Abstract: FR-PO145
Analysis of the Peritubular Capillaries and Lymphatics in a Model of Renal Injury and Repair
Session Information
- Molecular Mechanisms of CKD - II
October 26, 2018 | Location: Exhibit Hall, San Diego Convention Center
Abstract Time: 10:00 AM - 12:00 PM
Category: CKD (Non-Dialysis)
- 1903 CKD (Non-Dialysis): Mechanisms
Authors
- Monaghan, Marie-Louise, University of Edinburgh, Edinburgh, United Kingdom
- Cairns, Carolynn, University of Edinburgh, Edinburgh, United Kingdom
- Borthwick, Gary, University of Edinburgh, Edinburgh, United Kingdom
- Denby, Laura, University of Edinburgh, Edinburgh, United Kingdom
- Hughes, Jeremy, University of Edinburgh, Edinburgh, United Kingdom
- Conway, Bryan, University of Edinburgh, Edinburgh, United Kingdom
Background
Chronic kidney disease has a diverse range of aetiologies, but all are associated with alterations in the renal interstitial microvasculature. We hypothesised that renal injury promotes loss of peritubular capillaries leading to tubular cell hypoxia, but conversely lymphatic vessel density may increase as a compensatory response to reduce inflammation and facilitate repair.
Methods
C57Bl/6 mice underwent unilateral ureteric obstruction (UUO) or sham surgery. After 7 days of UUO, 8 mice were culled while in the remaining mice the ureter was re-implanted into the bladder to relieve obstruction (R-UUO) for 1, 2 and 4 weeks (n=6-8). Kidneys were harvested and analysed by flow cytometry, immunostaining, RNAseq, qPCR and electron microscopy.
Results
Flow cytometry demonstrated a 16-fold increase in the proportion of dead DAPI+/CD31+ endothelial cells (ECs) in UUO kidney v sham (p=<0.001). CD31 quantification by IHC revealed a doubling in areas of capillary rarefaction in UUO v sham (p=<0.001), with the degree of rarefaction falling by 31% following R-UUO (p=<0.001). Electron microscopy analysis demonstrated a reduction in peritubular capillary fenestrations and an increase in EC vacuolation, which partially reversed following R-UUO. While the density of Lyve-1+ lymphatic vessels remained similar in UUO kidney v sham, there was a doubling of lymphatic vessels after 4 weeks of relief of obstruction (p<0.05). RNAseq and qPCR analysis of renal cortex revealed a reduction in Vegfa in UUO v sham, which partially reversed following R-UUO. Conversely there was a significant up-regulation of Vegfc and Vegfd, most notably following R-UUO. Gene expression of Vegfr3 increased in UUO compared with sham (p<0.01), and this persisted through to week 4 after R-UUO. Dual immunofluorescence for VEGFR3 and LYVE-1 determined that the increase in VEGFR3 staining occurred largely in CD31+/Lyve1- vessels, most likely peritubular capillaries. This was confirmed by flow cytometry with the proportion of CD31+/podoplanin- cells that co-expressed VEGFR3 increasing from 7% in sham to 36% in UUO kidney.
Conclusion
Our data suggest that peritubular capillary rarefaction occurs following renal injury and only partially improves following cessation of injury. Therapies that prevent capillary rarefaction may reduce secondary renal injury and augment repair.
Funding
- Private Foundation Support