Deficiency of RNA helicase DDX5 alleviates renal interstitial fibrosis in mice with unilateral ureteral obstruction
10.3760/cma.j.cn441217-20240703-00709
- VernacularTitle:RNA解旋酶DDX5缺失改善单侧输尿管梗阻小鼠肾间质纤维化
- Author:
Xiaofei WANG
1
;
Lu ZHANG
1
;
Huiming WANG
1
Author Information
1. 武汉大学人民医院肾内科，武汉 430060
- Publication Type:Journal Article
- Keywords:
Kidney tubules, proximal;
Cell cycle;
Ureteral obstruction;
Renal interstitial fibrosis;
RNA helicase DDX5
- From:
Chinese Journal of Nephrology
2025;41(4):266-275
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To investigate the role of RNA helicase DDX5 in renal interstitial fibrosis and its potential mechanism, and provide possible molecular target for the diagnosis and treatment of renal interstitial fibrosis.Methods:The animal and cell models of unilateral ureteral obstruction (UUO) and DDX5-pharmacological and genetic blocking were established. Twenty-four male mice aged 6-8 weeks and weighting about 18-22 g were divided into sham operation group, UUO group, sham operation+RX5902 group and UUO+RX5902 group by random number table method, with 6 mice in each group. Supinoxin (RX5902, a phosphorylated DDX5 inhibitor, 75 mg/kg) was administered by gavage in mice in the sham operation+RX5902 group and UUO+RX5902 group on day 0 and day 3 after the operation, respectively. The mice were sacrificed to collect kidney specimens one week after the surgery. The human renal tubular epithelial cell line (HK-2 cells) was routinely cultured and divided into control group, scramble siRNA group, DDX5 siRNA group, TGF-β1 group, scramble siRNA+TGF-β1 group and DDX5 siRNA +TGF-β1 group. The dosage of RX5902 was 20 nmol/L, and the dosage of transforming growth factor-β1 (TGF-β1) was 10 ng/ml. HE staining and Masson staining were used to evaluate renal tubule injury and collagen fiber deposition. Western blotting, immunofluorescence and immunohistochemistry were used to detect the protein expression levels of DDX5 and fibrosis-related indexes such as fibronectin and α-smooth muscle actin (α-SMA). The flow cytometry was used to detect the cell cycle. Results:In comparison to sham operation group, UUO group exhibited significantly elevated protein expression levels of fibronectin, α-SMA and DDX5 in kidney tissues (all P<0.05), accompanied by notable nuclear translocation of DDX5. TGF-β1 stimulation facilitated the upregulation of fibronectin, α-SMA, and DDX5 expression, as well as DDX5 nuclear translocation in HK-2 cells (all P<0.05). Pharmacological and genetic inhibition of DDX5 attenuated UUO or TGF-β1-induced elevated protein levels of fibronectin and α-SMA. Compared with scramble siRNA+TGF-β1 group, DDX5 siRNA+TGF-β1 group exhibited reduced protein expression levels of fibronectin, α-SMA and DDX5 in HK-2 cells (all P<0.05). Compared with UUO group, UUO+RX5902 group showed alleviation of renal epithelial cell exfoliation, renal tubule atrophy or lumen dilation and decreased scores of both tubular injury and interstitial collagen deposition, along with reduced protein expression levels of fibronectin, α-SMA and DDX5 (all P<0.05). Similarly, RX5902 inhibited TGF-β1-induced fibrotic responses in HK-2 cells (all P<0.05). Flow cytometry analysis results revealed that TGF-β1 stimulation resulted in a significant increase in the proportion of HK-2 cells in the G2/M phase. Conversely, silencing of DDX5 led to a significant reduction in the proportion of HK-2 cells in the G2/M phase (all P<0.05). Conclusions:DDX5 is significantly elevated in the renal tissues of UUO mice and TGF-β1-induced HK-2 cells. The pharmacological and genetic blockade of DDX5 can delay renal fibrosis by reducing cell cycle arrest and alleviating cell damage.
