1,25(OH)(2)D(3) downregulates the Toll-like receptor 4-mediated inflammatory pathway and ameliorates liver injury in diabetic rats

BACKGROUND: Fatty acid deposition in the liver can activate a number of pro-inflammatory signaling pathways such as the Toll-like receptor 4 (TLR4) pathway, which may be important in the pathogenesis of nonalcoholic steatohepatitis. 1,25(OH)(2)D(3) downregulates the expression of TLR4 and may represent a novel treatment strategy for reducing hepatocyte injury. Therefore, in this study, we investigated the protective effects of 1,25(OH)(2)D(3) on diabetic liver injury in vivo. METHODS: Streptozotocin (STZ)-induced diabetic rats were randomly divided into five groups and treated with low-dose 1,25(OH)(2)D(3) (0.025 μg/kg/day), medium-dose 1,25(OH)(2)D(3) (0.15 μg/kg/day), high-dose 1,25(OH)(2)D(3) (0.3 μg/kg/day), insulin (protamine zinc insulin 16 U/kg/day, subcutaneous injection), or no intervention (the control group). Sixteen weeks later, the rats were killed, and blood samples were obtained to test lipid profiles and hepatic function. The infiltration of inflammatory cells, the level of fibrosis, and the expression levels of TLR4, nuclear factor-kappa B (NF-κB), and tumor necrosis factor-α (TNF-α) in the liver were analyzed. The hepatocytes were treated with vehicle control, LPS (100 ng), high fat [DMEM + FFA (0.1 mM: palmitic acid, oleic acid, 1:2)], LPS + high fat, vehicle + 1,25(OH)(2)D(3) (10(−7) M), LPS + 1,25(OH)(2)D(3), high fat + 1,25(OH)(2)D(3), or LPS + high fat + 1,25(OH)(2)D(3). RNA and protein were extracted to detect the expression of TLR4 and downstream inflammatory factors such as NF-ΚB, TNF-α, and IL-6. Groups of data were compared by single factor variance analysis. RESULTS: High-dose 1,25(OH)(2)D(3) administration for 16 weeks downregulated the expression of TLR4, NF-κB, and TNF-α in the liver tissue of diabetic rats and attenuated hepatic inflammation and fibrosis, as shown by immunohistochemical staining, hematoxylin and eosin staining, Masson’s trichrome staining, reverse transcription polymerase chain reaction (RT-PCR), and western blotting. In vitro, hepatocytes treated with high fat or LPS exhibited significantly increased expression of TLR4, NF-κB, and downstream inflammatory factors (P < 0.05). Intervention with 1,25(OH)(2)D(3) decreased the expression of TLR4, NF-κB, and inflammatory factors (P < 0.05). CONCLUSIONS: 1,25(OH)(2)D(3) exhibited protective effects against diabetes-related liver injury, possibly through downregulation of components of the TLR4 signaling pathway.