Excess 25-hydroxyvitamin D3 exacerbates tubulointerstitial injury in mice by modulating macrophage phenotype

Vitamin D hydroxylated at carbon 25 (25(OH)D) is generally recognized as a precursor of active vitamin D. Despite its low affinity for the vitamin D receptor (VDR), both deficient and excessive 25(OH)D levels are associated with poor clinical outcomes. Here we studied direct effects of 25(OH)D 3 on the kidney using 25(OH)D-1a-hydroxylase (CYP27B1) knockout mice. The effects of 25(OH)D 3 on unilateral ureteral obstruction were analyzed as proximal tubular cells and macrophages are two major cell types that take up 25(OH)D and contribute to the pathogenesis of kidney injury. Excess 25(OH)D 3 in obstructed mice worsened oxidative stress and tubulointerstitial fibrosis, whereas moderate levels of 25(OH)D 3 had no effects. The exacerbating effects of excess 25(OH)D 3 were abolished in CYP27B1/VDR double-knockout mice and in macrophage-depleted CYP27B1 knockout mice. Excess 25(OH)D3 upregulated both M1 marker (TNF-α) and M2 marker (TGF-β1) levels of kidney-infiltrating macrophages. In vitro analyses verified that excess 25(OH)D 3 directly upregulated TNF-α and TGF-β1 in cultured macrophages but not in tubular cells. TNF-α and 25(OH)D 3 cooperatively induced oxidative stress by upregulating iNOS in tubular cells. Aggravated tubulointerstitial fibrosis in mice with excess 25(OH)D 3 indicated that macrophage-derived TGF-β1 also had a key role in the pathogenesis of surplus 25(OH)D3. Thus, excess 25(OH)D 3 worsens tubulointerstitial injury by modulating macrophage phenotype.