结合计算目标优先级和B细胞成熟测定在系统性红斑狼疮的目标评估研究。

IF 5.9 2区 医学 Q1 MEDICAL LABORATORY TECHNOLOGY
Ming-Mei Shang , Zhuang Liu , Bogdan Knezevic , Christine Möller Westerberg , Sudeepta Kumar Panda , Hai Fang , Ning Xu Landén , Michael Sundström , Julian C. Knight , Louise Berg
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引用次数: 0

摘要

背景和目的:系统性红斑狼疮(SLE)是一种系统性自身免疫性疾病,涉及B细胞产生自身抗体。本研究旨在利用计算算法确定新的药物靶点,然后在原代B细胞成熟实验中通过siRNA敲低验证排名前11位的靶点。实验方法:通过优先指数(Pi)对SLE全基因组关联研究中排名前1%的基因(~ 150个基因)进行排序,Pi是一种整合基因组和网络信息的计算工具,可对疾病相关基因进行优先排序。这些药物通过网络连通性、可药物性、在自身免疫性疾病中的排名以及不直接干扰所使用的B细胞刺激鸡尾酒等因素进一步筛选。从中选择11个基因进行siRNA敲低验证:IFNGR1、IL-2、IRF4、IL-12A、IL-12B、VCAM-1、ATF6B、RELA、IKBKG、CHUK和MAPK14。流式细胞术分析其对原代血B细胞诱导成熟和活力的影响,ELISA法检测其对IgG分泌的影响。用siRNA处理的B细胞进行rna测序,以研究功能改变的分子机制。关键结果:实验结果表明,几个靶点(IFNGR1, IL-2, IL-12A, MAPK14, IRF4, CHUK, ATF6B, IKBKG和RELA)参与B细胞成熟,因为敲低导致IgG产生减少和/或B细胞成熟减慢。观察到的对IgG分泌和B细胞成熟影响的变异性表明,这些基因编码的蛋白质在机制作用上存在差异。对靶基因表达被调节的细胞进行的RNA-seq分析显示,参与对B细胞功能重要的细胞过程的数百个基因的表达水平受到影响。结论与意义:将靶标优先排序算法与基因敲低和全转录组学分析的实验功能验证研究相结合,是一种很有前途的方法,可以识别免疫疾病的潜在新药物靶点。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
Combining computational target prioritization and a B cell maturation assay for target evaluation studies in systemic lupus erythematosus

Background and Purpose

Systemic lupus erythematosus (SLE) is a systemic autoimmune disease involving production of autoantibodies by B cells. This study aimed at identifying novel drug targets using a computational algorithm to select targets and thereafter validate the top ranked 11 targets by siRNA knockdown in a primary B cell maturation assay.

Experimental Approach

The top 1 % genes (∼150 genes) from SLE genome-wide association studies were ranked by Priority index (Pi), a computational tool integrating genomic and network information to prioritize disease-relevant genes. These were further filtered by network connectivity, drugability, for ranking highly in autoimmune diseases and for not directly interfering with the B cell stimulation cocktail used. From this, 11 genes were selected for validation by siRNA knockdown: IFNGR1, IL-2, IRF4, IL-12A, IL-12B, VCAM-1, ATF6B, RELA, IKBKG, CHUK and MAPK14. Effects on induced maturation and viability of primary blood B cells were analyzed by flow cytometry, and effects on IgG secretion were investigated by ELISA. RNA-sequencing of B cells treated with siRNA was performed to investigate molecular mechanisms underlying the functional alterations.

Key Results

Experimental results show that several of the targets (IFNGR1, IL-2, IL-12A, MAPK14, IRF4, CHUK, ATF6B, IKBKG, and RELA) are involved in B cell maturation, as knockdown caused reduced IgG production and/or decreased maturation of B cells. The observed variability of effects on IgG secretion and B cell maturation suggests differences in the mechanistic roles of the proteins encoded by these genes. RNA-seq analysis of cells where expression of the targeted genes had been modulated showed effects on the expression level of hundreds of genes involved in cellular processes important for B cell functions.

Conclusion and Implications

Combining the target prioritization algorithm with experimental functional validation studies by gene knockdown and whole transcriptomics profiling constitutes a promising approach to identify potential novel drug targets in immune disorders.
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来源期刊
Translational Research
Translational Research 医学-医学:内科
CiteScore
15.70
自引率
0.00%
发文量
195
审稿时长
14 days
期刊介绍: Translational Research (formerly The Journal of Laboratory and Clinical Medicine) delivers original investigations in the broad fields of laboratory, clinical, and public health research. Published monthly since 1915, it keeps readers up-to-date on significant biomedical research from all subspecialties of medicine.
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