Plasma metabolomics in adults with cystic fibrosis during a pulmonary exacerbation: A pilot randomized study of high-dose vitamin D3 administration

Abstract

Background: Cystic fibrosis (CF) is a chronic catabolic disease often requiring hospitalization for acute episodes of worsening pulmonary exacerbations. Limited data suggest that vitamin D may have beneficial clinical effects, but the impact of vitamin D on systemic metabolism in this setting is unknown.

Objective: We used high-resolution metabolomics (HRM) to assess the impact of baseline vitamin D status and high-dose vitamin D₃ administration on systemic metabolism in adults with CF with an acute pulmonary exacerbation.

Design: Twenty-five hospitalized adults with CF were enrolled in a randomized trial of high-dose vitamin D₃ (250,000IU vitamin D₃ bolus) versus placebo. Age-matched healthy subjects served as a reference group for baseline comparisons. Plasma was analyzed with liquid chromatography/ultra-high resolution mass spectrometry. Using recent HRM bioinformatics and metabolic pathway enrichment methods, we examined associations with baseline vitamin D status (sufficient vs. deficient per serum 25-hydroxyvitamin D concentrations) and the 7-day response to vitamin D₃ supplementation.

Results: Several amino acids and lipid metabolites differed between CF and healthy control subjects, indicative of an overall catabolic state. In CF subjects, 343 metabolites differed (P<0.05) by baseline vitamin D status and were enriched within 7 metabolic pathways including fatty acid, amino acid, and carbohydrate metabolism. A total of 316 metabolites, which showed enrichment for 15 metabolic pathways-predominantly representing amino acid pathways-differed between the vitamin D₃- and placebo-treated CF subjects over time (P<0.05). In the placebo group, several tricarboxylic acid cycle intermediates increased while several amino acid-related metabolites decreased; in contrast, little change in these metabolites occurred with vitamin D₃ treatment.

Conclusions: Numerous metabolic pathways detected by HRM varied in association with vitamin D status and high-dose vitamin D₃ supplementation in adults with CF experiencing a pulmonary exacerbation. Overall, these pilot data suggest an anti-catabolic effect of high-dose vitamin D₃ in this clinical setting.

Keywords: Cholecalciferol; Cystic fibrosis; Metabolomics; Pulmonary exacerbation; Vitamin D.

Figure 1. LIMMA analysis discriminates between vitamin D sufficient and insufficient adult subjects with CF

Vitamin D sufficiency is defined as serum 25-hydroxyvitamin D [25(OH)D] ≥30 ng/mL. A total of 343 metabolites differed by vitamin D status at P < 0.05 using LIMMA. A) Type 2 Manhattan plot showing the negative log₁₀P-value of metabolite comparisons as a function of the chromatographic retention time. Metabolites above the dotted horizontal line were significant at a P-value <0.05. Metabolites shaded in blue and red are lower and higher, respectively, in the vitamin D insufficient group relative to the sufficient group. B) Two-way hierarchical cluster analysis of significant metabolites illustrating separation of the CF subjects as a function of baseline vitamin D status. Vitamin D sufficient CF subjects are depicted in green along the upper X-axis, while vitamin D insufficient CF subjects are depicted in red. C) Mummichog pathway enrichment analysis. The number of significant metabolites over the number of possible metabolites in each pathway is provided in parentheses. Pathways with less than 4 matches were excluded. Horizontal bars depict the negative log of pathway enrichment p values.

Figure 2. Metabolic effects of high-dose vitamin D₃ treatment in hospitalized adults with CF

Subjects were given 250,000 IU vitamin D₃ or placebo within 48 h of hospital admission for pulmonary exacerbation. High-resolution metabolomics analysis was performed in plasma collected before study drug administration and 1 week later. Linear mixed-effects model analyses, adjusted for age and genotype, were conducted on all detected metabolites. Out of 9,258 metabolites present in at least 80% of either the vitamin D or placebo groups across both points, 316 metabolites demonstrated a significant group-by-time interaction (P<0.05). A) Type 2 Manhattan plot showing the negative log₁₀P-value of the group-by-time interaction term of each metabolite as a function of the chromatographic retention time. Metabolites above dashed horizontal line were statistically significant. B) Mummichog pathway enrichment analysis of significant metabolites. The number of significant metabolites over the number of possible metabolites in each pathway are provided in parenthesis. Only pathways with 4 or more significant metabolites were included. The bar graphs indicate the negative log₁₀P-value. C) Representative metabolites related to amino acid pathways and the TCA cycle based on mummichog module analysis (P<0.001).