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Differential release of extracellular vesicle tRNA from oxidative stressed renal cells and ischemic kidneys.

ABSTRACT: While urine-based liquid biopsy has expanded to the analyses of extracellular nucleic acids, the potential of transfer RNA (tRNA) encapsulated within extracellular vesicles has not been explored as a new class of urine biomarkers for kidney injury. Using rat kidney and mouse tubular cell injury models, we tested if extracellular vesicle-loaded tRNA and their m¹A (N¹-methyladenosine) modification reflect oxidative stress of kidney injury and determined the mechanism of tRNA packaging into extracellular vesicles. We determined a set of extracellular vesicle-loaded, isoaccepting tRNAs differentially released after ischemia-reperfusion injury and oxidative stress. Next, we found that m¹A modification of extracellular vesicle tRNAs, despite an increase of the methylated tRNAs in intracellular vesicles, showed little or no change under oxidative stress. Mechanistically, oxidative stress decreases tRNA loading into intracellular vesicles while the tRNA-loaded vesicles are accumulated due to decreased release of the vesicles from the cell surface. Furthermore, Maf1-mediated transcriptional repression of the tRNAs decreases the cargo availability for extracellular vesicle release in response to oxidative stress. Taken together, our data support that release of extracellular vesicle tRNAs reflects oxidative stress of kidney tubules which might be useful to detect ischemic kidney injury and could lead to rebalance protein translation under oxidative stress.

SUBMITTER: Lee HK

PROVIDER: S-EPMC8803936 | biostudies-literature | 2022 Jan

REPOSITORIES: biostudies-literature

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Differential release of extracellular vesicle tRNA from oxidative stressed renal cells and ischemic kidneys.

Lee Hee Kyung HK Lee Byung Rho BR Lee Tae Jin TJ Lee Chang Min CM Li Chenglong C O'Connor Paul M PM Dong Zheng Z Kwon Sang-Ho SH

Scientific reports 20220131 1

While urine-based liquid biopsy has expanded to the analyses of extracellular nucleic acids, the potential of transfer RNA (tRNA) encapsulated within extracellular vesicles has not been explored as a new class of urine biomarkers for kidney injury. Using rat kidney and mouse tubular cell injury models, we tested if extracellular vesicle-loaded tRNA and their m<sup>1</sup>A (N<sup>1</sup>-methyladenosine) modification reflect oxidative stress of kidney injury and determined the mechanism of tRNA ...[more]

PMID: 35102218

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Project description:BackgroundExtracellular vesicles (EVs) contain bioactive cargo including miRNAs and proteins that are released by cells during cell-cell communication. Endothelial cells (ECs) form the innermost lining of all blood vessels, interfacing with cells in the circulation and vascular wall. It is unknown whether ECs release EVs capable of governing recipient cells within these 2 separate compartments. Given their boundary location, we propose ECs use bidirectional release of distinct EV cargo in quiescent (healthy) and activated (atheroprone) states to communicate with cells within the circulation and blood vessel wall.MethodsEVs were isolated from primary human aortic ECs (plate and transwell grown; ±IL [interleukin]-1β activation), quantified, visualized, and analyzed by miRNA transcriptomics and proteomics. Apical and basolateral EC-EV release was determined by miRNA transfer, total internal reflection fluorescence and electron microscopy. Vascular reprogramming (RNA sequencing) and functional assays were performed on primary human monocytes or smooth muscle cells±EC-EVs.ResultsActivated ECs increased EV release, with miRNA and protein cargo related to atherosclerosis. EV-treated monocytes and smooth muscle cells revealed activated EC-EV altered pathways that were proinflammatory and atherogenic. ECs released more EVs apically, which increased with activation. Apical and basolateral EV cargo contained distinct transcriptomes and proteomes that were altered by EC activation. Notably, activated basolateral EC-EVs displayed greater changes in the EV secretome, with pathways specific to atherosclerosis. In silico analysis determined compartment-specific cargo released by the apical and basolateral surfaces of ECs can reprogram monocytes and smooth muscle cells, respectively, with functional assays and in vivo imaging supporting this concept.ConclusionsDemonstrating that ECs are capable of polarized EV cargo loading and directional EV secretion reveals a novel paradigm for endothelial communication, which may ultimately enhance the design of endothelial-based therapeutics for cardiovascular diseases such as atherosclerosis where ECs are persistently activated.

Project description:Extracellular vesicles (EVs) are released by virtually all cells and may serve as intercellular communication structures by transmitting molecules such as proteins, lipids, and nucleic acids between cells. MicroRNAs (miRNAs) are an abundant class of vesicular RNA playing a pivotal role in regulating intracellular processes. In this work, we aimed to characterize vesicular miRNA profiles released in a side-directed manner by bronchial epithelial cells from healthy and asthmatic subjects using an air-liquid interface cell culture model. EVs were isolated from a culture medium collected from either the basolateral or apical cell side of the epithelial cell cultures and characterized by nano-flow cytometry (NanoFCM) and bead-based flow cytometry. EV-associated RNA profiles were assessed by small RNA sequencing and subsequent bioinformatic analyses. Furthermore, miRNA-associated functions and targets were predicted and miRNA network analyses were performed. EVs were released at higher numbers to the apical cell side of the epithelial cells and were considerably smaller in the apical compared to the basolateral compartment. EVs from both compartments showed a differential tetraspanins surface marker expression. Furthermore, 236 miRNAs were differentially expressed depending on the EV secretion side, regardless of the disease phenotype. On the apical cell side, 32 miRNAs were significantly altered in asthmatic versus healthy conditions, while on the basolateral cell side, 23 differentially expressed miRNAs could be detected. Downstream KEGG pathway analysis predicted mTOR and MAPK signaling pathways as potential downstream targets of apically secreted miRNAs. In contrast, miRNAs specifically detected at the basolateral side were associated with processes of T and B cell receptor signaling. The study proves a compartmentalized packaging of EVs by bronchial epithelial cells supposedly associated with site-specific functions of cargo miRNAs, which are considerably affected by disease conditions such as asthma.

Dataset Information

Differential release of extracellular vesicle tRNA from oxidative stressed renal cells and ischemic kidneys.

Publications

Differential release of extracellular vesicle tRNA from oxidative stressed renal cells and ischemic kidneys.

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