Background TH 17 cells are proposed to play a role in the pathology of asthma, including steroid-resistant (SR) disease. We previously identified a steroid-enhancing function of vitamin D in patients ...with SR asthma in restoring the impaired response to steroids for production of the anti-inflammatory cytokine IL-10. Objective We sought to investigate the production of the TH 17-associated cytokines IL-17A and IL-22 in culture in patients with moderate-to-severe asthma defined on the basis of their clinical response to steroids and the susceptibility of this response to inhibition by steroids and the active form of vitamin D, 1α,25-dihydroxyvitamin D3 (1,25OH2 D3). Methods PBMCs were stimulated in culture with or without dexamethasone and 1,25(OH)2 D3. A cytometric bead array, ELISA, and intracellular cytokine staining were used to assess cytokine production. The role of CD39 in inhibition of the TH 17 response was studied by using quantitative real-time PCR, flow cytometry, and addition of the antagonist POM-1 to culture. Results Asthmatic patients synthesized much higher levels of IL-17A and IL-22 than nonasthmatic control subjects, with patients with SR asthma expressing the highest levels of IL-17A. Glucocorticoids did not inhibit IL-17A cytokine expression in patients and enhanced production in cultures from control subjects. Treatment with 1,25(OH)2 D3 with or without dexamethasone significantly reduced both IL-17A and IL-22 levels. An antagonist of the ectonucleotidase CD39 reversed 1,25(OH)2 D3-mediated inhibition of the IL-17A response. Conclusion Patients with severe asthma exhibit increased levels of TH 17 cytokines, which are not inhibited by steroids. 1,25(OH)2 D3 inhibits TH 17 cytokine production in all patients studied, irrespective of their clinical responsiveness to steroids, identifying novel steroid-enhancing properties of vitamin D in asthmatic patients.
Calcidiol, the major circulating metabolite of vitamin D, supports induction of pleiotropic antimicrobial responses in vitro. Vitamin D supplementation elevates circulating calcidiol concentrations, ...and thus has a potential role in the prevention and treatment of infection. The immunomodulatory effects of administering vitamin D to humans with an infectious disease have not previously been reported. To characterize these effects, we conducted a detailed longitudinal study of circulating and antigen-stimulated immune responses in ninety-five patients receiving antimicrobial therapy for pulmonary tuberculosis who were randomized to receive adjunctive high-dose vitamin D or placebo in a clinical trial, and who fulfilled criteria for per-protocol analysis. Vitamin D supplementation accelerated sputum smear conversion and enhanced treatment-induced resolution of lymphopaenia, monocytosis, hypercytokinaemia, and hyperchemokinaemia. Administration of vitamin D also suppressed antigen-stimulated proinflammatory cytokine responses, but attenuated the suppressive effect of antimicrobial therapy on antigenstimulated secretion of IL-4, CC chemokine ligand 5, and IFN-α. We demonstrate a previously unappreciated role for vitamin D supplementation in accelerating resolution of inflammatory responses during tuberculosis treatment. Our findings suggest a potential role for adjunctive vitamin D supplementation in the treatment of pulmonary infections to accelerate resolution of inflammatory responses associated with increased risk of mortality.
A small population of patients with severe asthma does not respond to glucocorticoids (steroid resistant SR). They have high morbidity, highlighting an urgent need for strategies to enhance ...glucocorticoid responsiveness.
We investigated the immunologic differences between steroid-sensitive (SS) and SR asthmatic patients and the effect on immunophenotype of oral calcitriol treatment because it has been previously shown to beneficially modulate the clinical response to glucocorticoids in patients with SR asthma.
CD8-depleted PBMCs were isolated from 12 patients with SS and 23 patients with SR asthma and cultured for 7 days with anti-CD3 and IL-2 with or without dexamethasone. Cytokine production was assessed in supernatants by using the Cytometric Bead Array. Patients with SR asthma were subsequently randomized to oral calcitriol or placebo therapy, and identical studies were repeated.
Patients with SR asthma produced significantly increased IL-17A and IFN-γ levels compared with those in patients with SS asthma, although it was evident that cells from individual patients might overproduce one or the other of these cytokines. Production of IL-17A was inversely and production of IL-13 was positively associated with the clinical response to prednisolone. Oral calcitriol, compared with placebo, therapy of the patients with SR asthma significantly improved dexamethasone-induced IL-10 production in vitro while suppressing dexamethasone-induced IL-17A production. This effect mirrored the previously demonstrated improvement in clinical response to oral glucocorticoids in calcitriol-treated patients with SR asthma.
IL-17A(high) and IFN-γ(high) immunophenotypes exist in patients with SR asthma. These data identify immunologic pathways that likely underpin the beneficial clinical effects of calcitriol in patients with SR asthma by directing the SR cytokine profile toward a more SS immune phenotype, suggesting strategies for identifying vitamin D responder immunophenotypes.
Vitamin D deficiency is associated with susceptibility to tuberculosis (TB) in HIV-uninfected people in Europe, but it is not known whether such an association exists among HIV-infected people in ...subtropical Africa. We conducted a cross-sectional study to determine whether vitamin D deficiency was associated with susceptibility to active TB in HIV-uninfected (n = 196) and HIV-infected (n = 174) black Africans in Cape Town, South Africa. We also investigated whether there was evidence of seasonal variation in vitamin D status and TB notifications in this setting over an 8-y period. Vitamin D deficiency (serum 25-hydroxyvitamin D 25(OH)D <50 nmol/L) was present in 232 (62.7%) of 370 participants and was associated with active TB in both HIV-uninfected (odds ratio = 5.2, 95% confidence interval: 2.8–9.7; P < 0.001) and HIV-infected (odds ratio = 5.6, 95% confidence interval: 2.7–11.6; P < 0.001) people. Vitamin D status varied according to season: The mean serum 25(OH)D concentration was highest in January through March and lowest in July through September (56.8 vs. 30.7 nmol/L, respectively; P < 0.001). Reciprocal seasonal variation in TB notifications was observed: The mean number of TB notifications per quarter for Cape Town in 2003 to 2010 was lowest in April through June and highest in October through December (4,222 vs. 5,080; P < 0.001). Vitamin D deficiency is highly prevalent among black Africans in Cape Town and is associated with susceptibility to active TB both in the presence and absence of HIV infection. Reciprocal seasonal variation in serum 25(OH)D concentration and TB notifications suggests that seasonal variations in vitamin D status and TB incidence in this setting are causally related.
Vitamin D was used to treat tuberculosis in the pre-antibiotic era, and its metabolites induce antimycobacterial immunity in vitro. Clinical trials investigating the effect of adjunctive vitamin D on ...sputum culture conversion are absent.
We undertook a multicentre randomised controlled trial of adjunctive vitamin D in adults with sputum smear-positive pulmonary tuberculosis in London, UK. 146 patients were allocated to receive 2·5 mg vitamin D
3 or placebo at baseline and 14, 28, and 42 days after starting standard tuberculosis treatment. The primary endpoint was time from initiation of antimicrobial treatment to sputum culture conversion. Patients were genotyped for
TaqI and
FokI polymorphisms of the vitamin D receptor, and interaction analyses were done to assess the influence of the vitamin D receptor genotype on response to vitamin D
3. This trial is registered with
ClinicalTrials.gov number
NCT00419068.
126 patients were included in the primary efficacy analysis (62 assigned to intervention, 64 assigned to placebo). Median time to sputum culture conversion was 36·0 days in the intervention group and 43·5 days in the placebo group (adjusted hazard ratio 1·39, 95% CI 0·90–2·16; p=0.14).
TaqI genotype modified the effect of vitamin D supplementation on time to sputum culture conversion (p
interaction=0·03), with enhanced response seen only in patients with the
tt genotype (8·09, 95% CI 1·36–48·01; p=0·02).
FokI genotype did not modify the effect of vitamin D supplementation (p
interaction=0·85). Mean serum 25-hydroxyvitamin D concentration at 56 days was 101·4 nmol/L in the intervention group and 22·8 nmol/L in the placebo group (95% CI for difference 68·6–88·2; p<0·0001).
Administration of four doses of 2·5 mg vitamin D
3 increased serum 25-hydroxyvitamin D concentrations in patients receiving intensive-phase treatment for pulmonary tuberculosis. Vitamin D did not significantly affect time to sputum culture conversion in the whole study population, but it did significantly hasten sputum culture conversion in participants with the
tt genotype of the
TaqI vitamin D receptor polymorphism.
British Lung Foundation.
Summary Background Vitamin D was used to treat tuberculosis in the pre-antibiotic era, and its metabolites induce antimycobacterial immunity in vitro. Clinical trials investigating the effect of ...adjunctive vitamin D on sputum culture conversion are absent. Methods We undertook a multicentre randomised controlled trial of adjunctive vitamin D in adults with sputum smear-positive pulmonary tuberculosis in London, UK. 146 patients were allocated to receive 2·5 mg vitamin D3 or placebo at baseline and 14, 28, and 42 days after starting standard tuberculosis treatment. The primary endpoint was time from initiation of antimicrobial treatment to sputum culture conversion. Patients were genotyped for Taq I and Fok I polymorphisms of the vitamin D receptor, and interaction analyses were done to assess the influence of the vitamin D receptor genotype on response to vitamin D3 . This trial is registered with ClinicalTrials.gov number NCT00419068. Findings 126 patients were included in the primary efficacy analysis (62 assigned to intervention, 64 assigned to placebo). Median time to sputum culture conversion was 36·0 days in the intervention group and 43·5 days in the placebo group (adjusted hazard ratio 1·39, 95% CI 0·90–2·16; p=0.14). Taq I genotype modified the effect of vitamin D supplementation on time to sputum culture conversion (pinteraction =0·03), with enhanced response seen only in patients with the tt genotype (8·09, 95% CI 1·36–48·01; p=0·02). Fok I genotype did not modify the effect of vitamin D supplementation (pinteraction =0·85). Mean serum 25-hydroxyvitamin D concentration at 56 days was 101·4 nmol/L in the intervention group and 22·8 nmol/L in the placebo group (95% CI for difference 68·6–88·2; p<0·0001). Interpretation Administration of four doses of 2·5 mg vitamin D3 increased serum 25-hydroxyvitamin D concentrations in patients receiving intensive-phase treatment for pulmonary tuberculosis. Vitamin D did not significantly affect time to sputum culture conversion in the whole study population, but it did significantly hasten sputum culture conversion in participants with the tt genotype of the TaqI vitamin D receptor polymorphism. Funding British Lung Foundation.
Identification of CD8
T cell epitopes is critical for the development of immunotherapeutics. Existing methods for major histocompatibility complex class I (MHC class I) ligand discovery are time ...intensive, specialized and unable to interrogate specific proteins on a large scale. Here, we present EpiScan, which uses surface MHC class I levels as a readout for whether a genetically encoded peptide is an MHC class I ligand. Predetermined starting pools composed of >100,000 peptides can be designed using oligonucleotide synthesis, permitting large-scale MHC class I screening. We exploit this programmability of EpiScan to uncover an unappreciated role for cysteine that increases the number of predicted ligands by 9-21%, reveal affinity hierarchies by analysis of biased anchor peptide libraries and screen viral proteomes for MHC class I ligands. Using these data, we generate and iteratively refine peptide binding predictions to create EpiScan Predictor. EpiScan Predictor performs comparably to other state-of-the-art MHC class I peptide binding prediction algorithms without suffering from underrepresentation of cysteine-containing peptides. Thus, targeted immunopeptidomics using EpiScan will accelerate CD8
T cell epitope discovery toward the goal of individual-specific immunotherapeutics.
Patients with chronic obstructive pulmonary disease (COPD) often have vitamin D deficiency, which is associated with increased susceptibility to upper respiratory infection-a major precipitant of ...exacerbation. Multicentre trials of vitamin D supplementation for prevention of exacerbation and upper respiratory infection in patients with COPD are lacking. We therefore investigated whether vitamin D3 (colecalciferol) supplementation would reduce the incidence of moderate or severe COPD exacerbations and upper respiratory infections.
We did a randomised, double-blind, placebo-controlled trial of vitamin D3 supplementation in adults with COPD in 60 general practices and four Acute National Health Service Trust clinics in London, UK. Patients were allocated to receive six 2-monthly oral doses of 3 mg vitamin D3 or placebo over 1 year in a 1:1 ratio using computer-generated permuted block randomisation. Participants and study staff were masked to treatment assignment. Coprimary outcomes were time to first moderate or severe exacerbation and first upper respiratory infection. Analysis was by intention to treat. A prespecified subgroup analysis was done to assess whether effects of the intervention on the coprimary outcomes were modified by baseline vitamin D status. This trial is registered with ClinicalTrials.gov, number NCT00977873.
240 patients were randomly allocated to the vitamin D3 group (n=122) and placebo group (n=118). Vitamin D3 compared with placebo did not affect time to first moderate or severe exacerbation (adjusted hazard ratio 0·86, 95% CI 0·60-1·24, p=0·42) or time to first upper respiratory infection (0·95, 0·69-1·31, p=0·75). Prespecified subgroup analysis showed that vitamin D3 was protective against moderate or severe exacerbation in participants with baseline serum 25-hydroxyvitamin D concentrations of less than 50 nmol/L (0·57, 0·35-0·92, p=0·021), but not in those with baseline 25-hydroxyvitamin D levels of at least 50 nmol/L (1·45, 0·81-2·62, p=0·21; p=0·021 for interaction between allocation and baseline serum 25-hydroxyvitamin D status). Baseline vitamin D status did not modify the effect of the intervention on risk of upper respiratory infection (pinteraction=0·41).
Vitamin D3 supplementation protected against moderate or severe exacerbation, but not upper respiratory infection, in patients with COPD with baseline 25-hydroxyvitamin D levels of less than 50 nmol/L. Our findings suggest that correction of vitamin D deficiency in patients with COPD reduces the risk of moderate or severe exacerbation.
UK National Institute for Health Research.
Low-dose vitamin D supplementation is already recommended in older adults for prevention of fractures and falls, but clinical trials investigating whether higher doses could provide additional ...protection against acute respiratory infection (ARI) are lacking.
To conduct a clinical trial of high-dose versus low-dose vitamin D3 supplementation for ARI prevention in residents of sheltered-accommodation housing blocks ('schemes') and their carers in London, UK.
Fifty-four schemes (137 individual participants) were allocated to the active intervention (vitamin D3 2.4 mg once every 2 months +10 μg daily for residents, 3 mg once every 2 months for carers), and 54 schemes with 103 participants were allocated to control (placebo once every 2 months +vitamin D3 10 μg daily for residents, placebo once every 2 months for carers) for 1 year. Primary outcome was time to first ARI; secondary outcomes included time to first upper/lower respiratory infection (URI/LRI, analysed separately), and symptom duration.
Inadequate vitamin D status was common at baseline: 220/240 (92%) participants had serum 25(OH)D concentration <75 nmol/L. The active intervention did not influence time to first ARI (adjusted HR (aHR) 1.18, 95% CI 0.80 to 1.74, p=0.42). When URI and LRI were analysed separately, allocation to the active intervention was associated with increased risk of URI (aHR 1.48, 95% CI 1.02 to 2.16, p=0.039) and increased duration of URI symptoms (median 7.0 vs 5.0 days for active vs control, adjusted ratio of geometric means 1.34, 95% CI 1.09 to 1.65, p=0.005), but not with altered risk or duration of LRI.
Addition of intermittent bolus-dose vitamin D3 supplementation to a daily low-dose regimen did not influence risk of ARI in older adults and their carers, but was associated with increased risk and duration of URI.
clinicaltrials.gov NCT01069874.
A number of mechanisms have been proposed to contribute to glucocorticoid-resistant asthma, including increased expression of nuclear factor kappa B and activating protein 1 (AP-1), increased ...expression of histone deacetylase, polymorphisms in IL-10, increased expression of the dominant negative isoform of the glucocorticoid receptor beta (GRβ), and vitamin D insufficiency.2-4 Our earlier data showed that peripheral blood CD4+ T cells from glucocorticoid-resistant as compared with glucocorticoid-sensitive asthmatic patients failed to synthesize the anti-inflammatory cytokine IL-10 in response to glucocorticoid in vitro.5 The active form of vitamin D (calcitriol; 1,25-dihydroxyvitamin D3 1,25(OH)2D3) when used in combination with glucocorticoid restored this IL-10 response both in vitro and ex vivo following patient ingestion of calcitriol.6 These data, together with epidemiologic evidence linking vitamin D insufficiency/deficiency with a poor clinical response to treatment in asthma,3,4 provided the rationale for this proof-of-concept clinical trial. On the one hand, the decision to study a well-characterized cohort of glucocorticoid-resistant asthmatic patients produced significant challenges with recruitment and retention, but on the other hand may have facilitated our ability to observe a clinical effect that may be manifest most clearly in this small but important subset of patients.\n Study subjects were asked to report any adverse events from the day of commencement of the first course of oral prednisolone until 4 weeks after the second course by telephone or e-mail or at study visits.
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