Abstract: PO2492
Multi-Omic Analysis of Mouse Renal Tubule Cell Responses Following Unilateral Nephrectomy
Session Information
- CKD: Metabolism, Epigenetics, and Signaling
November 04, 2021 | Location: On-Demand, Virtual Only
Abstract Time: 10:00 AM - 12:00 PM
Category: CKD (Non-Dialysis)
- 2103 CKD (Non-Dialysis): Mechanisms
Authors
- Kikuchi, Hiroaki, National Institutes of Health, Bethesda, Maryland, United States
- Knepper, Mark A., National Institutes of Health, Bethesda, Maryland, United States
Background
The kidney increases in size following resection of the contralateral kidney. Modern ‘-omics’ methods provide an opportunity to understand this response at a cellular level.
Methods
Experiments were done in mice after unilateral nephrectomy (UNx) or sham nephrectomy. MRI was used to measure kidney volume. The earliest portion of the kidney proximal tubule (PCT) and the cortical collecting duct (CCD) were microdissected at different time points (24 hours and 72 hours). Microdissected tubules were analyzed by quantitative immunofluorescence microscopy to determine cell size and number, and by RNA-seq to identify gene expression changes. Quantitative protein mass spectrometry was used to identify proteomic changes.
Results
Increased kidney volume was already detectable at the 24 hour-time point after UNx (versus sham), and was increased further at 72 hours. Morphometry of microdissected PCT and CCD, labeled with apical and basolateral markers and DAPI, revealed a marked increase in total cell volume per unit length, but no significant change in mean cell volume in both PCT and CCD, revealing that the increase in total cell volume was due to cellular proliferation rather than hypertrophy of individual cells. Consistent with this observation, RNA-Seq at 72 hours after surgery showed significant increases in the abundance of transcripts associated with cell cycle regulation in both segments (Gene Ontology enrichment analysis) such as Cdk1 and Cdc20 among many others. To identify the earliest signaling events, RNA-seq was employed at 24 hours after UNx revealing that in PCT, UNx produced upregulation of numerous transcripts associated with free fatty acid generation and sterol metabolism including many genes that are known targets of the transcription factor PPARa such as Angptl4, Acot1 and Cyp4a14. Protein mass spectrometry of whole kidneys at 24 hours, composed chiefly of proximal tubule cells, confirmed upregulation of many PPARa-regulated proteins such as HMGCS2, CYP4A14 and ANGPTL4.
Conclusion
Increased kidney size in response to UNx was due to cellular proliferation not hypertrophy of individual cells. Many lipid-metabolism related mRNAs were highly upregulated that predict increased free fatty acid levels in proximal tubule cells. Lipid mediators, including those derived from the fatty acid arachidonic acid, may be involved the cellular proliferation.
Funding
- Other NIH Support