丹参对视网膜色素变性病理过程Müller细胞特征性基因变化及关键蛋白表达的影响
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摘要
目的基于网络药理学和生物信息学方法探讨丹参(SM)通过干预Müller细胞(MC)特征性基因和关键蛋白表达治疗视网膜色素变性(RP)的分子机制。方法运用中药系统药理学数据库和分析平台(TCMSP)检索、筛选SM的入血活性成分和作用靶点;通过GEO数据库获取正常和RP小鼠MC差异表达基因;通过疾病数据库检索RP相关的基因靶点;采用Cytoscape构建MC差异表达基因和疾病靶点、成分靶点的蛋白质-蛋白质交互作用网络并提取交集;运用DAVID数据库对特征性基因进行基因本体和KEGG通路分析;采用cytoHubba分析筛选关键蛋白靶点。结果检索得到与SM相关的化学成分202个,依据ADME参数筛得活性成分65个,其中入血活性成分13个,进一步检索配对得到这些成分可能作用的靶点117个;从芯片数据中分析得到正常和RP小鼠MC中差异表达基因242个;从疾病数据库获得与RP密切相关的靶点206个;提取交集得到85个SM影响RP病理过程MC的特征性基因;这些基因主要涉及转录调控、凋亡信号通路调控、DNA核内复制调节等生物学过程,分子功能主要包括转录辅激活子活性、蛋白激酶活性、核心启动子结合等,富集于细胞核、核质、转录因子复合物、Rb-E2F复合体等区域,主要与剪接体信号通路、肌动蛋白细胞骨架调控信号途径、细胞周期调控通路等有关;分析筛选出SM干预RP病理过程MC中的8个关键蛋白靶点。结论 SM药效作用的物质基础为隐丹参酮、木犀草素、丹参酮ⅡA等13个化学成分;其所干预的RP病理过程MC特征性基因与剪接体信号通路、肌动蛋白细胞骨架调控信号途径、细胞周期调控通路等机制相关,关键靶点包括RB1、E2F1、TFDP1等8个蛋白。
Objective To investigate the molecular mechanism of Salvia miltiorrhiza(SM) in the treatment of retinitis pigmentosa(RP) by interfering with the expression of characteristic genes and key protein in Müller cells(MC) based on the methods of network pharmacology and bioinformatics. Methods Retrieval and screening of active ingredients and therapeutic targets of SM in blood was performed by Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform(TCMSP). Differentially expressed genes of MC in normal and RP mice were obtained by searching GEO database. RP-related gene targets were retrieved through disease database. Cytoscape was used to construct protein-protein interaction networks of differentially expressed MC genes, disease targets and component targets and the intersection was extracted. Gene Ontology and KEGG signaling pathway analysis of characteristic genes were carried out by DAVID. CytoHubba was used to analyze and screen the key protein targets. Results A total of 202 chemical constituents related to SM were retrieved, 65 active ingredients were screened according to ADME parameters, of which 13 were active ingredients in blood. A total of 117 possible targets were obtained by further searching and matching. A total of 242 differentially expressed genes in MC of normal and RP mice were obtained from chip data. A total of 206 targets closely related to RP were obtained from disease databases. A total of 85 characteristic genes of SM affecting MC in RP pathological process were extracted and intersected. These genes were mainly involved in transcriptional regulation, apoptotic signaling pathway regulation, DNA nuclear replication regulation and other biological processes. Molecular functions mainly include transcriptional coactivator activity, protein kinase activity, core promoter binding, etc. They were enriched in nuclear, nucleoplasm, transcription factor complex, Rb-E2 F complex and other regions. The signaling pathways involved include splicer signaling pathway, actin cytoskeleton signaling pathway, cell cycle signaling pathway and so on. A total of eight key protein targets of SM on MC in RP pathological process were analyzed and screened. Conclusion The substance basis of the pharmacodynamics of SM is 13 chemical constituents, such as cryptotanshinone, luteolin, tanshinone ⅡA, etc. The MC characteristic genes involved in the pathological process of RP intervened by SM are related to spliceosome signaling pathway, actin cytoskeleton signaling pathway, cell cycle regulation pathway, etc. The key targets include eight protein such as RB1, E2 F1, TFDP1, etc.
Objective To investigate the molecular mechanism of Salvia miltiorrhiza(SM) in the treatment of retinitis pigmentosa(RP) by interfering with the expression of characteristic genes and key protein in Müller cells(MC) based on the methods of network pharmacology and bioinformatics. Methods Retrieval and screening of active ingredients and therapeutic targets of SM in blood was performed by Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform(TCMSP). Differentially expressed genes of MC in normal and RP mice were obtained by searching GEO database. RP-related gene targets were retrieved through disease database. Cytoscape was used to construct protein-protein interaction networks of differentially expressed MC genes, disease targets and component targets and the intersection was extracted. Gene Ontology and KEGG signaling pathway analysis of characteristic genes were carried out by DAVID. CytoHubba was used to analyze and screen the key protein targets. Results A total of 202 chemical constituents related to SM were retrieved, 65 active ingredients were screened according to ADME parameters, of which 13 were active ingredients in blood. A total of 117 possible targets were obtained by further searching and matching. A total of 242 differentially expressed genes in MC of normal and RP mice were obtained from chip data. A total of 206 targets closely related to RP were obtained from disease databases. A total of 85 characteristic genes of SM affecting MC in RP pathological process were extracted and intersected. These genes were mainly involved in transcriptional regulation, apoptotic signaling pathway regulation, DNA nuclear replication regulation and other biological processes. Molecular functions mainly include transcriptional coactivator activity, protein kinase activity, core promoter binding, etc. They were enriched in nuclear, nucleoplasm, transcription factor complex, Rb-E2 F complex and other regions. The signaling pathways involved include splicer signaling pathway, actin cytoskeleton signaling pathway, cell cycle signaling pathway and so on. A total of eight key protein targets of SM on MC in RP pathological process were analyzed and screened. Conclusion The substance basis of the pharmacodynamics of SM is 13 chemical constituents, such as cryptotanshinone, luteolin, tanshinone ⅡA, etc. The MC characteristic genes involved in the pathological process of RP intervened by SM are related to spliceosome signaling pathway, actin cytoskeleton signaling pathway, cell cycle regulation pathway, etc. The key targets include eight protein such as RB1, E2 F1, TFDP1, etc.
引文
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[18]Xu G,Li T,Chen J,et al.Autosomal dominant retinitis pigmentosa-associated gene PRPF8 is essential for hypoxia-induced mitophagy through regulating ULK1m RNA splicing[J].Autophagy,2018,14(10):1818-1830.
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[20]Dong E,Bachleda A,Xiong Y,et al.Reduced phosphoCREB in Müller glia during retinal degeneration in rd10 mice[J].Mol Vis,2017,23(1):90-102.
[21]Takeda M,Takamiya A,Jiao J W,et al.Alphaaminoadipate induces progenitor cell properties of Müller glia in adult mice[J].Invest Ophthalmol Vis Sci,2008,49(3):1142-1150.
[22]Arsenijevic Y.Cell cycle proteins and retinal degeneration:Evidences of new potential therapeutic targets[J].Adv Exp Med Biol,2016,854(1):371-377.
[2]Massengill M T,Ahmed C M,Lewin A S,et al.Neuroinflammation in retinitis pigmentosa,diabetic retinopathy,and age-related macular degeneration:Aminireview[J].Adv Exp Med Biol,2018,1074(1):185-191.
[3]Ebdali S,Hashemi B,Hashemi H,et al.Time and frequency components of ERG responses in retinitis pigmentosa[J].Int Ophthalmol,2018,38(6):2435-2444.
[4]Zhang S,Zhang S,Gong W,et al.Müller cell regulated microglial activation and migration in rats with N-methyl-N-nitrosourea-induced retinal degeneration[J].Front Neurosci,2018,doi:10.3389/fnins.2018.00890.
[5]彭清华,李传课.视网膜色素变性虚中夹瘀的机理研究小结[J].中国医药学报,1993,8(6):7-10.
[6]高丽娜,崔元璐,延阔,等.丹参红花配伍研究进展[J].中草药,2016,47(4):671-679.
[7]Ru J,Li P,Wang J,et al.TCMSP:A database of systems pharmacology for drug discovery from herbal medicines[J].J Cheminform,2014,6(1):13.
[8]Roesch K,Stadler M B,Cepko C L.Gene expression changes within Müller glial cells in retinitis pigmentosa[J].Mol Vis,2012,18(1):1197-1214.
[9]Zhou Q,Hong L,Wang J.Identification of key genes and pathways in pelvic organ prolapse based on gene expression profiling by bioinformatics analysis[J].Arch Gynecol Obstet,2018,297(5):1323-1332.
[10]Pang C P,Lam D S.Differential occurrence of mutations causative of eye diseases in the Chinese population[J].Hum Mutat,2002,19(3):189-208.
[11]Gardiner K L,Downs L,Berta-Antalics A I,et al.Photoreceptor proliferation and dysregulation of cell cycle genes in early onset inherited retinal degenerations[J].BMC Genomics,2016,doi:10.1186/s12864-016-2477-9.
[12]Kang M J,Chung J,Ryoo H D.CDK5 and MEKK1mediate pro-apoptotic signalling following endoplasmic reticulum stress in an autosomal dominant retinitis pigmentosa model[J].Nat Cell Biol,2012,14(4):409-415.
[13]Mc Murtrey J J,Tso M O M.A review of the immunologic findings observed in retinitis pigmentosa[J].Surv Ophthalmol,2018,63(6):769-781.
[14]Cyranoski D.Trials of embryonic stem cells to launch in China[J].Nature,2017,546(7656):15-16.
[15]Telegina D V,Kozhevnikova O S,Kolosova N G.Changes in retinal glial cells with age and during development of age-related macular degeneration[J].Biochemistry,2018,83(9):1009-1017.
[16]彭清华.继承朱文锋老师学术思想,开展眼部诊断研究[J].湖南中医药大学学报,2013,33(1):79-84.
[17]Wu Z,Li W,Liu G,et al.Network-based methods for prediction of drug-target interactions[J].Front Pharmacol,2018,doi:10.3389/fphar.2018.01134.
[18]Xu G,Li T,Chen J,et al.Autosomal dominant retinitis pigmentosa-associated gene PRPF8 is essential for hypoxia-induced mitophagy through regulating ULK1m RNA splicing[J].Autophagy,2018,14(10):1818-1830.
[19]Guen V J,Edvardson S,Fraenkel N D,et al.Ahomozygous deleterious CDK10 mutation in a patient with agenesis of corpus callosum,retinopathy,and deafness[J].Am J Med Genet A,2018,176(1):92-98.
[20]Dong E,Bachleda A,Xiong Y,et al.Reduced phosphoCREB in Müller glia during retinal degeneration in rd10 mice[J].Mol Vis,2017,23(1):90-102.
[21]Takeda M,Takamiya A,Jiao J W,et al.Alphaaminoadipate induces progenitor cell properties of Müller glia in adult mice[J].Invest Ophthalmol Vis Sci,2008,49(3):1142-1150.
[22]Arsenijevic Y.Cell cycle proteins and retinal degeneration:Evidences of new potential therapeutic targets[J].Adv Exp Med Biol,2016,854(1):371-377.