A Novel ß-Globin Variant, Hb Raklev [ß 75(E19) HBB:c.227T > A (Leu→Gln)].

We present a new hemoglobin variant, Hb Raklev, characterized by the substitution of leucine with glutamine at position 75 in the ß-globin chain. This variant was discovered inadvertently during an HbA1c evaluation using high performance liquid chromatography in a symptomless 54-year-old Caucasian woman, with the same variant also identified in her 16-year-old daughter. Purification of the hemoglobin revealed possibly diminished 2,3-bisphosphoglycerate (2,3-BPG) sensitivity, which may result in heightened oxygen affinity. Notably, two variants have been previously documented at this location: the unstable Hb Atlanta and the high-affinity Hb Pasadena.

Plasma fibulin-1 levels during pregnancy and delivery: a longitudinal observational study.

BACKGROUND: Fibulin-1 is an extracellular matrix protein expressed at high levels in the placenta. Elevated circulating fibulin-1 have been observed in women with severe pre-eclampsia, whereas low levels have been found in the fetal membranes, prior to membrane rupture. The aim of the study was primarily to evaluate plasma fibulin-1 during expected normal pregnancy and delivery, and secondarily to explore fibulin-1 levels in women developing pre-eclampsia or preterm premature rupture of fetal membranes (PPROM). METHODS: From the historical longitudinal cohort originally consisting of 801 healthy Danish women with a singleton pregnancy, 128 women (632 samples) were selected. Of these, 107 women had normal pregnancies, nine experienced PPROM, and 12 pre-eclampsia. All samples were analyzed for fibulin-1, and levels were compared with blood donors. Differences in mean fibulin-1 between groups were estimated using a linear mixed model. RESULTS: The mean concentration of fibulin-1 in 120 blood donors was 15.7 µg/mL, (25th-75th-percentiles, 12.3-18.2), with no significant difference in groups stratified by gender or age. Compared to baseline levels in week 12-20, fibulin-1 levels increased significantly from week 29-34 (estimated difference, 5.6 µg/mL; standard error, 1.7; p < 0.001) and 35-42 (12.5 µg/mL; 1.6; p < 0.001) and normalized after birth. The decrease at delivery tended to be more pronounced after elective (-7.0 µg/mL; 2.3; p = 0.002) and emergency (-5.6 µg/mL; 2.9; p = 0.05) cesarean section than after vaginal delivery (reference group). Women who developed PPROM had lower fibulin-1 levels throughout their pregnancies (-11.6 µg/mL; 4.2; p = 0.006). We did not observe a correlate between late pre-eclampsia and fibulin-1 (-0.2 µg/mL; 3.0; p = 0.9). CONCLUSIONS: Fibulin-1 was above non-pregnant levels at week 12 and increased significantly throughout pregnancy. We observed an association between low levels of fibulin-1 and PPROM. Further studies are needed to examine if fibulin-1 could serve as biomarker for the risk of PPROM. However, its role in late preeclampsia is doubtful. TRIAL REGISTRATION: The study was conducted in accordance with the Declaration of Helsinki. The participants provided written informed consent, including storage for future use. The study was approved on July 18, 2005 by The Danish National Committee on Bioethics (No. KA 05065 and S-20,090,061) and the Danish Data Protection Agency.

Thyroid function in COVID-19 and the association with cytokine levels and mortality.

The hypothalamic-pituitary-thyroid hormone axis might be affected in COVID-19, but existing studies have shown varying results. It has been hypothesized that hyperinflammation, as reflected by the secretion of cytokines, might induce thyroid dysfunction among patients with COVID-19. We explored thyroid hormone involvement in the acute phase of symptomatic COVID-19 and its possible associations with cytokine levels and mortality risk. This was a single-center study of 116 consecutive patients hospitalized for moderate-to-severe COVID-19 disease. Serum concentrations of thyroid-stimulating hormone (TSH), free thyroxine (T4), and 45 cytokines/chemokines were measured in all patients within 3 days of admission. Data were extracted retrospectively through a manual review of health records. At admission, 95 (81.9%) were euthyroid; while 21 (18.1%) had biochemically thyroid dysfunction including subclinical thyrotoxicosis (n = 11), overt thyrotoxicosis (n = 2), hypothyroidism (n = 1), non-thyroidal illness (n = 2), and normal TSH but high free T4 (n = 5). TSH levels were inversely correlated with IL-8 (rs = -0.248), IL-10 (rs = -0.253), IL-15 (rs = -0.213), IP-10 (rs = -0.334), and GM-CSF (rs = -0.254). Moreover, IL-8 levels, IP-10, and GM-CSF were significantly higher in patients with serum TSH < 0.4 mIU/L. Lastly, a two-fold increment of IL-8 and IL-10 was associated with significantly higher odds of having TSH < 0.4 mIU/L (odds ratio 1.86 (1.11-3.10) and 1.78 (1.03-3.06)). Serum TSH was not associated with 30- or 90-day mortality. In conclusion, this study suggests that fluctuations of TSH levels in patients with COVID-19 may be influenced by circulating IL-8, IL-10, IL-15, IP-10, and GM-CSF as previously described in autoimmune thyroid diseases.

Evaluation of the sysmex UF-5000 fluorescence flow cytometer as a screening platform for ruling out urinary tract infections in elderly patients presenting at the Emergency Department.

In this study, we evaluated the performance of the flow cytometer-based Sysmex UF-5000 automated urine analyzer as a screening tool for ruling out urinary tract infections in elderly patients presenting at the emergency department. A total of 1119 unselected patient samples (including 544 samples from elderly patients) submitted for urine culture were included in this study. Samples were measured on UF-5000 and dipsticks and the results were compared with interpretation of culture results, which is the gold standard. We obtained a diagnostic sensitivity of 99% and specificity of 51% with a low rate of false negatives (0.2%) and a negative predictive value of 99% at 108 colony forming bacteria/L (CFB/L). A bacterial count ≥ 50x106/L or yeast like cells ≥ 25x106/L was used as the cutoff value. At this cutoff value, 30% of the urine cultures would have been redundant. This resulted in 35% false positive samples, mainly due to particle contamination or nongrowing bacteria. In comparison, at best, the dipsticks have a diagnostic sensitivity of 89%, a specificity of 52% and a negative predictive value of 92% at 108 CFB/L.

Evaluation of Therapeutic Drug Monitoring in the Clinical Management of Patients with Rheumatic Diseases: Data from a Retrospective Single-Center Cohort Study.

PURPOSE: Treatment of rheumatic diseases with tumor necrosis factor inhibitors leads to improved clinical outcomes. Therapeutic drug monitoring (TDM) may assist in guiding clinical decisions. This study investigates the impact of TDM on clinical outcome, decision-making and biologics cost expenditure. PATIENTS AND METHODS: In a retrospective observational study of 306 patients with rheumatic diseases treated with four different tumor necrosis factor inhibitors, drug levels and antidrug antibodies were measured over a period of one year. Primary outcomes were the clinicians' response to each TDM result and the clinical outcome two years after TDM initiation. Outcomes were compared between the 111 TDM-guided patients and the 195 empirically guided patients. RESULTS: Treatment change occurred in 55% of the patients in the TDM group, but in only 38% in the empirically guided group. In the TDM group, 89 (79.5%) patients were in remission or had low disease activity after two years follow-up compared to 128 (65.6%) patients in the empirical group. The average cost of biologics per patient per year was lower in the TDM group than in the empirical group for patients receiving infliximab, adalimumab or etanercept at baseline but not for golimumab. CONCLUSION: TDM-guided decision-making is useful in rheumatic patients receiving TNFi and may optimize therapeutic decisions, leading to a better control of disease activity. Proactive TDM may support decisions on dose tapering, resulting in lower drug consumption and biologics cost expenditure.

Variation in plasma 25-hydroxyvitamin D2 and D3 in normal pregnancy with gestational age, sampling season, and complications: A longitudinal cohort study.

INTRODUCTION: Low levels of vitamin D in pregnancy have been associated with the risk of a variety of pregnancy outcomes. Few studies have investigated vitamin D concentrations throughout pregnancy in healthy women, and most guidelines recommend high vitamin D levels. In the present study, we investigated 25-hydroxyvitamin D concentrations in healthy Caucasian Danish women in relation to season, gestational age and possible vitamin D-linked complications. MATERIALS AND METHODS: Eight hundred and one healthy Caucasian Danish women with an expected normal pregnancy were recruited among 2147 women attending first trimester screening. Seven blood samplings were planned throughout the pregnancy and delivery period. The 25-hydroxyvitamin D2 (25(OH)D2) and 25-hydroxyvitamin D3 (25(OH)D3) concentrations were measured by LC-MS/MS and total 25-hydroxyvitamin D (25(OH)D) were calculated. RESULTS: A total of 3304 samples from 694 women were available for 25(OH)D measurements. The mean (25th-75th percentiles) concentrations of 25(OH)D, 25(OH)D3, and 25(OH)D2 were 54.6 (38.8-68.6) nmol/L, 52.2 (36.4-66.4) nmol/L, and 2.4 (2.2-2.2) nmol/L, respectively. Season was the strongest predictor of 25(OH)D concentration, with the lowest values observed in winter and spring, where only 42% and 41% of samples, respectively, were above 50 nmol/L. Nearly all women had values below the suggested optimal level of 75 nmol/L, independent of season. 25(OH)D peaked at gestational weeks 21-34. Plasma 25(OH)D2 levels were low in all seasons. Women with complications during pregnancy had higher 25(OH)D (estimated difference 9.8 nmol/L, standard error 2.7, p<0.001) than did women without complications, and women giving birth vaginally had lower 25(OH)D than did those delivering via elective (10.0 nmol/L, standard error 2.1, p<0.001) or emergency cesarean section (6.8 nmol/L, standard error 2.2, p<0.001). CONCLUSION: The 25(OH)D concentrations vary with both season and gestational age. Healthy women had lower 25(OH)D concentrations than recommended, without an association with an increased risk of pregnancy complications. Guidelines for vitamin D in pregnancy may require revision.

Bacterial colonization of seromas after breast cancer surgery with and without local steroid prophylaxis.

BACKGROUND: Seroma formation is a frequent postoperative sequela after mastectomy for primary breast cancer. We investigated the role of bacterial colonization of seroma fluid with three different culture methods and the effect of intracavitary steroids. METHODS: The study group consisted of 212 patients scheduled for mastectomy from a previously performed double-blind randomized placebo-controlled intervention trial. The patients were allocated to a single dose of 80 mg of steroids (methylprednisolone) or saline, and the effect on seroma formation was investigated. From each aspiration, an equal volume of seroma fluid (10 mL) was distributed into one sterile transport tube (conventional method), one aerobic blood culture bottle and one anaerobic blood culture bottle. RESULTS: There was significant variation in the number of bacterial species detected in seroma samples among the three culture methods, ranging from 18 species with the conventional culture tubes to 40 species with aerobic blood culture bottles. Patients receiving prophylactic steroids had significantly more frequent colonization than those in the saline group. Nevertheless, the clinical surgical site infection rate of 7.0% was equal between the two groups. CONCLUSIONS: In general, data analysis of the entire set of case material did not succeed in demonstrating a relationship between a specific bacterial species or a combination of species and seroma formation. However, in the few patients with growth of a pathogenic species, both the duration of seroma formation and volume of seroma fluid were more pronounced. TRIAL REGISTRATION: Ethics Committee of Copenhagen (H-4-2009-137), (EudraCT number 2009-016650-40), the Danish Data Protection Agency (code J. no. F.750.75-2), and the Danish Health and Medicines Authority (sponsor protocol code number 23837). Start date November 2010.

Urinary excretion of phenols, parabens and benzophenones in young men: Associations to reproductive hormones and semen quality are modified by mutations in the Filaggrin gene.

BACKGROUND: The filaggrin gene (FLG) encodes an epidermal protein, filaggrin, which is important for normal skin barrier functions. We previously showed that FLG loss-of-function mutation carriers have a higher internal exposure to some non-persistent chemicals such as certain phthalates and parabens, suggesting increased trans-epidermal penetration. Several groups of non-persistent chemicals are suspected endocrine disrupters with potential to affect testicular function. OBJECTIVES: To investigate associations between exposure to non-persistent chemicals and testicular function in young Danish men with and without FLG mutations. METHODS: We measured urinary concentrations of bisphenol A (BPA) and other simple phenols, parabens, and UV filters including benzophenones (BP-1, BP-3 and 4-HBP) in men genotyped for FLG R501X, 2282del4, and R2447X loss-of-function mutations; in total 65 mutation carriers and 130 non-carriers (controls) were included. Outcomes were markers of testicular function, assessed by serum reproductive hormones and semen quality. RESULTS: We found that associations between urinary chemical concentrations and outcomes were different in cases and controls. Within the group of FLG mutation carriers, higher urinary concentrations of BPA, BP-1 and BP-3 were associated with higher testosterone and estradiol serum levels and lower FSH. Similar trends in hormone levels were observed for FLG mutation carriers with measurable levels of 4-HBP compared to those who had no detectable levels of urinary 4-HBP. Furthermore, those in the highest urinary BPA quartile had lower sperm motility than those in the lower quartiles. None of these associations were evident in the control group. In the control group, however, lower sperm motility and sperm concentration were observed in the men with detectable urinary 4-HBP compared to the men non-detectable urinary 4-HBP. We found no association between any parabens and outcomes, nor for the other measured phenols or UV filters. CONCLUSIONS: Associations between male reproductive health parameters and urinary levels of BPA and benzophenones such as BP-3, BP-1 and 4-HBP were observed in FLG mutation carriers but not in controls from the same study population. This difference between FLG mutation carriers and non-carriers is not explained solely by differences in exposure levels of the examined compounds as e.g. BPA and 4-HBP urinary levels did not differ between the two groups. We hypothesise that effects of exposure to these compounds may be modulated in FLG mutation carriers by either different levels of co-exposures or by route of uptake, with a higher fraction of the uptake by dermal uptake.

Filaggrin gene loss-of-function mutations explain discordance of atopic dermatitis within dizygotic twin pairs.

OBJECTIVES: This study was designed to examine the association between loss-of-function mutations in the filaggrin gene (FLG) and atopic dermatitis (AD) and asthma in adult twins. METHODS: A previously well-characterized cohort of 575 adult twins were genotyped for the loss-of-function mutations in FLG (R501X, 2282del4 and R2447X) most common among northern Europeans. Subjects were examined for symptoms of atopic diseases as well as for lung function, airway responsiveness, and atopy. RESULTS: In the whole population of twins, the risk for AD was significantly increased in individuals with FLG mutations in comparison with wild-type carriers (34.3% vs. 21.8%) after adjustment for possible confounders (odds ratio [OR] 1.92, 95% confidence interval [CI] 1.07-3.41; P = 0.028). A significant association was also observed for persistent AD (OR 2.10, 95% CI 1.02-4.36; P = 0.046). There were no significant differences in risk for asthma by FLG mutation status in individuals with and without AD, respectively (P-value for interaction, 0.595). In 11 dizygotic twin pairs discordant for FLG mutation status, risk for AD was higher in the twin carrying the FLG mutation (five of 11 [45.5%] twins had developed AD) than in the non-carrier co-twin (two of 11 [18.2%] twins had developed AD) (OR 2.50, 95% CI 0.45-13.85; P = 0.293). FLG status did not explain a significant proportion of the variation in AD (P = 0.328) or asthma (P = 0.321). CONCLUSIONS: Filaggrin gene mutations are risk factors for the presence and persistence of AD and explain the discordance of AD within dizygotic twin pairs.

A comparison of anti-nuclear antibody quantification using automated enzyme immunoassays and immunofluorescence assays.

Anti-nuclear antibodies (ANA) have traditionally been evaluated using indirect fluorescence assays (IFA) with HEp-2 cells. Quantitative immunoassays (EIA) have replaced the use of HEp-2 cells in some laboratories. Here, we evaluated ANA in 400 consecutive and unselected routinely referred patients using IFA and automated EIA techniques. The IFA results generated by two independent laboratories were compared with the EIA results from antibodies against double-stranded DNA (dsDNA), from ANA screening, and from tests of the seven included subantigens. The final IFA and EIA results for 386 unique patients were compared. The majority of the results were the same between the two methods (n = 325, 84%); however, 8% (n = 30) yielded equivocal results (equivocal-negative and equivocal-positive) and 8% (n = 31) yielded divergent results (positive-negative). The results showed fairly good agreement, with Cohen's kappa value of 0.30 (95% confidence interval (CI) = 0.14-0.46), which decreased to 0.23 (95% CI = 0.06-0.40) when the results for dsDNA were omitted. The EIA method was less reliable for assessing nuclear and speckled reactivity patterns, whereas the IFA method presented difficulties detecting dsDNA and Ro activity. The automated EIA method was performed in a similar way to the conventional IFA method using HEp-2 cells; thus, automated EIA may be used as a screening test.

Associations of filaggrin gene loss-of-function variants with urinary phthalate metabolites and testicular function in young Danish Men.

BACKGROUND: Filaggrin is an epidermal protein that is crucial for skin barrier function. Up to 10% of Europeans and 5% of Asians carry at least one null allele in the filaggrin gene (FLG). Reduced expression of filaggrin in carriers of the null allele is associated with facilitated transfer of allergens across the epidermis. We hypothesized that these individuals may have increased transdermal uptake of endocrine disruptors, including phthalates. OBJECTIVES: We investigated urinary excretion of phthalate metabolites and testicular function in young men with and without FLG loss-of-function variants in a cross-sectional study of 861 young men from the general Danish population. METHODS: All men were genotyped for FLG R501X, 2282del4, and R2447X loss-of-function variants. We measured urinary concentrations of 14 phthalate metabolites and serum levels of reproductive hormones. We also evaluated semen quality. RESULTS: Sixty-five men (7.5%) carried at least one FLG-null allele. FLG-null carriers had significantly higher urinary concentrations of several phthalate metabolites, including a 33% higher concentration of MnBP (mono-n-butyl phthalate; 95% CI: 16, 51%). FLG-null variants were not significantly associated with reproductive hormones or semen quality parameters. CONCLUSION: This study provides evidence that carriers of FLG loss-of-function alleles may have higher internal exposure to phthalates, possibly due to increased transepidermal absorption. FLG loss-of-function variants may indicate susceptible populations for which special attention to transepidermal absorption of chemicals and medication may be warranted.

Local injection of methylprednisolonacetat to prevent seroma formation after mastectomy.

INTRODUCTION: This study served the following three purposes: To evaluate the prophylactic effect against seroma of a single dose of steroid in the mastectomy cavity, to evaluate the thesis that there is a connection between subclinical bacterial colonization and seroma formation and to evaluate if a simple urine stix test can detect postmastectomy infection. MATERIAL AND METHODS: This was a double-blinded and randomized study of injection of methylprednisolonacetate versus saline in the mastectomy cavity at the time of drain removal. A total of 160 females were enrolled after mastectomy. The study parameters were as follows: seroma volume, number of seroma punctures, frequency of clinical infections, degree and type of subclinical colonization, complications and evaluation of the microbiological results of the stix test with automatically read glucose, ketones, blood, pH, protein, nitrite and leucocytes. The degree of inflammation was monitored by measurement of 15 cytokines in each sample of seroma fluid. The study was initiated in August 2010 and is expected to run for three years. DISCUSSION: Some reports have concluded that seroma formation forms part of postsurgical inflammation. Steroids are effective against inflammation and accumulation of fluid at the surgical site after several types of surgery and have also proved valuable in the treatment of seroma formation. In the present study, the prophylactic effect of steroids on seroma formation is investigated. CONCLUSION: As the incidence of postmastectomy seroma formation is 80%, there is a need for improvement in the prophylaxis and treatment of this condition. FUNDING: not relevant. TRIAL REGISTRATION: Medicines Agency The EudraCT number 2009-016650-40 has been issued for your Sponsor's Protocol Code Number 23837. Data protection agency J.no. F.750.75-2. The study is perfomed in collaboration with the GCP Unit, capital Region, Bispebjerg Hospital under the EudraCT number: 2009-016650-40.

Interaction between filaggrin null mutations and tobacco smoking in relation to asthma.

BACKGROUND: The mechanisms underlying the association between filaggrin (FLG) deficiency and asthma are not known. It has been hypothesized that FLG deficiency leads to enhanced percutaneous exposure to environmental substances that might trigger immune responses. We hypothesized that interactions between FLG deficiency and environmental exposures play a role in asthma development. OBJECTIVE: We sought to investigate possible interactions between FLG null mutations and tobacco smoking in relation to asthma. METHODS: A total of 3471 adults from a general population sample participated in a health examination. Lung function and serum specific IgE levels to inhalant allergens were measured, and information on asthma and smoking was obtained by means of questionnaire. Participants were genotyped for the 2 most common FLG null mutations in white subjects: R501X and 2282del4. Another Danish population was used for replication. RESULTS: The FLG null mutation genotype was significantly associated with a higher prevalence of asthma and decreased FEV(1)/forced vital capacity ratio. In logistic regression analyses with asthma as the outcome, a significant interaction was found between FLG null mutations and smoking status (P = .02). This interaction was confirmed, although it was not statistically significant, in another Danish population study. Interactions between FLG genotype and cumulated smoking exposure were found in relation to asthma (P = .03) and decreased FEV(1)/forced vital capacity ratio (P = .03). A 3-way interaction was found among FLG genotype, smoking, and asthma, suggesting that the FLG-smoking interaction mainly played a role in nonatopic subjects. CONCLUSION: FLG null mutations modified the effects of smoking on the risk of asthma. This finding might have implications for risk stratification of the population.

Error tracking in a clinical biochemistry laboratory.

BACKGROUND: We report our results for the systematic recording of all errors in a standard clinical laboratory over a 1-year period. METHODS: Recording was performed using a commercial database program. All individuals in the laboratory were allowed to report errors. The testing processes were classified according to function, and errors were classified as pre-analytical, analytical, post-analytical, or service-related, and then further divided into descriptive subgroups. Samples were taken from hospital wards (38.6%), outpatient clinics (25.7%), general practitioners (29.4%), and other hospitals. RESULTS: A total of 1189 errors were reported in 1151 reports during the first year, corresponding to an error rate of 1 error for every 142 patients, or 1 per 1223 tests. The majority of events were due to human errors (82.6%), and only a few (4.3%) were the result of technical errors. Most of the errors (81%) were pre-analytical. Of the remainder, 10% were analytical, 8% were post-analytical, and 1% was service-related. Nearly half of the errors (n=550) occurred with samples received from general practitioners or clinical hospital wards. Identification errors were relatively common when non-technicians collected blood samples. CONCLUSIONS: Each clinical laboratory should record errors in a structured manner. A relation database is a useful tool for the recording and extraction of data, as the database can be structured to reflect the workflow at each individual laboratory.

[Retirement of creatinkinase MB]. / Pensionering af kreatininkinase myocardial band.

The introduction of a new test is often slow and obsolete tests are hard to get rid of. After nearly 20 years, the cardiac proteins troponin I and T have not yet replaced the old creatinin kinase myocardial band (CK-MB). International guidelines now recommend the use of troponin as marker for myocardial necrosis and only CK-MB when troponin is not available. However, 23 of 46 Danish laboratories still use CK-MB. The additional cost thus incurred could be used for other beneficial purposes. It is recommended to follow the international guidelines.