Inhibition of Xanthine Oxidase Protects against Sepsis-Induced Acute Kidney Injury by Ameliorating Renal Hypoxia

Figure 4

Hypoxia increased LPS-induced XO activity in HK-2 cells, but the inhibition of XO also improved hypoxia. (a) CCK8 showing the viability of HK-2 cells treated with LPS (10 μg/ml)±febuxostat (100 μM) under normoxic (21% O₂) and hypoxic (2% O₂) conditions for 6 h. (b) The XO activity of HK-2 cells treated with LPS±febuxostat under normoxic and hypoxic conditions for 6 h. (c, d) The intracellular hypoxia of HK-2 cells treated with LPS±febuxostat under normoxic and hypoxic conditions for 6 h was detected by an immunofluorescence assay using a hypoxia probe (800x magnification). (e, f) Knockdown of XO in HK-2 cells was confirmed using western blotting. (g) After the knockdown of XO with siRNA transfection, HK-2 cells were stimulated with LPS with 21% or 2% oxygen. Six hours later, the cells were harvested, and cell viability was assessed with the CCK8 method. (h) XO activity was assayed with the same method as in (b). (i, j) The intracellular hypoxia of HK-2 cells treated with LPS under normoxic and hypoxic conditions for 6 h was detected by an immunofluorescence assay using a hypoxia probe (800x magnification).  μm. , , and vs. control; ^#, ^##, and ^### vs. LPS+Veh or LPS+Ctr-siR (). Ctrl: control; Veh: vehicle; LPS: lipopolysaccharide; Feb: febuxostat; XO: xanthine oxidase.

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