Capturing the Kidney Transcriptome by Urinary Extracellular Vesicles—From Pre-Analytical Obstacles to Biomarker Research
(2023) In Genes 14(7).- Abstract
Urinary extracellular vesicles (uEV) hold non-invasive RNA biomarkers for genitourinary tract diseases. However, missing knowledge about reference genes and effects of preanalytical choices hinder biomarker studies. We aimed to assess how preanalytical variables (urine storage temperature, isolation workflow) affect diabetic kidney disease (DKD)—linked miRNAs or kidney—linked miRNAs and mRNAs (kidney-RNAs) in uEV isolates and to discover stable reference mRNAs across diverse uEV datasets. We studied nine raw and normalized sequencing datasets including healthy controls and individuals with prostate cancer or type 1 diabetes with or without albuminuria. We focused on kidney-RNAs reviewing literature for DKD-linked miRNAs from kidney... (More)
Urinary extracellular vesicles (uEV) hold non-invasive RNA biomarkers for genitourinary tract diseases. However, missing knowledge about reference genes and effects of preanalytical choices hinder biomarker studies. We aimed to assess how preanalytical variables (urine storage temperature, isolation workflow) affect diabetic kidney disease (DKD)—linked miRNAs or kidney—linked miRNAs and mRNAs (kidney-RNAs) in uEV isolates and to discover stable reference mRNAs across diverse uEV datasets. We studied nine raw and normalized sequencing datasets including healthy controls and individuals with prostate cancer or type 1 diabetes with or without albuminuria. We focused on kidney-RNAs reviewing literature for DKD-linked miRNAs from kidney tissue, cell culture and uEV/urine experiments. RNAs were analyzed by expression heatmaps, hierarchical clustering and selecting stable mRNAs with normalized counts (>200) and minimal coefficient of variation. Kidney-RNAs were decreased after urine storage at −20 °C vs. −80 °C. Isolation workflows captured kidney-RNAs with different efficiencies. Ultracentrifugation captured DKD -linked miRNAs that separated healthy and diabetic macroalbuminuria groups. Eleven mRNAs were stably expressed across the datasets. Hence, pre-analytical choices had variable effects on kidney-RNAs—analyzing kidney-RNAs complemented global correlation, which could fade differences in some relevant RNAs. Replicating prior DKD-marker results and discovery of candidate reference mRNAs encourages further uEV biomarker studies.
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- author
- Barreiro, Karina ; Dwivedi, Om Prakash ; Rannikko, Antti ; Holthöfer, Harry ; Tuomi, Tiinamaija LU ; Groop, Per Henrik and Puhka, Maija
- organization
- publishing date
- 2023
- type
- Contribution to journal
- publication status
- published
- subject
- keywords
- diabetic kidney disease, exosomes, miRNA, mRNA, reference genes, sequencing, urinary extracellular vesicles, urine
- in
- Genes
- volume
- 14
- issue
- 7
- article number
- 1415
- publisher
- MDPI AG
- external identifiers
-
- pmid:37510317
- scopus:85165977391
- ISSN
- 2073-4425
- DOI
- 10.3390/genes14071415
- language
- English
- LU publication?
- yes
- id
- 545cdb9e-2632-441e-90a6-39b04a16d963
- date added to LUP
- 2023-11-02 14:14:12
- date last changed
- 2024-04-19 03:29:04
@article{545cdb9e-2632-441e-90a6-39b04a16d963, abstract = {{<p>Urinary extracellular vesicles (uEV) hold non-invasive RNA biomarkers for genitourinary tract diseases. However, missing knowledge about reference genes and effects of preanalytical choices hinder biomarker studies. We aimed to assess how preanalytical variables (urine storage temperature, isolation workflow) affect diabetic kidney disease (DKD)—linked miRNAs or kidney—linked miRNAs and mRNAs (kidney-RNAs) in uEV isolates and to discover stable reference mRNAs across diverse uEV datasets. We studied nine raw and normalized sequencing datasets including healthy controls and individuals with prostate cancer or type 1 diabetes with or without albuminuria. We focused on kidney-RNAs reviewing literature for DKD-linked miRNAs from kidney tissue, cell culture and uEV/urine experiments. RNAs were analyzed by expression heatmaps, hierarchical clustering and selecting stable mRNAs with normalized counts (>200) and minimal coefficient of variation. Kidney-RNAs were decreased after urine storage at −20 °C vs. −80 °C. Isolation workflows captured kidney-RNAs with different efficiencies. Ultracentrifugation captured DKD -linked miRNAs that separated healthy and diabetic macroalbuminuria groups. Eleven mRNAs were stably expressed across the datasets. Hence, pre-analytical choices had variable effects on kidney-RNAs—analyzing kidney-RNAs complemented global correlation, which could fade differences in some relevant RNAs. Replicating prior DKD-marker results and discovery of candidate reference mRNAs encourages further uEV biomarker studies.</p>}}, author = {{Barreiro, Karina and Dwivedi, Om Prakash and Rannikko, Antti and Holthöfer, Harry and Tuomi, Tiinamaija and Groop, Per Henrik and Puhka, Maija}}, issn = {{2073-4425}}, keywords = {{diabetic kidney disease; exosomes; miRNA; mRNA; reference genes; sequencing; urinary extracellular vesicles; urine}}, language = {{eng}}, number = {{7}}, publisher = {{MDPI AG}}, series = {{Genes}}, title = {{Capturing the Kidney Transcriptome by Urinary Extracellular Vesicles—From Pre-Analytical Obstacles to Biomarker Research}}, url = {{http://dx.doi.org/10.3390/genes14071415}}, doi = {{10.3390/genes14071415}}, volume = {{14}}, year = {{2023}}, }