Effect of vitamin D on Epstein-Barr (EBV)-specific CD8+T cells in patients with early multiple sclerosis (MS)

Background: Infection with EBV and a lack in vitamin D may be important environmental triggers of MS. 1,25-(OH)2D3 mediates a shift of antigen presenting cells (APC) and CD4+ T cells to a less inflammatory profile. Although CD8+ T cells do express the vitamin D receptor, a direct effect of 1,25(OH)2D3 on these cells has not been demonstrated until now. Since CD8+ T cells are important immune mediators of the inflammatory response in MS, we examined whether vitamin D directly affects the CD8+ T cell response, and more specifically if it modulates the EBV-specific CD8+ T cell response. Material and Methods: To explore whether the vitamin D status may influence the pattern of the EBV-specific CD8+ T cell response, PBMC of 10 patients with early MS and 10 healthy controls (HC) were stimulated with a pool of immunodominant 8-10 mer peptide epitopes known to elicit CD8+ T cell responses. PBMC were stimulated with this EBV CD8 peptide pool, medium (negative control) or anti- CD3/anti-CD28 beads (positive control). The following assays were performed: ELISPOT to assess the secretion of IFN-gamma by T cells in general; cytometric beads array (CBA) and ELISA to determine whichcytokines were released by EBV-specific CD8+ T cells after six days of culture; and intracellular cytokine staining assay to determine by which subtype of T cells secreted given cytokines. To examine whether vitamin D could directly modulate CD8+ T cell immune responses, we depleted CD4+ T cells using negative selection. Results: We found that pre-treatment of vitamin D had an antiinflammatory action on both EBV-specific CD8+ T cells and on CD3/ CD28-stimulated T cells: secretion of pro-inflammatory cytokines (IFNgamma and TNF-alpha) was decreased, whereas secretion of antiinflammatory cytokines (IL-5 and TGF-beta) was increased. At baseline, CD8+ T cells of early MS patients showed a higher secretion of TNFalpha and lower secretion of IL-5. Addition of vitamin D did not restore the same levels of both cytokines as compared to HC. Vitamin D-pretreated CD8+T cells exhibited a decreased secretion of IFN-gamma and TNF-alpha, even after depletion of CD4+ T cells from culture. Conclusion: Vitamin D has a direct anti-inflammatory effect on CD8+ T cells independently from CD4+ T cells. CD8+ T cells of patients with earlyMS are less responsive to the inflammatory effect of vitamin D than HC, pointing toward an intrinsic dysregulation of CD8+ T cells. The modulation of EBV-specific CD8+T cells by vitaminDsuggests that there may be interplay between these twomajor environmental factors of MS. This study was supported by a grant from the Swiss National Foundation (PP00P3-124893), and by an unrestricted research grant from Bayer to RDP.

Vitamin D deficiency and a CYP27B1-1260 promoter polymorphism are associated with chronic hepatitis C and poor response to interferon-alfa based therapy

BACKGROUND: Vitamin D is an important immune modulator and preliminary data indicated an association between vitamin D deficiency and sustained virologic response (SVR) rates in patients with chronic hepatitis C. We therefore performed a comprehensive analysis on the impact of vitamin D serum levels and of genetic polymorphisms within the vitamin D cascade on chronic hepatitis C and its treatment. METHODS: Vitamin D serum levels, genetic polymorphisms within the vitamin D receptor and the 1α- hydroxylase were determined in a cohort of 468 HCV genotype 1, 2 and 3 infected patients who were treated with interferon-alfa based regimens. RESULTS: Chronic hepatitis C was associated with a high incidence of severe vitamin D deficiency compared to controls (25(OH)D3<10 ng/mL in 25% versus 12%, p<0.00001), which was in part reversible after HCV eradication. 25(OH)D3 deficiency correlated with SVR in HCV genotype 2 and 3 patients (63% and 83% SVR for patients with and without severe vitamin D deficiency, respectively, p<0.001). In addition, the CYPB27-1260 promoter polymorphism rs10877012 had substantial impact on 1-25- dihydroxyvitamin D serum levels and SVR rates in HCV genotype 1, 2 and 3 infected patients. CONCLUSIONS: Chronic hepatitis C virus infection is associated with vitamin D deficiency. Reduced 25- hydroxyvitamin D levels and CYPB27-1260 promoter polymorphism are associated with failure to achieve SVR in HCV genotype 1, 2, 3 infected patients.

Changing the awareness of low vitamin D status in a rheumatology population: a pre/post-study.

INTRODUCTION: In 2009 hypovitaminosis D was highly prevalent in a population of Swiss rheumatology patients (86%). We aimed to evaluate the evolution of vitamin D status in the same population two years later, after the results of the first study were disseminated to local physicians and patients, in order to determine the evolution of the problem and the impact of physician information. METHOD: Patients in our rheumatology clinic were screened for 25-OH vitamin D. Results were categorised as: deficient (<10 ng/ml or <25 nmol/l), insufficient (10 to 30 ng/ml or 25 to 75 nmol/l) or normal (>30 ng/ml or >75 nmol/l). We also used another cut-off of 20 ng/ml (50 nmol/l). We evaluated the evolution of 25-OH vitamin D dosages and vitamin D3 prescriptions between 2008 and 2011 in our institution and the number of publications on vitamin D in three important medical journals of the French speaking part of Switzerland. RESULTS: Compared with 2009, significantly more patients had normal results in 2011. Fifty-two percent of patients had levels >20 ng/ml in 2009 and 66% in 2011, difference statistically significant (p = 0.001). During the years separating the two study periods the number of 25-OH vitamin D dosages and the prescription of high doses of vitamin D3 increased in our hospital. In addition the number of publications on vitamin D increased between 2008 and 2011. CONCLUSION: We concluded that lower prevalence in hypovitaminosis D is certainly related to better adherence to daily supplements, and to better information and awareness of the physicians about hypovitaminosis D.

A-kinase anchoring protein-Lbc promotes pro-fibrotic signaling in cardiac fibroblasts

In response to stress or injury the heart undergoes an adverse remodeling process associated with cardiomyocyte hypertrophy and fibrosis. Transformation of cardiac fibroblasts to myofibroblasts is a crucial event initiating the fibrotic process. Cardiac myofibroblasts invade the myocardium and secrete excess amounts of extracellular matrix proteins, which cause myocardial stiffening, cardiac dysfunctions and progression to heart failure. While several studies indicate that the small GTPase RhoA can promote profibrotic responses, the exchange factors that modulate its activity in cardiac fibroblasts are yet to be identified. In the present study, we show that AKAP-Lbc, an A-kinase anchoring protein (AKAP) with an intrinsic Rho-specific guanine nucleotide exchange factor (GEF) activity, is critical for activating RhoA and transducing profibrotic signals downstream of type I angiotensin II receptors (AT1Rs) in cardiac fibroblasts. In particular, our results indicate that suppression of AKAP-Lbc expression by infecting adult rat ventricular fibroblasts with lentiviruses encoding AKAP-Lbc specific short hairpin (sh) RNAs strongly reduces the ability of angiotensin II to promote RhoA activation, differentiation of cardiac fibroblasts to myofibroblasts, collagen deposition as well as myofibroblast migration. Interestingly, AT1Rs promote AKAP-Lbc activation via a pathway that requires the α subunit of the heterotrimeric G protein G12. These findings identify AKAP-Lbc as a key Rho-guanine nucleotide exchange factor modulating profibrotic responses in cardiac fibroblasts.

Vitamin D levels and associated factors: a population-based study in Switzerland.

QUESTIONS UNDER STUDY: To update the prevalence of vitamin D insufficiency and to identify factors associated with vitamin D status in the Swiss adult population. METHODS: Data from the 2010-2011 Swiss Study on Salt intake, a population-based study in the Swiss population, was used. Vitamin D concentration in serum was measured by liquid chromatography- tandem mass spectrometry. Major factors that influence vitamin D levels were taken into account. Survey statistical procedures were used to estimate means and prevalences of vitamin D levels and status. Monthly-specific tertiles of vitamin D and ordinal logistic regression were used to determine the associations of covariates of interest with vitamin D status. RESULTS: The prevalences of vitamin D insufficiency (serum 25-hydroxyvitamin D: 20-29.9 ng/ml) and deficiency (<20 ng/ml) were the highest in the January-March period; 26.4% (95%CI: 21.6-31.7) and 61.6% (95%CI: 56.0-67.0), respectively. In the same period, more than 9 of ten men were vitamin D insufficient or deficient. Each unit increase of Body Mass Index was associated with an 8% decreased likelihood of being in a higher vitamin D tertiles. Oral contraceptive, altitude, urinary excretion of calcium, use of vitamin D supplement or treatment, high wine consumption, physical activity were associated with vitamin D tertiles. Compared to the French-speaking region, the Italian-speaking region was independently associated with a higher likelihood of being in higher vitamin D tertiles (OR: 1.66, 95%CI: 1.14-2.43). CONCLUSIONS: Low levels of vitamin D are common among Swiss adults, in particular during winter months and outside the Italian-speaking region.

Specific targeting of pro-death NMDA receptor signals with differing reliance on the NR2B PDZ ligand.

NMDA receptors (NMDARs) mediate ischemic brain damage, for which interactions between the C termini of NR2 subunits and PDZ domain proteins within the NMDAR signaling complex (NSC) are emerging therapeutic targets. However, expression of NMDARs in a non-neuronal context, lacking many NSC components, can still induce cell death. Moreover, it is unclear whether targeting the NSC will impair NMDAR-dependent prosurvival and plasticity signaling. We show that the NMDAR can promote death signaling independently of the NR2 PDZ ligand, when expressed in non-neuronal cells lacking PSD-95 and neuronal nitric oxide synthase (nNOS), key PDZ proteins that mediate neuronal NMDAR excitotoxicity. However, in a non-neuronal context, the NMDAR promotes cell death solely via c-Jun N-terminal protein kinase (JNK), whereas NMDAR-dependent cortical neuronal death is promoted by both JNK and p38. NMDAR-dependent pro-death signaling via p38 relies on neuronal context, although death signaling by JNK, triggered by mitochondrial reactive oxygen species production, does not. NMDAR-dependent p38 activation in neurons is triggered by submembranous Ca(2+), and is disrupted by NOS inhibitors and also a peptide mimicking the NR2B PDZ ligand (TAT-NR2B9c). TAT-NR2B9c reduced excitotoxic neuronal death and p38-mediated ischemic damage, without impairing an NMDAR-dependent plasticity model or prosurvival signaling to CREB or Akt. TAT-NR2B9c did not inhibit JNK activation, and synergized with JNK inhibitors to ameliorate severe excitotoxic neuronal loss in vitro and ischemic cortical damage in vivo. Thus, NMDAR-activated signals comprise pro-death pathways with differing requirements for PDZ protein interactions. These signals are amenable to selective inhibition, while sparing synaptic plasticity and prosurvival signaling.