[血小板反应蛋白1过表达后睑板腺癌细胞的蛋白质组学分析]。

Q3 Medicine
W Wang, H T Wang, X Liu, L M Zhu, T T Lin
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引用次数: 0

摘要

目的:通过蛋白组学分析,筛选并验证血栓反应蛋白1 (THBS1)过表达影响睑板腺癌(MGC)进展的关键蛋白。方法:实验研究时间为2023年2月~ 2024年6月。慢病毒转染后,将MGC细胞分为THBS1过表达组和对照组。提取两组蛋白进行4D无标记定量蛋白质组学分析。通过基因本体(GO)和京都基因与基因组百科全书(KEGG)途径富集分析,对差异表达蛋白(DEPs)及其调控的信号通路进行功能注释。采用实时定量聚合酶链反应(RT-qPCR)和Western blotting检测前5个DEPs的mRNA和蛋白表达水平。结果:THBS1基因在MGC细胞中成功过表达后,以|log 2 (fold change)、|>1.5、P0.05为标准筛选出666个蛋白。氧化石墨烯分析表明,DEPs主要定位于细胞质基质和外泌体(细胞组分),参与RNA结合和钙粘蛋白结合(分子功能),并与翻译和细胞内蛋白转运(生物学过程)相关。KEGG通路分析显示DNA复制和细胞周期通路显著富集。RT-qPCR结果显示,THBS1过表达组的天冬氨酸糖基化1同源基因(ALG1)、ap2相关蛋白激酶1 (AAK1)、Aladin WD重复核孔蛋白(AAAS)、suo特异性肽酶3 (SENP3)和Serrate RNA效应分子同源基因(SRRT) mRNA表达量分别为0.48±0.05、0.83±0.04、0.90±0.01、0.73±0.06和0.92±0.02,显著低于对照组(1.00±0.03、1.00±0.01、1.00±0.03、1.00±0.03和1.00±0.02);所有P0.05)。Western blotting证实,THBS1过表达组中ALG1、AAK1、AAAS、SENP3、SRRT蛋白表达水平分别为0.53±0.04、0.86±0.04、0.40±0.11、0.59±0.01、0.63±0.05,与对照组(1.00±0.01、1.00±0.03、1.00±0.19、1.00±0.14、1.00±0.01)相比,均显著降低;所有P0.05)。结论:在MGC进展中受THBS1过表达调控的关键蛋白中,DEPs前5位分别为ALG1、AAK1、AAAS、SENP3和SRRT。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
[Proteomic analysis of meibomian gland carcinoma cells after overexpression of thrombospondin 1].

Objective: To screen and validate key proteins involved in the progression of meibomian gland carcinoma (MGC) influenced by thrombospondin 1 (THBS1) overexpression through proteomic analysis. Methods: It was an experimental study conducted from February 2023 to June 2024. After lentiviral transfection, MGC cells were divided into the THBS1 overexpression group and the control group. Proteins were extracted from both groups for 4D label-free quantitative proteomic analysis. Functional annotation of differentially expressed proteins (DEPs) and their regulated signaling pathways was performed via Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses. Real-time quantitative polymerase chain reaction (RT-qPCR) and Western blotting were used to validate the mRNA and protein expression levels of the top 5 DEPs. Results: Following successful THBS1 gene overexpression in MGC cells, 666 proteins were screened based on the criteria of |log₂(fold change)|>1.5 and P<0.05. GO analysis showed that DEPs were mainly localized in the cytosolic matrix and exosomes (cellular component), involved in RNA binding and cadherin binding (molecular function), and associated with translation and intracellular protein transport (biological process). KEGG pathway analysis indicated significant enrichment in DNA replication and cell cycle pathways. RT-qPCR results showed mRNA expression levels of asparagine-linked glycosylation 1 homolog (ALG1), AP2-associated protein kinase 1 (AAK1), Aladin WD repeat nucleoporin (AAAS), SUMO-specific peptidase 3 (SENP3), and Serrate RNA effector molecule homolog (SRRT) in the THBS1 overexpression group were 0.48±0.05, 0.83±0.04, 0.90±0.01, 0.73±0.06, and 0.92±0.02, respectively, significantly lower than those in the control group (1.00±0.03, 1.00±0.01, 1.00±0.03, 1.00±0.03, and 1.00±0.02; all P<0.05). Western blotting confirmed protein expression levels of ALG1, AAK1, AAAS, SENP3, and SRRT in the THBS1 overexpression group were 0.53±0.04, 0.86±0.04, 0.40±0.11, 0.59±0.01, and 0.63±0.05, respectively, significantly reduced compared with the control group (1.00±0.01, 1.00±0.03, 1.00±0.19, 1.00±0.14, and 1.00±0.01; all P<0.05). Conclusion: Among the key proteins regulated by THBS1 overexpression in the MGC progression, the top 5 DEPs were ALG1, AAK1, AAAS, SENP3, and SRRT.

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来源期刊
中华眼科杂志
中华眼科杂志 Medicine-Ophthalmology
CiteScore
0.80
自引率
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发文量
12700
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