人肝癌Huh-7细胞培养模型缺乏载脂蛋白B分泌。

IF 4.1 2区 医学 Q1 BIOCHEMISTRY & MOLECULAR BIOLOGY
Journal of Lipid Research Pub Date : 2025-08-01 Epub Date: 2025-07-23 DOI:10.1016/j.jlr.2025.100867
Swati Valmiki, Sara Rosario, Ashley Mooring, Chandana Prakashmurthy, Florencia Schlamp, Binu Prakash, Atrayee Chattopadhyay, Abulaish Ansari, José O Alemán, Nathalie Pamir, M Mahmood Hussain
{"title":"人肝癌Huh-7细胞培养模型缺乏载脂蛋白B分泌。","authors":"Swati Valmiki, Sara Rosario, Ashley Mooring, Chandana Prakashmurthy, Florencia Schlamp, Binu Prakash, Atrayee Chattopadhyay, Abulaish Ansari, José O Alemán, Nathalie Pamir, M Mahmood Hussain","doi":"10.1016/j.jlr.2025.100867","DOIUrl":null,"url":null,"abstract":"<p><p>ApoB is an essential structural protein for the assembly and secretion of triglyceride-rich lipoproteins and therefore remains a potential target to lower plasma cholesterol levels in hypercholesterolemia patients. To understand the global consequences of APOB gene deficiency, we employed CRISPR-Cas9 system to generate apoB-deficient human hepatoma Huh-7 cells (Ako cells). ApoB was not detectable in the cells and media of the Ako cells. ApoB deficiency had no effect on microsomal triglyceride transfer protein expression and activity. These cells supported apoB48 secretion when transfected with plasmids for the expression of apoB48 suggesting that these cells retain all the lipoprotein assembly and secretion machinery except for apoB expression. APOB gene deficiency had no significant effect on cellular lipid levels, cell growth, and ER stress markers. Proteome analysis of secreted proteins revealed that the most upregulated protein was the vitamin D binding protein, while the most downregulated protein was apoB in Ako cells compared to control cells. This analysis also identified coagulation as an upregulated pathway. Total RNA transcriptome analysis identified DNA replication and complement and coagulation pathways as the most upregulated pathways in Ako cells. Further detailed studies are needed to establish how apoB regulates these pathways. These Ako cells may be useful in studying structure-function analysis of apoB peptides and to address the cellular consequences of disruptions in lipoprotein assembly and secretion.</p>","PeriodicalId":16209,"journal":{"name":"Journal of Lipid Research","volume":" ","pages":"100867"},"PeriodicalIF":4.1000,"publicationDate":"2025-08-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12396022/pdf/","citationCount":"0","resultStr":"{\"title\":\"Human hepatoma Huh-7 cell culture models deficient in apolipoprotein B secretion.\",\"authors\":\"Swati Valmiki, Sara Rosario, Ashley Mooring, Chandana Prakashmurthy, Florencia Schlamp, Binu Prakash, Atrayee Chattopadhyay, Abulaish Ansari, José O Alemán, Nathalie Pamir, M Mahmood Hussain\",\"doi\":\"10.1016/j.jlr.2025.100867\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p><p>ApoB is an essential structural protein for the assembly and secretion of triglyceride-rich lipoproteins and therefore remains a potential target to lower plasma cholesterol levels in hypercholesterolemia patients. To understand the global consequences of APOB gene deficiency, we employed CRISPR-Cas9 system to generate apoB-deficient human hepatoma Huh-7 cells (Ako cells). ApoB was not detectable in the cells and media of the Ako cells. ApoB deficiency had no effect on microsomal triglyceride transfer protein expression and activity. These cells supported apoB48 secretion when transfected with plasmids for the expression of apoB48 suggesting that these cells retain all the lipoprotein assembly and secretion machinery except for apoB expression. APOB gene deficiency had no significant effect on cellular lipid levels, cell growth, and ER stress markers. Proteome analysis of secreted proteins revealed that the most upregulated protein was the vitamin D binding protein, while the most downregulated protein was apoB in Ako cells compared to control cells. This analysis also identified coagulation as an upregulated pathway. Total RNA transcriptome analysis identified DNA replication and complement and coagulation pathways as the most upregulated pathways in Ako cells. Further detailed studies are needed to establish how apoB regulates these pathways. These Ako cells may be useful in studying structure-function analysis of apoB peptides and to address the cellular consequences of disruptions in lipoprotein assembly and secretion.</p>\",\"PeriodicalId\":16209,\"journal\":{\"name\":\"Journal of Lipid Research\",\"volume\":\" \",\"pages\":\"100867\"},\"PeriodicalIF\":4.1000,\"publicationDate\":\"2025-08-01\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12396022/pdf/\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Journal of Lipid Research\",\"FirstCategoryId\":\"99\",\"ListUrlMain\":\"https://doi.org/10.1016/j.jlr.2025.100867\",\"RegionNum\":2,\"RegionCategory\":\"医学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"2025/7/23 0:00:00\",\"PubModel\":\"Epub\",\"JCR\":\"Q1\",\"JCRName\":\"BIOCHEMISTRY & MOLECULAR BIOLOGY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Journal of Lipid Research","FirstCategoryId":"99","ListUrlMain":"https://doi.org/10.1016/j.jlr.2025.100867","RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"2025/7/23 0:00:00","PubModel":"Epub","JCR":"Q1","JCRName":"BIOCHEMISTRY & MOLECULAR BIOLOGY","Score":null,"Total":0}
引用次数: 0

摘要

载脂蛋白B (apoB)是富含甘油三酯的脂蛋白组装和分泌的必需结构蛋白,因此仍然是降低高胆固醇血症患者血浆胆固醇水平的潜在靶点。为了了解APOB基因缺乏的整体后果,我们采用CRISPR-Cas9系统生成APOB缺陷的人肝癌Huh-7细胞(Ako细胞)。在Ako细胞的细胞和培养基中未检测到ApoB。ApoB缺乏对MTP的表达和活性无影响。当转染表达apoB48的质粒时,这些细胞支持apoB48的分泌,这表明这些细胞保留了除apoB表达外的所有脂蛋白组装和分泌机制。APOB基因缺失对细胞脂质水平、细胞生长和内质网应激指标无显著影响。分泌蛋白的蛋白质组学分析显示,与对照细胞相比,Ako细胞中维生素D结合蛋白上调最多,apoB下调最多。该分析还确定了凝血是一种上调途径。总RNA转录组分析发现,DNA复制、补体和凝血途径是Ako细胞中上调最多的途径。需要进一步的详细研究来确定载脂蛋白ob如何调节这些途径。这些Ako细胞可能有助于研究载脂蛋白ob肽的结构-功能分析,并解决脂蛋白组装和分泌中断的细胞后果。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
Human hepatoma Huh-7 cell culture models deficient in apolipoprotein B secretion.

ApoB is an essential structural protein for the assembly and secretion of triglyceride-rich lipoproteins and therefore remains a potential target to lower plasma cholesterol levels in hypercholesterolemia patients. To understand the global consequences of APOB gene deficiency, we employed CRISPR-Cas9 system to generate apoB-deficient human hepatoma Huh-7 cells (Ako cells). ApoB was not detectable in the cells and media of the Ako cells. ApoB deficiency had no effect on microsomal triglyceride transfer protein expression and activity. These cells supported apoB48 secretion when transfected with plasmids for the expression of apoB48 suggesting that these cells retain all the lipoprotein assembly and secretion machinery except for apoB expression. APOB gene deficiency had no significant effect on cellular lipid levels, cell growth, and ER stress markers. Proteome analysis of secreted proteins revealed that the most upregulated protein was the vitamin D binding protein, while the most downregulated protein was apoB in Ako cells compared to control cells. This analysis also identified coagulation as an upregulated pathway. Total RNA transcriptome analysis identified DNA replication and complement and coagulation pathways as the most upregulated pathways in Ako cells. Further detailed studies are needed to establish how apoB regulates these pathways. These Ako cells may be useful in studying structure-function analysis of apoB peptides and to address the cellular consequences of disruptions in lipoprotein assembly and secretion.

求助全文
通过发布文献求助,成功后即可免费获取论文全文。 去求助
来源期刊
Journal of Lipid Research
Journal of Lipid Research 生物-生化与分子生物学
CiteScore
11.10
自引率
4.60%
发文量
146
审稿时长
41 days
期刊介绍: The Journal of Lipid Research (JLR) publishes original articles and reviews in the broadly defined area of biological lipids. We encourage the submission of manuscripts relating to lipids, including those addressing problems in biochemistry, molecular biology, structural biology, cell biology, genetics, molecular medicine, clinical medicine and metabolism. Major criteria for acceptance of articles are new insights into mechanisms of lipid function and metabolism and/or genes regulating lipid metabolism along with sound primary experimental data. Interpretation of the data is the authors’ responsibility, and speculation should be labeled as such. Manuscripts that provide new ways of purifying, identifying and quantifying lipids are invited for the Methods section of the Journal. JLR encourages contributions from investigators in all countries, but articles must be submitted in clear and concise English.
×
引用
GB/T 7714-2015
复制
MLA
复制
APA
复制
导出至
BibTeX EndNote RefMan NoteFirst NoteExpress
×
提示
您的信息不完整,为了账户安全,请先补充。
现在去补充
×
提示
您因"违规操作"
具体请查看互助需知
我知道了
×
提示
确定
请完成安全验证×
copy
已复制链接
快去分享给好友吧!
我知道了
右上角分享
点击右上角分享
0
联系我们:info@booksci.cn Book学术提供免费学术资源搜索服务,方便国内外学者检索中英文文献。致力于提供最便捷和优质的服务体验。 Copyright © 2023 布克学术 All rights reserved.
京ICP备2023020795号-1
ghs 京公网安备 11010802042870号
Book学术文献互助
Book学术文献互助群
群 号:604180095
Book学术官方微信