RÉSUMÉ
BACKGROUND:In recent years,the incidence of hyperuricemia caused by purine metabolism disorders has been increasing,which can induce inflammatory responses and lead to renal injury. OBJECTIVE:To explore the role and mechanism of solute carrier family 2 member 12(SLC2A12)in hyperuricemia-related renal injury. METHODS:Renal tubular cells(HK2 cells)were divided into five groups:HK2 group,HK2+uric acid group,HK2+uric acid+NC group,HK2+uric acid+siSLC2A12 group,and HK2+uric acid+siSLC2A12+MK-2206 group.HK2 cells were treated with uric acid and transfected with siRNA SLC2A12,followed by MK-2206 treatment to inhibit AKT expression.Cell proliferation was detected by CCK-8 assay.Apoptosis was detected by TUNEL assay.qRT-PCR and western blot assay were used to detect fibrogenic factors as well as activation of the AKT/FOXO3a pathway.The concentrations of inflammatory cytokines were measured by enzyme-linked immunosorbent assay. RESULTS AND CONCLUSION:(1)Uric acid treatment inhibited cell proliferation and promoted cell apoptosis in the HK2+uric acid group compared with the HK2 group.The proliferative ability of cells in the HK2+uric acid+siSLC2A12 group was further decreased and apoptotic cells were further increased compared with the HK2 group.Compared with the HK2+uric acid+siSLC2A12 group,the HK2+uric acid+siSLC2A12+MK-2206 group showed an increase in cell proliferation and a decrease in apoptotic cells.(2)Compared with the HK2 group,the connective tissue growth factor(CTGF),α-smooth muscle actin(α-SMA)and transforming growth factor beta(TGF-β)expressions increased in the HK2+uric acid group;CTGF,α-SMA and TGF-β expression further increased in the HK2+uric acid+siSLC2A12 group.Compared with the HK2+uric acid+siSLC2A12 group,the CTGF,α-SMA and TGF-β expressions decreased.(3)Compared with the HK2 group,the expression of p-AKT,FOXO3a,and p-FOXO3a elevated in the HK2+uric acid group;the expression of p-AKT further increased,while the expression of FOXO3a and p-FOXO3a decreased in the HK2+uric acid+siSLC2A12 group.Compared with the HK2+uric acid+siSLC2A12 group,p-AKT expression decreased;FOXO3a and p-FOXO3a expression increased in the HK2+uric acid+siSLC2A12+MK-2206 group.(4)Compared with the HK2 group,interleukin-6,interleukin-1 β,and tumor necrosis factor α levels increased in the HK2+uric acid group;interleukin-6,interleukin-1 β,and tumor necrosis factor α levels further increased in the HK2+uric acid+siSLC2A12 group.Compared with the HK2+uric acid+siSLC2A12 group,interleukin-6,interleukin-1 β,and tumor necrosis factor α levels diminished in the HK2+uric acid+siSLC2A12+MK-2206 group.(5)These findings indicate that SLC2A12 may protect against hyperuricemia-induced renal injury by counteracting uric acid-induced tubular fibrosis and inflammation through activation of the FOXO3a pathway.
RÉSUMÉ
Objective:To observe the effect of sodium fluoride (NaF) on the expression of phosphatidylinositol-3-kinase/protein kinase B (PI3K/Akt) signal and apoptosis in human osteosarcoma Saos-2 cells.Methods:Saos-2 cells were divided into 0 (control), 5, 10, 20 and 40 mg/L fluoride groups ( n= 3) according to the dose of NaF. The cells were collected after 24 and 48 h of culture in vitro. Western blotting was used to detect the expressions of PI3K/Akt signal and Forkhead transcription factor (FoxO) 1, and the apoptosis level of Saos-2 cells was detected by flow cytometry. Results:There were no significants differences in the expressions of PI3K and Akt in Saos-2 cells at 24 and 48 h ( P > 0.05). At 24 h, the expressions of phosphorylated PI3K (p-PI3K) in 5, 10, 20 and 40 mg/L fluoride groups (0.40 ± 0.06, 0.45 ± 0.02, 0.37 ± 0.06, 0.32 ± 0.06) were higher than that in the control group (0.28 ± 0.04, P < 0.05); at 48 h, the expressions of p-PI3K in 5 and 10 mg/L fluoride groups (0.46 ± 0.06, 0.58 ± 0.03) were higher than that in the control group (0.29 ± 0.04, P < 0.05), and that in the 40 mg/L fluoride group (0.21 ± 0.03) was lower than that in the control group ( P < 0.05). At 24 h, the expressions of phosphorylated Akt (p-Akt) in 5, 10 and 20 mg/L fluoride groups (0.27 ± 0.01, 0.30 ± 0.03, 0.27 ± 0.03) were higher than that in the control group (0.20 ± 0.02, P < 0.05); at 48 h, the expressions of p-Akt in 5 and 10 mg/L fluoride groups (0.42 ± 0.04, 0.60 ± 0.02) were higher than that in the control group (0.27 ± 0.01, P < 0.05), and that in the 40 mg/L fluoride group (0.18 ± 0.02) was lower than that in the control group ( P < 0.05). The expressions of FoxO1 in 10, 20 and 40 mg/L fluoride groups at 24 h (0.38 ± 0.07, 0.41 ± 0.06, 0.47 ± 0.08) were higher than that in the control group (0.34 ± 0.04, P < 0.05). At 48 h, the expressions of FoxO1 in 5, 10, 20 and 40 mg/L fluoride groups (0.36 ± 0.08, 0.37 ± 0.10, 0.54 ± 0.04, 0.73 ± 0.04) were higher than that in the control group (0.28 ± 0.04, P < 0.05). At 24 and 48 h, the apoptosis rates of control group and 5, 10, 20 and 40 mg/L fluoride groups were (2.867 ± 0.583)%, (3.647 ± 0.035)%, (3.773 ± 0.095)%, (5.440 ± 0.325)%, (7.203 ± 0.476)%; (3.707 ± 0.286)%, (4.023 ± 0.241)%, (4.970 ± 0.368)%, (12.473 ± 0.949)%, (19.387 ± 1.892)%, respectively. The apoptosis level of 40 mg/L fluoride group was higher than that of control group at 24 h ( P < 0.05), and that of 20 and 40 mg/L fluoride groups were higher than that of control group at 48 h ( P < 0.05). Conclusion:Fluoride can directly activate the PI3K/Akt signaling pathway in osteoblasts, and then has anti-apoptotic effect.