Exercise: A Possibly Effective Way to Improve Vitamin D Nutritional Status

Vitamin D deficiency has become a widespread public health problem owing to its potential adverse health effects. Generally, the nutritional status of vitamin D depends on sunlight exposure and dietary or supplementary intake. However, recent studies have found that exercise can influence circulating 25(OH)D levels; although, the results have been inconclusive. In this review, we focused on the effect of exercise on circulating vitamin D metabolites and their possible mechanisms. We found that endurance exercise can significantly increase serum 25(OH)D levels in vitamin D-deficient people but has no significant effect on vitamin D-sufficient people. This benefit has not been observed with resistance training. Only chronic endurance exercise training can significantly increase serum 1,25(OH)2D, and the effect may be sex-dependent. Exercise may influence 25(OH)D levels in the circulation by regulating either the vitamin D metabolites stored in tissues or the utilization by target tissues. The effects of exercise on 25(OH)D levels in the circulation may be dependent on many factors, such as the vitamin D nutritional status, exercise type and intensity, and sex. Therefore, further research on the effects and mechanisms of exercise on vitamin D metabolites is required.


Introduction
The recent increase in vitamin D-related research has led to the discovery of the vitamin D receptor (VDR) in many tissues. A growing body of literature has shown that the biological role of vitamin D goes beyond the traditionally understood duties dealing with muscles and bones and is important for energy metabolism, oxidative stress, maintenance, and improvement of physical fitness [1][2][3][4]. A study found that vitamin D 3 supplementation increased serum 25(OH)D levels; additionally, the expression of 291 genes, involving as many as 160 metabolic pathways, was significantly upregulated or downregulated [5]. This finding suggests that vitamin D plays an important role in health. Alarmingly, a survey found that vitamin D deficiency has become a global public health problem [6,7]. Vitamin D deficiency is closely associated with various chronic non-communicable diseases and functional disorders [1,[8][9][10]. Therefore, maintaining adequate vitamin D levels is significant for promoting health.
Vitamin D is mainly synthesized by the skin, and its sources in food are scarce [11,12]. In the epidermis, 7-dehydrocholesterol can be transformed into vitamin D 3 upon exposure to sunlight, while vitamin D 2/3 in foods/supplements is absorbed into the circulation through the intestines. Both skin-synthesized vitamin D 3 and food/supplement-derived D 2/3 are catalyzed by 25-hydroxylase [mainly cytochrome P450 family 27 subfamily A member 1 (CYP27A1) and cytochrome P450 family 2 subfamily R member 1 (CYP2R1)] to 25(OH)D in the liver. Due to its long half-life [13] and strong vitamin D binding protein (VDBP) binding ability [14], serum 25(OH)D is the most abundant and stable vitamin D metabolite in the circulation; hence, its serum concentration is used to evaluate the nutritional status of vitamin D [15]. Subsequently, 25 (Table 1). Mieszkowski et al. found that serum 25(OH)D levels in male runners [baseline serum 25(OH)D level >20 ng/mL] with and without vitamin D supplementation were significantly increased immediately and 24 h after an ultra-marathon race compared with those before the race [32]. Dzik also reported that serum 25(OH)D 3 levels in male soccer players (10-14 years old) significantly increased at 15 min and 1 h after a VO 2 max test [25(OH)D >70 nmol/L]. In an analysis of both pre-pubertal and pubertal boys, the concentration of 25(OH)D 3 increased 15 min after the VO 2 max test and dropped one hour after exercise, but these changes were not significantly different at specific time points [33]. However, Maimoun found that intensity exercise [47% Wmax, baseline serum 25(OH)D level: 79.4 ± 13.7 nmol/L; 64% Wmax, baseline serum 25(OH)D level: 83.4 ± 16 nmol/L] did not alter the concentration of 25(OH)D in male competitive road cyclists during cycling exercise or after 15 min of recovery [34]. Conversely, two studies involving non-athletes demonstrated that acute endurance exercise may increase circulating 25(OH)D levels. Maimoun et al. found that maximal incremental exercise can significantly increase the level of 25(OH)D in physically highly active elderly participants but not in moderately active elderly and young physically active adults [35]. Sun et al. found that serum 25(OH)D concentration significantly increased immediately and 1, 3, and 24 h after 30 min of cycling exercise at 70% VO 2 peak [36]. However, in the subgroup analysis, the 25(OH)D level increase of women [baseline serum 25(OH)D level:55.1 ± 15.6 nmol/L] was significant only at 24 h after exercise. The acute effect of exercise on 25(OH)D levels may be affected by intensity [33], sex [36], and age [33,35]. As for serum 1,25(OH) 2 D levels, no significant variation was observed in response to acute endurance exercise [34][35][36].

Animal Studies
Results from animal studies investigating the effect of acute endurance exercise on serum 25(OH)D have been inconsistent compared to those from human studies (Table 1). Makanae et al. found that acute endurance exercise (anaerobic threshold intensity) did not alter serum 25(OH)D levels in adult male Sprague-Dawley rats [37]. Moreover, serum 25(OH)D levels in horses were significantly reduced at 30 min, 1 week, and 3 weeks after high-intensity exercise [38]. However, only two experimental animal studies have been conducted to investigate the effect of acute endurance exercise on serum 25(OH)D levels.

Human Studies
Twelve human studies investigated the effect of endurance exercise training on serum 25(OH)D levels and did not yield consistent results (Table 1). Some studies found that chronic endurance exercise training can significantly increase serum 25(OH)D levels [39][40][41][42][43][44][45], but other studies have reported contradicting results [46][47][48][49][50]. However, when we sorted these studies, we found that in people with vitamin D deficiency {25(OH)D < 20 ng/mL or 50 nmol/L [57]}, endurance training can significantly improve serum 25(OH)D levels [39,40,42,43,45], and even severe vitamin D deficiency status (<10 ng/mL) improved to vitamin D deficiency status (10-20 ng/mL) in postmenopausal women [42]. However, endurance exercises had no significant effects on serum 25(OH)D levels in overweight and obese subjects, regardless of vitamin D nutritional status [48]. For participants with sufficient vitamin D levels {25(OH)D ≥ 20 ng/mL or 50 nmol/L [57]}, endurance training combined with vitamin D supplementation significantly increased serum 25(OH)D levels [40,44], while endurance training alone did not [40,46,49,50]. While Pilch et al. found that serum 25(OH)D levels were significantly reduced in postmenopausal obese women with sufficient vitamin D levels after endurance exercise intervention, the study was conducted in late autumn and had no control group; hence, it was impossible to determine whether the decrease in 25(OH)D was due to endurance exercise training or a seasonal decline [47]. When considering sun exposure, we found that studies providing relevant sun exposure information were all conducted in the morning [39][40][41]47] or evening [39] or autumn and winter [46,50]. During this time, sun exposure is weaker and has less effect on vitamin D. Taken together, the effect of chronic endurance exercise training on 25(OH)D levels in the circulation may be affected by the vitamin D status.

Animal Studies
Six animal studies investigated the effect of chronic endurance exercise training on 25(OH)D and 1,25(OH) 2 D (Table 1). Aly et al. found that a 4-week swimming regimen significantly increased serum 25(OH)D levels in diabetic mice. No significant change in serum 25(OH)D levels was observed in healthy mice; however, their serum 25(OH)D levels were significantly higher [51]. Buskermolen et al. found that although 6 weeks of endurance training increased serum 25(OH)D levels in female Wistar rats, the change was not significant [52]. The female Wistar rats in Buskermolen's study were fed 1.5 IU/g (>1000 IU/kg [58][59][60]) vitamin D 3 , which is sufficient to maintain an adequate vitamin D status. Some animal studies have shown that chronic endurance exercise training can significantly increase serum 1,25(OH) 2 D levels in healthy female rats [53,54]. However, while Wang et al. found that 12 weeks of treadmill endurance exercise training slightly increased serum 1,25(OH) 2 D 3 levels in aged male rats, the results were not significant [55]. Conversely, Xu et al. found that 8 weeks of swimming and downhill running significantly reduced serum 1,25(OH) 2 D 3 level in 5-week old male mice [56]. In all animal studies which reported increased 1,25(OH) 2 D levels [53,54], the mice were all female, while those that were unchanged or decreased were male [55,56]. The effect of exercise training on 1,25(OH) 2 D may therefore depend on sex.

Mechanism
Endurance exercise induces greater improvements in aerobic capacity and its associated cardiopulmonary and metabolic variables [61]. In terms of energy metabolism, endurance exercise can activate several secondary signal molecules, such as AMPK, CaMKII, and p38, which promote an increase in PGC-1α. Subsequently, PGC-1α promotes mitochondrial biogenesis, exercise-induced fast-to-slow fiber-type transformation, and exerciseinduced expression of important muscle antioxidant enzymes. Therefore, endurance exercise, especially submaximal endurance exercise, effectively increases fat metabolism [62]. Adipose tissue is one main storage depot for vitamin D [63]. Hengist et al. suggested that release of vitamin D stored in adipose tissue is a byproduct of lipolysis [64]. In other words, in the process of releasing triglycerides from adipocytes through the action of lipolytic enzymes, the stored vitamin D metabolites were also released. Lipolysis is regulated by various factors, such as atrial natriuretic peptides (ANPs), brain natriuretic peptides (BNPs), insulin, and beta adrenergic hormones [65]. Endurance exercise can promote the release of these hormones [66], promoting lipolytic processes and releasing vitamin D metabolites from the adipose tissue. Moreover, a systematic review showed that all exercise protocols (high-intensity interval exercise, moderate-intensity continuous exercise, and sprint interval exercise) can generate elevated energy expenditure through excessive post-exercise oxygen consumption (EPOC) [67]. Exercise-induced energy deficit has the most potent effect on endogenous lipid metabolism, elevating plasma triacylglycerol concentration and increasing plasma fatty acid mobilization and oxidation the day after performing endurance exercises [68]. The reason that endurance training can increase serum 25(OH)D levels may be attributed to lipolytic processes during exercise and EPOC.
Abboud et al. found that serum 25(OH)D levels in pasture sheep at the end of winter were significantly lower than those during the summer, but intramuscular 25(OH)D content at the end of winter was significantly higher [69].  [69]. In addition, vitamin D nutritional status is regulated by a variety of factors such as serum Ca 2+ , Pi, parathyroid hormone (PTH), and FGF23 (fibroblast growth factor-23 (FGF23)) [70,71]. PTH stimulates the expression of CYP27B1 in the kidney, while FGF23, high Ca 2+ or Pi levels, and 1,25(OH) 2 D downregulate it. In contrast, 1,25(OH) 2 D and FGF23 strongly induce the expression of CYP24A1, while PTH reduces its expression by stimulating its mRNA [72]. Moreover, PTH enhances the production of 1,25(OH) 2 D, which in turn activates an inhibitory loop regulating PTH production. Similarly, FGF23 regulates the production of 1,25(OH) 2 D, an inducer of FGF23 synthesis in the bones [70,73]. These factors work together to maintain vitamin D nutritional homeostasis, explaining why exercise cannot adequately elevate the level of 25(OH)D. Interestingly, serum calcium and PTH levels were significantly increased in the three groups [35]. Changes in PTH and calcium levels may therefore be responsible for the transient changes in 25(OH)D levels when there is no deficiency.
A summary of how endurance training may exert its effects on 25(OH)D in several ways can be seen in Figure 1.

Human and Animal Studies
One human study and one animal study have investigated the effect of acute resistance exercise intervention on 25(OH)D (  [37]. In the human study, subjects performed an intense-stretch shortening contraction (10 sets of 10 repetitive jumps), whereas rats were put through isometric exercise (5 sets of 10 contractions). The inconsistency in outcome between the two studies may partially be explained by the differences in resistance exercise intensity and volume. Table 2. Summary of the effect of resistance exercise intervention (human study and animal study).

The Effect of Acute Resistance Exercise Human and Animal Studies
One human study and one animal study have investigated the effect of acute resistance exercise intervention on 25(OH)D (  [37]. In the human study, subjects performed an intense-stretch shortening contraction (10 sets of 10 repetitive jumps), whereas rats were put through isometric exercise (5 sets of 10 contractions). The inconsistency in outcome between the two studies may partially be explained by the differences in resistance exercise intensity and volume.

Human Studies
Five human studies have investigated the effects of resistance exercise training on 25(OH)D levels ( Table 2). Resistance exercise training significantly increased circulating 25(OH)D levels in vitamin D-deficient post-stroke hemiplegia patients [75] and healthy participants [76]. Conversely, resistance exercise training had no effect on 25(OH)D lev-els in healthy vitamin D-deficient young men [77] and older adults without vitamin D supplementation [78]. However, Agergaard et al. found that resistance exercise training significantly reduced serum 25(OH)D levels in young and elderly participants without vitamin D supplementation [79]. Factors such as vitamin D supplementation, season, and experimental design should be considered when interpreting these findings. We found that all groups received vitamin D supplementation in Zhang's study [75]; however, serum 25(OH)D levels were higher at 3 months and 1 year following resistance exercise training combined with vitamin D, compared to only vitamin D supplementation. In Bass's study, there was a lack of control groups and seasonal information; hence, it is unclear whether the increase in serum 25(OH)D levels is due to resistance training or seasonal factors [76]. Detailed seasonal information was provided in Aschauer's study (from mid-February to mid-July) [78], Sun's study (from March to July) [77], and Agergaard's study (from November to December) [79]. We found a clear seasonal trend in mean serum 25(OH)D concentrations, suggesting that the change in 25(OH)D concentrations induced by resistance training may have been caused by large seasonal fluctuations [77,78].

Animal Studies
Two animal studies have investigated the effect of resistance exercise training on 25(OH)D and 1,25(OH) 2 D 3 ( Table 2). Buskermolen et al. found that 6 weeks of peak power training did not alter serum 25(OH)D levels in rats [52]. Conversely, Xu et al. found that 8 weeks of jumping training significantly reduced serum 1,25(OH) 2 D 3 levels in male mice [56].

Mechanisms
The predominant adaptation of resistance exercises is in the musculoskeletal system, including increases in muscle mass, muscle strength, and bone density [61]. Muscle mass is increased when resistance exercise triggers muscle signaling events that activate mTOR, leading to increased protein synthesis [62]. Therefore, resistance exercise can be effective in increasing muscle weight and hypertrophy. Mason et al. reported that circulating VDBP can be internalized into skeletal muscle cells to provide high-affinity intracellular binding sites for 25(OH)D [80]. The authors postulate that this intracellular VDBP enables 25(OH)D to diffuse into muscle cells where it is bound and retained until VDBP undergoes proteolysis [80]. The released 25(OH)D then diffuses from the skeletal muscle cells into the circulation and is immediately bound by VDBP in the circulation [80]. Thus, muscle tissue may be an important target tissue and extravascular storage pool for vitamin D. In Sun's study, fat-free mass and muscle mass were significantly increased [77]. Similarly, in Agergaard's study, the cross-sectional area of the quadriceps muscle had significant gains in the group who did not receive vitamin D supplements [79]. This result suggests that increased muscle mass from resistance training provides a reservoir of vitamin D, leading to reduced or unchanged serum 25(OH)D levels.
Moreover, 25(OH)D can be released from skeletal muscle [69,[81][82][83][84]. This release is regulated by the VDR, PTH, VDBP, and vitamin D nutritional status [69,83,84]. PTH reduces the net uptake of 25(OH)D 3 in C2 myotubes and mouse muscle fibers and reduces its retention in myotubes [69]. In Barker's study, PTH levels significantly increased after acute resistance exercise [74]. In Zhang's study, there was a significant increase in PTH levels at 3 months and 1 year following chronic resistance exercise combined with vitamin D supplements, compared to only receiving vitamin D supplements [75]. The increase in circulating 25(OH)D levels may be due to the effect of PTH on its uptake and retention in skeletal muscle cells [74,75]. However, chronic resistance training alone did not significantly alter the PTH levels [52,77]. This may also be the reason why vitamin D supplementation combined with resistance training, and not resistance exercise training alone, increases 25(OH)D levels.
Resistance training can increase the level of CYP27B1 [37], which can catalyze the conversion of 25(OH)D to 1,25(OH) 2 D 3 . Moreover, resistance training can increase target tissue VDR levels [37], increasing 1,25(OH) 2 D 3 utilization, which may explain why serum 25(OH)D levels are not altered in response to resistance training [37]. In addition, resistance training increases CYP24A1 levels [56], which can degrade 25(OH)D and 1,25(OH) 2 D; hence, the decrease in 1,25(OH) 2 D may be caused by increased degradation, while its synthesis remains unchanged [56]. These factors may individually or together contribute to reduction/unchanged 25(OH)D levels in the circulation in response to resistance training.
How resistance training may exert its effect on 25(OH)D in various ways is briefly illustrated in Figure 2.
mentation combined with resistance training, and not resistance exercise training alone, increases 25(OH)D levels.
Resistance training can increase the level of CYP27B1 [37], which can catalyze the conversion of 25(OH)D to 1,25(OH)2D3. Moreover, resistance training can increase target tissue VDR levels [37], increasing 1,25(OH)2D3 utilization, which may explain why serum 25(OH)D levels are not altered in response to resistance training [37]. In addition, resistance training increases CYP24A1 levels [56], which can degrade 25(OH)D and 1,25(OH)2D; hence, the decrease in 1,25(OH)2D may be caused by increased degradation, while its synthesis remains unchanged [56]. These factors may individually or together contribute to reduction/unchanged 25(OH)D levels in the circulation in response to resistance training.
How resistance training may exert its effect on 25(OH)D in various ways is briefly illustrated in Figure 2.

Others
The effect of endurance combined with resistance exercise training intervention on 25(OH)D was investigated in three human studies and one animal study, which did not yield consistent results (Table 3). In the human studies, chronic endurance combined with resistance exercise training intervention significantly increased serum 25(OH)D levels [85,86]. Evans et al. found that 4 months of recruit training significantly reduced serum 25(OH)D levels in healthy men with adequate vitamin D {25(OH)D ≥ 20 ng/mL or 50 nmol/L [57]}, while no significant change was observed in healthy women with adequate vitamin D levels [87]. Conversely, Buskermolen et al. reported that 6 weeks of peak power combined with endurance training did not alter serum 25(OH)D levels in Wistar rats [52].

Limitations and Perspectives
Aside from the small number of relevant studies, there are many limitations. First, while mass spectrometry, enzyme-linked immunosorbent assays, and other methods can detect 25(OH)D levels, their accuracies vary greatly. Moreover, 25(OH)D 2 levels are difficult to detect [24]. Second, the vitamin D nutritional status is affected by exposure to season/sunlight. Except for the Sun study and the Li study, which clearly stated that chronic exercise intervention was conducted indoors [77] or outdoors [86], the vast majority of studies did not provide relevant information. Some studies did not provide seasonal information or information on sunlight exposure. Third, some studies did not include a blank control group. These limitations should be addressed in future research. In this paper, there are also some strengths. First, we relatively comprehensively summarize the relevant research in recent years. Second, due to the different effects of different exercise types on health, we focus on the analysis of the effects of endurance training and resistance training exercise. Third, from the perspective of the two major extra-circulating depots and the regulatory factors of vitamin D, this review comprehensively explained the possible mechanism of exercise on vitamin D.
Because the nutritional status of vitamin D is influenced by various factors, we recommend incorporating the following considerations in future studies. First, due to their lipid solubility, vitamin D metabolites are sequestered in adipose tissue, leading to decreased bioavailability in obese subjects [88]. Moreover, Drincic believed that because of volumetric dilution, obese individuals have lower 25(OH)D concentrations [89]. Therefore, body fat is significantly negatively correlated with serum 25(OH)D levels [90] and obese individuals have a higher risk of vitamin D deficiency [91,92]. In addition, lipolysis may be impaired in obese individuals [93], and obesity affects the regulation of vitamin D metabolism enzymes [94], which may explain why 25(OH)D levels in overweight and obese adults were not altered in the Lithgow study [48]. Second, exercise in the fed and fasted states differed in terms of energy metabolism substrates. A study found that exercise performed in the fasted state induces higher fat oxidation than exercise performed in the fed state [95]. Moreover, fasting increases post-exercise circulating FFAs [96]. Therefore, the effects of exercise on serum 25(OH)D or 1,25(OH) 2 D levels may be influenced by whether it is performed under fed or fasted states. Third, vitamin D metabolites are primarily found in circulation, adipose tissue, and skeletal muscle [63]. Thus, adipose and muscle tissues are two major extra circulatory depots for vitamin D metabolites, which are not reflected in serum 25(OH)D levels. Therefore, when studying the effect of exercise on vitamin D, extravascular storage tissues should be included in the analysis.

Conclusions
In conclusion, endurance exercise can significantly increase serum 25(OH)D levels in vitamin D-deficient subjects but has no significant effect on vitamin D-sufficient subjects. Moreover, resistance training did not significantly increase 25(OH)D concentrations. Only chronic endurance exercise intervention significantly increased serum 1,25(OH) 2 D levels, and this effect may be sex-dependent. Exercise may influence 25(OH)D levels in circulation by regulating either the release of vitamin D metabolites from storage tissues or the utilization of target tissue (Figure 3). The effects of exercise on 25(OH)D levels may depend on the vitamin D nutritional status, exercise type, exercise intensity, and sex. The organism is a complex entity, and vitamin D is tightly regulated by a variety of factors. Exercise elicits various bodily responses, and the effects of exercise on vitamin D nutritional levels may be the result of a combination of these. Therefore, further research on the effects and mechanisms of exercise on 25(OH)D levels is needed.  Institutional Review Board Statement: Not applicable.
Informed Consent Statement: Not applicable.

Data Availability Statement:
No new data were created or analyzed in this study. Data sharing is not applicable to this article.