Abstract: TH-PO271

Mutation of the RORγC Gene in Rats Attenuates Th17 Cell Activity, Renal Injury, and Inflammation in Response to Ischemia Reperfusion

Session Information

Category: Acute Kidney Injury

  • 001 AKI: Basic

Authors

  • Mehrotra, Purvi, Indiana School of Medicine, Indianapolis, Indiana, United States
  • Collett, Jason Andrieu, Indiana University School of Medicine, Indianapolis, Indiana, United States
  • Basile, David P., Indiana University School of Medicine, Indianapolis, Indiana, United States
Background

T cells have been implicated in initiation of renal damage following I/R injury, while recent studies have also suggested that a specific T helper subset, Th17 cells, is associated with both initiation of AKI and the AKI-to-CKD progression. To evaluate the direct role of Th17 cells in AKI, RoRgc (RAR orphan receptor gamma), a transcription factor considered the master regulator for Th17 cell differentiation and IL-17 gene expression was targeted for mutation in Lewis rats.

Methods

Using a CRISPR-CAS9 approach, an 8 bp deletion was introduced into the rat RoRgc gene. The verified sequence analysis predicted a mis-sense mutation at a.a.31 and premature truncation relative to the 508 a.a. protein in wild-type rats. LewisRORγT-/- rats appear normal and healthy but show moderate growth retardation relative Lewis RORγT +/+ control rats.

Results

Under basal conditions, reduced percentages of CD4+ (50±2.4 vs 24.45±1.7; p<0.05), B cells (21.4±7.2 vs 13.85±0.85; p<0.05) and Macrophages (13.07±3.5 vs 7.7±0.97; p<0.05) in the spleen of 6-8-week old LewisRORγT -/- rats compared to wildtype controls. To evaluate the effect of RORgC mutation on the induction of Th17 cells during AKI, both mutant and wild type rats were subjected to bilateral renal I/R for 40 min. Renal Th17 cells were enhanced 2 days following I/R in wild type rats but were significantly reduced in LewisRORγT -/- rats (63094±10498 vs 24434±4830; p<0.05) post I/R with no effect on Th1 and Th2 cells. In addition, LewisRORγT -/- were resistant to AKI as measured by serum creatinine (4.5±0.4 vs 2.6±0.4; p<0.05) as well as reduced renal inflammation as indicated by total CD4+ (40%; p<0.05) and CD8+ (48%; p<0.05). After 7 days of recovery, AKI primed lymphocytes from wild type rats manifested an IL-17 mRNA induction in response to Ang II +170 mM Na+ in vitro, however AKI primed lymphocytes from LewisRORγT -/- rats showed no increase in IL-17 mRNA expression.

Conclusion

Taken together these data that mutation of RORγC in rats impairs Th17 cell activity in response to renal I/R and modulates the sensitivity to the development of AKI.

Funding

  • NIDDK Support