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Multiresolution imaging using bioluminescence resonance energy transfer identifies distinct biodistribution profiles of extracellular vesicles and exomeres with redirected tropism

ABSTRACT: Extracellular particles (EPs) including extracellular vesicles and exomeres play a significant role in diseases and therapeutic applications. However, the spatiotemporal dynamics of EPs in vivo remains to be elucidated with a suitable method. In this study, we developed a bioluminescence resonance energy transfer (BRET)-based reporter, PalmGRET, to visualize, track and quantify EPs both in vitro and in vivo. To explore the tropism effect of membrane proteins on EPs from lung metastatic hepatocellular carcimoma (HCC), we established PalmGRET in a mouse hepatocellular carcinoma cell line, HCA1, termed HCA1-PalmGRET. EP harvested from HCA1-PalmGRET cells were processed with gel electrophoresis, followed by in-gel digestion and LC-MS/MS analysis. The identified protein list was compared with published lung tropism databases and four membrane proteins were selected for subsequent gene knockdown experiments: solute carrier organic anion transporter family member 2A1 (Slco2a1), alanine aminopeptidase (Anpep/Cd13), chloride intracellular channel 1 (Clic1), and sodium-hydrogen antiporter 3 regulator 1 (Nherf1). Using PalmGRET, we revealed that knockdown of Slco2a1, Cd13 and Clic1 significantly reduced lung tropism of HCC-EPs with varying redirected delivery to other organs.

INSTRUMENT(S): Orbitrap Fusion Lumos

ORGANISM(S): Mus Musculus (mouse)

TISSUE(S): Hepatocyte, Cell Culture

DISEASE(S): Hepatocellular Carcinoma

SUBMITTER: Steven Chuo

LAB HEAD: Charles Pin-Kuang Lai

PROVIDER: PXD020446 | Pride | 2021-09-09

REPOSITORIES: Pride

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Multiresolution Imaging Using Bioluminescence Resonance Energy Transfer Identifies Distinct Biodistribution Profiles of Extracellular Vesicles and Exomeres with Redirected Tropism.

Wu Anthony Yan-Tang AY Sung Yun-Chieh YC Chen Yen-Ju YJ Chou Steven Ting-Yu ST Guo Vanessa V Chien Jasper Che-Yung JC Ko John Jun-Sheng JJ Yang Alan Ling AL Huang Hsi-Chien HC Chuang Ju-Chen JC Wu Syuan S Ho Meng-Ru MR Ericsson Maria M Lin Wan-Wan WW Cheung Chantal Hoi Yin CHY Juan Hsueh-Fen HF Ueda Koji K Chen Yunching Y Lai Charles Pin-Kuang CP

Advanced science (Weinheim, Baden-Wurttemberg, Germany) 20200816 19

Extracellular particles (EPs) including extracellular vesicles (EVs) and exomeres play significant roles in diseases and therapeutic applications. However, their spatiotemporal dynamics in vivo have remained largely unresolved in detail due to the lack of a suitable method. Therefore, a bioluminescence resonance energy transfer (BRET)-based reporter, PalmGRET, is created to enable pan-EP labeling ranging from exomeres (<50 nm) to small (<200 nm) and medium and large (>200 nm) EVs. PalmGRET emits ...[more]

PMID: 33042758

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Project description:Alcohol is a major risk factor for hepatocellular carcinoma (HCC) although the mechanisms underlying the alcohol-related liver carcinogenesis are still poorly understood. Alcohol is known to increase hepatocarcinogenesis possibly by inducing aberrant DNA methylation through the reduced provision of methyl groups within the hepatic one-carbon metabolism. Whether the epigenetically-regulated pathways in alcohol-associated HCC can be reversible or modifiable by nutritional factors is unknown. The aim of the present study was to investigate the genome wide promoter DNA methylation profiles along with array-based, gene expression profiles in non-viral, alcohol-associated HCC. From eight HCC patients the methylation status and transcriptional levels of all annotated genes were compared by analyzing HCC tissue and the cancer-free surrounding liver tissue, following curative surgery. After merging both the DNA methylation and gene expression data, we identified 159 hypermethylated-repressed, 30 hypomethylated-induced, 49 hypermethylated-induced and 56 hypomethylated-repressed genes. A number of potentially novel candidate tumor-suppressor genes (FAM107A, IGFALS, MT1G, MT1H, RNF180) demonstrated promoter hypermethylation and transcriptional repression in alcohol-associated HCC. Notably, promoter DNA methylation appeared as the regulatory mechanism for the transcriptional repression of genes controlling the retinol metabolic pathway (ADH1A, ADH1B, ADH6, CYP3A43, CYP4A22, RDH16) and SHMT1, a key gene within one-carbon metabolism. A genome-wide DNA methylation approach linked up with array-based gene expression profiles allowed identifying a number of novel, epigenetically-regulated candidate tumor-suppressor genes in alcohol-associated hepatocarcinogenesis. Retinol metabolism genes and SHMT1 are also epigenetically-regulated through promoter DNA methylation in alcohol-associated hepatocarcinogenesis. 16 samples (8 control samples from non-neoplastic liver tissue, 8 test samples from hepatocellular carcinoma) from 8 patients affected from hepatocellular carcinoma were analyzed.

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