Empagliflozin protected kidney function in CKD rat through suppressing hypoxic and fibrotic signalings mediated inflammation and EMT.

Abstract

BACKGROUND: This study tested the hypothesis that PI3K/Akt/GSK3&#x3b2; and TGF-&#x3b2;/Smad2/3 signaling play essential roles in mediating the epithelial-mesenchymal transition (EMT) and fibrosis, resulting in the deterioration of renal function and parenchyma in chronic kidney disease (CKD) rats, which is reversed by early empagliflozin treatment. METHODS AND RESULTS: NRK-52E cells were divided into the A1 (NRK-52E), A2 (NRK-52E + 200 &#x3bc;M p-Cresol), A3 (NRK-52E + 200 &#x3bc;M p-Cresol + 50 &#x3bc;M empagliflozin), B1 (NRK-52E), B2 (NRK-52E + 5 ng/mL TGF-&#x3b2;1) and B3 (NRK-52E + 5 ng/mL TGF-&#x3b2;1 + 50 &#x3bc;M empagliflozin) groups. Compared with those in the A1 group, the expression levels of proteins related to the EMT (TGF-&#x3b2;1/p-Smad2/p-Smad3/&#x3b1;-SMA), extracellular matrix (MMP2/9) and EMT (IGF-1) activators were significantly higher in the A2 group, but these changes were significantly reversed in the A3 group, whereas the protein expression levels of antifibrotic markers (TIMP1/TIMP2) exhibited the opposite pattern to the EMT-related proteins among the groups (all<0.001). The expression of these proteins, along with the other EMT markers (snail, fibronectin, and vimentin) related to cellular function/protein expression, also exhibited an identical pattern to the A1 to A3 groups among Groups B1 to B3 (all<0.001). Adult male SD rats were categorized into Groups 1 (sham-operated control), 2 (CKD) and 3 (CKD + empagliflozin). On Day 56 after CKD induction, the renal artery resistive index (RARI) was significantly higher in Group 2 than in Groups 1 and 3 and significantly higher in Group 3 than in Group 1 (all<0.0001). The expression of EMT (Snail/&#x3b1;-SMA/fibronectin/vimentin/TGF-&#x3b2;1/p-Smad2/3), apoptotic (cleaved caspase-3/cleaved-PARP), inflammatory (HIF-1&#x3b1;/IL-1&#x3b2;/TNF-&#x3b1;/MPO/MMP-2/MMP-9), and cell stress signaling (p-PI3K/p-Akt/GSK-3&#x3b2;) proteins and the cellular kidney injury score, expression of fibrosis and EMT markers (Snail/vimentin)/glomerular-hypercellularity/fibrocellular crescent formation displayed an identical pattern, whereas the cellular expression of podocyte components (podocin/synaptopodin/ZO-1) displayed the opposite pattern to the RARIs among the groups (all<0.0001). CONCLUSIONS: Empagliflozin protected kidney function and architecture mainly by suppressing fibrosis, cellular oxidative stress signaling, the EMT and inflammation.