Proof-of-concept study: Remote capillary blood collection for hCG analysis in early pregnancy.

INTRODUCTION: Capillary blood collection, a technique traditionally used in diabetes care, shows promise for many applications including pregnancy monitoring. Serial measurement of serum human Chorionic Gonadotrophin (hCG) is frequently necessary for managing early pregnancy, including molar pregnancy, requiring multiple visits to a maternity hospital for blood collection by venepuncture. This proof-of-concept study aimed to assess the clinical performance and user acceptability of capillary blood samples collected remotely, as an alternative to venous blood for hCG measurement. METHODS: Women attending the early pregnancy unit who required serum hCG measurement, were invited to participate. Following informed written consent, participants were shown how to collect capillary blood samples using the Mini-Collect® collection device. Matched venous and capillary blood samples were collected in clinic for hCG comparison purposes. Participants were also supplied with a home collection kit in a prepaid return envelope. They were asked to perform a finger-prick blood collection at home using the instructions provided and to return the capillary blood sample by post within 24 h of collection, along with a completed user-satisfaction questionnaire. Statistical analysis was performed using Analyse-it® software. RESULTS: The study enrolled 71 participants and over a third of these women collected a capillary blood sample at home. The median age of participants was 33 years (range 29-36). Passing-Bablok linear regression (y = -0.037 + 1.04x) and Spearman correlation (r = 0.999, p < 0.0001), demonstrated good agreement and strong correlation between venous and capillary samples, over a broad range of hCG values (1.2 to 224,0000 IU/L). The majority of capillary samples collected remotely (39%, 27/69) had sufficient blood volume for analysis (74%, 20/27). Respondents (77%, 18/25) found the collection device easy to use and expressed willingness to use a future service if available (80%, 20/25) CONCLUSION: The study demonstrated excellent agreement between the hCG results obtained from both collection methods, suggesting that capillary blood can serve as a reliable alternative for venous hCG measurement, particularly in clinical settings requiring frequent hCG monitoring. Feedback from the study questionnaire indicates a preference for this type of follow-up among women, indicating potential improvements in compliance for blood based diagnostic tests.

Novel scoring system provides high separation of diploidy and triploidy to aid partial hydatidiform mole diagnosis: an adaption of HER2 D-DISH for ploidy analysis.

AIMS: Diagnosis of hydatidiform mole or molar pregnancy based on morphology alone can be challenging, particularly in early gestation, necessitating the use of ancillary techniques for accurate diagnosis. We sought to adapt the VENTANA HER2 dual-colour dual-hapten in-situ hybridisation (D-DISH) assay by using the internal chromosome 17 enumeration probe to determine ploidy status. METHODS: We selected 25 products of conception, consisting of molar and non-molar cases, to validate the HER2 D-DISH assay. These cases had prior morphological assessment by a perinatal pathologist and ploidy analysis using molecular cytogenetics. Three independent observers, blinded to the original histopathological and genetic diagnosis, scored 10 representative areas on each slide. Interobserver variability was assessed by comparing the total scores of each observer using analysis of variance (ANOVA) and the kappa statistic. RESULTS: Our ploidy scoring system accurately determined the correct number of diploid and triploid conceptuses, demonstrating complete concordance with pre-existing ploidy status and the initial diagnosis. Interobserver agreement between three independent scorers was robust: ANOVA (p=0.36) and kappa statistic (0.812, p<0.001). We achieved clear separation of average nuclear signals for diploid and triploid conceptuses, which was statistically significant (p<0.05). Employing our innovative scoring system, known as the 'rule of 5', we established ploidy decision thresholds for all 25 cases. CONCLUSIONS: Our modified HER2 D-DISH ploidy assay simplifies the process of ploidy determination and improves the accuracy of morphological diagnosis of molar pregnancy. The HER2 D-DISH assay was selected for ploidy analysis due to the widespread availability of in-situ hybridisation in pathology laboratories.

Abnormal Glucose Tolerance in Women Diagnosed With Gestational Diabetes (WHO 2013) 10 Years After Index Pregnancy.

Context: It is not clear if the risk of abnormal glucose tolerance (AGT) is attenuated in the long-term in women diagnosed with gestational diabetes (GDM) using the World Health Organization (WHO) 2013 criteria and who have received appropriate treatment during pregnancy. Objective: We aimed to assess the long-term prevalence of AGT and other cardiovascular disease (CVD) risk factors in this cohort. Methods: A retrospective cohort follow-up study was conducted of 37 and 107 women diagnosed with and without GDM respectively using the WHO 2013 criteria between June 2010 and December 2010. Women were invited to attend our center, where they underwent a 75-g oral glucose tolerance test, blood and urine collection, body measurements, and electrocardiography. Main outcome measure included the development of AGT using the American Diabetes Association criteria. Results: Sixteen (43.2%) women with GDM compared to 5 (4.7%) women with normal glucose tolerance (NGT) at index pregnancy had AGT (P < .001). In the GDM group, 10 (27.0%), 7 (18.9%), and 4 (10.8%) women had impaired fasting glucose (IFG), impaired glucose tolerance (IGT), and type 2 diabetes mellitus (T2DM), respectively. In the NGT group, 2 (1.9%), 3 (2.8%), and 1 (0.9%) woman had IFG, IGT, and T2DM, respectively. Women with AGT also had an unfavorable metabolic profile including obesity, hypertension, insulin resistance, and dyslipidemia. Conclusion: Women treated for GDM (WHO 2013 criteria) remain at increased risk for developing AGT and adverse CVD risk factors as early as a decade after diagnosis. Continued efforts are needed to accurately follow this population to address modifiable risk factors.

Impact of the COVID-19 lockdown on the vitamin D status of people in the West of Ireland.

OBJECTIVE: Identify the impact of COVID-19 lockdown restrictions on the vitamin D status of individuals in the west of Ireland. DESIGN: Cross-sectional study. SETTING: Adults who had wintertime serum 25(OH)D analysis completed in Galway University Hospital. PARTICIPANTS: A total of 16,725 participants (2015-2020 (n = 13,449) and 2020-2021 (n = 3276)). Baseline demographics; sex, age, origin of the sample and the date of sample collection. RESULTS: Median serum vitamin D and serum vitamin D3 concentrations were higher in the 5-month period from October-February 2020-2021 (61 nmol/L (± 36-85 nmol/L) and 60 nmol/L (± 34-85 nmol/L)) respectively, than for the corresponding 5-month period (October-February) in 2015-2020 (53 nmol/L (± 32-78 nmol/L) and 51 nmol/L (± 30-77 nmol/L)) respectively. These changes coincided with a decline in the prevalence of deficiency. In the 5-month period October-February 2020-2021, 19.2% of the population were vitamin D deficient (< 30 nmol/L) compared to 22.5% in the corresponding 5-month period in 2015-2020, and 38.1% were vitamin D deficient (< 50 nmol/L) in the 5-month period October-February 2020-2021 compared to 46.6% in the corresponding 5-month period in 2015-2020. Males were more likely to be deficient at both thresholds (p < 0.001). For the total cohort, at the < 30 nmol/L threshold, inpatients (25.5%) and nursing home residents (34.1%) had higher prevalence of deficiency. CONCLUSIONS: Vitamin D levels were higher in the 5-month period of October-February 2020-2021, and this precipitated a decline in deficiency at both thresholds, indicating that lockdown coincided with enhanced vitamin D status. We postulate that it may be attributable to changes in diet and/or supplementation, or increased sun exposure, but further confirmatory studies are required.

Maintaining glucose integrity ex-vivo: Impact of preanalytical specimen handling.

BACKGROUND: Adopting the WHO protocol for glucose analysis is arguably impractical in the routine clinical setting. Deviations may develop due to a lack of understanding regarding the impact of glycolysis on the accuracy of results. AIM: We sought to assess the stability of glucose in two different blood collection tubes (BCT), BD Vacutainer® FX 'Fl-Ox' and Greiner Vacuette® FC-Mix 'FC-Mix' stored at room temperature (RT:18-22°C) and 4°C over 8.5 days. METHOD: Each participant provided venous whole blood collected into 51 BCTs; 'Fl-Ox' (n = 26) and 'FC-Mix' (n = 25). One Fl-Ox sample from each participant was handled according to the WHO recommended method. The remaining BCTs were stored at 4°C/RT prior to analyses at designated study timepoints. Glucose was measured using the hexokinase assay on the Cobas® 8000 platform. RESULTS: Participants (n = 8, Male = 2) were aged 24-56 years. Plasma glucose measured in FI-Ox BCTs according to the WHO sample-handling method had a median concentration of 5.73 mmol/L (Range: 5.39-10.37 mmol/L). Glucose decreased by greater than minimal difference (>0.26 mmol/L) in blood collected into Fl-Ox and stored @4°C/RT within 24 h of phlebotomy. FC-Mix BCT maintained glucose <0.26 mmol/L @4°C over a period of 8.5 days and up to 4 days @RT when compared to the WHO recommended method. CONCLUSION: Glucose in FC-Mix BCT stored @4°C demonstrated the best agreement with results determined using the WHO specifications. When FC-Mix tubes were stored @RT, glucose was stable for 4 days. These findings suggest that the FC-Mix BCT effectively inhibits glycolysis and should be introduced into routine clinical practice.

The utility of plasma glycated CD59 in predicting postpartum glucose intolerance: A prospective study of women diagnosed with GDM during a period of universal GDM screening.

AIMS: Gestational diabetes (GDM) is associated with the development of postpartum (PP) glucose intolerance. Plasma glycated CD59 (pGCD59) is an emerging biomarker for the detection of hyperglycaemia. The aim of this study was to assess the ability of PP pGCD59 to predict the development of PP GI as defined by the 2 h 75 g OGTT using the ADA criteria, in a cohort of women diagnosed with prior GDM in the index pregnancy using the 2 h 75 g OGTT at 24-28 weeks of gestation according to the World Health Organization (WHO) 2013 criteria. METHODS: Of the 2017 pregnant women recruited prospectively 140 women with gestational diabetes had samples for pGCD59 taken PP at the time of the OGTT. The ability of pGCD59 to predict the results of the PP OGTT was assessed using nonparametric receiver operating characteristic (ROC) curves. RESULTS: Women with PP glucose intolerance had significantly higher PP pGCD59 levels compared to women with normal glucose tolerance PP (3.8 vs. 2.7 SPU). PP pGCD59 identified women who developed glucose intolerance PP with an AUC of 0.80 (95% CI: 0.70-0.91). A PP pGCD59 cut-off value of 1.9 SPU generated a sensitivity of 100% (95% CI: 83.9-100), specificity of 16.9% (95% CI: 9.8-26.3), positive predictive value of 22.1% (95% CI: 21.0-22.6), and negative predictive value of 100% (95% CI: 87.4-100). PP fasting plasma glucose generated an AUC of 0.96 (95% CI: 0.89-0.99) for the identification of PP glucose intolerance. CONCLUSION: Our study found that PP pGCD9 may be a promising biomarker to identify women not requiring PP glucose intolerance screening using the traditional OGTT. While the diagnostic accuracy of pGCD59 is good, fasting plasma glucose remains a better test for the identification of PP glucose intolerance.

Sublethal Hyperthermia Transiently Disrupts Cortisol Steroidogenesis in Adrenocortical Cells.

Primary aldosteronism is the most common cause of secondary hypertension. The first-line treatment adrenalectomy resects adrenal nodules and adjacent normal tissue, limiting suitability to those who present with unilateral disease. Use of thermal ablation represents an emerging approach as a possible minimally invasive therapy for unilateral and bilateral disease, to target and disrupt hypersecreting aldosterone-producing adenomas, while preserving adjacent normal adrenal cortex. To determine the extent of damage to adrenal cells upon exposure to hyperthermia, the steroidogenic adrenocortical cell lines H295R and HAC15 were treated with hyperthermia at temperatures between 37 and 50°C with the effects of hyperthermia on steroidogenesis evaluated following stimulation with forskolin and ANGII. Cell death, protein/mRNA expression of steroidogenic enzymes and damage markers (HSP70/90), and steroid secretion were analyzed immediately and 7 days after treatment. Following treatment with hyperthermia, 42°C and 45°C did not induce cell death and were deemed sublethal doses while ≥50°C caused excess cell death in adrenal cells. Sublethal hyperthermia (45°C) caused a significant reduction in cortisol secretion immediately following treatment while differentially affecting the expression of various steroidogenic enzymes, although recovery of steroidogenesis was evident 7 days after treatment. As such, sublethal hyperthermia, which occurs in the transitional zone during thermal ablation induces a short-lived, unsustained inhibition of cortisol steroidogenesis in adrenocortical cells in vitro.

Major disparities in patient-reported adherence compared to objective assessment of adherence using mass spectrometry: A prospective study in a tertiary-referral hypertension clinic.

AIM: Many challenges exist in determining true rates of adherence to antihypertensive medications among individuals in a clinic setting. For the first time, we aimed to compare patient-reported antihypertensive adherence with objective evidence using mass spectrometry spot urinalysis in a tertiary referral clinic setting. METHODS: A prospective observational single-centre cohort study was performed in a tertiary referral hypertension clinic, encompassing antihypertensive initiation and persistence. Patients were referred with apparent treatment-resistant hypertension or for suspected secondary causes. Participants completed a self-reported assessment of antihypertensive adherence and provided a spot urine sample. The presence of antihypertensive medications and/or their respective metabolites was evaluated using high-performance liquid chromatography tandem mass spectrometry. Patients were determined to be adherent if they demonstrated both self-reported adherence and objective mass spectrometry evidence. RESULTS: Of all 105 eligible participants initially recruited, 73 (69.5%) met the eligibility criteria. Only 27.4% (95% confidence interval 0.2-0.4) of participants demonstrated true adherence to their self-reported antihypertensives, despite 75.3% (0.6-0.8) reporting adherence. Greatest medication adherence was achieved with angiotensin II receptor blockers (61%), with calcium-channel blockers and mineralocorticoid antagonists demonstrating least adherence (38%). CONCLUSION: In patients attending a tertiary hypertension clinic, the combined use of spot urine mass spectrometry and self-reporting identifies higher rates of nonadherence when compared to either modality alone. Both techniques should be combined for more accurate detection of medication adherence.

The ability of pGCD59 to predict adverse pregnancy outcomes: a prospective study of non-diabetic pregnant women in Ireland.

AIM: Even though most pregnancies are uneventful, occasionally complications do occur. Gestational diabetes is linked to an increased risk of adverse pregnancy outcomes. Early identification of women at risk of experiencing adverse outcomes, ideally through a single blood test, would facilitate early intervention. Plasma glycated CD59 (pGCD59) is an emerging biomarker which has shown promise in identifying hyperglycaemia during pregnancy and has been associated with the risk of delivering an LGA infant. The aim of this study was to explore the ability of the first- and second-trimester pGCD59 to predict adverse pregnancy outcomes. METHODS: This was a prospective study of 378 pregnant women. Samples for pGCD59 were taken at the first antenatal visit and at the time of the 2 h 75 g OGTT (24-28 weeks of gestation). Adjusted receiver operating characteristic curves were used to evaluate the ability of pGCD59 to predict maternal and neonatal outcomes. RESULTS: First-trimester pGCD59 levels were higher in women with gestational diabetes who delivered a macrosomic infant (4.2 ± 0.7 vs. 3.5 ± 1.0 SPU, p < 0.01) or an LGA infant (4.3 ± 0.3 vs. 3.6 ± 1.0 SPU, p = 0.01) compared to women with GDM that did not experience these outcomes. Second-trimester pGCD59 levels were higher in women that developed polyhydramnios (2.9 ± 0.4 vs. 2.5 ± 1.1 SPU, p = 0.03). First- and second-trimester pGCD59 predicted pregnancy-induced hypertension with good accuracy (AUC:0.85, 95%CI:0.78-0.91; AUC: 0.80, 95%CI: 0.73-0.88, respectively) and neonatal hypoglycaemia with fair to good accuracy (AUC:0.77, 95%CI: 0.54-0.99, AUC:0.81, 95%CI:0.62-0.99). CONCLUSIONS: This study has shown that pGCD59 has the potential to predict adverse pregnancy outcomes. Prospective studies with a larger number of cases are necessary to fully explore and validate the potential of this emerging biomarker in predicting adverse pregnancy outcomes.

Reference intervals for clinical biochemistry and haematology tests during normal pregnancy.

BACKGROUND: Pregnancy induces physiological changes which affect biochemical and haematological parameters. As the significance of laboratory test results change throughout pregnancy, the reference interval (RI) or key result interpretive guide should be specific to pregnancy. This study sought to establish trimester-specific-RIs for routine biochemical and haematological tests in healthy white European women with singleton pregnancies with comparison to RIs for non-pregnant European adults. METHODS: A retrospective analysis of a prospective longitudinal single-centre study of healthy pregnant women conducted between November 2018 and December 2020 in a tertiary academic hospital with approximately 3000 births annually. Inclusion criteria: signed informed consent, age ≥18 years, white European, body mass index (BMI) <25 kg/m2, blood pressure <140/90mmHg, non-smoker, no previous pathology or gestational diabetes. Trimester defined as T1: up to 13 weeks + 6 days, T2: 14-27 weeks + 6 days and T3: ≥28-41 weeks + 6 days. Baseline demographics, anthropometric and laboratory measurements were recorded. In total, 31 biochemical and 10 haematological ISO15189:2012 accredited tests were measured using Roche Cobas® and Sysmex XN-9100™ analysers, respectively. RIs were established according to the International Federation of Clinical Chemistry (IFCC) recommended method. RESULTS: Apparently healthy pregnant women (n = 124) with bio-banked serum samples in each trimester were recruited. At the booking visit, 49.2% (n = 61) of participants were nulliparous, with median age of 34.4 (IQR: 31.3-37.3) years, gestational age of 89 (IQR: 84-93) days, BMI of 22.5 (IQR: 21.0-23.7) kg/m2 and systolic and diastolic blood pressure of 116 (110-125) mmHg and 67 (61-75) mmHg, respectively. CONCLUSIONS: Normative trimester-specific biological intervals for routinely requested biochemical and haematological medical laboratory tests were established. These RIs will be invaluable to result interpretation and the management of pregnant women.

The Role of Early Pregnancy Maternal pGCD59 Levels in Predicting Neonatal Hypoglycemia-Subanalysis of the DALI Study.

CONTEXT: Neonatal hypoglycaemia (NH) is the most common metabolic problem in infants born of mothers with gestational diabetes. Plasma glycated CD59 (pGCD59) is an emerging biomarker that has shown potential in identifying women at risk of developing gestational diabetes. The aim of this study was to assess the association between early maternal levels of pGCD59 and NH. OBJECTIVE: The aim of this study was to assess the association between early pregnancy maternal levels of plasma glycated CD59 (pGCD59) and neonatal hypoglycemia (NH). METHODS: This is an observational study of pregnant women with a prepregnancy body mass index (BMI) greater than or equal to 29 screened for eligibility to participate in the Vitamin D and Lifestyle Intervention for Gestational Diabetes (DALI) trial. This analysis included 399 pregnancies. Levels of pGCD59 were measured in fasting maternal samples taken at the time of a 75-g, 2-hour oral glucose tolerance test performed in early pregnancy (< 20 weeks). NH, the study outcome, was defined as a heel-prick capillary glucose level of less than 2.6 mmol/L within 48 hours of delivery. RESULTS: We identified 30 infants with NH. Maternal levels of pGCD59 in early pregnancy were positively associated with the prevalence of NH (one-way analysis of variance, P < .001). The odds of NH were higher in infants from mothers in tertile 3 of pGCD59 levels compared to those from mothers in tertile 1 (odds ratio [OR]: 2.41; 95% CI, 1.03-5.63). However, this was attenuated when adjusted for maternal BMI (OR: 2.28; 95% CI, 0.96-5.43). The cross-validated area under the curve (AUC) was 0.64 (95% CI, 0.54-0.74), and adjusted for maternal BMI, age, and ethnicity, the AUC was 0.70 (95% CI, 0.56-0.78). CONCLUSION: Although pGCD59 levels in early pregnancy in women with BMI greater than or equal to 29 are associated with NH, our results indicate that this biomarker by itself is only a fair predictor of NH.

The utility of first trimester plasma glycated CD59 (pGCD59) in predicting gestational diabetes mellitus: A prospective study of non-diabetic pregnant women in Ireland.

AIMS: To evaluate the ability of first trimester plasma glycated CD59 (pGCD59) to predict gestational diabetes mellitus (GDM) at 24-28 weeks of gestation. METHODS: Prospectively, in 378 pregnant women, GDM was diagnosed using the one step 2 h 75 g oral glucose tolerance test adjudicated by the World Health Organisation (WHO) 2013 criteria. The ability of pGCD59 to predict GDM was assessed using receiver operating characteristic (ROC) curves adjusted for maternal age, body mass index (BMI), maternal ethnicity, parity, previous GDM, family history of diabetes mellitus and week of gestation at time of pGCD59 sampling. RESULTS: pGCD59 generated an adjusted area under the curve (AUC) of (a) 0.63 (95 %CI:0.56-0.70, p < 0.001) for predicting GDM, and (b) 0.71 (95 %CI:0.62-0.79, p < 0.001 for GDM diagnosed with a fasting plasma glucose (FPG) ≥ 5.1 mmol/L. Sensitivity analysis of BMI subgroups showed that pGCD59 generated the highest AUC in the 35 kg/m2 ≤ BMI < 40 kg/m2 (AUC:0.85, 95 %CI:0.70-0.98) and BMI ≥ 40 kg/m2 (AUC:0.88, 95 %CI:0.63-0.99) categories. CONCLUSIONS: Early in pregnancy, pGCD59 may be a good predictor of GDM in women with a high BMI and a fair predictor of GDM diagnosed by an elevated FPG independent of BMI.

The Diagnostic Accuracy of Second Trimester Plasma Glycated CD59 (pGCD59) to Identify Women with Gestational Diabetes Mellitus Based on the 75 g OGTT Using the WHO Criteria: A Prospective Study of Non-Diabetic Pregnant Women in Ireland.

The aim of this study was to evaluate the ability of second trimester plasma glycated CD59 (pGCD59), a novel biomarker, to predict the results of the 2 h 75 g oral glucose tolerance test at 24−28 weeks of gestation, employing the 2013 World Health Organisation criteria. This was a prospective study of 378 pregnant women. The ability of pGCD59 to predict gestational diabetes (GDM) was assessed using adjusted ROC curves for maternal age, BMI, maternal ethnicity, parity, previous GDM, and family history of diabetes. The pGCD59 levels were significantly higher in women with GDM compared to women with normal glucose tolerance (p = 0.003). The pGCD59 generated an adjusted AUC for identifying GDM cases of 0.65 (95%CI: 0.58−0.71, p < 0.001). The pGCD59 predicted GDM status diagnosed by a fasting glucose value of 5.1 mmol/L with an adjusted AUC of 0.74 (95%CI: 0.65−0.81, p < 0.001). Analysis of BMI subgroups determined that pGCD59 generated the highest AUC in the 35 kg/m2 ≤ BMI < 40 kg/m2 (AUC: 0.84 95%CI: 0.69−0.98) and BMI ≥ 40 kg/m2 (AUC: 0.96 95%CI: 0.86−0.99) categories. This study found that second trimester pGCD59 is a fair predictor of GDM status diagnosed by elevated fasting glucose independent of BMI and an excellent predictor of GDM in subjects with a very high BMI.

Remote capillary blood collection for HbA1c measurement during the COVID-19 pandemic: A laboratory and patient perspective.

AIMS: The purpose of this study was to assess the clinical performance and user acceptance of capillary blood samples prepared remotely using the MiniCollect® capillary blood collection device as an alternative to blood collection by venepuncture for glycated haemoglobin (HbA1c ) analysis. METHODS: Following written informed consent, a cross-sectional study was conducted in individuals aged ≥18 years with any type of diabetes who routinely self-monitor their blood glucose. Eligible participants recruited whilst attending their routine clinical appointments were required to provide a venous blood sample, prepare a capillary blood sample at home (remotely) and complete a bespoke questionnaire. HbA1c in whole blood collected in ethylenediaminetetraacetic acid was determined by capillary electrophoresis on the Sebia Capillary's 3 Tera analyser following standard operating procedure. RESULTS: HbA1c results from both venous and capillary collection demonstrated good agreement. Passing-Bablok regression: y = 0 + 1x (p = 0.18), Spearman correlation r = 0.986, p < 0.0001. The Bland-Altman difference plot provided a mean difference of 0.3 mmol/mol (2.2%). Over half of the participants found the MiniCollect device easy to use. The majority of participants were in favour of the remote capillary blood collection service and would use it if routinely available. CONCLUSION: The home collection of capillary blood for HbA1c determination is a valuable and convenient alternative to standard venous blood collection as it provides an opportunity to support routine HbA1c monitoring, whilst mitigating the transmission of SARS-CoV-2. This service would additionally allow individuals to attend clinic visits with a HbA1c value, ensuring optimal continuance of patient care for individuals with diabetes.

Title: Understanding a Low Vitamin D State in the Context of COVID-19.

While a low vitamin D state has been associated with an increased risk of infection by SARS-CoV-2 in addition to an increased severity of COVID-19 disease, a causal role is not yet established. Here, we review the evidence relating to i) vitamin D and its role in SARS-CoV-2 infection and COVID-19 disease ii) the vitamin D status in the Irish adult population iii) the use of supplemental vitamin D to treat a deficient status and iv) the application of the Bradford-Hill causation criteria. We conclude that reverse causality probably makes a minimal contribution to the presence of low vitamin D states in the setting of COVID-19. Applying the Bradford-Hill criteria, however, the collective literature supports a causal association between low vitamin D status, SARS-CoV-2 infection, and severe COVID-19 (respiratory failure, requirement for ventilation and mortality). A biologically plausible rationale exists for these findings, given vitamin D's role in immune regulation. The thresholds which define low, deficient, and replete vitamin D states vary according to the disease studied, underscoring the complexities for determining the goals for supplementation. All are currently unknown in the setting of COVID-19. The design of vitamin D randomised controlled trials is notoriously problematic and these trials commonly fail for a number of behavioural and methodological reasons. In Ireland, as in most other countries, low vitamin D status is common in older adults, adults in institutions, and with obesity, dark skin, low UVB exposure, diabetes and low socio-economic status. Physiological vitamin D levels for optimal immune function are considerably higher than those that can be achieved from food and sunlight exposure alone in Ireland. A window exists in which a significant number of adults could benefit from vitamin D supplementation, not least because of recent data demonstrating an association between vitamin D status and COVID-19. During the COVID pandemic, we believe that supplementation with 20-25ug (800-1000 IU)/day or more may be required for adults with apparently normal immune systems to improve immunity against SARS-CoV-2. We expect that higher monitored doses of 37.5-50 ug (1,500-2,000)/day may be needed for vulnerable groups (e.g., those with obesity, darker skin, diabetes mellitus and older adults). Such doses are within the safe daily intakes cited by international advisory agencies.

Reference intervals for commonly requested biochemical and haematological parameters in a healthy Irish adult Caucasian population.

INTRODUCTION: In laboratory medicine, reference intervals (RIs) are key decision support tools used to guide the clinical interpretation of numerical test results. Best practice suggests each laboratory establishes RIs in the local population prior to introducing an assay into routine clinical practice. AIM: The aim of this study was to define RIs for frequently requested biochemical/haematological parameters in a healthy adult Irish Caucasian population. METHODS: A cross-sectional study of non-pregnant apparently healthy volunteers was conducted. Baseline demographics, anthropometric and laboratory measurements were recorded. In total, 37 commonly requested biochemical (serum, n = 26) and haematological (venous blood, n = 11) ISO15189:2012 accredited tests were analysed, using the Roche Cobas® Sebia Capillarys 3 Tera and Siemens Advia® 2120i platforms following standard operating procedures. RIs were defined according to the International Federation of Clinical Chemistry (IFCC) recommended method. RESULTS: Of 208 apparently healthy volunteers, 76 failed to meet the study inclusion criteria. The reference population comprised of 132 participants (males: n = 65, 49.2%) with a median age of 29.7 (18.1-62.2) years. RIs for the majority of biochemical/haematological parameters were broadly in accord with those provided by Pathology Harmony (UK)/Irish RI Harmonisation Project and the manufacturer Roche Diagnostics. However, the established RI defined for HbA1c: 27-37 mmol/mol was markedly different from that quoted nationally, HbA1c: 20-42 mmol/mol. CONCLUSION: Normative biological intervals established in a healthy adult Irish population for 37 commonly requested biochemical/haematological parameters will be a valuable aid to result interpretation in clinical laboratories after appropriate verification in accordance with ISO 15189: 2012.

Patient-reported outcomes (PROs) in randomised controlled trials in diabetes and pregnancy: protocol for a systematic review.

INTRODUCTION: Diabetes mellitus is the most common metabolic complication of pregnancy and its prevalence worldwide is rising. The number of randomised controlled trials (RCTs) being conducted in people with diabetes is also increasing. Many studies preferentially publish findings on clinical endpoints and do not report patient-reported outcomes (PROs). In studies that do include PROs, PRO reporting is often of poor quality. METHODS: We will conduct this systematic review following the Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines. Using a combination of medical subject headings and keywords (combined using Boolean operators), we will search web-based databases (PubMed, Cochrane and EMBASE) for RCTs published in English between 2013 and 2021. Two reviewers will review titles and abstracts. We will review the full texts of any relevant abstracts and extract the following data: date of publication or recruitment period, journal of publication, country of study, multicentre or single centre, population and number of participants, type of intervention, frequency of PRO assessment and type of PRO (or PRO measurement) used. We will also record if the PRO was a primary, secondary or exploratory outcome. We will exclude reviews, observational studies, unpublished data for example, conference abstracts and trial protocols. Any published RCT that includes data on a PRO as a primary or secondary outcome will then be compared against the Consolidated Standards of Reporting Trials-Patient-Reported Outcome extension checklist, a structured and approved framework for the publication of results of PROs. ETHICS AND DISSEMINATION: Ethical approval to conduct this study was obtained from the ethics committee at Galway University Hospitals on 24 March 2021 (CA 2592). We aim to publish our findings in a peer-reviewed journal and present our findings at national and international conferences. SYSTEMATIC REVIEW REGISTRATION: This systematic review was registered prospectively with the International Prospective Register of Systematic Reviews (PROSPERO). Registration number CRD42021234917.

Making weight: acute muscle weakness and hypokalaemia exacerbated by thyrotoxicosis factitia in a bodybuilder.

SUMMARY: A 32-year-old Caucasian male presented to the emergency department with a one-day history of acute severe bilateral lower limb weakness, three days after competing in a bodybuilding competition. He consumed large quantities of carbohydrate-rich foods following the competition. His past medical history was significant for anxiety, and family history was non-contributory. Examination was normal except for reduced power and hyporeflexia in both legs, despite his muscular physique. He was noted to have severe hypokalaemia (K+= 1.9 mmol/L). His thyroid function tests were consistent with thyrotoxicosis. He reported taking thyroxine and several other agents to facilitate muscle mass generation before the bodybuilding competition. His presentation was reminiscent of thyrotoxic periodic paralysis, albeit uncommon with Caucasian ethnicity. He also had transient hyperglycaemia at presentation with concomitant hyperinsulinaemia, which could be attributed to the carbohydrate load and may have exacerbated his hypokalaemia through a transcellular shift. Urine toxicology screen subsequently ruled out the use of diuretics but confirmed the presence of a long-acting beta agonist (clenbuterol) which, along with other substances, may have aggravated the hypokalaemia further. After 12 h of i.v. replacement, the potassium level normalised and leg weakness resolved. The patient agreed to stop taking thyroxine and beta agonists and was well during the clinic visit at one month follow-up. This case highlights the potential for thyrotoxicosis factitia to exacerbate hypokalaemia and muscle weakness from other causes in bodybuilders presenting with acute severe weakness, irrespective of ethnicity. LEARNING POINTS: In patients presenting with muscle weakness and hypokalaemia, early consideration of thyrotoxicosis is essential, even in the absence of a past history of thyroid disease or specific symptoms of thyrotoxicosis, in order to allow prompt initiation of appropriate treatment and to prevent recurrence. Bodybuilders may constitute a uniquely 'at-risk' group for thyrotoxic periodic paralysis secondary to thyrotoxicosis factitia, especially where there is concomitant use of beta-adrenergic agonists, even in the absence of diuretic use. Although rare and usually described in patients of Asian or Polynesian ethnicity, this case highlights that thyrotoxic periodic paralysis secondary to thyrotoxicosis factitia can also occur in patients with Caucasian ethnicity. We speculate that consuming large quantities of carbohydrates may induce hyperinsulinaemia, which could theoretically contribute to worse hypokalaemia, though mechanistic studies would be needed to explore this further.

A case of adrenal Cushing's syndrome and primary hyperparathyroidism due to an atypical parathyroid adenoma.

Cushing's syndrome is a rare disorder of cortisol excess and is associated with significant morbidity and mortality. Hypercalcaemia due to hyperparathyroidism is a common condition; however, in 10% of young patients, it is associated with other endocrinopathies and occurs due to a genetic variant [e.g. multiple endocrine neoplasia (MEN) type 1 (MEN1), MEN2 or MEN4]. We report the case of a 31-year-old woman who was referred to the endocrinology out-patient service with an 8-month history of hirsutism, amenorrhoea and weight gain. Her biochemical work up was significant for adrenocorticotropic hormone (ACTH)-independent Cushing's syndrome. Radiological investigations revealed an adrenal adenoma. During investigation she was also found to have primary hyperparathyroidism due to a parathyroid adenoma. Pre-operatively, the patient was commenced on metyrapone and both her adrenal and parathyroid lesions were resected successfully. There were several concerning findings on initial examination of the parathyroid tumour, including possible extension of the tumour through the capsule and vascular invasion; however, following extensive review, it was ultimately defined as an adenoma. Given the unusual presence of two endocrinopathies in a young patient, she subsequently underwent genetic testing. Analysis of multiple genes did not reveal any pathogenic variants. The patient is currently clinically well, with a normal adjusted calcium and no clinical features of cortisol excess. She will require long-term follow up for recurrence of both hypercalcaemia and hypercortisolaemia.