Note: Very technical discussion of some of the ongoing research into the causes of Crohn's Disease and Ulcerative Colitis.

Vitamin D in IBD

Vitamin D deficiency is more common in adults and children with IBD, especially CD, than healthy controls, and correlates with a poorer health-related quality of life. Many factors are likely to contribute to this, including malabsorption secondary to mucosal disease or surgical resection, and reduced sunlight exposure, physical activity and dietary intake. An active inflammatory state, which results in reduced hepatic production of DBP, may cause a reduction in total 25(OH)D levels, but this has not specifically been studied in patients with IBD.

The relationship between the vitamin D axis and IBD appears to be a multi-faceted one, comprising maintenance of musculoskeletal health, and possibly control of disease activity through immunomodulation, and modification of the risk of IBD-associated malignancy.

Vitamin D and Musculoskeletal Health in IBD

The prevalence of low BMD is greater in patients with IBD than in healthy controls. It is estimated that 22–77% of patients with IBD have osteopenia, and 12–41% have osteoporosis. Most studies have reported that patients with CD have a greater prevalence of low BMD than those with UC, but some other studies have found similar rates. Bone loss occurs in both cortical and trabecular regions, though the former predominates in CD. Though glucocorticoid use is the most well-recognised risk factor for osteoporosis in IBD, reduced BMD is observed in patients with IBD in the absence of steroid use.  Other risk factors include older age, postmenopausal status, smoking, low body mass index, reduced physical activity, malnutrition and low vitamin D status. A chronic inflammatory state, with effects on osteoblast and osteoclast function mediated by cytokines such as TNF-α, IL-6 and IL-1β, likely contributes to bone loss.  The risk of low-trauma fractures in patients with CD is estimated to be increased by about 30%, and in UC about 20%, compared with controls.  The American Gastroenterology Association, the American College of Gastroenterology and British Society of Gastroenterology recommend BMD testing in patients with IBD aged 60 and above, cumulative exposure to glucocorticoids for ≥3 months, low BMI, family or personal history of low-trauma fractures, or hypogonadism. However, adherence to these recommendations by clinicians is suboptimal.

There is an absence of controlled prospective trials for fracture prevention specifically in patients with IBD. However, it is recommended that, along with control of disease activity, encouragement of physical activity and cessation of smoking, vitamin D and calcium supplementation should be given to those at moderate or high risk of fracture, and antiresorptive therapy in those at high risk. In one small prospective trial, 1,25(OH)2D2 supplementation reduced markers of bone turnover in patients with active CD. In another prospective trial in the early 1990s, 25(OH)D administration in 75 patients with CD for 1 year reduced BMD loss as measured by distal forearm absorptiometry. Bone protective therapies are underutilised in patients with IBD. One large review of over 2000 patients from 7 centres in USA found that only 59% and 75% of osteoporotic IBD patients received calcium/vitamin D supplementation and bisphosphonates, respectively.

Children and adults with IBD also have reduced muscle mass. Sarcopenia is reported in 60% of patients with CD. Increased apoptosis has been demonstrated in muscle biopsies in patients with IBD,[132] but the reasons for this have not yet been fully elucidated. Extrapolating from known effects of vitamin D insufficiency on muscle tissue, the optimisation of vitamin D status in patients with IBD may serve to preserve muscle health.

Vitamin D as an Immunomodulator in IBD

There are accumulating epidemiological, physiological, genetic and clinical data for a role of vitamin D in immunomodulation in IBD ( Table 1 ).

Epidemiological Associations There is a correlation between frequency of IBD and potential exposure to sunlight as indicated by distance from the equator. The incidence and prevalence of IBD is higher in northern Europe, North America, Australia and New Zealand than Asia. Also, the incidence of IBD in the Indian Subcontinent is low, but migrants to developed countries at Northern latitudes have a greatly increased risk of IBD. In the Northern hemisphere, the onset of UC and exacerbations of CD are noted to peak in winter months. Nerich et al. have recently reported that a graded relative risk of CD incidence, but interestingly not UC incidence, correlated with areas of low sunlight exposure within France as ascertained from a population wide health insurance system database. Furthermore, an analysis of 72 719 women aged 40–73 years enrolled in the Nurses' Health Study, using a predicted vitamin D level calculated from diet and lifestyle factors, has shown a reduced risk of CD (HR 0.54, 95% CI 0.30–0.99) and nonsignificantly reduced risk of UC (HR 0.65, 95% CI 0.34–1.25) in women with the highest quartile of vitamin D compared with the lowest quartile.

The interpretation of the associations between vitamin D levels and sunlight exposure on the one hand and the incidence of IBD on the other is confounded by numerous factors, and a causal relationship cannot be assumed. Nonetheless, the data do provide a foundation for the investigation of the physiological connection between the vitamin D axis and inflammation in IBD.

Evidence for Involvement in Inflammation and Immunomodulation Both UC and CD are characterised by a dysregulated mucosal immune response to intestinal microorganisms in a genetically susceptible host. Fascinating insights ascertained from characterisation of VDR and other vitamin D axis components in the gastrointestinal mucosa, as well as genetic associations, provide evidence for the potential involvement of vitamin D at several stages of initiation and perpetuation of inflammation in IBD (Figure 1) as follows.



Figure 1.

Postulated immunomodulatory role of vitamin D in IBD. (a) Crohn's disease. Vitamin D has been shown to promote transcription of the intracellular pattern recognition receptor NOD2 and inhibit pro-inflammatory cytokine production by macrophages and Th1 cells (TNF-α, IFN-γ and IL-21). (b) Ulcerative colitis. In both Crohn's disease and ulcerative colitis, vitamin D promotes intracellular bacterial degradation via cathelicidin, promotes the regulatory cytokine IL-10 secretion and regulatory T-cell function and potentially inhibits colonic crypt epithelial barrier pore-forming claudin-2 via PTPN2. A red '+' indicates a positive effect of vitamin D, and a red '−' indicates an inhibitory effect of vitamin D.

Maintenance of Epithelial Barrier The columnar epithelial monolayer lining the gastrointestinal tract from the stomach to the rectum acts as a crucial interface between the mucosa and lumen, serving as a physical barrier as well as antigen presenter and immune regulator. Epithelial cells are connected by intercellular junctions, comprising tight junctions and adherens junctions, collectively referred to as the apical junctional complex, and desmosomes. Altered intestinal permeability, resulting from defects in these junctions, may predispose to inflammation. Claudin-2, a pore-forming transmembrane protein that forms part of the tight junction, has been implicated in the pathogenesis of IBD. Phosphorylation and expression of claudin-2 is stimulated by signal transducer and activator of transmission (STAT) 1 and STAT3, which is induced by IFN-γ, which is in turn inhibited by the protein tyrosine phosphatise N2 (PTPN2). The gene coding for PTPN2 is a high-risk locus for IBD and type 1 diabetes mellitus and is particularly associated with colonic CD and UC. A recent ChIP-sequence genomic map has identified the gene bound by the VDR in CD as PTNP2. Hence, 1,25(OH)2D-VDR complex-induced PTPN2 expression may inhibit epithelial barrier pore formation and altered intestinal permeability.

In a mouse model of colitis, dextran sodium sulphate administration was demonstrated to reduce transepithelial electrical resistance and expression of the tight junction proteins Zo-1 and occludin prior to ulcers and clinical signs to a greater extent in VDR knockout (KO) mice than wild-type mice. Furthermore, administration of 1,25(OH)2D3 enhanced Zo-1 and E-Cadherin expression in CaCo-2 cell cultures and enhanced epithelial reconstitution following injury.

Innate Immune Response Intestinal epithelial, dendritic cells and macrophages (collectively referred to as antigen presenting cells) express pattern recognition receptors (PRRs), which enable continuous monitoring of luminal contents for commensal and pathogenic organisms through the recognition of conserved structures called pathogen-associated molecular patterns (PAMPs). Examples of PRRs include toll-like receptors (TLRs), nucleotide-binding domain and leucine-rich repeat-containing receptors (NLRs), C-type lectins and retinoic acid-inducible gene I-like receptors. The protein NOD2 (CARD15), part of the family of NLRs, recognises modified muramyl dipeptide, a PAMP which is the lysosomal breakdown product of bacterial peptidoglycan. More recently, viral elements have also been shown to induce NOD2. NOD2 has been associated with numerous regulatory roles in the mucosal immune system, with evidence for Paneth cell antimicrobial peptide generation, negative regulation of TLR signalling and hence induction of tolerance, and promotion of T-cell interleukin (IL)-10 expression. The NOD2 gene is the locus associated with the highest risk for CD so far identified and NOD2 mutations have been correlated with fibrostenosing CD. Significantly, dendritic cells, macrophages and intestinal epithelial cells express VDR, and 1,25(OH)2D3 has been shown to promote transcription of the NOD2 gene, highlighting an important link between the vitamin D axis and pathogenesis of IBD.

NOD2 has also been demonstrated to potentiate autophagy, the process by which damaged organelles, proteins and intracellular microorganisms are removed through engulfment into an autophagosome and lysosomally degraded. Autophagy-related 16-like 1 (ATG16L1) is a protein that forms an integral component of this process, and its recruitment is mediated by NOD2. Deficiency of ATG16L1 results in an exaggerated inflammatory response,[155] and the gene encoding it, ATG16L1, has been identified as another major CD susceptibility locus from genome wide association studies (GWAS). Hence, vitamin D signalling may indirectly promote regulation of inflammation through NOD2 and autophagy.

Vitamin D also directly participates in autophagy by potently stimulating cathelicidin, an antimicrobial peptide produced by macrophages. Cathelicidin plays an important role in defence against intracellular organisms, particularly mycobacteria. The expression of cathelicidin is significantly increased in inflamed and non-inflamed mucosal biopsies from patients with UC, but unaltered in patients with CD, which may imply a defect in mucosal defence in the latter condition. In human monocyte cultures, E. coli DNA induced expression of cathelicidin, a process dependent on TLR9 and MyD88 signalling. Furthermore, cathelicidin expression is increased in mice administered intra-colonic bacterial DNA, and cathelicidin KO mice develop more severe DSS-induced colitis than wild-type mice. In addition, intra-rectal cathelicidin administration ameliorated DSS-colitis.

TNF-α is a major cytokine implicated in inflammation and cytotoxicity in both CD and UC and is increased in the lamina propria in both conditions. TNF-α is believed to be primarily derived from macrophages in UC, but other cells within the lamina propria including T helper (Th) 1 cells contribute in CD. In peripheral blood mononuclear cells derived from patients with IBD, proliferation and secretion of TNF-α are reported to be significantly reduced by administration of the VDR agonist KH 1060.

The influence of vitamin D in the proliferation and maturation of APCs also extends to dendritic cells, the maturation of which is inhibited by both 1,25(OH)2D3[162] and its synthetic analogues. Recent data from microarray studies show that 1,25(OH)2D upregulates the expression of immune tolerogenic genes in dendritic cells.

Adaptive T-cell Response Perhaps, the largest body of in vitro and animal in vivo evidence for an immunoregulatory role for vitamin D in IBD regards the adaptive T-cell response. APCs activate T-cell responses through direct interaction between MHC class I and II receptors and the T-cell receptor (TCR), in the presence of co-stimulatory signals (CD 80/86 and CD40 on APCs and CD28 and CD40L on T cells). CD is characterised by a Th1 and Th17 CD4+ response, whereas UC comprises a Th2-like response. The Th1 cell response and secretion of IFN-γ is induced by IL-12, which is derived from dendritic cells secondary to PRR signalling. The Th17 response and production of IL-17 is stimulated by TGF-β and IL-6, in the presence of IL-23, which are also derived from APCs. In the absence of IL-23, Th17 cells produce IL-10, an anti-inflammatory cytokine. In contrast, the UC mucosal immune response predominantly consists of natural killer T cell production of IL-13 and, to a lesser extent, IL-5. Th1 and Th17 cells are also noted to be increased in number in the mucosa in UC. Recently, IL-21, produced by Th1 and T follicular helper cells, has been recognised to promote Th17 cells and autoimmunity, as well as germinal centre B cells. IL-21 is overexpressed in inflamed mucosa in CD and UC.

VDR is expressed on activated T cells. 1,25(OH)2D3 has been shown to inhibit dendritic cell production of IL-12, and CD4+ T-cell production of IFN-γ, IL-17 and IL-21. In addition, 1,25(OH)2D3 stimulates expression of dendritic cell production of IL-10, and T-cell levels of CTLA-4 (an inhibitory co-stimulatory signal) and FoxP3 (a lineage specification factor of regulatory T cells), further enhancing its anti-inflammatory effect.

Pathogen-free VDR/IL-10 double KO mice develop fulminant DSS-induced colitis, compared with lack of colitis seen in pathogen-free IL-10 KO mice. VDR KO mice also have reduced IL-10 and anti-inflammatory intra-epithelial CD8αα lymphocyte levels. IL-10 KO mice develop worse colitis and have a lower survival when fed a vitamin D deficient diet. Vitamin D supplementation ameliorates and blocks the progression of colitis in IL-10 KO mice. In 2,4,6-trinitrobenzenesulfonic acid-induced colitis mouse models, treatment with 1,25(OH)2D3 reduces expression of IL-6, IL-17, IL-12p70 and IL-23p19 and increases expression of regulatory T-cell markers IL-10, TGF-β, FoxP3, CTLA4 and Th2 markers IL-4 and GATA3. The experimental VDR agonist, BXL-62, inhibits DSS-induced colitis in mice.

In PBMCs of humans with IBD, BXL-62 reduces pro-inflammatory cytokines TNF-α, IL-12/23p40, IL-6 and IFN-γ, both at mRNA and protein level. In peripheral blood CD4+ T cells isolated from patients with IBD, 1,25(OH)2D3 reduces IFN-γ and increases IL-10 production, alone and in combination with dexamethasone.

Similar benefits have been reported in experimental models of other Th1-mediated autoimmune diseases, including multiple sclerosis, type 1 diabetes and rheumatoid arthritis.

Genetic Associations Genetic polymorphisms in components of the vitamin D axis have been associated with IBD risk. Genome screening in Caucasians suggests the TaqI tt genotype is the VDR genotype over-represented in CD, with a prevalence of 22% and an odds ratio of 1.99 (95% CI 1.14–3.47; P = 0.017). This increased frequency was replicated but limited to males with IBD in two other cohorts. Interestingly, immune modulation by vitamin D may be associated with the VDR polymorphisms. VD3 supplementation of antituberculosis therapy was only significantly superior in patients with the TaqI tt genotype. A study in Iranian IBD patients, contrastingly, revealed an association with the FokI f allele. A further large study in Irish IBD patients found no statistically significant association with VDR genotype.

Recently, a reduced frequency of Gc-2 alleles was reported in 636 IBD patients compared with 248 non-IBD controls, with a significant association for both CD and UC patients.[182] The mechanism by which this allele may reduce risk of IBD is uncertain. However, given that the Gc-2 allele confers a lower affinity for 25(OH)D and 1,252(OH)D than Gc-1f and Gc-1s alleles, one may speculate that vitamin D metabolites are less freely available for immunoregulatory functions with the latter alleles in patients with IBD.

Clinical Evidence for Vitamin D as an Immunomodulator in IBD Small human clinical trials have suggested that vitamin D supplementation may have immunomodulatory activity in IBD. In a nonblinded trial of 37 patients with CD in clinical remission as defined by Crohn's Disease Activity Index (CDAI) <150, 18 patients administered 0.5 μg alfacalcidol (1(OH)D3, a vitamin D analogue) had a superior improvement in CDAI and CRP over a 6-week period compared with 17 patients given 2000 IU cholecalciferol over a 6-week period.

A randomised controlled trial of 94 CD patients with steroid-free remission (CDAI < 150, normal CRP and normal albumin) recently demonstrated a nonsignificant reduction in risk of relapse (13% vs. 29% at 12 months, P = 0.06) with 1200 IU cholecalciferol daily compared with placebo. Somewhat surprisingly, a subgroup analysis of peripheral blood mononuclear cells isolated from CD patients treated for 26 weeks with cholecalciferol (n = 10) demonstrated increased IL-6 production and CD4+ T-cell proliferation compared with placebo (n = 10). This apparently paradoxical finding in the presence of clinical improvement may be explained by a dual function of the IL-6 cytokine family, which may also be cytoprotective via downstream effects under certain conditions as well as being pro-inflammatory.

More recently, findings from a small trial of 15 patients demonstrated a significantly improved clinical symptom score in patients with CD given 10 000 IU oral vitamin D daily compared with patients given 1000 IU vitamin D daily at week 26.

Interestingly, VDR mRNA expression is reduced in colonic biopsy specimens in patients with CD and UC, and immunohistochemically localised VDR protein is less frequently noted in colonic specimens from patients with UC than normal controls. Furthermore, vitamin D3 supplementation has been shown to induce VDR mRNA expression in a variety of tissues.



(reproduced from Medscape with permission)