PMC6476011_01
(a) Sequences of the CYP24A1 exon 2 obtained by proband and her parents. CYP24A1 gene amplification and sequencing were performed as reported. Sequences of the exon 2 obtained by proband and her parents were shown. Arrows indicate the position of c.428_430delAAG heterozygous and homozygous variant in the parents and the proband, respectively. (b) Family tree.
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fig1
The genetic analysis in the proband was made using High Resolution Melting Analysis (HRMA) and further confirmed using gene amplification and sequencing, revealing a known homozygous PV (c.428_430delAAG, rs777676129, p.Glu143del) in the CYP24A1 gene (Figure 1(a)). The same heterozygous variant was detected in the parents and the siblings (Figure 1(b)).
LC-MS/MS chromatogram of dihydroxylated vitamin D metabolites. Serum samples were prepared by immunoextraction and derivatized with DMEQ-TAD. Peaks corresponding to 6S and 6R isomers of DMEQ-TAD adducts of 24,25(OH)2D3 and 1,25(OH)2D3 were observed using the multiple reaction monitoring (MRM) transition of mass/charge (m/z) 762.3- ->468.1+484.1, from which serum concentrations of these metabolites were determined. The figure reveals the dramatically reduced concentration of 24,25(OH)2D3 in the proband II.3 as well as elevated 1,25(OH)2D3, in comparison to unaffected family members.
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fig2
The other family members, who present as heterozygous variants, exhibited essentially normal serum 24,25(OH)2D3 concentrations and 25(OH)D3:24,25(OH)2D3 ratios (Table 3 and Figure 2).