The improvement of homocysteine-induced myocardial inflammation by vitamin D depends on activation of NFE2L2 mediated MTHFR.

Objectives: Myocardial inflammation underlies a broad spectrum of conditions that cause damage to the myocardium and lead to structural and functional defects. Homocysteine (Hcy) is closely related to the occurrence and development of cardiovascular diseases. We investigated the mechanism underlying the effects of vitamin D as a prophylactic treatment for Hcy-induced cardiac inflammation.

Methods: The levels of 25(OH)D and Hcy were assessed using ELISA kits. Expression levels of the vitamin D receptor (VDR), NFE2 like bZIP transcription factor 2 (NFE2L2), methylenetetrahydrofolate reductase (MTHFR) and inflammatory factors were examined by Western blotting, immunohistochemistry and real time polymerase chain reaction. NFE2L2/MTHFR-knockdown HL-1 cells and NFE2L2 mouse were used to test the effects of vitamin D.

Results: We found the levels of Hcy in the serum and myocardial tissue of mice in the Hcy + CCE group were lower than in the Hcy groups, which was opposed to the trend exhibited by the serum 25(OH)D level of mice. The mRNA and protein expression levels of the inflammatory factors in cardiac tissues and cardiomyocytes were strongly decreased by the Hcy treatment, compared to the Hcy + CCE/Hcy + 1,25(OH)D groups. Moreover, the results revealed that the level of nuclear NFE2L2 in Hcy + CCE/Hcy + 1,25(OH)D group was increased compared to Hcy group with a reciprocal decrease in the level of cytosolic NFE2L2 in vivo and in vitro. Similarly, the MTHFR mRNA and protein expression in the Hcy + CCE group was higher than the Hcy group. We determined that NFE2L2 promoted the expression of MTHFR. However, based on Hcy treatment, the combination of 1,25(OH)2D3 and MTHFR reversed the decline in IL-6 and TNFα expression caused by 1,25(OH)2D3 alone. Chromatin immunoprecipitation and luciferase reporter assays showed the up-regulation effect of VDR on NFE2L2 and NFE2L2 on MTHFR.

Conclusions: Our findings indicate that vitamin D/VDR could improve Hcy-induced myocardial inflammation through activation of NFE2L2 mediated MTHFR.