Abstract: PO1973
RNA Sequencing and ATAC-Seq Reveal Gene Profiles in Injured Podocytes in Mice, and Podocyte-Specific Hypoxia Inducible Factor 2α Deletion Protects from Adriamycin-Induced Podocyte Injury
Session Information
- Podocyte Biology
October 22, 2020 | Location: On-Demand
Abstract Time: 10:00 AM - 12:00 PM
Category: Glomerular Diseases
- 1204 Podocyte Biology
Authors
- Liang, Xiaoyan, Ann and Robert H Lurie Children's Hospital of Chicago, Chicago, Illinois, United States
- Schnaper, H. William, Northwestern University Feinberg School of Medicine, Chicago, Illinois, United States
- Hayashida, Tomoko, Northwestern University Feinberg School of Medicine, Chicago, Illinois, United States
Background
Roles for hypoxia-inducible factor (HIF) in kidney diseases have been controversial. Upregulating HIF expression is beneficial in protecting kidney from acute injury, while we and others showed that HIF aggravates chronic fibrosis. In podocytes, deletion of VEGFR caused progressive glomerular damage, prevented by overexpression of hypoxia inducible factor 2α.
Here, using podocyte-specific HIF-2α deletion mice, we tested effects of HIF-2a deletion on gene profiles in injured podocytes.
Methods
HIF-1αf/f or HIF-2αf/f mice were crossed with either NPHS1 or 2-Cre mice. HIF-2αf/f or WT; NPHS1/2-Cre mice were further crossed with Z/EG mice that express GFP only in cells where Cre is active. At 8 weeks old, the mice were given Adriamycin (12 mg/kg BW, i.v.). Two weeks later, kidneys, urine and blood were harvested. Creatinine and albumin levels in urine and serum, and blood urine urea levels in serum were measured with ELISA kits. Podocytes were isolated from HIF-2αf/f or WT; NPHS2-Cre; Z/EG mice by flor sorting, and RNAseq and ATACseq were performed.
Results
HIF-2αf/f; NPHS2-Cre showed preservation of foot processes and kidney functions, and significantly less proteinuria compared to the WT littermates after being subjected to Adriamycin, while HIF-1α f/f; NPHS2-Cre mice developed a similar degree of proteinuria to that of WT. HIF-1 or 2f/f; NPHS1-Cre mice did not show any protective effect. HIF-1 or 2αf/f; NPHS1-Cre; Z/EG mice showed little GFP expression in podocytes, suggesting that weak penetrance of NPHS1-Cre led to minimal functional effects. This group was therefore not further evaluated. Podocytes isolated from HIF-2αf/f or WT; NPHS2-Cre; Z/EG were subjected to RNAseq and ATACseq. In RNAseq, Ndufa12 [NADH:ubiquinone oxidoreductase subunit A12] was most significantly upregulated in podocytes subjected to ADR, while Slc22a30 [solute carrier family 22, member 30], Slc7a13 [solute carrier family 7, (cationic amino acid transporter, y+ system) member 13] and Pzp [PZP, alpha-2-macroglobulin like] were most significantly downregulated. Correlation with ATAC-seq results are being processed.
Conclusion
HIF-2α deletion in podocytes protects podocyte from acute adriamycin injury. Novel genes associated with podocyte injury were discovered by RNAseq and ATACseq of podocytes.
Funding
- NIDDK Support