(DCP016) NRF2 DELETION ATTENUATES LIPID DROPLET ACCUMULATION AND PROXIMAL TUBULAR INJURY VIA INHIBITION CD36 AND FABP4 EXPRESSION IN TYPE 2 DIABETIC (T2D) DB/DB MICE

Background: Lipid droplet deposition in the kidney induces oxidative stress, which can worsen kidney function in diabetes. Scavenger receptor CD36 and fatty acid binding protein 4 (FABP4) are highly expressed in renal proximal tubules (RPTs) in diabetes and mediate lipid deposition by uptake free-fatty acid (FFA). The role of nuclear factor erythroid 2-related factor 2 (Nrf2) in diabetic kidney disease (DKD) progression is controversial.
To investigate whether genetic deletion of Nrf2 could attenuate lipid droplet deposition and oxidative stress as well as ameliorate tubular injury in db/db mice via down-regulation of CD36 and FABP4 expression in RPTCs.

METHODS AND RESULTS: Db/db and db/db Nrf2 knockout (KO) mice at the age of 16 weeks and human RPTCs (HK2) with and without NRF2 KO by CRISPR-Cas9 genome editing were studied. Physiological and kidney morphological changes were assessed. Oil Red O staining was used to assess the lipid droplet accumulation in the kidney. DHE staining and Mitosox staining were performed to detect ROS and superoxide generation in RPTs and mitochondria, respectively. Western blot (WB) and real-time qPCR (RT-qPCR) were employed to assess protein and gene expression in isolated RPTs and HK2.

Results: Nrf2 deletion lowered fasting blood glucose and ameliorated tubular injury score in db/db Nrf2 KO mice vs db/db mice. Lipid droplet accumulation and ROS level were significantly increased in kidneys of db/db mice and attenuated in kidneys of db/db Nrf2 KO mice. Sodium-glucose cotransporter 2 (SGLT2), CD36 and FABP4 protein and gene expression were also significantly increased in RPTs of db/db mice and attenuated in db/db Nrf2 KO mice. In vitro, high glucose (HG, 35 mM D-glucose) and FFA (palmitate/oleate-BSA, 200 µM) acted synergistically enhancing the lipid droplet accumulation and mitochondria superoxide level in HK2 but their effects were attenuated in HK2 with NRF2 KO. Furthermore, HG and FFA stimulated SGLT2 and CD36 expression in HK2 but not in HK2 with NRF2 KO.

Conclusion: Deletion of Nrf2 in db/db mice attenuated hyperglycemia, ROS generation, renal tubular lipid droplet accumulation and kidney injury via inhibition of SGLT2, CD36 and FABP4 expression.