A Comparison of the Performance of SARS-CoV-2 Antibody Assays in Healthcare Workers with COVID-19.

Aims Since its emergence, significant interest surrounds the use of SARS-CoV-2 serological tests as an alternative or as an adjunct to molecular testing. However, given the speed of this pandemic, paralleled with the pressure to develop and provide serological tests in an expediated manner, not every assay has undergone the rigorous evaluation that is usually associated with medical diagnostic assays. We aimed to examine the performance of several commercially available SARS-CoV-2 IgG antibody assays among participants with confirmed COVID-19 disease and negative controls. Methods Serum taken between day 17 and day 40 post onset of symptoms from 41 healthcare workers with RT-PCR confirmed COVID-19 disease, and pre-pandemic serum from 20 negative controls, were tested for the presence of SARS-CoV-2 IgG using 7 different assays including point-of-care (POC) and laboratory-based assays. Results Assay performance varied. The lab-based Abbott diagnostics SARS-CoV-2 IgG assay proved to be the assay with the best positive and negative predictive value, and overall accuracy. The POC Nal von Minden GmbH and Biozek assays also performed well. Conclusion Our research demonstrates the variations in performance of several commercially available SARS-CoV-2 antibody assays. These findings identify the limitations of some serological tests for SARS-CoV-2. This information will help inform test selection and may have particular relevance to providers operating beyond accredited laboratories.

The influence of dietary supplementation with cranberry tablets on the urinary risk factors for nephrolithiasis.

PURPOSE: Cranberry supplements are commonly used as a natural deterrent to urinary tract infection. However, one small study (n = 5) which showed an increase in urinary oxalate levels following cranberry supplementation has led to its use with caution among patients susceptible to nephrolithiasis. Furthermore, most commonly available cranberry tablet preparations contain vitamin C, which has been independently shown to increase urinary oxalate excretion. The aim of this study is to investigate the influence of cranberry supplementation on urinary oxalate excretion. METHODS: Fifteen participants were randomised to receive cranberry tablets alone or cranberry tablets containing vitamin C. Tablets were taken at the manufacturers recommended dosage for a period of 14 days. Participants provided a 24 h urine collection at trial entry and day 14. Urinary variables were compared to assess for changes in oxalate levels. RESULTS: The median age was 27 years (21-43). There was no difference in the 24 h urine volume pre or post commencement of cranberry tablets (1.7 vs 2 L, p = 0.07). An increase in median urinary oxalate excretion was observed in participants taking both cranberry-only tablets (0.10 mmol/day) and tablets containing vitamin C (1.15 mmol/day). CONCLUSION: Dietary supplementation with cranberry increases urinary oxalate excretion and should be avoided in patients at risk of urolithiasis.

The benefits of utilising geo-mapping for visualising the vitamin D status of Dublin city and the surrounding urban districts.

CONTEXT: There have been few published reports of visualising vitamin D status at a micro level, i.e., within large individual urban centres of countries. OBJECTIVE: To produce a visual map of the vitamin D status [25-hydroxy vitamin D-25(OH)D] of a large urban centre (n > 350,000) incorporating the regions of Dublin city that constitute the general practitioner catchment area of a large academic teaching adult hospital. DESIGN, SETTING AND PARTICIPANTS: An observational investigation of 5287 free living Irish adults (>18 years). RESULTS: Approximately, 15.2 % of those sampled in the winter period (October-February) were vitamin D deficient (<30 nmol/L) compared with 10.8 % of those sampled in the summer period (March-September). Vitamin D tests requested from the most social economically deprived urban locations (Dublin 8 and Lucan postal districts) had the highest rates of deficiency (23.5 and 20.4 %, respectively, both seasons). On average, females had a significantly higher 25(OH)D concentration compared with males (57.9 vs 52.3 nmol/L, respectively), while the younger participants (18-50 years) mean 25(OH)D concentration was 27 % lower in winter and 20.7 % lower in summer in comparison with the older participants (>50 years) (P < 0.0001). CONCLUSIONS: For the first time in Ireland, a visual depiction of data can be used to aid in the rapid identification of vitamin D status trends within a major urban area. These findings provide useful data to help inform public health policy regarding endemic vitamin D insufficiency to help target the population groups and resident location areas most at risk.

Stability and accuracy of total and free PSA values in samples stored at room temperature.

INTRODUCTION: In 2010, an estimated 476,076 total PSA tests were performed in Ireland, at a cost of €3.6 million with the majority ordered by general practitioners. We aimed to replicate storage conditions at room temperature and see if prolonged storage affected total and free PSA values. METHODS: Blood samples were taken from 20 male patients in four VACUETTE® Serum Separator tubes (Greiner-Bio-One, Austria) and stored at room temperature (22 °C) for different time intervals (4, 8, 24, 48 h) before being centrifuged and analyzed. Total PSA (tPSA) and free PSA (fPSA) values were determined using the Tosoh AIA 1800 assay (Tokyo, Japan). RESULTS: Mean tPSA values were measured at 4, 8, 24 and 48 h with values of 7.9, 8.1, 7.8 and 8.0 µg/L, respectively. Values ranged from -1.26 to +2.53 % compared to the initial 4 h interval reading, indicating tPSA remained consistent at room temperature. The tPSA showed no significance between groups (ANOVA, p = 0.283). Mean fPSA values at 4, 8, 24 and 48 h were 2.05, 2.04, 1.83, 1.82 µg/L, respectively. At 24 and 48 h there was 10.73 and 11.22 % reduction, respectively, in fPSA compared to the 4-h time interval, indicating prolonged storage resulted in reduced fPSA values. After 24 h, there was an 8.8 % reduction in the free/total PSA %. The fPSA showed significant differences between groups (ANOVA, p = 0.024). CONCLUSIONS: Our recommendation is that samples that have been stored for prolonged amounts of time (greater than 24 h) should not be used for free PSA testing.

Metabolic syndrome, central obesity and insulin resistance are associated with adverse pathological features in postmenopausal breast cancer.

AIMS: Obesity is associated with both an increased risk of postmenopausal breast cancer and increased mortality rates. The mechanism is unclear, and central (visceral) obesity, insulin resistance, altered sex steroids and altered adipokines are mooted as possible factors. These features may cluster in the so-called metabolic syndrome. The relevance of metabolic syndrome to the biology of breast cancer is unknown, and this was the focus of the present study. MATERIALS AND METHODS: All postmenopausal women with newly diagnosed breast cancer (n=105) were recruited. A detailed clinical history was carried out, as well as a body composition analysis, metabolic screen and measurement of adipokines and inflammatory markers. RESULTS: The median age was 68 years (40-94 years) and the mean body mass index was 28.3+/-5.2 kg/m2, with 87% of patients centrally obese. Metabolic syndrome was diagnosed in 39% of patients, and was significantly associated with central obesity (P<0.005) and increased inflammation, with C-reactive protein levels doubling in metabolic syndrome patients compared with non-metabolic syndrome patients (10.3 vs 5.8 mg/l; P=0.084). Patients with a later pathological stage (II-IV) were significantly more likely to be obese (P=0.007), centrally obese (P=0.009), hyperglycaemic (P=0.047) and hyperinsulinaemic (P=0.026); 51% had metabolic syndrome compared with 12% for early stage disease. Patients with node-positive disease were significantly more likely to be hyperinsulaemic (P=0.030) and have metabolic syndrome (P=0.028) than patients with node-negative disease. DISCUSSION: The data suggest that metabolic syndrome and central obesity are common in postmenopausal breast cancer patients, and that metabolic syndrome may be associated with a more aggressive tumour biology.

The Leu262Val polymorphism of presenilin associated rhomboid like protein (PARL) is associated with earlier onset of type 2 diabetes and increased urinary microalbumin creatinine ratio in an Irish case-control population.

AIMS: Environmental and genetic factors contribute to the evolution of type 2 diabetes (T2DM). Presenilin associated rhomboid like protein (PARL) is a mitochondrial protein that has been implicated in T2DM in both the rodent Psammomys obesus and in humans. The SNP variant (Leu262Val) in PARL has been shown to be associated with hyperinsulinaemia in an age-dependent manner in a US non-diabetic, cohort. However, this finding has not been replicated in UK cohorts. We studied Leu262Val associations in an Irish Caucasian T2DM case-control population. METHODS: An RFLP-PCR assay using BstN I was used to assess Leu262Val genotype in a total of 613 subjects, 421 with T2DM and 192 controls. RESULTS: In the control group genotype frequencies were as follows 27.37% (GG), 51.58% (CG) and 21.05% (CC), while in the group with T2DM 30.64% (GG), 47.74% (CG) and 21.62% (CC). We observed no association between Leu262Val variant and T2DM nor was there an association with plasma insulin concentrations or BMI. There was no interaction between age and fasting plasma insulin concentration. However, in the group with T2DM the C allele was associated with higher urinary albumin to creatinine ratio while the GG genotype was associated with an earlier age of onset of T2DM. CONCLUSION: The Leu262Val polymorphism of PARL is not associated with markers of insulin resistance. However, in subjects with T2DM, genetic variation at this locus may indicate earlier onset of T2DM and increased susceptibility to nephropathy and cardiovascular complications.

Physiological effects of sleeping with the head of the bed elevated 18 in. in young healthy volunteers.

BACKGROUND: Sleeping with the head of bed tilted upwards (SHU) is recommended as a treatment of orthostatic hypotension though the supporting evidence is weak. AIM: To investigate the physiological effects of SHU amongst a group of young healthy volunteers. METHODS: Twenty-nine volunteers, mean age 22 years, underwent 1-week of SHU at 18-in. elevation. Before and after hemodynamic and non-haemodynamic parameters were recorded. RESULTS: After SHU, there were reductions in the systolic blood pressure drop on standing, upright total peripheral resistance, haemoglobin, nocturnal urinary volume, orthostatic dizziness and increases in weight, standing cardiac output and ankle circumference. There were no differences in heart rate, stroke volume, renin, aldosterone, pro-atrial natriuretic peptide or 24-h blood pressure. CONCLUSIONS: In these healthy subjects, SHU for 1 week had a nocturnal antidiuretic effect with both intra- and extra-vascular accumulation of fluid and was associated with reduced postural drop in SBP and improved orthostatic tolerance.

Adverse metabolic profiles in a cohort of obese Irish children.

BACKGROUND: The metabolic characteristics of obese Irish children are not well defined. We prospectively examined the relationship between the degree of obesity and glucose metabolism, insulin sensitivity and suspected non-alcoholic steatohepatosis (NASH) in a pilot study of obese Irish children. METHODS: We measured height, weight, body mass index (BMI), blood pressure, waist and hip circumference in 18 participants (mean age 15.5 years). Fasting blood glucose, insulin, lipid profile and alanine aminotransferase (ALT) concentrations were also measured. A standard 75 g oral glucose tolerance test was performed and insulin sensitivity was derived from this using a mathematical model--oral glucose insulin sensitivity. RESULTS: There were significant associations between the degree of obesity, insulin sensitivity and markers of liver steatosis. For example, when adjusted for pubertal status, there were significant associations between standardized BMI and insulin sensitivity (regression coefficient, beta = -70.1, P = 0.018) and ALT (beta = 20.7, P = 0.007). CONCLUSION: This study suggests that the degree of obesity is associated with lower insulin sensitivity and possible NASH in obese Irish children.

Meta-analysis of the Gly482Ser variant in PPARGC1A in type 2 diabetes and related phenotypes.

AIMS/HYPOTHESIS: Peroxisome proliferator-activated receptor-gamma co-activator-1alpha (PPARGC1A) is a transcriptional co-activator with a central role in energy expenditure and glucose metabolism. Several studies have suggested that the common PPARGC1A polymorphism Gly482Ser may be associated with risk of type 2 diabetes, with conflicting results. To clarify the role of Gly482Ser in type 2 diabetes and related human metabolic phenotypes we genotyped this polymorphism in a case-control study and performed a meta-analysis of relevant published data. MATERIALS AND METHODS: Gly482Ser was genotyped in a type 2 diabetes case-control study (N=1,096) using MassArray technology. A literature search revealed publications that examined Gly482Ser for association with type 2 diabetes and related metabolic phenotypes. Meta-analysis of the current study and relevant published data was undertaken. RESULTS: In the pooled meta-analysis, including data from this study and seven published reports (3,718 cases, 4,818 controls), there was evidence of between-study heterogeneity (p<0.1). In the fixed-effects meta-analysis, the pooled odds ratio for risk of type 2 diabetes per Ser482 allele was 1.07 (95% CI 1.00-1.15, p=0.044). Elimination of one of the studies from the meta-analysis gave a summary odds ratio of 1.11 (95% CI 1.04-1.20, p=0.004), with no between-study heterogeneity (p=0.475). For quantitative metabolic traits in normoglycaemic subjects, we also found significant between-study heterogeneity. However, no significant association was observed between Gly482Ser and BMI, fasting glucose or fasting insulin. CONCLUSIONS/INTERPRETATION: This meta-analysis of data from the current and published studies supports a modest role for the Gly482Ser PPARGC1A variant in type 2 diabetes risk.

Expression of the thermogenic nuclear hormone receptor coactivator PGC-1alpha is reduced in the adipose tissue of morbidly obese subjects.

Peroxisome proliferator-activated receptor gamma coactivator 1alpha (PGC1alpha) is an accessory protein which can potentiate the transcriptional activation function of many nuclear hormone receptors. Its tissue distribution and physiological studies suggest that its principal in vivo roles are to promote cold-induced thermogenesis, mitochondrial biogenesis, hepatic gluconeogenesis, and fatty acid beta-oxidation. It is expressed in the white adipose tissue of both humans and rodents, and in rodents it has been suggested to mediate in part the leptin-induced conversion of white adipocytes from fat storing to fat oxidising cells. In this study, quantitative real-time PCR has been used in human tissue to demonstrate that (1) PGC1alpha mRNA levels in subcutaneous fat are three-fold lower in morbidly obese than in slim subjects; (2) there are no differences in PGC1alpha mRNA between omental and subcutaneous mature adipocytes; (3) there is a robust induction of PGC1alpha expression during subcutaneous human preadipocyte differentiation ex vivo. Whether low PGC1alpha expression is a prelude to the development of obesity, or a consequence of that obesity, attempts to upregulate endogenous white adipose tissue expression may prove a valuable new avenue to explore in obesity therapy.

Mitochondrial uncoupling proteins (UCPs) and obesity.

Obesity is now regarded as major public health problem worldwide. Research into this condition has been increasingly focussed on elucidating the cellular and molecular mechanisms regulating mammalian energy intake and expenditure. It is widely acknowledged that the brown adipose tissue (BAT) mitochondrial uncoupling protein (UCP1) plays a pivotal role in adaptive thermogenic responses. Two homologues of UCP1 (UCP2 and UCP3) have recently been identified and population-based genetic studies have linked them with basal metabolic rate, while in vitro studies report that both have proton transport activity and may thus be involved in regulation of energy homeostasis and hence obesity. However, evidence from genetically modified animal models indicates that UCP2 and UCP3 have no specific physiological thermogenic function in vivo, though they may still be useful therapeutic targets for obesity. Furthermore, their role in modulating levels of reactive oxygen species and glucose homeostasis is also being investigated.

Depot-related and thiazolidinedione-responsive expression of uncoupling protein 2 (UCP2) in human adipocytes.

OBJECTIVES: Uncoupling protein 2 (UCP2) is a recently described homologue of the uncoupling protein of brown adipocytes (UCP1), which is expressed at high levels in human white adipose tissue. Studies were undertaken (1) to establish whether the expression of UCP2 mRNA varies in a depot-related manner in isolated human adipocytes, (2) to determine whether thiazolidinedione exposure influences the expression of UCP2 mRNA in cultured human pre-adipocytes, and (3) to determine whether human UCP2 is targeted to mitochondria when transfected into mammalian cells. SUBJECTS: Abdominal subcutaneous and omental adipose tissue biopsies were obtained from adult patients undergoing elective intra-abdominal surgical procedures. MEASUREMENTS: A competitive reverse transcriptase-polymerase chain reaction (RT-PCR) was used to quantify UCP2 mRNA expression in human omental and subcutaneous adipocytes, and in cultured human preadipocytes differentiated in vitro using the thiazolidinedione, BRL49653. Chinese hamster ovary cells were transfected with a vector expressing human UCP2, and its cellular localization was determined by confocal immunofluorescence microscopy. RESULTS: Adipocytes isolated from human omentum consistently expressed more UCP2 mRNA than did subcutaneous adipocytes from the same subjects (mean fold difference 2.92+/-0.44 P<0.001, n=11) with no effect of gender or body mass index being seen. BRL49653 treatment of subcutaneously, but not omentally, derived preadipocytes stimulated expression of UCP2 mRNA (5.1+/-1.1 fold). Transfected human UCP2 was detected exclusively in mitochondria of CHO cells. CONCLUSIONS: Increased expression of UCP2 in human omental adipose tissue relative to subcutaneous adipose tissue is related to the expression levels in adipocytes per se, a finding which may relate to the particular functional attributes of this sub-population of adipocytes. Furthermore, BRL 49653 has site-specific effects of on the expression of UCP2 in human preadipocytes, a finding which may be relevant to the therapeutic effects of such compounds. Finally we present evidence for the mitochondrial localisation of human UCP2.

Bone loss in long-term renal transplantation: histopathology and densitometry analysis.

BACKGROUND: There is little information of the spectrum and factors implicated in the bone loss in long-term renal transplantation, and virtually no data using both histomorphometric and densitometric analysis. METHODS: Twenty-three males and 22 females (13 postmenopausal) were studied with a bone biopsy and densitometry. Sixteen patients were on cyclosporine A monotherapy, 20 on azathioprine + prednisolone, and 9 on cyclosporine A + prednisolone or triple therapy. The mean time after transplantation was 127 +/- 70 months. RESULTS: No group had a significant decrease in bone mineral density (BMD) of the axial skeleton compared with an age- and sex-matched normal population. Compared with sex-matched young controls, osteopenia was observed in all groups at the femoral neck (except premenopausal women and triple therapy) and in the triple-therapy group at the L1-L4 spine region. At the distal radius, osteopenia was found in all the groups. Histopathological diagnosis was mixed uremic osteodystrophy in 46.5%, adynamic bone in 23.2%, hyperparathyroid disease in 13.9%, and normal bone in 16.3%. The diagnosis was not different according to immunosuppressive therapy, but men tended to show more mixed uremic bone disease. There was no significant difference in BMD between histopathological subtypes. In general, patients showed slight osteoclast function increase, osteoblast function decrease, and marked retardation of dynamic parameters. The cyclosporine A monotherapy group had a significantly lower appositional rate than azathioprine + prednisolone. Men had a significantly lower bone volume than women, and premenopausal women had a significantly lower mineralizing surface than postmenopausal women and men. In the multivariate analysis, male gender, time after transplantation, old age, and time on dialysis prior to transplantation were significant predictive factors for a negative effect on bone mass. CONCLUSIONS: Long-term renal transplant-patients showed reduced BMD in both trabecular and cortical bone. This reduction in BMD was not as severe as in short-term reports and was associated with osteoclast stimulation, osteoblast suppression, and retardation of mineral apposition and bone formation rates. Bone mass loss was not different between the immunosuppression therapy groups. Male gender and age were the strongest predictive factors for low bone mass.

Indications for cholesterol-lowering medication: comparison of risk-assessment methods.

BACKGROUND: Recommendations for the prescription of lipid-lowering drugs emphasise the importance of an assessment of absolute coronary heart disease (CHD) risk based on all risk factors, rather than simply the serum cholesterol concentration. If, however, the methods recommended for risk assessment are inaccurate, recommended prescribing will not occur. We compared several sets of guidelines for such treatment in a series of patients referred to a lipid clinic, to assess the difference in degree of risk of CHD at which lipid-lowering medication is recommended by each set of guidelines. METHODS: For a series of 570 patients (50% men) without pre-existing clinical evidence of atherosclerosis referred to a lipid clinic, we compared the algorithms, charts, and tables used by the US National Cholesterol Education Program (NCEP), the joint guidelines of the European Society of Cardiology, the European Atherosclerosis Society, and the European Society of Hypertension, and the report of the UK Standing Medical Advisory Committee with the Framingham risk equation programmed into a computer. FINDINGS: In 386 patients for whom the NCEP and UK guidelines could be compared, 62% of the men and 72% of the women met NCEP criteria for lipid-lowering medication, whereas only 9% of the men and less than 1% of the women met the UK criteria. The Framingham equation estimated a CHD risk of more than 3% per year in 22% of the men and 7% of the women, which shows that the UK tables underestimated CHD risk. European guidelines could be applied to only 261 patients, and were reasonably accurate in assessment of a CHD risk of 2% per year. INTERPRETATION: Guidelines for the use of statin treatment in patients with CHD differ in their assessment of CHD risk. The method of risk assessment recommended in future guidelines for CHD prevention should be critically tested in relevant groups of patients.

Dominant negative mutations in human PPARgamma associated with severe insulin resistance, diabetes mellitus and hypertension.

Thiazolidinediones are a new class of antidiabetic agent that improve insulin sensitivity and reduce plasma glucose and blood pressure in subjects with type 2 diabetes. Although these agents can bind and activate an orphan nuclear receptor, peroxisome proliferator-activated receptor gamma (PPARgamma), there is no direct evidence to conclusively implicate this receptor in the regulation of mammalian glucose homeostasis. Here we report two different heterozygous mutations in the ligand-binding domain of PPARgamma in three subjects with severe insulin resistance. In the PPARgamma crystal structure, the mutations destabilize helix 12 which mediates transactivation. Consistent with this, both receptor mutants are markedly transcriptionally impaired and, moreover, are able to inhibit the action of coexpressed wild-type PPARgamma in a dominant negative manner. In addition to insulin resistance, all three subjects developed type 2 diabetes mellitus and hypertension at an unusually early age. Our findings represent the first germline loss-of-function mutations in PPARgamma and provide compelling genetic evidence that this receptor is important in the control of insulin sensitivity, glucose homeostasis and blood pressure in man.