Introduction

Inflammatory bowel disease (IBD), whose major subtypes include Crohn’s disease (CD) and ulcerative colitis (UC), is characterized by chronic relapsing and remitting inflammation of the gastrointestinal tract. The etiology of IBD is yet unknown, although its pathogenesis is believed to partly stem from a dysregulated immune response to commensal intestinal bacteria and environmental factors in a genetically predisposed individual. Much progress has been made in identifying genetic risk factors for IBD [1], although the precise environmental triggers are still unclear.

Vitamin D is traditionally known for its involvement in calcium and phosphorus homeostasis but has more recently been implicated in immune regulation. Animal, in vitro, and epidemiologic studies have since associated vitamin D deficiency with diverse autoimmune disorders, while emerging data now suggest that vitamin D may also be an environmental factor in IBD pathogenesis. In this review, we will discuss the mechanisms of vitamin D action, its putative effects on the immune system, and its proposed involvement in IBD pathogenesis.

Vitamin D Synthesis and Action

Vitamin D comprises a group of fat-soluble secosteroid hormones that primarily includes vitamin D2 (ergocalciferol) and vitamin D3 (cholecalciferol). As humans cannot synthesize ergosterol, the vitamin D2 precursor, ergocalciferol largely derives from exogenous sources, such as the diet and pharmacologic supplementation [2]. Cholecalciferol, on the other hand, derives from the endogenous conversion of 7-dehydrocholesterol to pre-vitamin D3, and subsequently vitamin D3, during ultraviolet B irradiation (280 to 315 nm) of the epidermis [3, 4]. While bound to the vitamin D binding protein (DBP) or albumin [5], vitamin D is transported to the liver where it is hydroxylated to form 25-hydroxyvitamin D (25[OH]D) [6]; it is then transported to the renal tubules where it is converted to the active metabolite: 1,25-dihydroxyvitamin D (1,25[OH]2D) [7]. At the target tissues, 1,25(OH)2D binds to the vitamin D receptor (VDR) in the cytosol [8], translocates into the nucleus, selectively binds the vitamin D response element (VDRE), and facilitates transcription of target genes [9–11].

Vitamin D and Immunity

For decades, vitamin D had uniquely been associated with calcium, phosphorus, and bone metabolism. However, an expanded role for vitamin D was considered after the discovery of its possible effects on leukemia cell differentiation in 1981 [12]. Not long thereafter, molecular receptors for vitamin D were found in peripheral mononuclear leukocytes [13, 14], later followed by their identification in diverse hematopoietic cell lines [15]. The potential involvement of vitamin D in immune regulation was further bolstered by the identification of macrophage- and dendritic cell-mediated production of the active 1,25(OH)2D metabolite, suggesting the intracellular and extrarenal presence of 25(OH)D-1-α-hydroxylase [16, 17].

In macrophages, vitamin D can promote immune function, such as monocyte differentiation, surface antigen presentation, production of reactive oxygen metabolites, and lysosomal enzyme activity [18, 19]; these processes are impaired in vitamin D deficiency. In monocytes, vitamin D downregulates Toll-like receptor (TLR) expression, leading to an impaired inflammatory response to lipopolysaccharide (LPS) [20]. There is additionally dose-dependent suppression of LPS-induced tumor necrosis factor-alpha (TNF-α) production. This downregulation of TLR expression and hyporesponsiveness to bacterial cell wall antigens are reversible through VDR antagonism. In lymphocytes, vitamin D has been demonstrated to impair proliferation and differentiation [21–23]. Vitamin D additionally interferes with dendritic cell differentiation, maturation, and IL-12 secretion, thus leading to inhibition of alloreactive T lymphocyte activation and relative hyporesponsiveness [24, 25]. Vitamin D and its analog calcipotriol have been demonstrated to promote regulatory T cell production [26–28]. Vitamin D promotes synthesis of the anti-inflammatory cytokine IL-10, while inhibiting synthesis of pro-inflammatory cytokines [20, 21, 28–32]. It also directly inhibits transcription of the granulocyte-macrophage colony-stimulating factor (GMCSF) gene [33].

Vitamin D and Autoimmune Disease

Vitamin D has been implicated in the pathogenesis of several autoimmune diseases. For one, experimental autoimmune encephalomyelitis (EAE) is an inducible murine model of multiple sclerosis (MS) that involves a Th1 response against myelin basic protein and other autoantigens in the central nervous system. Administration of 1,25(OH)2D was found to reversibly prevent development or progression of EAE [34, 35]. Interestingly, severe vitamin D deficiency interferes with the pathogenesis of EAE in mice [36]. This seeming paradox may stem from the impairment of cell-mediated immunity in a state of severe vitamin D deficiency [37].

In humans, a prospective evaluation of dietary vitamin D intake in the Nurses’ Health Study (NHS) and Nurses’ Health Study II (NHS II) revealed that women in the highest quintile of vitamin D intake had a 33 % lower age-adjusted risk of MS [38]. Likewise, intake of vitamin D supplements was associated with a 41 % reduction in risk of MS. These findings were later corroborated in another prospective study that showed a dose-response trend, where higher serum 25(OH)D levels were associated with decreased odds of incident MS [39]. Restriction site polymorphisms on the VDR gene have also been linked with an increased risk of MS [40, 41]. Similar to the NHS for MS, an analysis of 29,520 women (ages 55–69) in the Iowa Women’s Health Study revealed that women with greater intake of dietary and supplementary vitamin D had a lower risk of rheumatoid arthritis (RA) [42]. Vitamin D supplementation has also been shown to inhibit progression of infection- or collagen-induced arthritis in mice [43].

The first reported observation of vitamin D deficiency in systemic lupus erythematosus (SLE) included 7 out of 12 prednisone-treated adolescents who were found to have low 1,25(OH)2D levels [44]. Since that report in 1979, there have been over 20 additional published studies, mostly correlating vitamin D deficiency with SLE and/or disease activity [45]. The studies were all small, which may have limited the power in the few that did not find statistical significance. Nonetheless, the largest study had a multi-center European and Israeli cohort of 378 SLE patients in whom the investigators found an inverse relationship between 25(OH)D levels and disease activity scores [46].

The evidence of vitamin D’s role in other autoimmune diseases is less robust, largely due to a dearth of data, but still suggests a relationship between vitamin D deficiency and disease pathogenesis. For autoimmune diabetes, 1,25(OH)2D treatment was shown to arrest development or progression of diabetes in non-obese diabetic (NOD) mice, an animal model for human autoimmune diabetes (insulin-dependent diabetes mellitus or juvenile diabetes) [47–49]. There are also emerging data for a role of vitamin D in other immunologic conditions, such as asthma and atopy [50–52]. Taken together, the accumulating in vitro, in vivo, and epidemiologic data implicate vitamin D in immune-related disease.

Vitamin D and Inflammatory Bowel Disease

The relationship between vitamin D and IBD has been explored in animal models, in vitro assays, and human observational studies. Although none of the individual studies confirm that vitamin D deficiency leads to IBD in humans, the totality of evidence is compelling.

Animal Models

In the IL-10 knockout mouse model, the mice may spontaneously develop enterocolitis. Vitamin D deficiency in these mice accelerates the development of enterocolitis, while also leading to more severe diarrhea, wasting disease, and death [53]. However, treatment with 1,25(OH)2D leads to amelioration of these symptoms, and treatment of calcitriol in 2,4,6-trinitrobenzene sulfonic acid (TNBS)-induced colitis is associated with a decrease in Th1/Th17 cells in favor of Th2 and regulatory T cells [28]. Similarly, administration of a VDR agonist attenuates colitis in IL-10 knockout mice or TNBS-induced colitis [54, 55]. VDR agonism also reduces expression of pro-inflammatory cytokines and lymphocyte infiltration in the lamina propria of mice with dextran sodium sulfate (DSS)-induced colitis [56].

Genetic Evidence

VDR knockouts serve as another model of vitamin D deficiency by ablating a step along the putative pathway of vitamin D’s mechanism of action. As such, VDR and IL-10 double knockout mice have been observed to develop severe colitis, although these symptoms are not as readily apparent in either VDR or IL-10 single knockouts [57–59]. These findings illustrate that functional vitamin D deficiency can trigger IBD in a susceptible mouse model and that vitamin D deficiency alone may not be sufficient for pathogenesis. This would be consistent with observations in humans where the widespread prevalence of vitamin D deficiency far exceeds the prevalence of IBD.

In humans, polymorphisms in the VDR gene have been associated with IBD but appear to vary according to population. In European Caucasian patients, the TaqI polymorphism on the VDR gene is significantly more common in CD patients than UC patients or controls [60]. In Iranian patients, the FokI polymorphism was more common in CD and UC patients [61]. In Jewish Ashkenazi patients, the BsmI polymorphism is associated with UC, but not CD [62]. A larger study of 1359 Irish participants (660 IBD patients) did not find any significant association between VDR gene polymorphisms in FokI, BsmI, ApaI, or TaqI and IBD [63]. While this finding may contradict prior studies, the population was homogenously Irish and prior studies have demonstrated variations in significant polymorphisms based on racial backgrounds. Relevant associations differed among studies, but they were limited by small cohorts. However, in the highly powered IBD immunochip study, there was genome wide evidence of association establishing a VDR intronic polymorphism (rs11168249) as a risk factor for IBD, although the effect size was small (OR 1.05) [1]. Homozygotes for the Thr420Lys mutation in the vitamin D binding protein (DBP) have also been associated with CD and UC, although the true functional effect of these variations remains unclear [64].

A link between vitamin D and CD risk genetics has additionally been demonstrated through in vitro experiments of monocytic and epithelial cells, where 1,25(OH)2D induced expression of NOD2/CARD15, defensin β2, and cathelicidin in the presence of muramyl dipeptide, but not in cells with homozygous non-functional NOD2 [65]. Inadequate recognition or clearance of bacterial antigens may contribute to the abnormal immunologic response in CD, although the specific mechanisms for NOD2- or ATG16L1-mediated risk of CD are still unclear. These findings nonetheless indicate a possible mechanism of gene-environment interaction in CD pathogenesis.

Epidemiologic Evidence

On a population level, epidemiologic observations have revealed a north-south gradient for IBD, where more northern geographic regions with less sunlight exposure have a higher incidence of disease [66–70]. The European Collaborative Study on Inflammatory Bowel Disease (EC-IBD) reported 80 % and 40 % greater incidence of CD and UC, respectively, when comparing northern and southern European centers [71]. Similar latitudinal trends have been noted in individual European countries, such as in France [68, 69, 72] and Scotland [66]. One French study went further to compare surface UV radiation intensity and IBD incidence [69], and found that low sunlight exposure was associated with an increased incidence of CD, but not UC. Retrospective analyses of the Caris Life Science database and the Nurses’ Health Study (NHS), a prospective cohort of 72,719 women (ages 40 or above), revealed similar north-south incidence trends in the USA [73, 74]. A separate study of the NHS additionally found an inverse correlation between predicted 25(OH)D levels (indirectly estimated in regression models from measured factors, such as dietary and supplement intake, sunlight exposure, race, and body mass index) and CD incidence [75•].

Beyond its putative role in IBD pathogenesis, vitamin D may additionally exert effects on IBD severity. These findings do not prove that vitamin D deficiency increases IBD risk but provides some insight into the potential role it plays in IBD pathophysiology. A large retrospective study of 403 CD and 101 UC patients found that vitamin D deficiency was associated with increased disease activity and decreased quality of life in CD, but not UC [76]. Nonetheless, a separate retrospective study of 34 UC patients found an inverse association between vitamin D levels and disease activity, measured using the Mayo index [77]. An Indian study compared vitamin D levels and disease characteristics between 34 CD patients and 34 age- and sex-matched controls with irritable bowel syndrome [70]. The study demonstrated that patients with mild disease appeared to have similar vitamin D levels as the controls, but patients with more moderate to severe disease activity appeared to have significantly lower vitamin D levels. Similarly, vitamin D deficiency prior to the initiation of anti-TNF therapy had a threefold greater risk of earlier loss of treatment response among CD patients, but not UC patients [78].

The temporal relationship between vitamin D and IBD activity was, however, unclear in these studies. Nonetheless, using sunlight exposure as a surrogate for vitamin D status, low ultraviolet exposure was ecologically associated with increased risk of hospitalization, prolonged length of stay, and IBD-related surgery among CD and UC patients [79]. These trends were not observed for non-IBD surgeries. On a patient level, a collaboration among several medical centers in Boston retrospectively evaluated the effects of vitamin D deficiency and normalization of deficient levels in 3217 IBD patients [80•]. Consistent with earlier findings, vitamin D level was associated with a twofold risk of IBD-related surgery and hospital admissions among CD and UC patients. Vitamin D-deficient CD patients who had normalization of their 25(OH)D levels had a reduced risk of surgery, but not hospitalization. However, vitamin D-deficient UC patients who had normalization of 25(OH)D levels did not experience a change in their risk of surgery or hospitalization.

Intervention Trials

If vitamin D clinically influenced the natural course of IBD, one would surmise that vitamin D supplementation could serve as an adjunctive therapy for induction and/or maintenance of remission. A randomized placebo-controlled trial of 1200 IU of cholecalciferol daily in 94 total CD patients showed improvement in serum 25(OH) levels after 3 months [81•]. The relapse rate in the cholecalciferol arm was lower but did not reach statistical significance (13 vs. 29 %; P = 0.06). The study may have been underpowered and/or the effect of vitamin D may be not as strong as hoped. Similarly, a Hungarian trial of 1,25(OH)2D supplementation in CD patients found improvement in disease activity indices and C-reactive protein (CRP) at week 6 [82]. Another study that used adaptive methods for vitamin D3 supplementation (either 1000 IU daily with progressive escalation or a fixed 5000 IU daily) in CD patients with mild-moderate disease found that vitamin D repletion to 40 ng/mL was accompanied by improvement in 25(OH)D levels, disease activity indices, and quality of life scores [83]. Most patients required the maximum allotted 5000 IU of daily cholecalciferol to reach a serum 25(OH)D level of 40 ng/mL.

These studies highlight the value of repleting and maintaining sufficient vitamin D levels among IBD patients to augment medical therapies and to reduce the risk of flares, need for surgical intervention, or hospitalization.

Cause or Effect?

Vitamin D deficiency can be commonly found in children and adults with IBD [76, 84–89]. After adjusting for dietary intake and seasonal variation, one study reported significantly lower 25(OH)D3 levels in CD patients when compared with healthy controls [89]. While inflammation and diarrhea can explain malabsorption of the fat-soluble vitamin D [90, 91], absorption and bioavailability are still on average 30 % lower than normal in patients with quiescent disease [91]. Therefore, unlike with other immune-related diseases (e.g., MS, RA, SLE) that have been linked to vitamin D deficiency, the nature of vitamin malabsorption in IBD confounds the interpretation of current evidence. It is therefore unclear whether vitamin D deficiency is a cause or effect of IBD in humans.

This question of cause or effect becomes particularly important for observational studies that attempt to link vitamin D deficiency with disease pathogenesis. On one hand, the large Nurses’ Health Study showed convincing evidence of lower predicted serum 25(OH)D levels prior to CD diagnosis [75•]. A major caveat is the fact that serum 25(OH)D was indirectly predicted based on surveyed data of dietary choices, supplemental D intake, sunlight exposure, regional UVB intensity, body mass index, and race. Direct measurement of serum 25(OH)D prior to the onset of IBD is yet absent due to significant logistical and financial challenges in prospectively identifying individuals in the general population at risk for developing IBD and collecting pre-disease serum samples.

Moreover, the temporal causation of vitamin deficiency and actual disease onset has not yet been established. Subclinical disease of unclear duration can exist prior to formal diagnosis. Among children, a change in growth velocity can serve as a surrogate marker for disease onset and has been found to typically precede diagnosis by 3 to 78 months [92]. Similarly, two cross-sectional studies of asymptomatic siblings of CD patients revealed elevated fecal markers of intestinal inflammation compared to healthy controls, suggesting the presence of subclinical disease [93, 94]. Even if pre-diagnosis serum were available, it would be important to determine whether this was collected before the onset of subclinical disease, which in itself can lead to vitamin D malabsorption and subsequent deficiency. Therefore, given the currently available evidence, the question remains as to whether vitamin D deficiency is truly a cause or effect of IBD in humans.

Conclusions

The serendipitous discovery of VDR in immune cells opened the doors for investigation of the relationship between vitamin D and immunoregulation. Vitamin D deficiency has since been shown to influence pathogenesis of autoimmune disorders. Animal studies and epidemiologic data suggest that the same relationship may exist in IBD, more consistently for CD than UC. However, unlike other non-gastrointestinal autoimmune disorders, IBD can itself reduce vitamin D levels due to malabsorption. The dilemma of cause or effect is a pertinent challenge in establishing vitamin D deficiency as an environmental risk factor of IBD. Direct vitamin D measurement of pre-disease samples is needed. Clinical trials on the effects of vitamin D supplementation on disease severity are also lacking. There are intriguing signals that vitamin D influences IBD pathogenesis and severity, thus motivating the need for further investigation into the role of vitamin D in IBD and its potential use as a therapeutic adjunct for IBD.