It is now well established that the physiological importance of the vitamin D status extends far beyond the regulation of bone metabolism. According to its manifold functions in immune homeostasis, increasing evidence relates serum vitamin D levels as well as polymorphisms in enzymes involved in vitamin D metabolism to the incidence of chronic inflammatory diseases like asthma, atherosclerosis and autoimmune diseases (Stojanovic et al., 2011; Summerday et al., 2012; Szekely and Pataki, 2012). However, whether vitamin D exerts a salutatory or deteriorating role in such diseases is still under debate. This review will focus on the knowledge regarding the role of vitamin D in inflammatory diseases by the examples of asthma, atherosclerosis and autoimmune diseases.

According to the World Health Organization (WHO), asthma is the most common chronic disease among children (http://www.who.int/mediacentre/factsheets/fs307/en/index.html). In this context, several studies addressed the interrelationship of the maternal as well as infant vitamin D status and the prevalence and severity of asthma. Three studies by Brehm et al. analyzed the relationship between vitamin D levels and asthma severity in Costa Rican, North American and Puerto Rican children, respectively (Brehm et al., 2009, 2010, 2012). Collectively, they found high prevalences of vitamin D insufficiency in asthmatic children and vitamin D insufficiency was correlated with severe asthma exacerbations. However, the prevalence of vitamin D insufficiency was high in Puerto Rican children irrespective of the indisposition from asthma, with roughly comparable percentages between asthma patients and otherwise healthy children (Brehm et al., 2012). Although few studies showed no correlation between serum vitamin D levels and the presence of asthma (Menon et al., 2012; Gergen et al., 2013), many studies state a higher prevalence of vitamin D deficiency in asthmatic children (Freishtat et al., 2010; Chinellato et al., 2011a,b; Ehlayel et al., 2011; Hollams et al., 2011; Bener et al., 2012; Krobtrakulchai et al., 2013) and adults (Li et al., 2011b). Additionally, in many cases a relation between low vitamin D levels and reduced asthma control is found. Furthermore, metabolomic analysis of breath condensates revealed reduced levels of vitamin D metabolites in children with asthma (Carraro et al., 2013). Similarly, enhanced vitamin D binding protein levels were found in bronchoalveolar lavage fluid of asthmatic children (Gupta et al., 2012b). Interestingly, one study describes an age-dependent association between serum vitamin D level and asthma prevalence in children (Van Oeffelen et al., 2011).

A different relationship between the vitamin D status and asthma has been brought up by a northern Finland birth cohort study, which revealed an increased risk of asthma in adults who received high dose vitamin D supplementation in their childhood (Hypponen et al., 2004). In accordance with these findings, a prospective study by Tolppanen et al. revealed an increased risk of wheezing in association with higher vitamin D levels, but no correlation of lower vitamin D levels to respiratory sicknesses (Tolppanen et al., 2013). Another study reinforces the finding of increased susceptibility to asthma after vitamin D supplementation, yet only regarding supplementation of water soluble formulations and not in connection with vitamin D supplementation in peanut oil (Kull et al., 2006).

There is debate as to whether maternal vitamin D levels during the pregnancy influence the susceptibility to asthma of the progeny. Whereas some reports showed no correlation between maternal or cord blood vitamin D levels and an increased risk of childhood asthma (Camargo et al., 2011; Rothers et al., 2011; Morales et al., 2012; Pike et al., 2012; Magnus et al., 2013), another report indicates that high maternal vitamin D levels correlate with enhanced probability of asthma development in children (Gale et al., 2008). In contrast, some reports associate higher vitamin D intake during pregnancy with reduced risk of childhood wheezing and asthma (Camargo et al., 2007; Devereux et al., 2007; Erkkola et al., 2009).

Mechanistically, vitamin D induced protection against airway inflammation has been related to a modulated T cell response to allergens as well as induction of the immunoglobulin-like anti-inflammatory cell surface protein CD200 on T cells, that acts on target immune cells which express the CD200 receptor (CD200R) (Dimeloe et al., 2012; Gorman et al., 2012; Urry et al., 2012). Many authors suggest that the beneficial effect of sufficient vitamin D levels on asthma development results from the immune enhancing effect of vitamin D and the simultaneous prevention of respiratory infections (Ginde et al., 2009; Camargo et al., 2011; Majak et al., 2011; Morales et al., 2012).

Furthermore, there is evidence that the serum vitamin D level has also an influence on asthma therapy, as vitamin D has been demonstrated to enhance glucocorticoid (GC) action and lower serum vitamin D levels are associated with higher corticosteroid requirement, at least in children, or even therapy-resistance (Searing et al., 2010; Goleva et al., 2012; Gupta et al., 2012a; Wu et al., 2012). Additionally, the therapeutic effect of specific allergen immunotherapy has been correlated to serum vitamin D levels (Majak et al., 2012).

Besides serum vitamin D levels also polymorphisms of genes of the vitamin D pathway such as the vitamin D receptor (VDR) have been associated with asthma (Poon et al., 2004; Raby et al., 2004; Saadi et al., 2009; Li et al., 2011a; Pillai et al., 2011; Maalmi et al., 2013), yet, not all studies revealed a correlation between vitamin D pathway polymorphisms and asthma prevalence (Vollmert et al., 2004; Fang et al., 2009).

Another chronic inflammatory disease that is more prevalent in the elderly population is atherosclerosis. Early studies on atherosclerosis development in several animal models revealed an accelerating effect of high doses of vitamin D. Vascular calcification was observed in some of these studies, but not all (Zemplenyi and Mrhova, 1965; Kudejko, 1968; Taura et al., 1979; Kunitomo et al., 1981; Toda et al., 1983, 1985). Moreover, 1α,25-dihydroxyvitamin D₃, the active form of vitamin D, stimulated vascular calcification by in vitro by reducing the expression of parathyroid hormone-related peptide as well as stimulating alkaline phosphatase activity in bovine vascular smooth muscle cells (Jono et al., 1998). On the other hand, there is a large body of research from clinical studies in humans indicating that low levels of serum 25-hydroxy vitamin D are associated with atherosclerosis (Reis et al., 2009; Carrelli et al., 2011; Shanker et al., 2011; Cheraghi et al., 2012). In line with this, the incidence of osteoporosis, a disease known to be related to vitamin D inadequacy, correlates with the incidence of atherosclerosis (Stojanovic et al., 2011). Therefore, different mechanisms may account for the promotion of atherogenesis by high and low vitamin D levels, respectively, and calcification may be crucial in the case of hypervitaminosis. Moreover, differences between the animal and human system may account for the conflicting results.

With respect to atherogenesis, 1α,25-dihydroxyvitamin D₃ has been demonstrated to reduce macrophage adhesion and migration as well as foam cell formation in monocytes isolated from type 2 diabetic patients (Oh et al., 2012; Riek et al., 2013a,b). Mechanistic investigations in the context of these studies attributed the beneficial effects of vitamin D to a reduction of endoplasmatic reticulum stress in macrophages. This has been investigated in two mouse models, where vitamin D deficiency facilitated atherosclerosis, which could be reversed in the course of macrophage endoplasmatic reticulum stress suppression (Weng et al., 2013). Further evidence on beneficial effects of calcitriol treatment on atherosclerosis development has been obtained from an investigation with apolipoprotein E knock-out mice. In this study, oral calcitriol treatment decreased the production of proinflammatory chemokines, led to a reduced amount of inflammatory effector cells in atherosclerotic plaques and simultaneously increased amounts of regulatory T cells (Takeda et al., 2010). A similar link between vitamin D, T cell modulation, and atherosclerosis has also been established in humans with chronic kidney disease (CKD) (Yadav et al., 2012).

The renin-angiotensin-system is known for its detrimental effects on the cardiovascular system and has been shown to play an important role in the development of atherosclerosis. Interestingly, numerous studies in mice document that vitamin D signaling suppresses the renin-angiotensin-system and that vitamin D deficiency is associated with an increased activity of the renin-angiotensin-system (Li et al., 2002; Zhou et al., 2008; Szeto et al., 2012; Weng et al., 2013). Moreover, the inverse associations which are described for vitamin D and the occurrence of inflammatory cytokines, C-reactive protein, and adhesion molecules suggest a inhibitory role for vitamin D in the genesis of atherosclerosis (Brewer et al., 2011). Additionally, there is experimental evidence that vitamin D reduces the expression of matrix metalloproteinases that are involved in vascular calcification (Nakagawa et al., 2005; Qin et al., 2006).

However, there are also studies that found no evidence for an association between low vitamin D and atherosclerosis in patients suffering from different autoimmune diseases (Mok et al., 2012; Sachs et al., 2013). Similarly, there was no evidence for an association of BsmI polymorphism, an intronic single nucleotide variation of the VDR gene, with atherosclerosis (El-Shehaby et al., 2013). Yet, it has been shown that atherosclerosis in monkeys is associated with low levels of VDR expression in coronary arteries even in the presence of higher plasma vitamin D concentrations (Schnatz et al., 2012a,b). Moreover, the activation of vitamin D can occur locally in macrophages that infiltrate atherosclerotic lesions and local vitamin D response might thus not necessarily correlate with serum vitamin D levels (Richart et al., 2007).

It is well established that vitamin D plays an important role in the regulation of immune functions (Schwalfenberg, 2011; Zhang et al., 2013a). Accordingly, several inflammatory autoimmune diseases like rheumatic disorders and type 1 diabetes have been associated with vitamin D deficiency (Adorini and Penna, 2008; Shapira et al., 2010). Inflammatory processes in the central nervous system are a hallmark of the autoimmune disease multiple sclerosis (MS) (Deckx et al., 2013). Several studies indicate that MS patients have lower levels of vitamin D and that higher vitamin D levels as well as vitamin D supplementation have a protective effect against MS (Munger et al., 2004, 2006; Ozgocmen et al., 2005). Moreover, vitamin D levels have been shown to vary in concordance with MS exacerbations (Correale et al., 2009) and it is possible that low vitamin D levels are rather a consequence of the sun avoidance of MS patients and not a direct cause of the disease (Munger et al., 2006). Regarding the effectiveness of vitamin D supplementation in the course of MS treatment, there are studies in mice and humans that suggest a beneficial effect of treatment (Goldberg et al., 1986; Wingerchuk et al., 2005; Pedersen et al., 2007; Burton et al., 2010). Interestingly, a gender specific effect of vitamin D has been demonstrated in mice and humans, which points to greater effects of vitamin D in females (Spach and Hayes, 2005; Correale et al., 2010).

Overall, there have been only a few controlled trials documenting the outcome of vitamin D supplementation on disease activity in rheumatic conditions, and the role of vitamin D in rheumatoid arthritis is therefore controversially discussed (Gatenby et al., 2013). Yet, a metaanalysis of observational studies on the vitamin D intake and vitamin D serum levels suggests an inverse association with rheumatoid arthritis (Song et al., 2012). Additionally, in vitro experiments with macrophages from healthy donors and rheumatoid arthritis patients indicate an enhanced anti-inflammatory potential of vitamin D in macrophages from the latter group (Neve et al., 2013).

It has been shown that the onset of autoimmunity in type 1 diabetes is preceded by a proinflammatory metabolic serum profile (Knip and Simell, 2012). Concurrently, a study in Italian children revealed reduced vitamin D serum levels in children at the onset of type 1 diabetes compared to children hospitalized for other reasons (Franchi et al., 2013). In conformity with these findings, metaanalyses suggest an association between vitamin D intake in early life and susceptibility for type 1 diabetes (Zipitis and Akobeng, 2008; Dong et al., 2013).

For inflammatory bowel disease (IBD), another autoimmune disorder, similar associations to that described above regarding vitamin D status and sunlight exposure have been reported (Garg et al., 2012; Ananthakrishnan, 2013). Animal studies in vitamin D deficient and VDR knockout (KO) mice reveal a dysregulation of T cells that might be of importance in the pathogenesis of IBD (Ooi et al., 2012).

In summary, there is considerable evidence for an association between vitamin D deficiency and inflammatory diseases. However, regarding the causality of this association and the benefit of vitamin D supplementation, only limited information is available and the existing data are still inconsistent.

Cell type specific up-regulation of proinflammatory genes and down-regulation of anti-inflammatory genes is a hallmark of the onset of an inflammatory reaction. Depending on the cell type, up-regulation of certain cytokines or enzymes which generate mediators of inflammation can occur at the transcriptional or posttranscriptional level. In addition, there is considerable crosstalk between various pathways which allows adaptation of the host defense reactions to the environment. According to their functions, the regulators of inflammatory reactions can be receptors such as toll like receptors, signal transducers as well as transcription factors which translate the activation of certain signal cascades into gene transcription. Additionally, regulation of gene expression during inflammatory processes can also occur on posttranslational level which is not focus of this review.

At the level of intracellular signal transduction, MAP kinases such as JNK or p38 have been identified as central signal transducers of inflammatory signals. Interestingly, it has been observed that there is a cross talk between VDR/RXR and MAP kinase signaling on many levels and the outcome, e.g., stimulation or inhibition, depends on the stimulus, cell type and the response (Miodovnik et al., 2012). Regarding inflammation, it has become obvious that vitamin D inhibits production of proinflammatory cytokines like IL-6 or TNFα in monocytes via the inhibition of p38 MAP kinase (Zhang et al., 2012). Inhibition of p38 in monocytes was found to be due to induction of MAPK phosphatase-1 (MKP1) which dephosphorylates p38 and thus reduces p38 activation (Figure 1). A similar mechanism was found in prostate cells where induction of MKP5 by 1α,25(OH)₂D₃ was responsible for down-regulation of IL-6 mRNA expression (Nonn et al., 2006). 1α,25(OH)₂D₃ increases MKP5 transcription by induction of VDR/RXR binding to a VDRE in the MKP5 promoter. Beside this indirect modulation of signaling cascades, 1α,25(OH)₂D₃ and its receptor complex VDR/RXR can interact with other transcription factors such as NF-κB, nuclear factor of activated T-cells (NFAT), or the glucocorticoid receptor (GCR) which leads to anti-inflammatory effects (Figure 2). Activation of VDR inhibits NF-κB activation and signaling. NFκB is a ubiquitously expressed transcription factor which represents a heterodimer. In the inactive state it interacts with IκB which keeps it in the cytosol (Karin and Lin, 2002). Upon cell activation by proinflammatory stimuli, IκB is phosphorylated and subsequently ubiquitinylated, which leads to proteasomal degradation of the IκB protein. Free NFκB translocates to the nucleus where it activates transcription of proinflammatory cytokines, antiapoptotic factors as well as of enzymes involved in the generation of proinflammatory mediators such as COX-2 (Karin and Lin, 2002; Tsatsanis et al., 2006). It has been shown that 1α,25(OH)₂D₃ down-regulates NF-κB levels in lymphocytes (Yu et al., 1995) and that the vitamin D analog TX 527 prevents NF-κB activation in monocytes (Stio et al., 2007). Inhibition of NFκB activation by 1α,25(OH)₂D₃-mediated up-regulation of IκB expression was reported in human peritoneal macrophages (Cohen-Lahav et al., 2006) (Figure 2). Additionally, interference of vitamin D signaling with DNA binding of NFκB was found (Harant et al., 1998). It was shown that 1α,25(OH)₂D₃ inhibits NF-κB activity in human MRC-5 fibroblasts but not translocation of its subunits p50 and p65. The partial inhibition of NFκB DNA binding by 1α,25(OH)₂D₃ was dependent on de novo protein synthesis, suggesting that 1α,25(OH)₂D₃ may regulate expression of cellular factors which contribute to reduced DNA binding of NFκB (Harant et al., 1998). Thus, it seems that vitamin D is able to inhibit NFκB activation as well DNA binding (Figure 2).

Another interesting target for the anti-inflammatory signaling of vitamin D is NFAT (Figure 2). This transcription factor is activated by dephosphorylation by calcineurin which leads to translocation of this protein and transcriptional activation of proinflammatory genes such as interleukin 2 and cyclooxygenase-2 (Duque et al., 2005; Muller and Rao, 2010). In T-lymphocytes, it was shown for the interleukin 2 promoter that VDR-RXR heterodimers bind to an NFAT binding site and thus inhibit NFAT activity (Takeuchi et al., 1998). Similar data were obtained for interleukin 17 where 1α,25(OH)₂D₃ blocked NFAT activity which contributed to repression of interleukin17A expression in inflammatory CD4⁺ T cells by the hormone (Joshi et al., 2011).

Another interesting finding was that vitamin D enhances the anti-inflammatory activities of GCs (Figure 1). The strong anti-inflammatory activities of GCs are mediated by the GCR. It belongs to the nuclear receptor family. Upon ligand binding the receptor dimerizes and translocates into the nucleus where it binds to GC-responsive elements (GRE) and modulates gene expression (Barnes, 1998). In general, GCs down-regulate expression of pro-inflammatory genes and up-regulate anti-inflammatory genes. It was found in asthmatics that dexamethasone-induced MKP-1 expression as a marker for GC responsiveness is significantly increased when serum vitamin D levels increase suggesting that vitamin D may enhance GC responsiveness (Sutherland et al., 2010). It is interesting to note that MKP-1 is also a vitamin D target gene as mentioned above (Zhang et al., 2012). Vitamin D enhancement of GC-induction of MKP1 was abolished both in purified CD14⁺ and CD14⁻ cells and it was found that the synergism depends on vitamin D-induced GM-CSF release from CD14⁻ cells and GM-CSF-dependent MED14 induction in CD14⁺ cells (Zhang et al., 2013b). MED14 is part of the mediator complex involved in the regulation of transcriptional initiation and it was found to form a complex with VDR and mediate ligand-dependent enhancement of transcription by the VDR (Rachez et al., 1999) (Figure 1). Interestingly, MED14 also enhances gene activation by the GCR in a gene-specific manner (Chen et al., 2006). For MKP1 it was found in human monocytes that VDR and GCR bind to a corresponding VDRE and two GREs after ligand stimulation (Figure 1). After GM-CSF treatment, MED14 was recruited to the promoter after addition of 1α,25(OH)₂D₃ but not dexamethasone indicating that MED14 recruitment depends on the VDR (Zhang et al., 2013b). 1α,25(OH)₂D₃ enhanced the binding of the GCR to the GRE in close proximity to the VDRE in the presence of GM-CSF and ChIP analysis suggest a MED14-VDR-GCR complex at the MKP1 promoter with bridges the crosstalk between vitamin D and GCs (Zhang et al., 2013b). The data from single gene analyses such as MKP1 suggest that the VDR interacts with other signaling pathways.

At present there are genome-wide data available from immortalized lymphoblastoid cell lines (Ramagopalan et al., 2010), undifferentiated and LPS stimulated THP-1 cells (Heikkinen et al., 2011; Tuoresmäki et al., 2014), LS180 colorectal cancer cells (Meyer et al., 2012) and LX2 hepatic stellate cells (Ding et al., 2013). These six ChIP-seq data sets showed 21,776 non-overlapping VDR binding sites whereas only 54 sites were common in all six data sets. The data suggest that, apart from a few sites, VDR binding is strongly cell and stimulus specific. In the non-overlapping binding sites, only 17.5% contain a DR3-type VDRE whereas the percentage of DR3-type response elements is enriched in highly ligand-responsive loci. All these data suggest that the VDR interacts with other transcription factors and that these interactions might only be in part ligand dependent. Regarding inflammation, the genome-wide effects of LPS on VDR location in THP-1 cells are of special interest (Tuoresmäki et al., 2014). From the 805 VDR binding sites, only 462 overlap in untreated and LPS-treated THP cells which were stimulated with 1α,25(OH)₂D₃. Thus, LPS treatment leads to a considerable change in VDR location. In THP-1 cells, bioinformatic searches for shared binding sites revealed motifs for CEBP1, PU.1 in stimulated THP-1 cells whereas NFYA, LHX3-like and NANOG were found for unstimulated cells but no transcription factor has been identified in conjunction with LPS treatment. Of note, binding sites for JUN, a component of the AP1 transcription factor, were found to be enriched at VDR loci in LX2 hepatic stellate cells. This is of interest regarding inflammation as AP1 is known to be a transcription factor that regulates expression of many proinflammatory genes. At present, there are many data available on single gene levels but there is still a missing link between these data and the genome-wide observations. Since VDR signaling seems to be strongly cell type and stimulus-dependent, more genome-wide data with different cell types and stimuli are required to understand the mechanisms how 1α,25(OH)₂D₃ modulates gene expression under inflammatory conditions.

Arachidonic acid derived eicosanoids which comprise prostaglandins and leukotrienes play an important role in inflammatory processes (Harizi et al., 2008). Of the enzymes involved in prostaglandin synthesis, cyclooxygenase-2 (COX-2) and microsomal prostaglandin E synthase 1 (mPGES-1) have been shown to be induced in many inflammatory conditions (Tomasoni et al., 1998; Murakami et al., 2000; Cipollone and Fazia, 2006; Petrovic et al., 2006) and inhibition of both enzymes is a common approach in the treatment of inflammatory diseases (Fahmi, 2004; Ramalho et al., 2009; Dallaporta et al., 2010).

In prostate cancer cells it has been demonstrated that 1α,25(OH)₂D₃ inhibits the expression of COX-2 on mRNA and protein level as well as the expression of prostaglandin receptors on mRNA level and simultaneously upregulates prostaglandin catabolism via 15-hydroxyprostaglandin dehydrogenase (Moreno et al., 2005). In addition, the combination of calcitriol with COX-inhibitors led to synergistic growth inhibition (Moreno et al., 2005). Similar results were obtained with the combination of 1α,25(OH)₂D₃ and COX-inhibitors in different leukemia cells (Jamshidi et al., 2008). In accordance with the previous findings, treatment with the vitamin D analog elocalcitol resulted in decreased COX-2 expression and diminished PGE₂ synthesis in prostate cells (Penna et al., 2009). The COX-2/PGE₂-pathway was also identified as the mediator of the growth inhibitory effect of calcitriol in breast cancer cells (Yuan et al., 2012). Furthermore, COX-2 upregulation in placental trophoblasts in response to oxidative stress and in myometrial cells in response to interleukin-1β was inhibited by 1α,25(OH)₂D₃ (Sun et al., 2013; Thota et al., 2013).

Thill et al. found correlations between VDR expression and expression of COX-2 as well as 15-hydroxy PG dehydrogenase in malignant breast cells and in cells from female reproductive tissues (Thill et al., 2009, 2010, 2012).

In human lung fibroblasts inhibition of PGE₂-production by vitamin D was found which was not due to altered COX-expression. Yet, vitamin D inhibited IL-1β-induced mPGES-1 expression and simultaneously stimulated 15-hydroxy PG dehydrogenase (Liu et al., 2014).

5-lipoxygenase (5-LO) accounts for the first two steps in leukotriene biosynthesis. Leukotrienes exert potent proinflammatory actions and have been associated with several chronic inflammatory diseases (Haeggstrom and Funk, 2011).

In the myeloid cell line HL-60, treatment with 1α,25(OH)₂D₃ triggers differentiation into monocytic cells. Simultaneously, 1α,25(OH)₂D₃ has been shown to induce 5-LO expression on mRNA and protein level as well as to increase 5-LO enzyme activity (Bennett et al., 1993; Brungs et al., 1994). A similar effect was also observed in the monocytic cell line Mono Mac 6. Additionally, this effect was strongly enhanced by the combination of 1α,25(OH)₂D₃ with transforming growth factor β (TGF-β) (Brungs et al., 1995; Harle et al., 1998). Mechanistically, the effect of 1α,25(OH)₂D₃ on 5-LO expression was related to VDR binding sites in the 5-LO promoter and distal parts of the 5-LO gene (Sorg et al., 2006; Stoffers et al., 2010) and is due to stimulation of 5-LO transcript elongation (Stoffers et al., 2010).

Previous results suggest a modulatory role of vitamin D in the inflammatory response of cells of the monocyte/macrophage lineage, which is again modulated by TGF-β. In this context, it is interesting that macrophages contain 1α-hydroxylase and therefore are capable of autocrine or paracrine activation of vitamin D (Lagishetty et al., 2011). Moreover, in keratinocytes autocrine TGF-β production is induced by vitamin D (Kim et al., 1992). Crucial participation of monocytes/macrophages in diverse inflammatory processes has been demonstrated (Cutolo, 1999; Yoon and Jun, 1999; Moore et al., 2013). Besides induction of 5-lipoxygenase, the combination of TGF-β and 1α,25(OH)₂D₃ has been shown to induce the differentiation antigen CD69 in monocytic cells (Wobke et al., 2013). Overexpression of CD69 again, has been shown in the context of local dermal inflammation, systemic lupus erythematosus, hyperthyroid Graves' disease and autoimmune thyroiditis (Fernandez-Herrera et al., 1996; Portales-Perez et al., 1997; Crispin et al., 1998; Gessl and Waldhausl, 1998).

It is well established that 1α,25(OH)₂D₃ influences cytokine gene expression and signaling in several different cell types. Firstly, this is the case for the pleiotropic mediator TGF-β, for which it has been shown that either the expression of the cytokine itself or expression of associated signaling components is downregulated by 1α,25(OH)₂D₃. In hepatocytes, 1α,25(OH)₂D₃ has been found to influence TGF-β signaling in a genome wide scale by directing binding of Smad proteins to target genes. These actions of 1α,25(OH)₂D₃ on TGF-β expression or signaling were able to inhibit fibrosis and associated inflammation. Second, the interleukins are a vast group of inflammatory cytokines that are clearly regulated by 1α,25(OH)₂D₃ in a cell-specific manner. However, for several members of this family (e.g., IL-1, IL-6, and IL-8), both positive or negative regulation by 1α,25(OH)₂D₃ has been observed. A closer look at the parameters that determine the outcome of 1α,25(OH)₂D₃ action on the expression of these genes is warranted. This applies in particular to the time-scale of changes in gene expression, as different responses may occur during separate stages of 1α,25(OH)₂D₃ action. Regarding the mechanisms, recruitment of VDR to the respective genomic regions, as well as interaction of 1α,25(OH)₂D₃ signaling with other transcription factors involved in IL expression (NFAT, NF-κB, Runx1), seem to occur. Concerning the p38 MAP kinase phosphatase MKP1, it was found that GCR and VDR/RXR act in a synergistic manner to induce MKP1 expression in monocytes. This results in reduced p38 activation and reduced formation of proinflammatory cytokines. As a further cytokine, the proinflammatory mediator TNFα has been identified as a 1α,25(OH)₂D₃ target gene. Also in this case, the vitamin D effects are cell-specific: With cell samples that mainly contain T-cells, downregulation of TNFα has been observed, whereas for monocytic cells, either positive or negative regulation occurred depending on the differentiation state. Finally, gene expression of the proinflammatory mediator IFNγ has been described to be suppressed by 1α,25(OH)₂D₃ in T-cells. Altogether, the influence of 1α,25(OH)₂D₃ on the expression of interleukins, TNFα, and IFNγ by different cell types, and the consequences for the cellular interplay that are to be anticipated, amounts to a complex picture. In Figure 3, the influence of 1α,25(OH)₂D₃ on the expression of these cytokines is summarized for the major immune cells (monocytes, DCs, and different T-cell subsets). The resulting pattern supports a shift of T-cell responses from a Th1 type toward Th2 reactions and a suppression of Th17 responses. The effect of 1α,25(OH)₂D₃ on cytokine expression in antigen presenting cells (monocytes, DCs) remains unclear and seems to depend on the time of stimulation, the differentiation state and other factors.

Modulation of GCR, NFκB, NFAT as well as SMAD signaling plays a central role in the immunomodulatory activities of 1α,25(OH)₂D₃. Mechanistic studies on individual genes gave some mechanistic insights into the mechanisms involved in the interaction between VDR/RXR and the above mentioned transcription factors. These mechanisms include competitive binding as well as a crosstalk between the signaling pathways on multiple levels including the promoter level. However, by using ChIP seq and other techniques which allow a genome-wide view, we are just starting to understand the signaling network which is responsible for cell-type-specific and locus-dependent gene activation by ligand-regulated transcription factors such as VDR/RXR. For example, intersecting VDR/SMAD regulatory circuits have just been unraveled and it was shown that TGFβ signaling facilitates VDR binding to certain gene loci. More such data are required to increase our understanding of the complex gene regulatory network that is affected by 1α,25(OH)₂D₃. Especially, genome-wide data on VDR loci in conjunction with analyses of other, inflammation-related key transcription factors in different cell types and various stimuli are necessary to understand the complex regulation of gene transcription during inflammation.