Jaak Billen

Progesterone increases airway eosinophilia and hyper‐responsiveness in a murine model of allergic asthma

Background Sex hormones might affect the severity and evolution of bronchial asthma. From existing literature, there exists, however, no convincing evidence for either exacerbation or improvement of allergic symptoms by progesterone.

The role of inhibins B and antimüllerian hormone for diagnosis and follow-up of granulosa cell tumors.

The peptide hormones inhibin and antimüllerian hormone (AMH), both produced by the granulosa cells, are potential candidates for diagnosis and follow-up of granulosa cell tumors (GCTs). The objective was to evaluate the usefulness of serum levels of inhibin B and AMH in the diagnosis and follow-up of GCT. The review summarizes and discusses the value and limitations of the laboratory tests of these hormones by investigating the performance characteristics of the serum analyses. A search in PubMed database was accomplished to find articles describing serum inhibin and/or AMH as a diagnostic test or for follow-up of GCT. The literature search included articles published between 1989 and September 2008. The sensitivity of inhibin B and AMH for diagnosing patients with a progressive disease is rather equivalent. Antimüllerian hormone is a more specific serum parameter than inhibin, because inhibin may also increase in some (mucinous) epithelial ovarian tumors. Nowadays, specific and ultrasensitive assays are commercially available as well for inhibin B as for AMH, so that early detection of GCT might be possible. For patients with elevated levels of inhibin B and/or AMH at initial diagnosis of GCT, inhibin B and/or AMH seemed to be reliable markers during follow-up for early detection of residual or recurrent disease. Elevated concentrations of these hormones predict relapse earlier than clinical symptoms, which leads to less morbidity of the patients. In conclusion, inhibin B and AMH are both useful serum markers for diagnosis and especially for follow-up of patients with a GCT. Currently, there is no evidence-based preference for inhibin B or AMH as tumor marker.

Active vitamin D (1,25-dihydroxyvitamin D) and bone health in middle-aged and elderly men: The European Male Aging Study (EMAS)

CONTEXT There is little information on the potential impact of serum 1,25-dihydroxyvitamin D [1,25(OH)2D] on bone health including turnover. OBJECTIVE The objective of the study was to determine the influence of 1,25(OH)2D and 25-hydroxyvitamin D [25(OH)D] on bone health in middle-aged and older European men. DESIGN, SETTING, AND PARTICIPANTS Men aged 40-79 years were recruited from population registers in 8 European centers. Subjects completed questionnaires that included questions concerning lifestyle and were invited to attend for quantitative ultrasound (QUS) of the heel, assessment of height and weight, and a fasting blood sample from which 1,25(OH)2D, 25(OH)D, and PTH were measured. 1,25(OH)2D was measured using liquid chromatography tandem mass spectrometry. Bone markers serum N-terminal propeptide of type 1 procollagen (P1NP) and crosslinks (β-cTX) were also measured. Dual-energy x-ray absorptiometry (DXA) of the hip and lumbar spine was performed in 2 centers. MAIN OUTCOME MEASURE(S) QUS of the heel, bone markers P1NP and β-cTX, and DXA of the hip and lumbar spine were measured. RESULTS A total of 2783 men, mean age 60.0 years (SD 11.0) were included in the analysis. After adjustment for age and center, 1,25(OH)2D was positively associated with 25(OH)D but not with PTH. 25(OH)D was negatively associated with PTH. After adjustment for age, center, height, weight, lifestyle factors, and season, 1,25(OH)2D was associated negatively with QUS and DXA parameters and associated positively with β-cTX. 1,25(OH)2D was not correlated with P1NP. 25(OH)D was positively associated with the QUS and DXA parameters but not related to either bone turnover marker. Subjects with both high 1,25(OH)2D (upper tertile) and low 25(OH)D (lower tertile) had the lowest QUS and DXA parameters and the highest β-cTX levels. CONCLUSIONS Serum 1,25(OH)2D is associated with higher bone turnover and poorer bone health despite being positively related to 25(OH)D. A combination of high 1,25(OH)2D and low 25(OH)D is associated with the poorest bone health.

Small-dose oral iron absorption test in anaemic and non-anaemic elderly hospitalized patients

Abstract: The small‐dose (20 mg) oral iron absorption test (OIAT) was performed in 76 hospitalized elderly patients and 30 healthy adults. Of the elderly patients, 34 were considered as iron deficient (serum ferritin level <20 μg/L) of whom 23 were anaemic and 11 not anaemic, 21 had the anaemia of chronic disorders (ACD) and another 21 were non‐anaemic patients with a normal serum ferritin level. There was a significant inverse correlation between the serum ferritin level as a measure of iron store and the maximum increase in serum iron during a 3‐h test (Cmax), in the elderly as well as in the healthy adult group. A decision limit of 80 μg/dL for Cmax is a good discriminant between absent (serum ferritin <20 μg/L) and adequate body iron stores. Sixty‐eight per cent of the patients with a serum ferritin level <20 μg/L but virtually none of the ACD patients, non‐anaemic elderly inpatients with normal serum ferritin levels and healthy adults had a Cmax level >80 μg/L. Although further investigation is needed before the OIAT can be recommended as a valuable test for evaluating iron absorption, predicting mild iron deficiency and differentiating between different categories of anaemia, it seems worthwile that more effort should be done to validate this simple and safe test.

Calcium and bone homeostasis in heterozygous carriers of CYP24A1 mutations: A cross-sectional study.

BACKGROUND Bi-allelic CYP24A1 mutations can cause idiopathic infantile hypercalcemia (IIH), adult-onset nephrocalcinosis, and possibly bone metabolism disturbances. It is currently unclear if heterozygous carriers experience clinical problems or biochemical abnormalities. Our objective is to gain insight in the biochemical profile and health problems in CYP24A1 heterozygotes. STUDY DESIGN Cross-sectional evaluation of participants. Data of previously reported carriers are reviewed. SETTING AND PARTICIPANTS Outpatient clinic of a tertiary care hospital. Participants were eight family members of an infant with a well-characterized homozygous CYP24A1 mutation c.1186C>T p.(Arg396Trp). OUTCOMES Serum vitamin D metabolites. Symptoms or biochemical signs of hypercalcemia, hypercalciuria or nephrocalcinosis. Bone health in heterozygous as compared to wild type (WT) subjects. MEASUREMENTS Genotyping by Sanger sequencing; vitamin D metabolites by liquid chromatography tandem mass spectrometry; renal, calcium and bone markers by biochemical analyses; presence of nephrocalcinosis by renal ultrasound; bone health by dual-energy X-ray absorptiometry and peripheral quantitative computed tomography. RESULTS Six participants were heterozygous carriers of the mutation. None of the heterozygous subjects had experienced IIH. One had a documented history of nephrolithiasis, two others had complaints compatible with this diagnosis. No major differences between WT and heterozygous subjects were found regarding bone health, serum or urinary calcium or 25OHD/24,25(OH)2D ratio. Literature reports on three out of 33 heterozygous cases suffering from IIH. In all three, the 25OHD/24,25(OH)2D ratio was highly elevated. Nephrocalcinosis was frequently reported in family members of IIH cases. LIMITATIONS Small sample size, lack of a large control group. CONCLUSIONS Our and literature data suggest that most heterozygous CYP24A1 mutation carriers have a normal 25OHD/24,25(OH)2D ratio, are usually asymptomatic and have a normal skeletal status but may possibly be at increased risk of nephrocalcinosis. A review of the available literature suggests that an elevated 25OHD/24,25(OH)2D ratio may be associated with symptoms of IHH, irrespective of carrier status.

Need for Estradiol Assays With a Lower Functional Sensitivity in Clinical Studies Examining Postmenopausal Women Treated With Aromatase Inhibitors

TO THE EDITOR: In their recently published study, Folkerd et al found that in postmenopausal women with early estrogen receptor– positive breast cancer who are treated with aromatase inhibitors (AIs), suppressed levels of estradiol and estrone sulfate are related to body mass index. If confirmed, this observation is of great interest, not only because it would explain why treatment with an AI results in a better outcome compared with tamoxifen in lean patients but not in obese patients, but also because it might open the way toward individualized AI treatment by adapting the dose of AIs on the basis of the plasma estradiol and estrone sulfate concentration. Although the measurement of AIs using mass spectrometry is not so difficult, the measurement of estrogens, particularly estradiol, is quite challenging. As reported by the authors, most on-treatment values for estradiol were below 3 pmol/L, which was the “conventional sensitivity” limit of the assay used. Conventional sensitivity was defined by the authors as the 95% CI of the zero standard, which corresponds more or less to the limit of blank (LoB) as defined by the Clinical and Laboratory Standards Institute. The LoB is the highest apparent analyte concentration that is expected to be found when replicates of a sample containing no analyte are tested. The limit of detection is the lowest analyte concentration likely to be reliably distinguished from the LoB and at which detection is feasible. The limit of quantitation, which is defined as the lowest concentration at which the analyte can not only be reliably detected but also measured with an acceptable degree of imprecision, is usually significantly higher (and cannot be lower) than the LoB and the limit of detection. Functional sensitivity is usually defined as the concentration that results in a coefficient of variation (CV) of 20%. The authors reported a between-run CV of 12% for a concentration of 37 pmol/L for their estradiol assay, suggesting that the functional sensitivity of their assay is significantly higher than 3 pmol/L, given that the CV increases exponentially around the limit of quantitation. Such a functional sensitivity is insufficient to allow a reliable quantification of estradiol concentrations in individual postmenopausal women who are treated with AIs. For estrone sulfate, there is a similar problem, although to a lesser extent. The functional sensitivity is most likely also significantly higher than 15 pmol/L, given that the authors reported a CV of 11% at 130 pmol/L, but in contrast to estradiol, a majority of on-treatment values were above the conventional sensitivity defined by the authors. Toexaminetherelationbetweenestrogenconcentration,bodymass index, and outcome, and to open the way toward individualized AI treatment, clinical studies should use estrogen assays with a better functional sensitivity, particularly for estradiol. Unfortunately, the only estradiol assays that currently have a functional sensitivity of less than 3 pmol/L are labor-intensivemassspectrometrymethodsthatrequireextensivesample pretreatmentincludingliquid/liquidextraction.Asimplemassspectrometrymethodnotrequiring liquid/liquidextraction,derivatization,or large volumes of serum (eg, 500 L) is therefore needed.

Lanreotide Reduces Liver Volume, But Might Not Improve Muscle Wasting or Weight Loss, in Patients With Symptomatic Polycystic Liver Disease

BACKGROUND & AIMS Polycystic liver disease (PCLD) can induce malnutrition owing to extensive hepatomegaly and patients might require liver transplantation. Six months of treatment with the somatostatin analogue lanreotide (120 mg) reduces liver volume. We investigated the efficacy of a lower dose of lanreotide and its effects on nutritional status. METHODS We performed an 18-month prospective study at 2 tertiary medical centers in Belgium from January 2011 through August 2012. Fifty-nine patients with symptomatic PCLD were given lanreotide (90 mg, every 4 weeks) for 6 months. Patients with reductions in liver volume of more than 100 mL (responders, primary end point) continued to receive lanreotide (90 mg) for an additional year (18 months total). Nonresponders were offered increased doses, up to 120 mg lanreotide, until 18 months. Liver volume and body composition were measured by computed tomography at baseline and at months 6 and 18. Patients also were assessed by the PCLD-specific complaint assessment at these time points. RESULTS Fifty-three patients completed the study; 21 patients (40%) were responders. Nineteen of the responders (90%) continued as responders until 18 months. At this time point, they had a mean reduction in absolute liver volume of 430 ± 92 mL. In nonresponders (n = 32), liver volume increased by a mean volume of 120 ± 42 mL at 6 months. However, no further increase was observed after dose escalation in the 24 patients who continued to the 18-month end point. All subjects had decreased scores on all subscales of the PCLD-specific complaint assessment, including better food intake (P = .04). Subjects did not have a mean change in subcutaneous or visceral fat mass, but did have decreases in mean body weight (2 kg) and total muscle mass (1.06 cm(2)/h(2)). Subjects also had a significant mean reduction in their level of insulin-like growth factor 1, from 19% below the age-adjusted normal range level at baseline to 50% at 18 months (P = .002). CONCLUSIONS In a prospective study, we observed that low doses of lanreotide (90 mg every 4 weeks) reduced liver volumes and symptoms in patients with PCLD. However, patients continued to lose weight and muscle mass. The effects of somatostatin analogues on sarcopenia require investigation. Clinicaltrials.gov: NCT01315795.

Determination of human reference values for serum total 1,25-dihydroxyvitamin D using an extensively validated 2D ID-UPLC-MS/MS method.

BACKGROUND To assess a patients vitamin D status the precursor metabolite 25-hydroxyvitamin D can be determined. However, measurement of 1,25-dihydroxyvitamin D is required when disorders of 1a-hydroxylation, extrarenal 1a-hydroxylation, or vitamin D receptor defects are suspected. METHODS The aim of this study was to determine reference values for 1,25-dihydroxyvitamin D3 and D2 using a 2D ID-UPLC-MS/MS method. RESULTS The LC-MS/MS method, able to measure picomolar concentrations of both 1,25-dihydroxyvitamin D3 and D2 in human serum, was extensively validated. Intra-assay variations were <5% and 8.5% and <7.5% and 11%, for 1,25-dihydroxyvitamin D3 and D2, respectively, over the whole dynamic range (3.1-376 and 3.1-652pmol/L). Limit of quantitation was 3.4pmol/L for both compounds. Our method correlated well with a published LC-MS/MS method (r=0.87) and with the average 1,25-dihydroxyvitamin D3 results of the vitamin D External Quality Assessment Scheme (DEQAS) determined with LC-MS/MS (r=0.93). Reference ranges, determined in 96 plasma samples of healthy volunteers were 59-159pmol/L and <17pmol/L for respectively 1,25-dihydroxyvitamin D3 and D2. The female part of the reference group showed a statistically significant decrease of 1,25-dihydroxyvitamin D3 concentrations with age. The presence of significantly higher average 1,25-dihydroxyvitamin D3 levels in premenopausal women taking oral contraceptive pills compared to postmenopausal women suggests that this effect is estrogen-related, as estrogens lead to a higher vitamin D binding protein. CONCLUSIONS The major finding of the present study is a reference interval of 59-159pmol/L for 1,25-dihydroxyvitamin D3 determined with a highly sensitive and precise LC-MS/MS method.

Arthralgia induced by endocrine treatment for breast cancer: A prospective study of serum levels of insulin like growth factor-I, its binding protein and oestrogens

BACKGROUND Aromatase inhibitors (AIs) frequently induce or enhance musculoskeletal problems (AI-induced musculoskeletal syndrome (AIMSS)) which sometimes are debilitating. Apart from low oestrogen levels, underlying mechanisms are unknown and likely multiple. We previously hypothesised a role for the growth hormone/insulin like growth factor-I (IGF-I) axis. Here, we report the effect of tamoxifen and AI on IGF-I, IGF binding protein-3 (IGFBP-3) and oestrogen levels from a prospective study. MATERIALS AND METHODS Postmenopausal women with an early breast cancer scheduled to start adjuvant endocrine therapy with an AI or tamoxifen were recruited. A rheumatologic questionnaire was completed and serum was collected for assessment of IGF-I, IGFBP-3 and oestrogen levels. Re-evaluation was done after 3, 6 and 1 2months of therapy. RESULTS 84 patients started on tamoxifen (n=42) or an AI (n=42). 66% of the latter group experienced worsening of pre-existing or de novo complaints in joint and/or muscle, compared to 29% of tamoxifen-treated patients. AI therapy resulted in elevated IGF-I levels with a statistically significant increase at 6months (p=0.0088), whereas tamoxifen users were characterised by a decrease in IGF-I levels at all follow-up times (p<0.0004). No effect on IGFBP-3 was seen in the latter group. AI-users, however, showed decreased IGFBP-3 levels at 12 months (p=0.0467). AIMSS was characterised by a decrease in IGFBP-3 levels (p=0.0007) and a trend towards increased IGF-I/IGFBP-3 ratio (p=0.0710). CONCLUSION These findings provide preliminary evidence that AI-induced musculoskeletal symptoms are associated with changes in the growth hormone (GH)/IGF-I axis.

1β,25-Dihydroxyvitamin D3: A new vitamin D metabolite in human serum

BACKGROUND The measurement of 1α,25(OH)2D3 in human serum poses a true challenge as concentrations are very low and structurally similar metabolites can interfere. MATERIALS AND METHODS During optimization of our in-house LC-MSMS method for serum 1α,25(OH)2D3 a previously co-eluting isobaric interference was separated. The isobar was identified as 1β,25(OH)2D3 by comparing retention time and fragmentation spectra to standards (other isobaric dihydroxylated vitamin D3 analogs). 1β,25(OH)2D3 showed specific cluster formation (water), not present in 1α,25(OH)2D3. 1β,25(OH)2D3 was measured in serum of apparently healthy human volunteers (n=20), patients with high serum 25-hydroxyvitamin D [25(OH)D] concentrations (>50ng/mL) (n=33 among which 4 with very high levels (>150ng/mL)) and patients with kidney failure (n=68; 39 stage 1-3, 29 stage 4-5). Pearsons r was calculated for correlations and Mann-Whitney statistic to compare group medians. RESULTS Median serum 1β,25(OH)2D3 was 11pg/mL in apparently healthy volunteers and increased to 20pg/mL for serum 25(OH)D concentrations above 80ng/mL (n=22) (p<0.0001). 1β,25(OH)2D3 concentrations were significantly correlated to serum 25(OH)D concentrations (r=0.85) for the combined results from healthy volunteers and patient sera (n=53) (p<0.0001). For patients with kidney failure, median serum 1β,25(OH)2D3 was 7pg/mL and not different from the median level in healthy volunteers (p=0.06). The median concentration did not vary with different stages. CONCLUSIONS We present evidence for the widespread presence of 1β,25(OH)2D3, a new vitamin D metabolite, in human serum. The level increases with rising serum 25(OH)D concentrations and is particularly high in patients with very high 25(OH)D levels. We previously demonstrated that 1β,25(OH)2D3 is a poor genomic agonist but a potent non-genomic antagonist of 1α,25(OH)2D3. The clinical implications of the presence of this analog therefore require further exploration.