Vitamin D Binding Protein, Total and Free Vitamin D Levels in Different Physiological and Pathophysiological Conditions

Abstract

This review focuses on the biologic importance of the vitamin D binding protein (DBP) with emphasis on its regulation of total and free vitamin D metabolite levels in various clinical conditions. Nearly all DBP is produced in the liver, where its regulation is influenced by estrogen, glucocorticoids and inflammatory cytokines but not by vitamin D itself. DBP is the most polymorphic protein known, and different DBP alleles can have substantial impact on its biologic functions. The three most common alleles-Gc1f, Gc1s, Gc2-differ in their affinity with the vitamin D metabolites and have been variably associated with a number of clinical conditions. Although DBP has a number of biologic functions independent of vitamin D, its major biologic function is that of regulating circulating free and total levels of vitamin D metabolites. 25 hydroxyvitamin D (25(OH)D) is the best studied form of vitamin D as it provides the best measure of vitamin D status. In a normal non-pregnant individual, approximately 0.03% of 25(OH)D is free; 85% is bound to DBP, 15% is bound to albumin. The free hormone hypothesis postulates that only free 25(OH)D can enter cells. This hypothesis is supported by the observation that mice lacking DBP, and therefore with essentially undetectable 25(OH)D levels, do not show signs of vitamin D deficiency unless put on a vitamin D deficient diet. Similar observations have recently been described in a family with a DBP mutation. This hypothesis also applies to other protein bound lipophilic hormones including glucocorticoids, sex steroids, and thyroid hormone. However, tissues expressing the megalin/cubilin complex, such as the kidney, have the capability of taking up 25(OH)D still bound to DBP, but most tissues rely on the free level. Attempts to calculate the free level using affinity constants generated in a normal individual along with measurement of DBP and total 25(OH)D have not accurately reflected directly measured free levels in a number of clinical conditions. In this review, we examine the impact of different clinical conditions as well as different DBP alleles on the relationship between total and free 25(OH)D, using only data in which the free 25(OH)D level was directly measured. The major conclusion is that a number of clinical conditions alter this relationship, raising the question whether measuring just total 25(OH)D might be misleading regarding the assessment of vitamin D status, and such assessment might be improved by measuring free 25(OH)D instead of or in addition to total 25(OH)D.

Keywords: free 25(OH)D; free hormone hypothesis; liver cirrhosis; megalin; polymorphisms; pregnancy; vitamin D; vitamin D binding protein.

Figure 1

The major DBP alleles. The amino acid differences between the three major DBP alleles are depicted. These differences affect not only their electrophoretic properties but also their glycosylation pattern. In particular Gc2 is not glycosylated, which prevents it from forming the DBP-macrophage activating factor (DBP-MAF). Other biologic differences are discussed in the text.

Figure 2

The Free Vitamin D hypothesis. As noted in the text, vitamin D (OH) metabolites are bound to D Binding Protein (DBP) and to a lesser extent albumin in the circulation. These cross the cell membrane as the free (unbound) metabolite in most tissue. However, In the kidney, parathyroid gland, and placenta, the megalin/cubilin complex can transport bound D (OH) metabolites into cells.

Figure 3

Distribution of free 25(OH)D in Adults and Selected Patient Groups. Distribution of directly-measured free 25(OH)D in normal adults (in green), pregnant women (pink), cirrhotics (orange), and nursing home residents (gray). Distributions are shifted leftward toward lower free 25(OH)D concentrations in pregnant women in the 2nd and 3rd trimesters concordant with increased DBP while decreased synthetic function and DBP in cirrhotics shifts free 25(OH) concentrations to the right toward higher levels. The mechanism for higher free 25(OH) concentrations in Nursing home residents is likely related to D supplementation, somewhat lower, albumin, and the pro-inflammatory state of frailty. Figure generated form data in Schwartz et al. (40).

Figure 4

Percent free 25(OH)D in adults by clinical condition or DBP Haplotype. Percent free 25(OH)D concentrations for selected clinical groups on the left panel (community outpatients, NH=nursing home patients, cirrhotics, pregnant women, prediabetics, and normal individuals) and by DBP haplotype on the right . Boxplots show 10th, 25th, median, 75th, and 90th percentile values. Individual points represent values above the 90th and below the 10th percentiles. Both clinical subgroup and DBP genotype significantly effect percentage free 25(OH)D. Between group comparisons for clinical conditions were significant for all but healthy persons compared with pregnant women or outpatients, or for pregnant women compared with outpatients. For DBP haplotypes, smaller but significant differences were detected between the 1s/1s haplotype and the 1s/1f,1f/2, 1f/1f, and 1s/2 haplotypes and between the 1s/2 and 1f/2 and 1f/1f haplotypes and between the 1s/1f and 1f/1f haplotypes. Data are reproduced with permission from Schwartz et al. (40).