Generation of Rapid and High-Quality Serum by Recombinant Prothrombin Activator Ecarin (RAPClot™).

We recently reported the potential application of recombinant prothrombin activator ecarin (RAPClot™) in blood diagnostics. In a new study, we describe RAPClot™ as an additive to develop a novel blood collection prototype tube that produces the highest quality serum for accurate biochemical analyte determination. The drying process of the RAPClot™ tube generated minimal effect on the enzymatic activity of the prothrombin activator. According to the bioassays of thrombin activity and plasma clotting, γ-radiation (>25 kGy) resulted in a 30-40% loss of the enzymatic activity of the RAPClot™ tubes. However, a visual blood clotting assay revealed that the γ-radiation-sterilized RAPClot™ tubes showed a high capacity for clotting high-dose heparinized blood (8 U/mL) within 5 min. This was confirmed using Thrombelastography (TEG), indicating full clotting efficiency under anticoagulant conditions. The storage of the RAPClot™ tubes at room temperature (RT) for greater than 12 months resulted in the retention of efficient and effective clotting activity for heparinized blood in 342 s. Furthermore, the enzymatic activity of the RAPClot™ tubes sterilized with an electron-beam (EB) was significantly greater than that with γ-radiation. The EB-sterilized RAPClot™ tubes stored at RT for 251 days retained over 70% enzyme activity and clotted the heparinized blood in 340 s after 682 days. Preliminary clinical studies revealed in the two trials that 5 common analytes (K, Glu, lactate dehydrogenase (LD), Fe, and Phos) or 33 analytes determined in the second study in the γ-sterilized RAPClot™ tubes were similar to those in commercial tubes. In conclusion, the findings indicate that the novel RAPClot™ blood collection prototype tube has a significant advantage over current serum or lithium heparin plasma tubes for routine use in measuring biochemical analytes, confirming a promising application of RAPClot™ in clinical medicine.

Oral glucose tolerance test to diagnose gestational diabetes mellitus: Impact of variations in specimen handling.

To improve birth outcomes, all pregnant women without known diabetes are recommended for an oral glucose tolerance test (OGTT) to screen for hyperglycaemia in pregnancy (diabetes in pregnancy or gestational diabetes mellitus (GDM)). This narrative review presents contemporary approaches to minimise preanalytical glycolysis in OGTT samples with a focus on GDM diagnosis using criteria derived from the Hyperglycemia and Adverse Pregnancy Outcomes (HAPO) study. The challenges of implementing each approach across a diverse Australian healthcare setting were explored. Many Australian sites currently collect and transport OGTT samples at ambient temperature in sodium fluoride (NaF) tubes which is likely to lead to missed diagnosis of GDM in a significant proportion of cases. Alternative preanalytical solutions should be pragmatic and tailored to individual settings and as close as possible to the preanalytical conditions of the HAPO study for correct interpretation of OGTT results. Rapid centrifugation of barrier tubes to separate plasma could be suitable in urban settings provided time to centrifugation is strictly controlled. Tubes containing NaF and citrate could be useful for remote or resource poor settings with long delays to analysis but the impact on the interpretation of OGTT results should be carefully considered. Testing venous blood glucose at the point-of-care bypasses the need for glycolytic inhibition but requires careful selection of devices with robust analytical performance. Studies to evaluate the potential error of each solution compared to the HAPO protocol are required to assess the magnitude of misdiagnosis and inform clinicians regarding the potential impact on patient safety and healthcare costs.

Potential Application of Recombinant Snake Prothrombin Activator Ecarin in Blood Diagnostics.

We describe here the purification and cloning of a codon-optimized form of the snake prothrombin activator ecarin from the saw scaled viper (Echis carinatus) expressed in mammalian cells. Expression of recombinant ecarin (rEcarin) was carried out in human embryonic kidney cells (HEK) cells under conditions for the development and performance of a novel and scalable recombinant snake ecarin to industry standards. Clotting performance of the rEcarin was established in recalcified citrated whole blood, plasma, and fresh whole blood and found to be comparable to native ecarin (N-Ecarin). Furthermore, hemolysis was observed with N-Ecarin at relatively high doses in both recalcified citrated and fresh whole blood, while clotting was not observed with rEcarin, providing an important advantage for the recombinant form. In addition, rEcarin effectively clotted both recalcified citrated whole blood and fresh whole blood containing different anticoagulants including heparin, warfarin, dabigatran, Fondaparinux, rivaroxaban and apixaban, forming firm clots in the blood collection tubes. These results demonstrate that rEcarin efficiently clots normal blood as well as blood spiked with high concentrations of anticoagulants and has great potential as an additive to blood collection tubes to produce high quality serum for analyte analysis in diagnostic medicine.

Pre-dialysis levels of inflammation and endotoxin in people receiving hemodialysis are not associated with blood pressure variability.

There are multiple risk factors for inflammation in dialysis. One potential cause is the presence of circulating levels of Gram-negative bacteria-derived endotoxin which is a strong inducer of inflammation. Gut-associated endotoxin may enter the circulation via a defective blood-gut barrier during episodes of hypotension or reduced perfusion. MATERIALS AND METHODS: In this study, 165 patients receiving outpatient-based hemodialysis in a facility (FHD) or at home (HHD), were studied. Levels of inflammation were quantified by developing an inflammatory score derived from the measurement of pro-inflammatory cytokines, high-sensitivity C-reactive protein (hsCRP), tumor necrosis factor alpha (TNF-α) and interleukin 6 (IL-6). Intradialytic blood pressure (BP) variability and hypotension events were recorded. This included the final session of dialysis, at the commencement of which the blood samples were drawn, as well as the five preceding sessions. RESULTS: The median inflammatory score was 2 (range 0 - 3), and 30% of patients had an inflammatory score of three suggesting significant levels of inflammation. Only 8.5% had an inflammatory score of 0. Endotoxin was measured in all participants and was only positive in N = 3. The mean systolic blood pressure (SBP) was 134 ± 20 mmHg and the BP variability was 11.7 ± 3.5. In a multivariable ordinal regression model, a higher inflammatory score was significantly associated with younger age (OR 0.95, 95% CI 0.95 - 0.99, p = 0.03), higher ultrafiltration volume (OR 1.62, CI 1.04 - 2.54, p = 0.03) and lower body mass index (OR 0.9, CI 0.86 - 0.96, p = 0.01). There was no association between inflammatory score and dialysis modality, access type, kidney replacement therapy (KRT), BP variability, or endotoxin. Endotoxin was detected in only 3 of 165 patients and was not associated with inflammation. CONCLUSION: Pre-dialysis levels of inflammation are prevalent in the hemodialysis population after the long break but are not related to intradialytic BP variability or hypotension in the preceding 2 weeks. However, endotoxemia is uncommon and unlikely to be a significant driver of inflammation.

Evaluation of the Atellica TnIH cardiac troponin I assay and assessment of biological equivalence.

OBJECTIVES: We evaluated the analytical performance characteristics and the biological equivalence of the Atellica TnIH assay. METHODS: Precision, detection capability, linearity, and sex specific 99th percentiles were determined de novo. Classification of patients relative to the 99th percentiles was used to assess biological equivalence. RESULTS: Analytical precision and detection capability of the Atellica TnIH assay is excellent with a limit of blank <1 ng/L and 62.5% of women and 93% of men had results above the limit of detection. The 99th percentiles (90% CI) in women were 49 ng/L (31-67) and 70 ng/L (48-121) in men. An asymmetrical distribution involving 5% of results was notable. Agreement was moderate (Kappa 0.58, 95% CI 0.53-0.63) with 20% of patients discordantly classified with Atellica TnIH below and Access hsTnI above the 99th percentiles. Serial results in 195 patients demonstrated good agreement (Kappa 0.84, 95% CI 0.77-0.90). Differences greater than the assay specific reference change values (z≥±1.96) occurred in 65% (95% CI 53-76%) of 99th percentile discordant patients compared to 2.7% (p<0.001) and 76% (p=0.17) of the concordant low and high cTnI groups respectively. CONCLUSIONS: The 99th percentile discordant and the concordantly elevated groups are more alike with respect to their z≥±1.96 rates. This favours an overestimated Atellica TnIH 99th percentile as more likely, and we hypothesize that antibody interference resulting in asymmetric scatter of nearly 5% samples may be the underlying mechanism. Analytical accuracy and interferences in cardiac troponin assays should be investigated and resolved with high priority.

Copeptin is increased by nausea and vomiting during hypertonic saline infusion in healthy individuals.

OBJECTIVE: Measurement of hypertonic saline-stimulated copeptin has recently been described for the differentiation of polyuria-polydipsia syndrome. This study aims to determine the copeptin response to intravenous 3% hypertonic saline, including evaluation of adverse effects, in a local cohort of healthy adults >18 years in Australia. DESIGN: Prospective clinical study. METHODS: Twenty healthy volunteers (10 males and 10 females) were recruited. Participants underwent infusion of 3% hypertonic saline via a previously described standardized protocol, until the plasma sodium was ≥150 mmol/L, with measurement of plasma copeptin. RESULTS: Mean peak sodium was 152 mmol/L ± SD 1.4 with osmolality 315 mmol/kg ± SD 3.9. Median volume of hypertonic saline infused to reach target sodium ≥ 150 mmol/L was 1536 mL (IQR 1362, 1992). Mean rate of plasma sodium rise was 5.9 mmol/L/hour ± SD 1.5. Hypertonic saline-stimulated copeptin had non-parametrical distribution with median of 33.8 pmol/L (IQR 27.6, 63.6). Overall median symptom burden was 6/10 (range 3/10-9/10). Copeptin was significantly higher for those who experienced nausea and/or vomiting (n = 13) (median 39.0 pmol/L; IQR 32.5, 90), compared to those participants who did not experience either (median 20.0 pmol/L; IQR 13.0, 31.0) (P = 0.003). There were no serious adverse events. CONCLUSION: Hypertonic saline-stimulated copeptin measurements were similar in our population compared with previously reported reference intervals in healthy volunteers. There is a wide range of stimulated copeptin measurements in the healthy population. Nausea and vomiting are common adverse effects which enhance the copeptin response.

The relationship between early post-operative ACTH / cortisol following pituitary surgery and long-term glucocorticoid requirement - Do ultradian rhythms matter?

OBJECTIVE: To determine whether early (4-8h) post-operative ACTH after trans-sphenoidal surgery (TSS) predicts long-term hypothalamic-pituitary-adrenal (HPA) axis function and to investigate early morning day 1 ACTH/cortisol variability using rapid sampling. DESIGN: Prospective observational study. METHODS: Participants undergoing TSS were included; those treated with glucocorticoids pre-operatively received 100 mg intravenous hydrocortisone on anaesthetic induction. ACTH and cortisol were measured post-operatively at + 4h and + 8h after induction and on day 1 every 10 minutes between 0700h and 0900h. PRIMARY OUTCOME: glucocorticoid requirement at 6 months. RESULTS: Nineteen participants (10F, 9M): 6/19 (32%) were treated with replacement glucocorticoids pre-operatively; 4 had ceased by 6 weeks post-operatively. One patient developed new hypopituitarism post-operatively meaning 3/19 (16%) required glucocorticoids at 6 months. Post-operative + 4h ACTH < 14.3 pmol/L (65 ng/L) predicted secondary adrenal insufficiency (SAI) (sensitivity 100%, specificity 75%), whilst no participant with a post-operative + 4h ACTH ≥ 14.3 pmol/L (65 ng/L) required glucocorticoids at 6 months. Day 1 ACTH and cortisol showed a significant circadian fall between 0700h-0900h; ACTH 4.2 pmol/L (IQR 2.9-5.9) to 3.7 pmol/L (IQR 2.9-5.1) P = .006 and cortisol 549 nmol/L (IQR 337-618) to 439 nmol/L (IQR 315-606) P < .001, with clinically insignificant ultradian secretory pulses. CONCLUSIONS: No participant with a post-operative + 4h ACTH ≥ 14.3 pmol/L (65 ng/L) required glucocorticoids at 6 months; however, given only 3/19 participants had the primary outcome of interest, this must be confirmed in a larger cohort. The timing of a day 1 morning cortisol between 0700h and 0900h influences the accuracy of a single cut-off to diagnose SAI after pituitary surgery.

Interference by piperacillin/tazobactam in the measurement of creatinine with the Jaffe method and of total protein with the biuret method.

Serum creatinine and total protein are routinely measured biochemical parameters used in clinical medicine. An abnormal result caused by interference with the assay does not accurately reflect a patient's clinical state and therefore risks misleading clinicians. We report the case of a patient who had unexplainable high creatinine and total protein results. The blood collection was contaminated with intravenous fluid and the patient was receiving piperacillin/tazobactam. Additional laboratory studies demonstrated piperacillin/tazobactam was the cause of the false positive results and the elevation in both serum creatinine and protein level was dependent on the concentration of antibiotic present.

The Effect of High-Flow Nasal Oxygen on Carbon Dioxide Accumulation in Apneic or Spontaneously Breathing Adults During Airway Surgery: A Randomized-Controlled Trial.

BACKGROUND: High-flow nasal oxygen (HFNO) is an emerging technology that has generated interest in tubeless anesthesia for airway surgery. HFNO has been shown to maintain oxygenation and CO2 clearance in spontaneously breathing patients and is an effective approach to apneic oxygenation. Although it has been suggested that HFNO can enhance CO2 clearance during apnea, this has not been established. The true extent of CO2 accumulation and resulting acidosis using HFNO during prolonged tubeless anesthesia remains undefined. METHODS: In a single-center trial, we randomly assigned 20 adults undergoing microlaryngoscopy to apnea or spontaneous ventilation (SV) using HFNO during 30 minutes of tubeless anesthesia. Serial arterial blood gas analysis was performed during preoxygenation and general anesthesia. The primary outcome was the partial pressure of CO2 (Paco2) after 30 minutes of general anesthesia, with each group compared using a Student t test. RESULTS: Nineteen patients completed the study protocol (9 in the SV group and 10 in the apnea group). The mean (standard deviation [SD]) Paco2 was 89.0 mm Hg (16.5 mm Hg) in the apnea group and 55.2 mm Hg (7.2 mm Hg) in the SV group (difference in means, 33.8; 95% confidence interval [CI], 20.6-47.0) after 30 minutes of general anesthesia (P < .001). The average rate of Paco2 rise during 30 minutes of general anesthesia was 1.8 mm Hg/min (SD = 0.5 mm Hg/min) in the apnea group and 0.8 mm Hg/min (SD = 0.3 mm Hg/min) in the SV group. The mean (SD) pH was 7.11 (0.04) in the apnea group and 7.29 (0.06) in the SV group (P < .001) at 30 minutes. Five (55%) of the apneic patients had a pH <7.10, of which the lowest measurement was 7.057. No significant difference in partial pressure of arterial O2 (Pao2) was observed after 30 minutes of general anesthesia. CONCLUSIONS: CO2 accumulation during apnea was more than double that of SV after 30 minutes of tubeless anesthesia using HFNO. The use of robust measurement confirms that apnea with HFNO is limited by CO2 accumulation and the concomitant severe respiratory acidosis, in contrast to SV. This extends previous knowledge and has implications for the safe application of HFNO during prolonged procedures.

Progress Curve Analysis of the one stage chromogenic assay for ecarin.

Snake venom prothrombin activators such as Ecarin are readily assayed by continuous spectrophotometric monitoring of p-nitroaniline production in a one step assay containing prothrombin and a p-nitroanilide peptide substrate for thrombin. The coupled reactions result in accelerating p-nitroaniline (pNA) production over the course of the assay giving non-linear progress curves, from which initial velocities are not readily obtained. Most studies therefore resort to approximate estimates of activity, based on the absorbance reached at an arbitrary time. A simple kinetic analysis of the coupled reactions shows that the early points of such curves should be fitted by second order polynomials, representing the accelerating reaction rate in µmol pNA/min/min. The first derivative of the polynomial then gives the increasing velocity of pNA production in µmol pNA/min over the time course of the assay. We demonstrate here that, with the substrate S2238, these rates can be converted to absolute thrombin concentrations using the Michaelis-Menten equation, substituted with values for kcat and Km. These thrombin concentrations increase linearly over the time course of the assay allowing the activity to be expressed in units, defined as µmol product/min, most commonly used to report enzyme activity.

Methamphetamine intoxication and acute kidney injury: A prospective observational case series.

AIM: The effects of methamphetamine intoxication on the kidney are not well reported. We aimed to investigate acute kidney injury (AKI) associated with methamphetamine intoxication, in particular its severity, duration and association with rhabdomyolysis. METHODS: This is a prospective observational series of methamphetamine-intoxicated patients presenting to an Emergency Department. Patients self-reporting recent methamphetamine use, with a positive urine drug screen and an elevated creatinine, were eligible for the study. Urinary neutrophil gelatinase-associated lipocalin (NGAL) was measured, and serum creatinine, creatine kinase and cystatin C concentrations were performed on arrival and at several time points until discharge from hospital. Demographic and clinical data were obtained from the medical records. RESULTS: There were 634 presentations with methamphetamine intoxication over a 10-month period, with 73/595(12%) cases having an elevated serum creatinine concentration on arrival. Fifty presentations in 48 patients were included in the study. Most patients (85%) were male with a median age of 32 years. The median serum creatinine concentration on presentation was 125 µmol/L (IQR:113-135 µmol/L) with 45 (90%) presentations meeting diagnostic criteria for AKI. Concurrent rhabdomyolysis occurred in 22 (44%) presentations with a median CK of 2695 U/L (IQR:1598-5060 U/L). Cystatin C was elevated (> 0.98 mg/L) in 18 cases. An elevated NGAL concentration (>150 µg/L) was present in five (10%) cases. No patients required dialysis. The median length of stay was 19 hours (IQR 14-24 hours). CONCLUSION: AKI is common in methamphetamine intoxication. The kidney injury is relatively mild and short-lived, resolving with crystalloid therapy.

Rapid serum tube technology overcomes problems associated with use of anticoagulants.

INTRODUCTION: Failure to obtain complete blood clotting in serum is a common laboratory problem. Our aim was to determine whether snake proth-rombin activators are effective in clotting blood and producing quality serum for analyte measurement in anticoagulated patients. MATERIALS AND METHODS: Whole blood clotting was studied in a total of 64 blood samples (41 controls, 20 Warfarin patients, 3 anticoagulated patients using snake venom prothrombin activator (OsPA)) with plain tubes. Coagulation was analysed using a visual assay, Hyland-Clotek and thromboelastography. Healthy control blood was spiked with a range of anticoagulants to determine the effectiveness of OsPa-induced clotting. A paired analysis of a Dabigatran patient and a control investigated the effectiveness of the OsPA clotting tubes. Biochemical analytes (N = 31) were determined for 7 samples on chemistry and immunoassay analysers and compared with commercial tubes. RESULTS: Snake venom prothrombin activators efficiently coagulated blood and plasma spiked with heparin and commonly used anticoagulants. Clotting was observed in the presence of anticoagulants whereas no clotting was observed in BDRST tubes containing 3 U/mL of heparin. Snake venom prothrombin activator enhanced heparinised blood clotting by shortening substantially the clotting time and improving significantly the strength of the clot. Comparison of 31 analytes from the blood of five healthy and two anticoagulated participants gave very good agreement between the analyte concentrations determined. CONCLUSIONS: Our results showed that the snake venom prothrombin activators OsPA and PtPA efficiently coagulated recalcified and fresh bloods with or without added anticoagulants. These procoagulants produced high quality serum for accurate analyte measurement.

Biochemical evaluation of kidney disease.

Different biochemical markers exist in both blood and urine for assessing renal function. Most of these biomarkers have advantages and limitations associated with their use, which is important to consider when ordering and utilising them in the clinical setting. The ideal marker should be able to detect acute kidney injury (AKI) at the onset and be used for the diagnosis and ongoing monitoring and management of kidney disease. The search for such a marker is ongoing, as all potential candidates thus far are associated with certain limitations. This article will attempt to compare and contrast established and emerging kidney disease markers.

Elevated insulin-like growth factor-1 in Cushing's disease.

OBJECTIVE: To determine if patients with untreated Cushing's disease have higher serum insulin-like growth factor-1 (IGF-1) compared to matched controls, and if IGF-1 decreases following remission of Cushing's disease. DESIGN: Retrospective case-control study matching Cushing's disease cases to control patients for adenoma size, age, sex, diabetic and gonadal status, body mass index and serum IGF-1 measured within one year. Paired analysis of pre-operative (untreated) and >3 months post-operative (remission) serum IGF-1 for cases. PATIENTS AND MEASUREMENTS: All patients were investigated at the Princess Alexandra Hospital Endocrine Unit between 2005 and 2017. Serum IGF-1 was measured in 25 cases and 49 controls, 23 case-control pairs and 13 cases pre- and post-operatively. RESULTS: Mean serum IGF-1 in cases was significantly higher compared to controls-32 ± 12 nmol/L compared to 25 ± 8 nmol/L, (P = 0.005). The proportion of cases with elevated serum IGF-1 above an age-adjusted reference range was higher compared to 1:1 matched controls (8/23 (35%) vs 1/23 (4%), P = 0.02). In 13 cases in remission post-operatively, serum IGF-1 decreased significantly from 31 (IQR 29-40.5) nmol/L to 23 (IQR 15-28.5) nmol/L, (P < 0.001), despite no difference in the prevalence of pre- vs post-operative pituitary hormone dysfunction (P = 0.47). CONCLUSION: Patients with untreated Cushing's disease may have elevated IGF-1, which decreases following remission. Mildly elevated IGF-1 in Cushing's disease does not imply pathological growth hormone (GH) excess.

Urea treatment in fluid restriction-refractory hyponatraemia.

OBJECTIVE: Hyponatraemia in hospitalized patients is common and associated with increased mortality. International guidelines give conflicting advice regarding the role of urea in the treatment of SIADH. We hypothesized that urea is a safe, effective treatment for fluid restriction-refractory hyponatraemia. DESIGN: Review of urea for the treatment of hyponatraemia in patients admitted to a tertiary hospital during 2016-2017. Primary end-point: proportion of patients achieving a serum sodium ≥130 mmol/L at 72 hours. PATIENTS: Urea was used on 78 occasions in 69 patients. The median age was 67 (IQR 52-76), 41% were female. Seventy (89.7%) had hyponatraemia due to SIADH-CNS pathology (64.3%) was the most common cause. The duration was acute in 32 (41%), chronic in 35 (44.9%) and unknown in the rest. RESULTS: The median nadir serum sodium was 122 mmol/L (IQR 118-126). Fluid restriction was first-line treatment in 65.4%. Urea was used first line in 21.8% and second line in 78.2%. Fifty treatment episodes (64.1%) resulted in serum sodium ≥130 mmol/L at 72 hours. In 56 patients who received other prior treatment, the mean sodium change at 72 hours (6.9 ± 4.8 mmol/L) was greater than with the preceding treatments (-1.0 ± 4.7 mmol/L; P < 0.001). Seventeen patients (22.7%) had side effects (principally distaste), none were severe. No patients developed hypernatraemia, overcorrection (>10 mmol/L in 24 hours or >18 mmol/L in 48 hours), or died. CONCLUSIONS: Urea is safe and effective in fluid restriction-refractory hyponatraemia. We recommend urea with a starting dose of ≥30 g/d, in patients with SIADH and moderate to profound hyponatraemia who are unable to undergo, or have failed fluid restriction.

Chronic kidney cortical damage is associated with baseline kidney function and albuminuria in patients managed with radical nephrectomy for kidney tumours.

This study evaluated the relationship between histological markers of chronic kidney damage in patients undergoing radical nephrectomy for kidney tumours and preoperative kidney function, degree of albuminuria, and changes in glomerular volume. A schema to grade chronic kidney damage could be used to identify patients at risk of developing CKD following nephrectomy. Non-neoplastic cortical tissue was sourced from 150 patients undergoing radical nephrectomy for suspected kidney cancer. This tissue was evaluated for indicators of chronic damage, specifically: glomerulosclerosis, arteriosclerosis, interstitial fibrosis, and tubular atrophy. Glomerular volume was determined using the Weibel and Gomez method. Associations between these parameters and both estimated glomerular filtration rate (eGFR) and albumin-creatinine ratio (ACR) were determined using either a Mann-Whitney U-test or a Kruskal-Wallis ANOVA. Associations between both eGFR and ACR and glomerular volume were assessed using linear regression. eGFR was inversely associated with the degree of glomerulosclerosis (p < 0.001), vascular narrowing (p = 0.002), tubular atrophy (p < 0.001), and interstitial fibrosis (p < 0.001). ACR was associated only with the degree of interstitial fibrosis (p = 0.02) and tubular atrophy (p = 0.02). Glomerular volume was greater for males, diabetics, hypertensive patients, and patients with a greater degree of interstitial fibrosis. Glomerular volume was positively associated with ACR. A schema to grade chronic damage was developed. The proposed schema is associated with baseline clinical indices of kidney function and damage. Longitudinal validation is necessary to determine the prognostic utility of this schema.

Mineralocorticoid antagonism enhances brown adipose tissue function in humans: A randomized placebo-controlled cross-over study.

AIM: To investigate whether mineralocorticoid (MC) antagonism enhances brown adipose tissue (BAT) function in humans. MATERIALS AND METHODS: In a randomized double-blind, cross-over designed trial, 10 healthy adults (two men, eight women) underwent 2 weeks of spironolactone (100 mg/d) treatment and placebo, with an intervening 2-week wash-out period. BAT function was assessed in response to cooling and to a mixed meal. Metabolic activity was measured by fluoro-deoxyglucose (FDG) uptake (maximal standardized uptake value, SUVmax ) using PET-CT. Thermogenic activity was measured by skin temperatures overlying supraclavicular (SCL) BAT depots using infrared thermography. Postprandial metabolism was measured by energy production rate (EPR) and lipid synthesis using indirect calorimetry. RESULTS: During cooling, BAT metabolic activity (SUV 6.30 ± 2.16 vs 3.98 ± 1.34; P < 0.05) and volume (54.9 ± 22.8 vs 21.6 ± 11.8 cm3 ; P < 0.05) were significantly higher, and mean SCL temperature decreased by a smaller degree (-0.3°C°± 0.2°C vs -0.9°C ± 0.2°C; P = 0.05) with spironolactone treatment. A mixed meal increased SCL temperature and EPR. The postprandial rise in SCL temperature (+0.4°C ± 0.1°C vs +0.1°C ± 0.1°C; P < 0.05) but not in EPR was greater during spironolactone treatment. Postprandial lipid synthesis occurred in three participants with placebo but in none with spironolactone treatment (P = 0.06). CONCLUSION: MC antagonism enhanced human BAT function in response to cooling and to a meal during which lipid synthesis was suppressed. As postprandial EPR comprises energy dissipated as heat and energy required to store nutrients, the reduction in lipid synthesis during MC antagonism is a probable consequence of concurrent stimulation of BAT thermogenesis. The shift in energy usage from storage to heat dissipation indicates that MC antagonists may have therapeutic benefit for obesity.

Next-generation rapid serum tube technology using prothrombin activator coagulant: fast, high-quality serum from normal samples.

Background Incomplete blood clotting or latent clotting in serum is a common laboratory problem, especially for patients on anticoagulant therapy or when serum tubes are centrifuged before clotting is completed. We describe a novel approach to producing high-quality serum using snake venom prothrombin activator complex (OsPA) as an additive in blood collection tubes for non-anticoagulated (normal) individuals. Methods Plasma clotting assays were performed using a Hyland-Clotek instrument. Blood clotting was visually observed, and thromboelastography was also performed to determine the important parameters of coagulation. Thrombin generation was assayed using the chromogenic substrate S-2238, and biochemical analytes in the serum were determined on chemistry and immunoassay analysers. Fibrinogen was determined by either ELISA or Clauss fibrinogen assay. Results We initially showed that OsPA had strong coagulation activity in clotting not only recalcified citrated plasma and recalcified citrated whole blood, but also fresh whole blood in a clinical setting. The use of TEG clearly showed improved speed of clotting and generation of a firmer clot. We also showed that the use of OsPA to produce serum did not interfere with the determination of commonly measured biochemical analytes. The underlying clotting mechanism involves a burst of thrombin production at the initial stages of the clotting process upon contact with prothrombin in blood. Conclusions These results demonstrate rapid generation of high-quality serum, contributing to faster turnaround times with standardised quality samples, for accurate analyte determinations in normal individuals.

Investigation and management of moderate to severe inpatient hyponatraemia in an Australian tertiary hospital.

BACKGROUND: Hyponatraemia is the most common electrolyte disturbance amongst hospitalised patients. Both American and European guidelines recommend fluid restriction as first line treatment for SIADH, however differ on second line recommendations. The objective of this study was to examine investigation and management of hyponatraemia in hospitalised patients in an Australian tertiary hospital. METHODS: A retrospective audit was conducted of electronic medical records and laboratory data of inpatients with serum sodium (Na) ≤125 mmol/L, admitted over a 3 month period to the Princess Alexandra Hospital, Brisbane, Australia. The main outcomes measured included: demographic characteristics, investigations, accuracy of diagnosis, management strategy, change in Na and patient outcomes. RESULTS: The working clinical diagnosis was considered accurate in only 37.5% of cases. Urine Na and osmolality were requested in 72 of 152 patients (47.4%) and in 43 of 70 euvolaemic patients (61.4%). Thyroid function tests (67.1%) and morning cortisol (45.7%) were underutilized in the euvolaemic group. In the SIADH cohort, fluid restriction resulted in a median (IQR) 7.5 mmol/L (4-10.5) increase in Na after 3 days; no treatment resulted in a median 0 mmol/L (- 0.5-1.5) change. Oral urea was utilized in 5 SIADH patients where Na failed to increase with fluid restriction alone. This resulted in a median 10.5 mmol/L (3.5-13) increase in Na from baseline to day 3. There were no cases of osmotic demyelination. The median length of stay was 8 days (4-18.5). Mortality was 11.2% (17 patients). There was a weak but significant correlation between nadir serum Na and mortality (R = 0.18, P = 0.031). CONCLUSION: Inpatient hyponatraemia is often inadequately investigated, causing errors in diagnosis. Treatment is heterogeneous and often incorrect. In cases with hyponatraemia refractory to fluid restriction, oral urea presents an effective alternative treatment.