To explore the potential molecular mechanism of astaxanthin (AST) in renal interstitial fibrosis and provide data for clinical prevention and treatment of renal fibrosis.

In animal experiment, unilateral ureteral ligation (UUO) was used to establish renal interstitial fibrosis model. 60 male C57BL/6 mice were randomly divided into six groups with 10 mice each, including sham operation group, sham operation+ AST 200mg/kg group, UUO group, UUO+ AST 50, 100, 200 mg/kg groups. In cell experiments, the HK-2 cell line epithelial-mesenchymal transition (EMT) model was induced by TGF-β1. Plasmid transfection experiments divided HK-2 cells into 7 groups: control group, blank group, CCN1 overexpression group, CCN1 knockdown group, TGF-β1 group, TGF-β1+ CCN1 overexpression group, and TGF-β1+ CCN1 knockdown group. The AST treatment experiments divided the HK-2 cells into 5 groups: control group, TGF-β1 group, TGF-β1+ AST group, TGF-β1+ CCN1 knockdown group, and TGF-β1+ CCN1 knockdown+ AST group. Real time-PCR was used to detect the biomarkers of kidney injury in kidney tissue: neutrophil gelatinase-associated lipocalin (NGAL), kidney injury molecule-1 (KIM-1), serum cystatin-C (Cys-C), and interleukin-18 (IL-18). Western blot was used to determine expression of fibrosis markers: type I collagen (Col-Ⅰ), fibronectin (FN), α-smooth muscle actin (α-SMA), vimentin, and E-cadherin (E-cad) in the cells supernatant and kidney tissues, as well as expression of TGF-β1/Smad and Notch signaling pathway molecules including pSmad3, Notch1, Notch3, and Jagged-1.

Compared with the sham operation group, the UUO group showed significant higher levels of kidney injury biomarkers as NGAL, KIM-1, Cys-C, and IL-18. The UUO group also displayed significant higher expression level of renal fibrosis marker proteins as well as lower expression level of E-cad. Besides, in the UUO group, the protein expression levels of pSmad3, Notch1, Notch3, and Jagged-1 also increased significantly, and the expression of Smad7 decreased significantly. AST treatment significantly decreased the expression of biomarkers of renal injury (NGAL, KIM-1, Cys-C and IL-18) and protein expression of fibrosis markers, but increased the expression of E-cad protein in a dose-dependent manner. The cell experiments showed that overexpression of CCN1 inhibited the activation of TGF-β1/Smad and Notch signaling pathways, while knocking down CCN1 not only intensified the activation of TGF-β1/Smad and Notch signaling pathways, but also weakened the protective effect of AST in the EMT model induced by TGF-β1.

AST may regulate TGF-β1/Smad and Notch signaling pathways through CCN1, so as to reduce renal interstitial fibrosis and play a role in renal protection.