25-Hydroxyvitamin D3 and its C-3 epimer are elevated in the skin and serum of Skh-1 mice supplemented with dietary vitamin D3

Abstract

Scope: UV exposure is a risk factor for keratinocyte carcinoma (KC) while critical for endogenous vitamin D production. We investigated dietary modulation of skin and serum 25-hydroxyvitamin D₃ (25OHD₃ ) and its C-3 epimer (C3epi) in a mouse model of KC. C3epi is an under-investigated metabolite of vitamin D with respect to its biological implications.

Methods and results: Male and female Skh-1 mice were supplemented with 25, 150 or 1000 IU/kg diet vitamin D₃ for 25 weeks, with some exposed to UV light. Skin and serum vitamin D metabolites were quantitated using HPLC-MS/MS (n = 3 per dose/sex/UV treatment). Serum and skin 25OHD₃ and C3epi significantly increased with dose (P<0.0001), but with different response patterns. UV exposure significantly attenuated serum, but not skin, levels of both metabolites (P<0.001, P = 0.0287), while up-regulating expression of renal Cyp24a1 (P < 0.01). A dose by sex interaction trended toward significance with serum and skin levels of C3epi, wherein male mice attained higher levels of C3epi with higher dietary vitamin D₃ . This reflected a similar, but non-significant pattern in average tumor size.

Conclusion: The complex relationship between vitamin D and KC requires further investigation. This study provides insight into modulation of local and systemic vitamin D status with dietary supplementation.

Keywords: 3-epi-25-hydroxyvitamin D; Keratinocyte carcinoma; Skin cancer; Vitamin D.

Figure 1

Structures of 25OHD₃ and C3epi.

Figure 2

Dietary supplementation with vitamin D₃ significantly (P<0.0001 for all) raised 25OHD₃ and C3epi levels in serum (A) and skin (B), regardless of sex or UV exposure. Different letters indicate significant differences, n=12 per dietary level (including 3 mice from each sex/UV exposure, per dietary level).

Figure 3

Effects of UV exposure on vitamin D₃ metabolite levels at each dietary supplementation level, regardless of sex. There was no significant dose by UV exposure interaction, but there was a significant effect of UV exposure on serum 25OHD₃ levels (A) (P<0.001; n=18 per UV exposure group including 3 male and 3 female mice at each dietary level). There was a dose by UV interaction for serum C3epi levels (B) (P=0.0287; n=6, including 3 male and female per UV/dose group). No significant effects of either UV exposure by dose or UV exposure alone were detected for skin 25OHD₃ (C) or C3epi (D) levels (n=6 per UV/dose group or n=18 per UV group).

Figure 4

Expression of Cyp24a1 in kidney. UV exposure significantly upregulated expression (No UV, n=39; UV, n=57; including males and females at all doses). A significant effect of dose was also observed (25IU, n=13 no UV & 20 UV; 150IU, n=13 no UV & 17 UV; 1000IU, n=13 no UV & 20 UV).

Figure 5

Effects of mouse sex on vitamin D₃ metabolite levels with respect to dietary supplementation levels. To look at the effect of sex and dietary vitamin D3 dose on metabolite levels in skin and serum, metabolite data for both UV groups was combined. Dose by sex interactions trended towards significance for C3epi levels in serum (B) (P=0.0562; n=6 including 3 no UV and 3 UV mice) and skin (D) (P=0.0510; n=6 including 3 no UV and 3 UV mice). No significant effects of mouse sex were detected for 25OHD₃ levels in serum (A) or skin (C) (n=6 per sex/dose group or n=18 per sex group).

Figure 6

Correlations between skin and serum 25OHD₃ (A) and C3epi (B). Coefficients in figure were calculated across all samples (n=36). No significant differences were detected between coefficients calculated according to UV exposure or sex (n=18 per group).