| 報告時間：2025-6-20 |
| 報告地點：Room 407 |
| 指導老師：Shuen-Ei Chen |
| 學生：YING-TENG CHEN |
| 摘要 |
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In essence, vitamin D functions as a steroid hormone precursor with multiple functions. It plays a critical role in calcium and phosphorus absorption and metabolism, significantly influencing bone formation. In recent years, it has also been implicated in immune regulation. However, past reports showed distinctive results and divergent conclusions and perspectives on the its immunomodulatory effects. Promoters are DNA sequences located at the upstream of genes to initiate transcription and regulate downstream gene expressions. Genes regulated by vitamin D typically contain a consensus sequence known as the vitamin D response element (VDRE) within their promoter regions. VDRE is a hexameric direct repeat with a specific spacing that can be recognized by a heterodimer of the vitamin D receptor (VDR) and the retinoid X receptor (RXR). Upon ligand binding, this complex activates transcription, and the strength of transcriptional activation depends on ligand affinity, which is influenced by conformational changes in the VDR. This study hypothesizes that various vitamin D compounds with different transcriptional activities may lead to distinct immunoregulatory outcomes. Therefore, THP-1 monocytic cells were used to investigate vitamin D–mediated regulation of cytokines. Meanwhile, DNA fragments containing VDRE from regulatory gene regions were constructed into Luciferase reporter vector and transfected into HEK293 kidney cells to assess transcriptional activity. The results demonstrated that in vitro, kidney cells were capable of converting 25-hydroxyvitamin D₃ (25(OH)D₃) into 1,25-dihydroxyvitamin D₃ (1,25(OH)₂D₃). At 6 and 24 hour post-treatment, 1,25(OH)₂D₃ at concentrations above 100nM significantly enhanced luciferase reporter activity. In contrast, 25(OH)D₃ showed no significant activation at 6 hours, but the doses ≥500nM at 24 and 48 hours yielded significant increases in transcriptional activity. However, nuclear VDR abundance were significantly decreased after treatment with 1,25(OH)₂D₃ at 1, 6, 24, and 48 hours. 25(OH)D₃ treatment showed no significant effects on nuclear VDR abundance despite a downward trend. Under the same conditions, both forms of vitamin Ds showed an increased trend of total VDR protein, but not significant. In THP-1 monocytic cells, 1,25(OH)₂D₃ alone did not promote IL-1β expression. However, when combined with LPS stimulation, 1,25(OH)₂D₃ enhanced IL-1β expression in a dose-dependent manner, even under identical LPS concentrations. This effect may be attributed to vitamin D–mediated enhancement of CD14 receptor activity. CD14 directly binds LPS and facilitates its delivery to the TLR4/MD-2 signaling complex, playing a crucial role in sensitizing cells to low-dose LPS. As for 25(OH)D₃, pre-treatment with LPS was necessary to elicit an immunomodulatory effect. Under these conditions, expression of 1α-hydroxylase (CYP27B1) was significantly upregulated 18 hours post-LPS treatment, supporting the notion that different forms of vitamin D can be interconverted in vitro and that their differential transcriptional activities may underlie their varied immunoregulatory effects. |
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