Abstract: FR-PO254
Sources and Carriers of Circulating MicroRNAs Associated with ESKD in Diabetes
Session Information
- Diabetic Kidney Disease: Basic - 1
October 25, 2024 | Location: Exhibit Hall, Convention Center
Abstract Time: 10:00 AM - 12:00 PM
Category: Diabetic Kidney Disease
- 701 Diabetic Kidney Disease: Basic
Authors
- Satake, Eiichiro, Joslin Diabetes Center, Boston, Massachusetts, United States
- Krolewski, Bozena, Joslin Diabetes Center, Boston, Massachusetts, United States
- Md Dom, Zaipul, Joslin Diabetes Center, Boston, Massachusetts, United States
- Hartley, Maryalice, Eli Lilly and Company, Indianapolis, Indiana, United States
- Willency, Jill, Eli Lilly and Company, Indianapolis, Indiana, United States
- Niewczas, Monika A., Joslin Diabetes Center, Boston, Massachusetts, United States
- Wilson, Jonathan Matthew, Eli Lilly and Company, Indianapolis, Indiana, United States
- Duffin, Kevin L., Eli Lilly and Company, Indianapolis, Indiana, United States
- Krolewski, Andrzej S., Joslin Diabetes Center, Boston, Massachusetts, United States
Background
Our previous study identified 17 (8 risk and 9 protective) circulating miRNAs associated with end-stage kidney disease (ESKD) in diabetes (Satake et al, JASN 2021). The current study investigated tissues/sources and carriers of these miRNAs.
Methods
We measured 2,083 human mature miRNAs in plasma and urine obtained from patients enrolled in the Joslin Kidney Study using the HTG EdgeSeq platform, which requires a small sample volume (15 μl) and does not require RNA extraction. Seventeen miRNAs associated with ESKD from these studies were quantified in blood exosomes, urine, lysates, and supernatants of HUVECs (human umbilical vein endothelial cells), RPTECs (renal proximal tubular epithelial cells), and fibroblasts to identify potential sources of miRNA expression and secretion. Plasma from patients was fractionated by FPLC and by size exclusion spin filters to help identify miRNA carriers in blood.
Results
Out of 17 ESKD miRNAs, none were derived from platelets. Risk miRNAs were predominantly found in plasma rather than cell components, while protective miRNAs were more abundant in cellular components. Several risk miRNAs were more abundant in urine than in plasma; most protective ones were abundant in plasma and almost absent in urine. In vitro experiments showed that risk miRNAs were more abundant in supernatants than in cell lysates in HUVECs and RPTECs, but not in fibroblasts. Regarding carriers, most ESKD miRNAs had significantly higher levels in plasma than in exosomes. Size exclusion chromatography identified that ESKD miRNAs were more abundant in the plasma free fraction that included smaller proteins than in lipoprotein particle fractions.
Conclusion
Our findings suggest that endothelial and proximal tubular cells could be a significant source of circulating ESKD miRNAs, and most of these miRNAs were bound to non-exosomal carriers that included proteins smaller than HDL cholesterol. It is important to understand the regulation of circulating ESKD miRNAs secretion, trafficking, and function to advance our understanding of the mechanisms of the progression of diabetic kidney disease to ESKD, so new therapeutic strategies can be developed.
Funding
- NIDDK Support