R. Norton

Vitamin D and respiratory health

Vitamin D is now known to be of physiological importance outside of bone health and calcium homeostasis, and there is mounting evidence that it plays a beneficial role in the prevention and/or treatment of a wide range of diseases. In this brief review the known effects of vitamin D on immune function are described in relation to respiratory health. Vitamin D appears capable of inhibiting pulmonary inflammatory responses while enhancing innate defence mechanisms against respiratory pathogens. Population‐based studies showing an association between circulating vitamin D levels and lung function provide strong justification for randomized controlled clinical trials of vitamin D supplementation in patients with respiratory diseases to assess both efficacy and optimal dosage.

Sulforaphane represses matrix-degrading proteases and protects cartilage from destruction in vitro and in vivo.

Objective Sulforaphane (SFN) has been reported to regulate signaling pathways relevant to chronic diseases. The aim of this study was to investigate the impact of SFN treatment on signaling pathways in chondrocytes and to determine whether sulforaphane could block cartilage destruction in osteoarthritis. Methods Gene expression, histone acetylation, and signaling of the transcription factors NF-E2–related factor 2 (Nrf2) and NF-κB were examined in vitro. The bovine nasal cartilage explant model and the destabilization of the medial meniscus (DMM) model of osteoarthritis in the mouse were used to assess chondroprotection at the tissue and whole-animal levels. Results SFN inhibited cytokine-induced metalloproteinase expression in primary human articular chondrocytes and in fibroblast-like synovial cells. SFN acted independently of Nrf2 and histone deacetylase activity to regulate metalloproteinase expression in human articular chondrocytes but did mediate prolonged activation of JNK and p38 MAPK. SFN attenuated NF-κB signaling at least through inhibition of DNA binding in human articular chondrocytes, with decreased expression of several NF-κB–dependent genes. Compared with cytokines alone, SFN (10 μM) abrogated cytokine-induced destruction of bovine nasal cartilage at both the proteoglycan and collagen breakdown levels. An SFN-rich diet (3 μmoles/day SFN versus control chow) decreased the arthritis score in the DMM model of osteoarthritis in the mouse, with a concurrent block of early DMM-induced gene expression changes. Conclusion SFN inhibits the expression of key metalloproteinases implicated in osteoarthritis, independently of Nrf2, and blocks inflammation at the level of NF-κB to protect against cartilage destruction in vitro and in vivo.

Response of lung epithelial cells to inflammatory stimuli following exposure to the active form of vitamin D

The active form of vitamin D (1,25(OH)2D3) is now of huge interest to the scientific community with numerous beneficial effects beyond Ca homoeostasis and rickets. Inflammation plays a central role in the pathogenesis of numerous pulmonary diseases and recent evidence suggests vitamin D exerts immunomodulatory effects in the lung. Serum concentrations of inactive 25(OH)D3 have been shown to directly correlate with FEV1 and inversely with upper respiratory tract infection. More recent studies have shown potential beneficial effects of high dose vitamin D improving treatment for some tuberculosis patients. It is clear from this evidence that vitamin D may play an important role in the lung, but due to the low levels of vitamin D available from the diet, it is unlikely that levels of vitamin D required to be of benefit are achievable through diet alone. However, on a cellular level, respiratory epithelial cells have been shown to be capable of activating vitamin D but as yet it is unknown what concentrations of active vitamin D can be achieved locally given sufficient serum concentrations of 25(OH)D3. Vitamin D inadequacy is a global problem, especially among elderly patients (5) and with an ever increasing ageing population and lifestyles that reduce exposure to sunlight, it is crucial to identify mechanisms by which vitamin D improves function and to identify optimal levels. In this study, the effects of 1,25(OH)2D3 on pro-inflammatory mediator production were investigated in primary human small airway epithelial cells (SAEC). Cells were incubated with 100 nM 1,25(OH)2D3 for up to 48 h and the effects on basal mRNA expression of cytokines (IL-1b, IL-6, IL-8, TNFa, IL-10 and IL-12p70) were investigated using qRT-PCR. 1,25(OH)2D3 treatment significantly reduced the expression of IL-6 mRNA from just 1 h of treatment (results at least in triplicate, one way ANOVA, P<0.001), but had no effect on basal mRNA of the other cytokines. IL-10 and IL-12p70 mRNA and protein were below the limit of detection. Both 10 ng/ml TNFa and 10 mg/ml lipopolysaccharide (LPS)-induced expression of IL-1b, IL-6, IL-8 and TNFa mRNA. Pre-treatment of SAEC with 1, 10 or 100 nM 1,25(OH)2D3 for 1 h before stimulation with TNFa or LPS led to a dose-dependent reduction of IL-6 mRNA induction that was significant at 100 nM with TNFa treatment (results at least in triplicate, t test, P<0.05). Induction of IL-6 protein expression by TNFa was significantly reduced by 1 h pre-incubation with 100 nM 1,25(OH)2D3 as assessed by flow cytometry (results in triplicate, one way ANOVA, P<0.01). IL-6 is a key cytokine involved in the initiation and extension of the inflammatory response both locally and systemically in lung diseases. This data suggests that vitamin D may be of benefit in reducing IL-6 production by primary human small airway epithelial cells. More research is required into further effects and the mechanism of action of vitamin D on lung epithelium, to determine potential benefits for preventing or treating lung diseases and to define optimum levels.

200 SULFORAPHANE REPRESSES MATRIX-DEGRADING PROTEASES AND PROTECTS CARTILAGE FROM DESTRUCTION IN VITRO

Purpose: Increased expression of inducible NO synthase (iNOS) and cyclooxygenase (COX)-2 plays a key role in the pathogenesis of osteoarthritis (OA). Methylation of lysine 4 on histone H3 (H3K4) was shown to be of fundamental importance in the regulation of gene expression. In the present study, we investigated the role of H3K4 methylation in interleukin-1β (IL-1)-induced COX-2 and iNOS expression in human OA chondrocytes Methods: Chondrocytes were stimulated with IL-1 for various time periods and the expression of iNOS and COX-2 mRNAs and proteins were evaluated using real-time reverse transcriptase-polymerase chain reaction (RT-PCR) and Western blotting, respectively. H3K4 methylation at the iNOS and COX-2 promoters was evaluated using chromatin immunoprecipitation (ChIP) assays. The role of histone methylation was further evaluated using the methyltransferase inhibitor, 5’-deoxy-5’(methylthio) adenosine (MTA). Results: IL-1 induced iNOS and COX-2 mRNA and protein in a doseand time-dependent manner. The induction of iNOS and COX-2 expression by IL-1 was associated with H3K4 diand trimethylation at the iNOS and COX-2 promoters, whereas the levels of H3K4 monomethylation remained unchanged. Treatment with MTA inhibited IL-1-induced H3K4 methylation as well as IL-1-induced iNOS and COX-2 expression. Conclusions: These results indicate that H3K4 methylation contributes to IL-1-induced iNOS and COX-2 expression and suggest that this pathway could be a potential target for pharmacological intervention in the treatment of OA.