Tulk Sarah E, Liao Kuo-Chieh, Muruve Daniel A, Li Yan, Beck Paul L, MacDonald Justin A
Department of Biochemistry & Molecular Biology, University of Calgary, 3280, Hospital Drive, NW, Calgary, AB, T2N 4Z6, Canada.
J Cell Biochem. 2015 May;116(5):711-20. doi: 10.1002/jcb.24985.
Vitamin D3 has emerged as an important regulator of the immune system. With metabolic enzymes for vitamin D3 activation and vitamin D receptors (VDR) now identified in a variety of immune cells, the active vitamin D3 metabolite 1,25(OH)2D3, is thought to possess immunomodulatory properties. We examined whether 1,25(OH)2D3 might also enhance the NLRP3-dependent release of mature IL-1β from macrophages. PMA-differentiated THP-1 cells were stimulated with vitamin D3 metabolites and assessed for CYP27, CYP24, NLRP3, ASC, pro-caspase-1 expression by western blot and real-time qPCR as well as inflammasome activation with pro-inflammatory cytokine IL-1β release measured by ELISA. Exposure to 1,25(OH)2D3 had no effect on the basal expression levels of VDR; however, CYP27A1 transcript was suppressed and CYP24A1 transcript was substantively elevated. Both 1,25(OH)2D3 - and 25(OH)D3 induced IL-1β release from THP-1 cells, and these effects were blocked with application of the caspase-1 inhibitor YVAD and the NLRP3 inhibitors glyburide and Bay 11-7082. Interestingly, 1,25 (OH)2D3 exposure reduced NLRP3 protein expression but had no effect on ASC or pro-caspase-1 protein levels. The increase in mature IL-1β elicited by 1,25(OH)2D3 was modest compared to that found for ATP or C. difficile toxins. However, co-treatment of THP-1 cells with ATP and 1,25(OH)2D3 resulted in more IL-1β secretion than ATP or 1,25(OH)2D3 alone.
维生素D3已成为免疫系统的重要调节因子。随着现已在多种免疫细胞中鉴定出维生素D3激活代谢酶和维生素D受体（VDR），活性维生素D3代谢物1,25(OH)2D3被认为具有免疫调节特性。我们研究了1,25(OH)2D3是否也可能增强巨噬细胞中依赖NLRP3的成熟IL-1β释放。用维生素D3代谢物刺激经佛波酯（PMA）分化的THP-1细胞，并通过蛋白质印迹法、实时定量PCR评估CYP27、CYP24、NLRP3、ASC、前半胱天冬酶-1的表达，以及通过酶联免疫吸附测定（ELISA）测量促炎细胞因子IL-1β释放来评估炎性小体激活。暴露于1,25(OH)2D3对VDR的基础表达水平没有影响；然而，CYP27A1转录本受到抑制，CYP24A1转录本显著升高。1,25(OH)2D3和25(OH)D3均诱导THP-1细胞释放IL-1β，并且这些作用被半胱天冬酶-1抑制剂YVAD以及NLRP3抑制剂格列本脲和Bay 11-7082阻断。有趣的是，暴露于1,25(OH)2D3会降低NLRP3蛋白表达，但对ASC或前半胱天冬酶-1蛋白水平没有影响。与ATP或艰难梭菌毒素相比，1,25(OH)2D3引起的成熟IL-1β增加幅度较小。然而，THP-1细胞用ATP和1,25(OH)2D3共同处理导致的IL-1β分泌比单独使用ATP或1,25(OH)2D3更多。
