeutic effects on diabetic nephropathy via alleviating oxidative stress and apoptosis.the experienced prescription Jiangtang decoction treatment significantly inhibited the D-ribose-stimulated JNK and p38 MAPK activation.展开更多
Background:To investigate the pharmacological effects of Fangshi Jiangtang decoction(FSJT)on type 2 diabetes mellitus(T2DM)model rats and explore its mechanism of action from the perspective of mitochondrial autophagy...Background:To investigate the pharmacological effects of Fangshi Jiangtang decoction(FSJT)on type 2 diabetes mellitus(T2DM)model rats and explore its mechanism of action from the perspective of mitochondrial autophagy.Methods:Sixty Sprague Dawley rats were randomly divided into six groups after one week of adaptive feeding:Control group,T2DM model group,metformin group(0.2 g/kg by gavage),and FSJT low,medium,and high dose groups(9.5,19,38 g/kg by gavage).Except for the Control group,the other five groups were given a high-fat diet.The treatment lasted for 8 weeks,and blood glucose levels were measured weekly.Eight weeks later,blood samples were collected from the rats,and serum was separated for the determination of HbA1c,oral glucose tolerance test,and homeostatic model assessment for insulin resistance index.The pancreas of the rats was collected,weighed,and fixed.The same part of the pancreas was used for hematoxylin-eosin.Kits were used to detect triglycerides,total cholesterol,interleukin-1β,interleukin-6,tumor necrosis factor-α,malondialdehyde,glutathione peroxidase,and superoxide dismutase in pancreatic tissue to assess the effects of FSJT on inflammation and oxidative stress in T2DM rats.Western blot analysis was performed to detect the expression of VDAC1,TOM20,COXⅣ,PINK1,Parkin,beclin1,light chain 3,and selective autophagy adaptor protein P62 to evaluate the effects of FSJT on mitochondrial autophagy in T2DM model rats.Results:Compared with the T2DM model group,FSJT intervention significantly reduced blood glucose,HbA1c,oral glucose tolerance test,and homeostatic model assessment for insulin resistance index in T2DM model rats,alleviated pancreatic tissue lesions,reduced levels of total cholesterol,triglycerides,interleukin-1β,interleukin-6,tumor necrosis factor-α,and malondialdehyde,increased glutathione peroxidase and superoxide dismutase activities,downregulated the expression of VDAC1,TOM20,COXⅣ,and P62 proteins,and upregulated the expression of PINK1,Parkin,Beclin1,and light chain 3 proteins.Conclusion:FSJT can improve insulin resistance in T2DM by promoting the activation of mitochondrial autophagy.展开更多
目的探讨在高糖作用下,体外培养的小鼠足细胞凋亡及内质网应激凋亡因子Caspase-12 m RNA和蛋白表达水平的变化及左归降糖益肾方含药血浆的调控作用。方法将体外培养的小鼠足细胞随机分为空白组(A组,25 mmol/L葡萄糖,10%空白血浆)、高糖...目的探讨在高糖作用下,体外培养的小鼠足细胞凋亡及内质网应激凋亡因子Caspase-12 m RNA和蛋白表达水平的变化及左归降糖益肾方含药血浆的调控作用。方法将体外培养的小鼠足细胞随机分为空白组(A组,25 mmol/L葡萄糖,10%空白血浆)、高糖组(B组,200 mmol/L葡萄糖,10%空白血浆)、左归降糖益肾方含药血浆组(C组,200 mmol/L葡萄糖,10%含药血浆),4-苯基丁酸含药血浆组(D组,200 mmol/L葡萄糖,10%含药血浆)。培养48 h后,采用免疫荧光细胞化学法、流式细胞术检测足细胞凋亡情况;Western blot法、RT-PCR法分别检测Caspase-12蛋白或m RNA表达水平的变化。结果与A组比较:在高糖状态下B组足细胞凋亡率、Caspase-12 m RNA和蛋白的表达水平均升高,差异有统计学意义(P<0.05)。与B组比较:C组、D组足细胞凋亡率,Caspase-12 m RNA和蛋白的表达水平均降低,差异均有统计学意义(P<0.05)。与C组比较:D组足细胞凋亡率降低,差异有统计学意义(P<0.05);但Caspase-12 m RNA和蛋白的表达水平,差异均无统计学意义(P>0.05)。结论 Caspase-12表达上调,是足细胞凋亡的分子机制之一;10%左归降糖益肾方含药血浆可能通过降低Caspase-12的表达水平,从而保护足细胞。展开更多
目的:探讨活血降糖饮治疗糖尿病肾病的分子机制。方法:通过高脂饲料喂养加尾静脉注射链脉佐菌素(STZ)等建立糖尿病肾病大鼠模型,并将模型大鼠随机分为糖尿病肾病(DN)组、模型+双胍+中药组(DN+Met+ZY)、模型+双胍+卡托普利组(DN+Met+CP)...目的:探讨活血降糖饮治疗糖尿病肾病的分子机制。方法:通过高脂饲料喂养加尾静脉注射链脉佐菌素(STZ)等建立糖尿病肾病大鼠模型,并将模型大鼠随机分为糖尿病肾病(DN)组、模型+双胍+中药组(DN+Met+ZY)、模型+双胍+卡托普利组(DN+Met+CP),给予相应药物进行灌胃治疗,并另设正常对照组,每组10只,干预16周后,观察各组大鼠空腹血糖(FBG)、血脂、血清尿素氮(BUN)、血肌酐(Scr)、24 h尿总蛋白(24 h UTP)及尿微量清蛋白(24 h mALB)的变化。观察肾组织病理改变,并对各组肾组织进行RNA-seq测序。结果:与模型组比较,中药组和卡托普利组可以显著下调糖尿病肾病大鼠FBG、TC、TG、LDL-C、Scr、BUN、24 h UTP、24 h mALB等水平,而HDL-C无明显变化;与卡托普利组之间比较,中药组FBG、TC、TG、LDL-C明显下降,HDL-c、Scr、BUN、24 h UTP、24 h mALB均无明显差别。与模型组比较,中药组和卡托普利组均可以显著改善糖尿病肾病大鼠的肾脏病理学变化。转录组测序结果显示模型组对比正常组与中药组对比模型组转录本中基因总量接近,其中表达差异最明显的双向调控基因共筛选出10个;京都基因与基因组百科全书(KEGG)前20条显著富集通路分析显示模型组对比正常组与中药组对比模型组共同作用的信号通路有代谢相关通路及cAMP信号通路。结论:活血降糖饮联合二甲双胍能改善糖尿病肾病大鼠的糖脂代谢紊乱,减少尿白蛋白的排泄,改善肾功能和肾脏病理改变,延缓糖尿病肾病病情进展,其作用机制可能是通过Cftr、Pdk4、Angptl4、Hmgcs2等多靶点介导多通路实现的。展开更多
基金This study was supported by the National Natural Science Foundation of China(grants 81573763)Beijing Municipal Natural Science Foundation(grants 7172221)Beijing Traditional Chinese Medicine Science and Technology Development Project(JJ-2020-03).
文摘Background:The purpose of this research is to predict the mechanisms of the experienced prescription Jiangtang decoction for treating diabetic nephropathy based on network pharmacology,the predicted targets and pathways were validated by celluar experiments.Methods:The active ingredients of the experienced prescription Jiangtang decoction and their putative targets were collected from TCMSP Database and SwissTargetPrediction platform.Diabetic nephropathy-related targets were excavated from GeneCards and DisGeNET database.Then,the interactions of potential targets of the experienced prescription Jiangtang decoction with well-known diabetic nephropathy targets were used to construct the protein-protein interaction network by STRING database and Cytoscape,and screened the core targets via topological analysis.Gene Ontology and Kyoto Encyclopedia of Genes and Genomes(KEGG)enrichment analysis were performed by Metascape platform.Finally,we conducted in vitro experiments to verify this prediction of the network pharmacology.A diabetic nephropathy model was established by treating mesangial cells with D-ribose,in which the therapeutic effects of the experienced prescription Jiangtang decoction were evaluated.CCK-8 and LDH assay were used to test cell viability and cell toxicity,cell apoptosis was evaluated by Hoechst 33258 staining,AO/EB staining,levels of ROS were detected by fluorescent probe,the expression levels of MAPK signaling pathway-associated proteins and apoptosis-related proteins Bax were measured by western blotting assay.Results:A total of 74 active ingredients contained and 871 potential identified targets were retrieved from databases.Simultaneously,803 diabetic nephropathy-associated targets were also obtained,180 overlapped targets were considered as potential therapeutic targets of the experienced prescription Jiangtang decoction against diabetic nephropathy.By constructing a protein-protein interaction network and topological analysis,57 core targets were screened.Gene Ontology analysis highlighted 1655 Gene Ontology terms main including apoptotic signaling pathway,regulation of reactive oxygen species metabolic process and positive regulation of cell migration.KEGG enrichment analysis revealed that core targets were enriched mainly in MAPK,AGE-RAGE,TNF,PI3K-Akt signaling pathways.Cellular experiments further demonstrated D-ribose decreased mesangial cells viability,increased LDH release and the ROS level,induced apoptosis and activated the p38/JNK MAPK signal pathways,while the experienced prescription Jiangtang decoction could be useful in attenuation of D-ribose-induced damage.Conclusion:Network pharmacology intuitively shows the multi-component,multi-target and multi-pathway therapeutic effects of the experienced prescription Jiangtang decoction on diabetic nephropathy.By in vitro experiment,it revealed that the experienced prescription Jiangtang decoction has potential therapeutic effects on diabetic nephropathy via alleviating oxidative stress and apoptosis.the experienced prescription Jiangtang decoction treatment significantly inhibited the D-ribose-stimulated JNK and p38 MAPK activation.
基金supported by Applied Basic Research Project of Jiaxing Science and Technology Bureau(2023AD11047)the Joint Project of Yunnan Provincial Science and Technology Department and Yunnan University of Chinese Medicine(202101AZ070001-064).
文摘Background:To investigate the pharmacological effects of Fangshi Jiangtang decoction(FSJT)on type 2 diabetes mellitus(T2DM)model rats and explore its mechanism of action from the perspective of mitochondrial autophagy.Methods:Sixty Sprague Dawley rats were randomly divided into six groups after one week of adaptive feeding:Control group,T2DM model group,metformin group(0.2 g/kg by gavage),and FSJT low,medium,and high dose groups(9.5,19,38 g/kg by gavage).Except for the Control group,the other five groups were given a high-fat diet.The treatment lasted for 8 weeks,and blood glucose levels were measured weekly.Eight weeks later,blood samples were collected from the rats,and serum was separated for the determination of HbA1c,oral glucose tolerance test,and homeostatic model assessment for insulin resistance index.The pancreas of the rats was collected,weighed,and fixed.The same part of the pancreas was used for hematoxylin-eosin.Kits were used to detect triglycerides,total cholesterol,interleukin-1β,interleukin-6,tumor necrosis factor-α,malondialdehyde,glutathione peroxidase,and superoxide dismutase in pancreatic tissue to assess the effects of FSJT on inflammation and oxidative stress in T2DM rats.Western blot analysis was performed to detect the expression of VDAC1,TOM20,COXⅣ,PINK1,Parkin,beclin1,light chain 3,and selective autophagy adaptor protein P62 to evaluate the effects of FSJT on mitochondrial autophagy in T2DM model rats.Results:Compared with the T2DM model group,FSJT intervention significantly reduced blood glucose,HbA1c,oral glucose tolerance test,and homeostatic model assessment for insulin resistance index in T2DM model rats,alleviated pancreatic tissue lesions,reduced levels of total cholesterol,triglycerides,interleukin-1β,interleukin-6,tumor necrosis factor-α,and malondialdehyde,increased glutathione peroxidase and superoxide dismutase activities,downregulated the expression of VDAC1,TOM20,COXⅣ,and P62 proteins,and upregulated the expression of PINK1,Parkin,Beclin1,and light chain 3 proteins.Conclusion:FSJT can improve insulin resistance in T2DM by promoting the activation of mitochondrial autophagy.
文摘目的探讨在高糖作用下,体外培养的小鼠足细胞凋亡及内质网应激凋亡因子Caspase-12 m RNA和蛋白表达水平的变化及左归降糖益肾方含药血浆的调控作用。方法将体外培养的小鼠足细胞随机分为空白组(A组,25 mmol/L葡萄糖,10%空白血浆)、高糖组(B组,200 mmol/L葡萄糖,10%空白血浆)、左归降糖益肾方含药血浆组(C组,200 mmol/L葡萄糖,10%含药血浆),4-苯基丁酸含药血浆组(D组,200 mmol/L葡萄糖,10%含药血浆)。培养48 h后,采用免疫荧光细胞化学法、流式细胞术检测足细胞凋亡情况;Western blot法、RT-PCR法分别检测Caspase-12蛋白或m RNA表达水平的变化。结果与A组比较:在高糖状态下B组足细胞凋亡率、Caspase-12 m RNA和蛋白的表达水平均升高,差异有统计学意义(P<0.05)。与B组比较:C组、D组足细胞凋亡率,Caspase-12 m RNA和蛋白的表达水平均降低,差异均有统计学意义(P<0.05)。与C组比较:D组足细胞凋亡率降低,差异有统计学意义(P<0.05);但Caspase-12 m RNA和蛋白的表达水平,差异均无统计学意义(P>0.05)。结论 Caspase-12表达上调,是足细胞凋亡的分子机制之一;10%左归降糖益肾方含药血浆可能通过降低Caspase-12的表达水平,从而保护足细胞。
文摘目的:探讨活血降糖饮治疗糖尿病肾病的分子机制。方法:通过高脂饲料喂养加尾静脉注射链脉佐菌素(STZ)等建立糖尿病肾病大鼠模型,并将模型大鼠随机分为糖尿病肾病(DN)组、模型+双胍+中药组(DN+Met+ZY)、模型+双胍+卡托普利组(DN+Met+CP),给予相应药物进行灌胃治疗,并另设正常对照组,每组10只,干预16周后,观察各组大鼠空腹血糖(FBG)、血脂、血清尿素氮(BUN)、血肌酐(Scr)、24 h尿总蛋白(24 h UTP)及尿微量清蛋白(24 h mALB)的变化。观察肾组织病理改变,并对各组肾组织进行RNA-seq测序。结果:与模型组比较,中药组和卡托普利组可以显著下调糖尿病肾病大鼠FBG、TC、TG、LDL-C、Scr、BUN、24 h UTP、24 h mALB等水平,而HDL-C无明显变化;与卡托普利组之间比较,中药组FBG、TC、TG、LDL-C明显下降,HDL-c、Scr、BUN、24 h UTP、24 h mALB均无明显差别。与模型组比较,中药组和卡托普利组均可以显著改善糖尿病肾病大鼠的肾脏病理学变化。转录组测序结果显示模型组对比正常组与中药组对比模型组转录本中基因总量接近,其中表达差异最明显的双向调控基因共筛选出10个;京都基因与基因组百科全书(KEGG)前20条显著富集通路分析显示模型组对比正常组与中药组对比模型组共同作用的信号通路有代谢相关通路及cAMP信号通路。结论:活血降糖饮联合二甲双胍能改善糖尿病肾病大鼠的糖脂代谢紊乱,减少尿白蛋白的排泄,改善肾功能和肾脏病理改变,延缓糖尿病肾病病情进展,其作用机制可能是通过Cftr、Pdk4、Angptl4、Hmgcs2等多靶点介导多通路实现的。
