Shah Anisha B, DiMartino Stephen J, Trujillo Glenda, Kew Richard R
Department of Pathology, Health Sciences Center, Stony Brook University School of Medicine, Stony Brook, NY 11794-8691, USA.
Mol Immunol. 2006 Mar;43(8):1109-15. doi: 10.1016/j.molimm.2005.07.023. Epub 2005 Aug 22.
The Vitamin D binding protein (DBP) is a multifunctional plasma protein that can significantly enhance the chemotactic response to complement fragment C5a. The chemotactic cofactor function of DBP requires cell surface binding in order to mediate this process. The goal of this study was to investigate the effect of ligating DBP with its two primary physiological ligands, Vitamin D and G-actin, on both binding to neutrophils and the ability to enhance chemotaxis to C5a. There was no difference in neutrophil binding between of the holo (bound) forms versus the apo (unbound) form of radioiodinated DBP, indicating that the cell binding region of DBP is likely distinct from the Vitamin D sterol and G-actin binding sites. Likewise, G-actin, 25(OH)D3, and G-actin plus 25(OH)D3 bound to DBP did not alter its capacity to enhance chemotaxis toward C5a. However, the active form of Vitamin D (1,25(OH)2D3) completely eliminated the chemotactic cofactor function of DBP. Dose-response curves demonstrated that as little as 1pM 1,25(OH)2D3 significantly inhibited chemotaxis enhancement. Moreover, at physiological concentrations 1,25(OH)2D3 needs to be bound to DBP to mediate the inhibitory effect. Neutrophil chemotaxis to optimal concentrations of C5a, formyl peptide, CXCL8 or leukotriene B4 was not altered by 1,25(OH)2D3, indicating that the active vitamin does not have a global inhibitory effect on neutrophil chemotaxis. Finally, inhibition of cell surface alkaline phosphatase (AP) with sodium orthovanadate completely reversed the inhibitory effect of 1,25(OH)2D3. These results indicate that the cell binding and co-chemotactic functions of DBP are not altered when the protein binds G-actin and/or Vitamin D. Furthermore, the co-chemotactic signal from DBP can be eliminated or counteracted by 1,25(OH)2D3.
维生素D结合蛋白（DBP）是一种多功能血浆蛋白，它能显著增强对补体片段C5a的趋化反应。DBP的趋化辅因子功能需要细胞表面结合才能介导这一过程。本研究的目的是探讨将DBP与其两种主要生理配体维生素D和G-肌动蛋白连接，对其与中性粒细胞结合以及增强对C5a趋化性能力的影响。放射性碘化DBP的全（结合）形式与脱辅基（未结合）形式在与中性粒细胞结合方面没有差异，这表明DBP的细胞结合区域可能与维生素D固醇和G-肌动蛋白结合位点不同。同样，与DBP结合的G-肌动蛋白、25（OH）D3以及G-肌动蛋白加25（OH）D3并没有改变其增强对C5a趋化性的能力。然而，维生素D的活性形式（1,25（OH）2D3）完全消除了DBP的趋化辅因子功能。剂量反应曲线表明，低至1pM的1,25（OH）2D3就能显著抑制趋化增强。此外，在生理浓度下，1,25（OH）2D3需要与DBP结合才能介导抑制作用。1,25（OH）2D3不会改变中性粒细胞对最佳浓度的C5a、甲酰肽、CXCL8或白三烯B4的趋化性，这表明活性维生素对中性粒细胞趋化性没有全面的抑制作用。最后，用原钒酸钠抑制细胞表面碱性磷酸酶（AP）可完全逆转1,25（OH）2D3的抑制作用。这些结果表明，当该蛋白与G-肌动蛋白和/或维生素D结合时，DBP的细胞结合和共趋化功能不会改变。此外，来自DBP的共趋化信号可被1,25（OH）2D3消除或抵消。
