Effects of High Doses of Cholecalciferol in Normal Subjects: A Randomized Double-Blinded,Placebo-Controlled Trial

Background

Vitamin D repletion with high doses of vitamin D is often recommended to patients and healthy subjects. The safety, especially concerning changes in urinary calcium excretion is of great importance.

Methods

In a double-blinded, placebo-controlled study in 40 healthy volunteers, we examined the changes in mineral metabolism during supplementation with 3000 IU of oral cholecalciferol daily during 4 months.

Results

Both 25(OH)vitamin D and 1,25(OH)₂vitamin D increased significantly in the active treated group as compared to the placebo group (186% versus 14% (P<0.001) and 28% versus – 8% (P<0.001)). No change was observed in urinary calcium excretion in the active group compared to the placebo group (P = 0.891). Fibroblast growth factor 23 increased significantly by 10% (P<0.018) in the active group. However, there was no difference in changes in FGF23 between treatment groups (P = 0.457).

Conclusion

High dose cholecalciferol significantly increases 25(OH)vitamin D and 1,25(OH)₂vitamin D levels compared to placebo. No changes in urinary calcium excretion or other measured components of the mineral metabolism were found between groups.

Trial Registration

ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT00952562","term_id":"NCT00952562"}}NCT00952562.     

Influence of Hepatitis C Virus and IL28B Genotypes on Liver Stiffness

Objective

Liver fibrosis has been associated with hepatitis C virus (HCV) genotype and genetic variation near the interleukin 28B (IL28B) gene, but the relative contribution is unknown. We aimed to investigate the relation between HCV genotypes, IL28B and development of liver stiffness.

Patients and Methods

This cross-sectional study consists of 369 patients with chronic hepatitis C (CHC). Liver stiffness was evaluated using transient elastograhy (TE). Factors associated with development of liver fibrosis were identified by logistic regression analysis.

Results

We identified 369 patients with CHC. 235 were male, 297 Caucasians, and 223 had been exposed to HCV through intravenous drug use. The overall median TE value was 7.4 kPa (interquartile range (IQR) 5.7–12.1). HCV replication was enhanced in patients carrying the IL28B CC genotype compared to TT and TC (5.8 vs. 5.4 log₁₀ IU/mL, p = 0.03). Patients infected with HCV genotype 3 had significantly higher TE values (8.2 kPa; IQR, 5.9–14.5) compared to genotype 1 (6.9 kPa; IQR, 5.4–10.9) and 2 (6.7 kPa; IQR, 4.9–8.8) (p = 0.02). Within patients with genotype 3, IL28B CC genotype had the highest TE values (p = 0.04). However, in multivariate logistic regression, using various cut-off values for fibrosis and cirrhosis, only increasing age (odds ratio (OR) 1.09 (95% confidence interval (CI), 1.05–1.14 per year increment)), ALT (OR 1.01 (95% CI, 1.002–1.011), per unit increment) and HCV genotype 3 compared to genotype 1 (OR 2.40 (95% CI, 1.19–4.81), were consistently associated with cirrhosis (TE>17.1 kPa).

Conclusions

Age, ALT and infection with HCV genotype 3 were associated with cirrhosis assessed by TE. However, IL28B genotype was not an independent predictor of fibrosis in our study.     

Serum levels of the cytokines IL-1beta, IL-6 and ICAM-1 after 131I-treatment of Graves' disease and nodular goiter.

Cytokines might be involved in the immunological flare up, seen in some patients after 131I-treatment. Therefore, we measured serum levels of interleukin-6 (IL-6), interleukin-1beta (IL-1beta), interleukin-6 soluble receptor (IL-6sR) and Intercellular-adhesion-molecule-1 (ICAM-1) as well as tumor necrosis factor (TNF-alpha) after 131I-treatment of Graves' disease and nodular goiter. Seven patients with Graves' disease, eight with toxic nodular goiter and seven with non-toxic nodular goiter, were followed after 131I-treatment. The patients were treated in the euthyroid state. Blood samples were drawn at day 0, 4, 7, 21 and after 3 months. Significant increases were seen in free T4 index (FT4I), free T3 index (FT3I) and thyroglobulin (Tg) within the first weeks, and TSH simultaneously decreased. None of the cytokines demonstrated any change during follow-up, neither in the entire group nor in subgroups. FT4I and FT3I correlated significantly to ICAM-1. In conclusion, our data suggest that there does not seem to be prolonged cytokine activation after 131I-treatment for thyroid disorders.     

Environmental variation partitioned into separate heritable components

Trait variation is normally separated into genetic and environmental components, yet genetic factors also control the expression of environmental variation, encompassing plasticity across environmental gradients and within‐environment responses. We defined four components of environmental variation: plasticity across environments, variability in plasticity, variation within environments, and differences in within‐environment variation across environments. We assessed these components for cold tolerance across five rearing temperatures using the Drosophila melanogaster Genetic Reference Panel (DGRP). The four components were found to be heritable, and genetically correlated to different extents. By whole genome single marker regression, we detected multiple candidate genes controlling the four components and showed limited overlap in genes affecting them. Using the binary UAS‐GAL4 system, we functionally validated the effects of a subset of candidate genes affecting each of the four components of environmental variation and also confirmed the genetic and phenotypic correlations obtained from the DGRP in distinct genetic backgrounds. We delineate selection targets associated with environmental variation and the constraints acting upon them, providing a framework for evolutionary and applied studies on environmental sensitivity. Based on our results we suggest that the traditional quantitative genetic view of environmental variation and genotype‐by‐environment interactions needs revisiting.     

Immunochip analysis identifies association of the RAD50/IL13 region with human longevity

Human longevity is characterized by a remarkable lack of confirmed genetic associations. Here, we report on the identification of a novel locus for longevity in the RAD50/IL13 region on chromosome 5q31.1 using a combined European sample of 3208 long‐lived individuals (LLI) and 8919 younger controls. First, we performed a large‐scale association study on 1458 German LLI (mean age 99.0 years) and 6368 controls (mean age 57.2 years) by targeting known immune‐associated loci covered by the Immunochip. The analysis of 142 136 autosomal single nucleotide polymorphisms (SNPs) revealed an Immunochip‐wide significant signal (P_Immunochip = 7.01 × 10^–9) for the SNP rs2075650 in the TOMM40/APOE region, which has been previously described in the context of human longevity. To identify novel susceptibility loci, we selected 15 markers with P_Immunochip < 5 × 10^–4 for replication in two samples from France (1257 LLI, mean age 102.4 years; 1811 controls, mean age 49.1 years) and Denmark (493 LLI, mean age 96.2 years; 740 controls, mean age 63.1 years). The association at SNP rs2706372 replicated in the French study collection and showed a similar trend in the Danish participants and was also significant in a meta‐analysis of the combined French and Danish data after adjusting for multiple testing. In a meta‐analysis of all three samples, rs2706372 reached a P‐value of P_{Immunochip+Repl} = 5.42 × 10^?7 (OR = 1.20; 95% CI = 1.12–1.28). SNP rs2706372 is located in the extended RAD50/IL13 region. RAD50 seems a plausible longevity candidate due to its involvement in DNA repair and inflammation. Further studies are needed to identify the functional variant(s) that predispose(s) to a long and healthy life.     

Purification and some properties of uracil phosphoribosyltransferase from Escherichia coli K12

Uracil phosphoribosyltransferase from Escherichia coli K12 was purified to homogeneity as determined by polyacrylamide gel electrophoresis. For this purpose a pyrimidine-requiring strain harboring the upp gene on a ColE1 plasmid was used, which showed 15-times higher uracil phosphoribosyltransferase activity in a crude extract. When this strain was grown under conditions of uracil starvation, an additional 10-times elevation of the enzyme activity was obtained. The molecular weight of uracil phosphoribosyltransferase was determined to be 75000; the enzyme consists of three subunits with a molecular weight of 23500. Uracil phosphoribosyltransferase is specific for uracil and some uracil analogues. The apparent Km values for uracil and PRib-PP were 7 microM and 300 microM, respectively. As an effector of enzyme activity, GTP lowered the Km for PRib-PP to 90 microM and increased the Vmax value 2-fold, but had no effect on the Km for uracil. The effect of GTP was found to be pH-dependent. The enzymatic characterization of uracil phosphoribosyltransferase and the observed regulation of its synthesis emphasizes the role of the enzyme in pyrimidine salvage.     

Genetic evidence for a repressor of synthesis of cytosine deaminase and purine biosynthesis enzymes in Escherichia coli.

Addition of purines to the growth medium of Escherichia coli represses synthesis of cytosine deaminase (codA) and enzymes of purine de novo synthesis. After Tn10 mutagenesis, mutants displaying derepressed levels of cytosine deaminase in the presence of hypoxanthine were isolated. One of these had simultaneously acquired resistance to the hypoxanthine analog 6-mercaptopurine. The mutation purR6::Tn10 was shown to affect de novo synthesis of the purine enzymes glutamine phosphoribosylpyrophosphate amidotransferase (purF) and phosphoribosyl glycinamide synthetase (purD). The mutation was mapped by P1 transduction at 36 min on the E. coli linkage map. A plasmid containing the purR region was obtained by complementation of the purR6::Tn10 mutation. By comparing the restriction maps of the cloned fragment and the E. coli chromosome, the purR gene was found to be located very close to the lpp gene (36.3 min).