急性内皮应激确定了微小RNA let-7b-5p和非编码SLC11A2（NRAMP2/DMT1）外显子作为生物标志物，这些生物标志物与在恶性和非恶性疾病中检测到的生物标志物重叠。

Acute endothelial stresses identify microRNA let-7b-5p and non-coding SLC11A2 (NRAMP2/DMT1) exon as biomarkers that overlap with those detected in malignant and non-malignant diseases.

作者信息

Bielowka Adrianna M, Patel Dilip, Li Dongyang, Bernabeu-Herrero Maria E, Game Laurence, Aldred Micheala A, Mollet Inês G, Shovlin Claire L

机构信息

National Heart and Lung Institute, Imperial College London, London, UK.

NIHR Imperial Biomedical Research Centre, London, UK.

出版信息

QJM. 2024 Dec 10. doi: 10.1093/qjmed/hcae235.

DOI:10.1093/qjmed/hcae235

PMID:39658243

Abstract

BACKGROUND

Disease biomarkers are often identified long after initiating pathologies, hampering mechanistic understanding and development of preventative strategies. We hypothesised that aberrant cellular responses to normally-encountered stresses may be relevant to later disease states.

AIM

To model two under-explored acute cellular stresses for blood-exposed cells, and cross-reference to known biomarkers of disease.

METHODS

Normal primary human endothelial cells (ECs) were treated for 1-6 h with cycloheximide 100 μg/mL to inhibit protein translation and nonsense mediated decay (modelling the integrated stress response), or 10 μmol/L ferric citrate (modelling diurnal variation in serum iron that can be augmented by treatments prescribed 8 million times/year in England). Directional whole transcriptome RNA-seq identified differentially expressed genes and micro(mi)RNAs. Customized novel scripts examined expression of 517,225 exons to predict 1 h cycloheximide-stabilized exons. Validations were by cel-miR-39-spiked qRT-PCR and RNA-seq in other endothelial cell types, peripheral blood mononuclear cells (PBMCs), and plasma.

RESULTS

miRNAs fell transiently at 1 h after 10 μmol/L ferric citrate (p < 0.01), specifically in let-7 family member pre-miRNAs ('let-7', p < 0.05), where there was an accompanying differential 6 h increased expression of 570 let-7-target mRNAs identified through TargetScan (p < 0.0001). qRT-PCR and RNA-seq validations in other normal endothelial cells, plasma and PBMCs confirmed up to 80% falls in pre-let-7b/let-7b-5p after 1 h iron, and exon 3B of the SLC11A2 (NRAMP2/DMT1)-encoded iron/copper transporter as a novel exon most consistently stabilized following 1 h treatment with cycloheximide. Overlaps with disease biomarkers for cancer, growth retardation, and multiple organ-specific diseases were identified.

CONCLUSIONS

Biomarkers for normal, acute cellular responses overlap with disease-state biomarkers, warranting further study.

摘要

背景

疾病生物标志物通常在引发病理状态很久之后才被发现，这阻碍了对发病机制的理解以及预防策略的制定。我们推测，细胞对正常遇到的应激的异常反应可能与后期的疾病状态相关。

目的

对血液暴露细胞的两种未充分研究的急性细胞应激进行建模，并与已知的疾病生物标志物进行交叉对照。

方法

将正常原代人内皮细胞（ECs）用100μg/mL放线菌酮处理1 - 6小时，以抑制蛋白质翻译和无义介导的衰变（模拟综合应激反应），或用10μmol/L柠檬酸铁处理（模拟血清铁的昼夜变化，在英国每年有800万次治疗会增加这种变化）。定向全转录组RNA测序确定差异表达的基因和微小（mi）RNA。定制的新脚本检查了517,225个外显子的表达，以预测1小时放线菌酮稳定的外显子。通过cel-miR-39加标的qRT-PCR以及在其他内皮细胞类型、外周血单核细胞（PBMCs）和血浆中的RNA测序进行验证。

结果

在10μmol/L柠檬酸铁处理后1小时，miRNA短暂下降（p < 0.01），特别是在let-7家族成员前体miRNA（“let-7”，p < 0.05）中，通过TargetScan鉴定出570个let-7靶标mRNA伴随有6小时的差异表达增加（p < 0.0001）。在其他正常内皮细胞、血浆和PBMCs中的qRT-PCR和RNA测序验证证实，铁处理1小时后前体let-7b/let-7b-5p下降高达80%，并且SLC11A2（NRAMP2/DMT1）编码的铁/铜转运蛋白的外显子3B是用放线菌酮处理1小时后最一致稳定的新外显子。确定了与癌症、生长发育迟缓及多种器官特异性疾病的疾病生物标志物的重叠。