Wang H, Zhang Q, Chai Y, Liu Y, Li F, Wang B, Zhu C, Cui J, Qu H, Zhu M
Department of Endocrinology, Tianjin Medical University General Hospital, Tianjin, 300052, China.
Division of Epidemiology, Human Genetics and Environmental Sciences, University of Texas School of Public Health, Houston, TX, 77030, USA.
J Endocrinol Invest. 2015 Oct;38(10):1083-91. doi: 10.1007/s40618-015-0287-6. Epub 2015 Apr 24.
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)2D3 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)2D3 on diabetic liver injury in vivo.
Streptozotocin (STZ)-induced diabetic rats were randomly divided into five groups and treated with low-dose 1,25(OH)2D3 (0.025 μg/kg/day), medium-dose 1,25(OH)2D3 (0.15 μg/kg/day), high-dose 1,25(OH)2D3 (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)2D3 (10(-7) M), LPS + 1,25(OH)2D3, high fat + 1,25(OH)2D3, or LPS + high fat + 1,25(OH)2D3. 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.
High-dose 1,25(OH)2D3 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)2D3 decreased the expression of TLR4, NF-κB, and inflammatory factors (P < 0.05).
1,25(OH)2D3 exhibited protective effects against diabetes-related liver injury, possibly through downregulation of components of the TLR4 signaling pathway.
肝脏中的脂肪酸沉积可激活多种促炎信号通路，如Toll样受体4（TLR4）通路，这在非酒精性脂肪性肝炎的发病机制中可能起重要作用。1,25(OH)2D3可下调TLR4的表达，可能代表一种减轻肝细胞损伤的新治疗策略。因此，在本研究中，我们在体内研究了1,25(OH)2D3对糖尿病肝损伤的保护作用。
将链脲佐菌素（STZ）诱导的糖尿病大鼠随机分为五组，分别用低剂量1,25(OH)2D3（0.025μg/kg/天）、中剂量1,25(OH)2D3（0.15μg/kg/天）、高剂量1,25(OH)2D3（0.3μg/kg/天）、胰岛素（精蛋白锌胰岛素16U/kg/天，皮下注射）治疗，或不进行干预（对照组）。16周后，处死大鼠，采集血样检测血脂和肝功能。分析肝脏中炎症细胞浸润、纤维化程度以及TLR4、核因子-κB（NF-κB）和肿瘤坏死因子-α（TNF-α）的表达水平。肝细胞分别用溶剂对照、脂多糖（LPS，100ng）、高脂[DMEM+游离脂肪酸（FFA，0.1mM：棕榈酸、油酸，1:2）]、LPS+高脂、溶剂+1,25(OH)2D3（10⁻⁷M）、LPS+1,25(OH)2D3、高脂+1,25(OH)2D3或LPS+高脂+1,25(OH)2D3处理。提取RNA和蛋白质以检测TLR4及下游炎症因子如NF-ΚB、TNF-α和IL-6的表达。数据组间比较采用单因素方差分析。
高剂量1,25(OH)2D3给药16周可下调糖尿病大鼠肝组织中TLR4、NF-κB和TNF-α的表达，并减轻肝脏炎症和纤维化，免疫组织化学染色、苏木精和伊红染色、Masson三色染色、逆转录聚合酶链反应（RT-PCR）和蛋白质印迹法均显示了这一点。在体外，用高脂或LPS处理的肝细胞中TLR4、NF-κB和下游炎症因子的表达显著增加（P<0.05）。用1,25(OH)2D3干预可降低TLR4、NF-κB和炎症因子的表达（P<0.05）。
1,25(OH)2D3对糖尿病相关肝损伤具有保护作用，可能是通过下调TLR4信号通路的成分来实现的。
