Role of vitamin D

The role of vitamin D in skeletal health is robust; the major biological function of the active form of vitamin D (1,25(OH)₂D) is to maintain serum calcium and phosphorus homeostasis, essential for bone mineralisation⁽ Reference DeLuca ^{1 )} and neuromuscular function⁽ Reference Holick, Binkley and Bischoff-Ferrari ^{2 )}. The discovery that the vitamin D receptor is expressed in virtually all cells in the human body⁽ Reference Norman ^{3 )} has led to the recognition that vitamin D may have a role to play in many more processes than previously thought. This concurs with the growing body of observational data showing associations between 25-hydroxyvitamin D (25(OH)D) concentrations and chronic diseases and conditions such as CVD, diabetes and colorectal cancer⁽ Reference Autier, Boniol and Pizot ^{4 )}. However, whether low 25(OH)D concentrations are a cause or effect of such conditions has not been established⁽ Reference Autier, Boniol and Pizot ^{4 )}.

Sources of vitamin D

Vitamin D is the generic term used for both vitamin D₂ and vitamin D₃, also known as ergocalciferol and cholecalciferol, respectively. Vitamin D₂ is synthesised by the exposure of ergosterol in plants to UVB radiation, whereas vitamin D₃ is synthesised in the skin of human subjects and animals by the action of UVB radiation.

There are therefore two sources of vitamin D; diet and exposure to sunlight. In human subjects, the action of direct sunlight, containing UVB radiation of wavelengths 290–315 nm, on skin results in synthesis of vitamin D. The first step in this synthesis is the conversion of 7-dehydrocholesterol to pre-vitamin D₃, which is then converted to vitamin D₃ by a temperature-dependent isomerisation reaction.

Naturally occurring dietary sources of vitamin D include both vitamin D₂, found in plants and fungi, and vitamin D₃, found in meat, fish and eggs. However, few naturally occurring food sources of vitamin D are considered a rich source of vitamin D. The few foods considered a good source of vitamin D are mostly of animal origin and therefore contain vitamin D₃ such as oily fish, egg yolks and meat, liver and kidney.

Other sources include supplements or fortified foods, which will be discussed later in this review. Both vitamins D₂ and D₃ can be commercially synthesised for use as supplements and to fortify foods, by UVB irradiation of ergosterol from plants and fungi and 7-dehydrocholesterol from sheep's wool, respectively.

In most countries, exposure of skin to UVB radiation or supplement use are the main source of vitamin D and in the UK skin synthesis is the primary source. The latest UK National Diet and Nutrition Survey data collected in 2012/13 and 2013/14 showed mean dietary intakes of 3·1 and 2·5 µg/d in adult men and women, respectively⁽ Reference Bates, Cox and Nicholson ^{5 )}, although lower dietary intakes of 1·25–1·6 µg/d have been reported for South Asian women in the UK⁽ Reference Macdonald, Mavroeidi and Fraser ^{6 )}.

Rates of vitamin D deficiency

Data from across the globe have shown that vitamin D deficiency is a worldwide issue. Although different cut-off points have been used to define deficiencies, these data are based on 25(OH)D concentrations below 25 nm/l. Studies have reported the prevalence of deficiency to range from 2 to 30 % across Europe⁽ Reference Spiro and Buttriss ^{7 )}, 38 to 80 % in women of child-bearing age in the Middle East⁽ Reference Bassir, Laborie and Lapillonne ^{8 ,} Reference Molla, Al Badawi and Hammoud ^{9 )}, and 3·5 % in the USA, although when ethnicity was considered the prevalence of deficiency were highest among non-Hispanic black individuals at 15 %⁽ Reference Jain ^{10 )}.

In the UK, the latest National Diet and Nutrition Survey⁽ Reference Bates, Cox and Nicholson ^{5 )} reported that 22 % of men and 15 % of women aged 19–64 years had total 25(OH)D concentrations below 25 nm/l all year-round. Furthermore, seasonal variation in 25(OH)D concentrations exists⁽ Reference Hyppönen and Power ^{11 )}; previous National Diet and Nutrition Survey data reported rates of deficiency increasing to 39 % in men and women between January and March⁽ Reference Bates, Lennox and Prentice ^{12 )}. Sub-groups of the population, particularly those considered at-risk of vitamin D deficiency, have shown even higher rates of deficiency; a longitudinal study (D-FINES) looking at dietary and sunlight contribution to seasonal vitamin D status in South Asian and Caucasian women of child-bearing age reported that 51·4 % of the South Asian women had 25(OH)D concentrations <25 nm/l in the summer months, and this rose to 64·5 % in the winter⁽ Reference Darling, Hart and Macdonald ^{13 )}.

Vitamin D dietary recommendations

In July 2016, the Scientific Advisory Committee on Nutrition published the new UK recommendations for vitamin D^{( 14 )}. Based on the health outcome of musculoskeletal health, the Scientific Advisory Committee on Nutrition concluded that the estimated average nutrient intake required to maintain 25(OH)D concentrations at or above 25 nm/l in winter for 97·5 % of the population aged between 11 years and older is 10 µg/d, and a reference nutrient intake (RNI) of 10 µg/d was set for everyone aged 1 year and above. Prior to this update, there was no RNI for vitamin D set for those aged 4–64 years^{( 15 )}, unless the individual was considered at-risk of deficiency. This was based on the assumption that sufficient vitamin D was made, through skin synthesis, and stored during the summer months to sustain 25(OH)D concentrations during the winter months, which is now known not to be the case⁽ Reference Hyppönen and Power ^{11 ,} Reference Darling, Hart and Macdonald ^{13 )}.

This new RNI brings the UK in-line with other European and international recommendations being previously the only country in Europe without a dietary recommendation for all aged 4–6 years⁽ Reference Spiro and Buttriss ^{7 )}.

Comparative metabolism

The chemical structures of vitamins D₂ and D₃ are similar but not identical; vitamin D₃ has an additional double bond and methyl group, and it is postulated that this different structure results in vitamin D₃ being the preferred substrate at several stages of the pathway of metabolism of vitamin D.

Both vitamins D₂ and D₃, irrespective of source, undergo the same metabolism process from the venous circulatory system, a two-step hydroxylation process. Firstly, vitamins D₂ and D₃ are transported by the vitamin D-binding protein to the liver where they are converted to 25(OH)D₂ and 25(OH)D₃, respectively, by the action of 25-hydroxylase. These are then transported, again by the vitamin D-binding protein, to the kidneys where they are converted to the biologically active form of vitamin D (1,25(OH)₂D), by the action of 1-α-hydroxlyases.

There is data to suggest that the differences in the side chains of the two forms of vitamin D directly affect the rate of vitamin D hydroxylation at the liver, with vitamin D₃ thought to be the preferred substrate for hepatic 25-hydroxylase⁽ Reference Holmberg, Berlin and Ewerth ^{16 ,} Reference Cheng, Levine and Bell ^{17 )}. Vitamin D₃ and its metabolites also have a higher binding affinity to the vitamin D-binding protein than vitamin D₂ ⁽ Reference Houghton and Vieth ^{18 )}. In addition to these metabolic differences between the two forms of vitamin D, the degradation of vitamin D₃ requires an addition step/process to that of vitamin D₂, which suggest that the degradation rate of vitamin D₂ may be higher than that of vitamin D₃ ⁽ Reference Horst, Reinhardt and Ramberg ^{19 )}.

Therefore, there are several biologically plausible mechanisms by which vitamin D₃ may have a greater capacity than vitamin D₂ to raise and maintain 25(OH)D concentrations, as reviewed by Houghton and Vieth⁽ Reference Houghton and Vieth ^{18 )}, although randomised-controlled trial (RCT) data have not conclusively supported this theory, as discussed below.

Comparative efficacy at raising 25-hydroxyvitamin D concentrations

Historically, vitamins D₂ and D₃ were considered as equally effective at raising 25(OH)D concentrations⁽ Reference Park ^{20 )}. However, since the 1980s there have been a number of intervention trials, including RCT, published that have investigated specifically this, as shown in Table 1, and these have shown conflicting results. Although the majority of intervention trials comparing the two forms of vitamin D have provided data suggesting that vitamin D₃ is superior to vitamin D₂ in raising 25(OH)D concentrations⁽ Reference Armas, Hollis and Heaney ^{21 –} Reference Trang, Cole and Rubin ^{36 )}, there have also been four trials that have produced data supportive of vitamins D₂ and D₃ being equally effective⁽ Reference Biancuzzo, Young and Bibuld ^{37 –} Reference Nimitphong, Saetung and Chanprasertyotin ^{40 )}. There are no studies which have shown that vitamin D₂ is more effective than vitamin D₃.

Table 1. Study characteristics and outcomes of intervention trials comparing the effects of vitamins D₂ and D₃ on 25-hydroxyvitamin D (25(OH)D) concentrations

Key: vit, vitamin; M, males; F, females; d, days; wk, weeks; mo, months; OJ, orange juice; 25(OH)D, 25-hydroxyvitamin D; IM, intramuscular.

In 2012, Tripkovic et al. conducted a systematic review and meta-analysis of RCT data comparing the efficacy of vitamins D₂ and D₃ in raising 25(OH)D concentrations⁽ Reference Tripkovic, Lambert and Hart ^{41 )}. Although ten studies were identified within the systematic review, only seven studies had sufficient and available data to be included within the meta-analysis. The primary analysis, of all seven studies regardless of dosing frequency, showed that vitamin D₃ led to a greater absolute change in 25(OH)D concentrations than vitamin D₂, with a weighted mean difference of 15·23 (95 % CI 6·12, 24·34; Z = 3·28; I ² = 81 %; P = 0·001). To determine any confounding effect of dosing frequency, separate analyses were performed on: (a) studies giving bolus doses and (b) studies with daily supplementation. For bolus studies alone vitamin D₃ remained significantly more effective than vitamin D₂, with a weighted mean difference of 34·10 (95 % CI 16·39, 51·83; Z = 3·77; I ² = 77 %; P = 0·0002). However, when the analysis was completed on the daily dosing RCT data alone, the differentiation between the two forms of vitamin D was moderated with a non-significant weighted mean difference of 4·83 (95 % CI −0·98, 10·64; Z = 1·63; I ² = 41 %; P = 0·10).

Key limitations of the meta-analysis were identified and discussed by the authors. Firstly, there were few studies for inclusion within the analysis, and of the studies that were available these were small and unpowered with respect to study population size (n 19–89). There was also substantial between-study heterogeneity, with diverse intervention strategies, including varied doses of vitamin D, frequencies of supplementation and methods of administration, and all the studies used supplementation doses in excess of current recommendations^{( 14 )}. Taking these factors into consideration, the authors concluded that far larger, more robust trials are required to not only measure 25(OH)D concentrations in response to vitamins D₂ and D₃, but also to explore potential mechanisms behind any differences seen.

Since the 2012 meta-analysis, there have been at least another ten further intervention trials comparing the efficacy of vitamins D₂ and D₃ in raising 25(OH)D concentrations⁽ Reference Cipriani, Romagnoli and Pepe ^{23 ,} Reference Itkonen, Skaffari and Saaristo ^{26 ,} Reference Lehmann, Hirche and Stangl ^{27 ,} Reference Logan, Gray and Peddie ^{29 –} Reference Oliveri, Mastaglia and Brito ^{31 ,} Reference Shieh, Chun and Ma ^{33 ,} Reference Stepien, O'Mahony and O'Sullivan ^{34 ,} Reference Fisk, Theobald and Sanders ^{38 ,} Reference Nimitphong, Saetung and Chanprasertyotin ^{40 )}. However, findings remain equivocal with eight of the studies showing vitamin D₃ to be more effective at raising or maintaining 25(OH)D concentrations compared to vitamin D₂ ⁽ Reference Cipriani, Romagnoli and Pepe ^{23 ,} Reference Itkonen, Skaffari and Saaristo ^{26 ,} Reference Lehmann, Hirche and Stangl ^{27 ,} Reference Logan, Gray and Peddie ^{29 –} Reference Oliveri, Mastaglia and Brito ^{31 ,} Reference Shieh, Chun and Ma ^{33 ,} Reference Stepien, O'Mahony and O'Sullivan ^{34 )}, and two of these trials reporting no significant difference between the two forms⁽ Reference Fisk, Theobald and Sanders ^{38 ,} Reference Nimitphong, Saetung and Chanprasertyotin ^{40 )}. However, a direct comparison between the total change in 25(OH)D concentrations in response to vitamins D₂ and D₃ was not reported in the analysis by Fisk et al. which returned neutral findings⁽ Reference Fisk, Theobald and Sanders ^{38 )}.

These more recent studies have addressed some, but not all, of the limitations identified from the 2012 systematic review and meta-analysis. Specifically, over half of the studies since 2012 examined supplementation doses closer to the range of current global recommendations. However, underpowered sample sizes remain an issue; the largest published RCT comparing the efficacy of vitamins D₂ and D₃ on raising 25(OH)D concentrations had a total of 107 subjects across three intervention groups⁽ Reference Lehmann, Hirche and Stangl ^{27 )} as shown in Table 1. There remains a need for a large, robust RCT to provide more conclusive evidence in which confidence in results can be sought and the publication of the BBSRC Diet and Health Research Industry Club (DRINC) funded D2–D3 Study (BBSRC DRINC: BB/I006192/1, ISRCTN23421591); an RCT in n 335 healthy white Caucasian and South Asian women, should provide just that⁽ Reference Tripkovic, Wilson and Hart ^{42 )}.

Potential mechanisms from randomised-controlled trial data

Within some of these intervention trials the mechanisms by which vitamins D₂ and D₃ might lead to different effects on total 25(OH)D concentrations have been explored. Where methods such as LC-MS/MS for measuring 25(OH)D concentration are implemented, 25(OH)D₂ and 25(OH)D₃ concentrations are measured, which are added together to determine total 25(OH)D concentrations. In studies where these two metabolites have been measured, the vitamin D₂ interventions have led to an increase in 25(OH)D₂ concentrations and the vitamin D₃ interventions have led to an increase in 25(OH)D₃ concentrations⁽ Reference Armas, Hollis and Heaney ^{21 ,} Reference Glendenning, Chew and Seymour ^{24 ,} Reference Lehmann, Hirche and Stangl ^{27 ,} Reference Logan, Gray and Peddie ^{29 ,} Reference Biancuzzo, Young and Bibuld ^{37 ,} Reference Fisk, Theobald and Sanders ^{38 )}, as would be expected. However, a decreasing effect of vitamin D₂ interventions on 25(OH)D₃ concentrations has been noted in several intervention trials⁽ Reference Armas, Hollis and Heaney ^{21 ,} Reference Binkley, Gemar and Engelke ^{22 ,} Reference Lehmann, Hirche and Stangl ^{27 ,} Reference Tjellesen, Hummer and Christiansen ^{35 )}, although not all⁽ Reference Glendenning, Chew and Seymour ^{24 ,} Reference Biancuzzo, Young and Bibuld ^{37 ,} Reference Fisk, Theobald and Sanders ^{38 )}. The same has not been shown for vitamin D₃; vitamin D₃ interventions have not shown a decreasing effect on 25(OH)D₂ concentrations; however, baseline 25(OH)D₂ concentrations tend to be far lower at baseline (typically <5 nm/l) and so the opportunity for a decreasing effect is not available. Although even where mean baseline 25(OH)D₂ concentrations were slightly higher (13·3 nm/l) no change was shown in the vitamin D₃ intervention group⁽ Reference Glendenning, Chew and Seymour ^{24 )}.

This decline in 25(OH)D₃ concentrations reported in those taking vitamin D₂ could explain why vitamin D₃ is more effective at raising total 25(OH)D concentrations, and although the exact mechanisms are unknown this could reflect either competitive binding for 25-hydroxylase or the vitamin D-binding protein, or changes in degradation rate as discussed previously. Further research is needed to elucidate the exact mechanisms and to understand the impact of these changes on overall health, and not just total 25(OH)D concentrations.

Supplementation

Supplementation is another potential strategy to support the UK population in achieving the dietary recommendation and subsequently improving vitamin D status. Although there are no data showing the number of people in the UK currently taking vitamin D supplements specifically, National Diet and Nutrition Survey data have shown that 23 % of adults aged 19–64 years and 39 % of adults over 65 years take at least one dietary supplement⁽ Reference Bates, Lennox and Prentice ^{12 )}. Currently the UK advise daily supplements for those considered at-risk of vitamin D deficiency such as young children, pregnant and breastfeeding women, housebound elderly and those with darker skin tones. However, the awareness and adherence to this current guidance is poor⁽ Reference Buttriss ^{50 )} and so the impact of this advice, and any potential future advice, will only be realised if individuals are willing to take supplements and remember to do so. Universal provision of supplements has been identified as a successful strategy in some European countries where they are recommended for infants (Norway, Germany, Austria and Switzerland) and children up to the age of 5 years (Sweden). The potential universal use of vitamin D supplementation in the UK is therefore worthy of further research, as discussed in greater detail in two recent reviews⁽ Reference Spiro and Buttriss ^{7 ,} Reference Buttriss ^{50 )}.