Enhanced detection and Characterization of M-proteins in multiple myeloma patients using An Agilent AssayMAP Bravo liquid handling system coupled to an LC-QTOF.

Background: Mass spectrometry methods are emerging as tools to detect M-proteins in the serum of multiple myeloma patients with increased sensitivity and specificity compared to traditional electrophoretic methods.

Methods: A liquid handling system, the Agilent AssayMAP Bravo, with liquid chromatography high-resolution quadrupole-time-of-flight (LC-QTOF) mass spectrometry to analyze intact light chains was compared to immunofixation electrophoresis (IFE) for M-protein analysis. 210 patient serum samples were analyzed in a split sample comparison (LC-QTOF vs.

Lack of interference of metronidazole with Roche cobas c502 and c702 chemistry tests.

Objective: The antibiotic metronidazole has been suggested to absorb light at a wavelength range commonly used in spectrophotometric assays. We sought to determine if any of the spectrophotometric assays used in our core laboratory would be susceptible to clinically significant interference from metronidazole in blood-based patient specimens.

Methods: Following characterization of the absorbance spectrum for metronidazole, spectrophotometric assays involving either main or subtraction wavelengths that might be susceptible to interference from metronidazole were identified.

Lot verification practices in Ontario clinical chemistry laboratories - Results of a patterns-of-practice survey.

Objectives: Verifying new reagent or calibrator lots is crucial for maintaining consistent test performance. The Institute for Quality Management in Healthcare (IQMH) conducted a patterns-of-practice survey and follow-up case study to collect information on lot verification practices in Ontario.

Methods: The survey had 17 multiple-choice questions and was distributed to 183 licensed laboratories.

Survey of renin and aldosterone testing practices by Ontario laboratories - Providing insight into best practices.

Objectives: Testing for renin and aldosterone in clinical laboratories is complicated by pre-analytical considerations such as the posture for blood collection and susceptibility to cryoactivation of renin. From an analytical perspective, there are both renin activity and renin mass or concentration assays available. There can also be variability in result reporting practices and the aldosterone-renin ratio (ARR) cut-off applied to screen for primary aldosteronism (PA).

Biochemistry tests in hospitalized COVID-19 patients: Experience from a Canadian tertiary care centre.

Background: Coronavirus Disease 2019 (COVID-19) has variable clinical presentation, from asymptomatic to severe disease leading to death. Biochemical markers may help with management and prognostication of COVID-19 patients; however, their utility is still under investigation.

Methods: A retrospective study was conducted to evaluate alanine aminotransferase, C-reactive protein (CRP), ferritin, lactate, and high sensitivity troponin T (TnT) levels in 67 patients who were admitted to a Canadian tertiary care centre for management of COVID-19.

Reducing overutilisation of serum vitamin D testing at a tertiary care centre.

Introduction: Testing of 25-hydroxy (25-OH) vitamin D serum levels has increased drastically in recent years and much of it is considered inappropriate based on current guidelines.

Methods: In consultation with our physician groups (experts and frequent orderers), we modified existing guidelines and implemented a rational policy for 25-OH vitamin D testing and 1,25 dihydroxy (1,25 di-OH) vitamin D testing at a tertiary care centre. A computer decision support tool requiring selection of one of five acceptable testing indications was created for each test as part of a computerised physician order entry system.

Reducing red blood cell folate testing: a case study in utilisation management.

Mandatory enrichment of wheat flour in Canada with folic acid since 1998 has caused folate deficiency to be rare. There were 3019 red blood cell (RBC) folate tests performed during an 18-month period at London Health Sciences Centre (LHSC)/St. Joseph's Healthcare London (SJHC) without any folate deficiency detected.

Vitamin B deficiency and hyperhomocysteinaemia in outpatients with stroke or transient ischaemic attack: a cohort study at an academic medical centre.

Objective: We sought to assess the current magnitude of the opportunity for secondary stroke prevention with B vitamins.

Design: A cohort study.

Setting: The Urgent TIA (Transient Ischaemic Attack) Clinic at an academic medical centre.

Activation of the dsRNA-Activated Protein Kinase PKR in Mitochondrial Dysfunction and Inflammatory Stress in Metabolic Syndrome.

Background: The double stranded RNA (dsRNA)-activated protein kinase PKR is a well-established protein kinase that is activated by dsRNA during viral infection, and it inhibits global protein synthesis by phosphorylating the alpha subunit of eukaryotic initiation factor 2α (eIF2α). Recent studies have greatly broadened the recognized physiological activities of PKR by demonstrating its fundamental role in inflammatory signaling, particularly in chronic, low-grade inflammation induced by metabolic disorders, known as metaflammation. Metaflammation is initiated by the activation of the NOD-like receptor (NLR), leucine-rich repeat, pyrin domaincontaining 3 (NLRP3) gene by mitochondrial reactive oxygen species (ROS).

Assessing the necessity of including a crossover period with dual reporting when changing total prostate-specific antigen methods.

Objectives: Options are limited for the laboratory when changing analytical methods for tumor markers. Standardization between methods should alleviate this burden. Our objective was to assess the necessity of dual reporting for total prostate-specific antigen (tPSA) when changing methods that have been calibrated against the World Health Organization PSA reference material.

Role of cysteine-protease CGHC motifs of ER-60, a protein disulfide isomerase, in hepatic apolipoprotein B100 degradation.

Apolipoprotein B100 (apoB), the structural component of very low density lipoproteins (VLDL), is susceptible to misfolding and subsequent degradation by several intracellular pathways. ER-60, which has been implicated in apoB degradation, is a protein disulfide isomerase (PDI) that forms or rearranges disulfide bonds in substrate proteins and also possesses cysteine protease activity. To determine which ER-60 function is important for apoB degradation, adenoviruses encoding wild-type human ER-60 or a mutant form of human ER-60 (C60A, C409A) that lacked cysteine protease activity were overexpressed in HepG2 cells.

Lysosomal disruption preferentially targets acute myeloid leukemia cells and progenitors.

Despite efforts to understand and treat acute myeloid leukemia (AML), there remains a need for more comprehensive therapies to prevent AML-associated relapses. To identify new therapeutic strategies for AML, we screened a library of on- and off-patent drugs and identified the antimalarial agent mefloquine as a compound that selectively kills AML cells and AML stem cells in a panel of leukemia cell lines and in mice. Using a yeast genome-wide functional screen for mefloquine sensitizers, we identified genes associated with the yeast vacuole, the homolog of the mammalian lysosome.

Inhibition of mitochondrial translation as a therapeutic strategy for human acute myeloid leukemia.

To identify FDA-approved agents targeting leukemic cells, we performed a chemical screen on two human leukemic cell lines and identified the antimicrobial tigecycline. A genome-wide screen in yeast identified mitochondrial translation inhibition as the mechanism of tigecycline-mediated lethality. Tigecycline selectively killed leukemia stem and progenitor cells compared to their normal counterparts and also showed antileukemic activity in mouse models of human leukemia.

Fructose: a highly lipogenic nutrient implicated in insulin resistance, hepatic steatosis, and the metabolic syndrome.

As dietary exposure to fructose has increased over the past 40 years, there is growing concern that high fructose consumption in humans may be in part responsible for the rising incidence of obesity worldwide. Obesity is associated with a host of metabolic challenges, collectively termed the metabolic syndrome. Fructose is a highly lipogenic sugar that has profound metabolic effects in the liver and has been associated with many of the components of the metabolic syndrome (insulin resistance, elevated waist circumference, dyslipidemia, and hypertension).

Apolipoprotein B100 biogenesis: a complex array of intracellular mechanisms regulating folding, stability, and lipoprotein assembly.

Apolipoprotein B100 (apoB) is a large amphipathic lipid-binding protein that is synthesized by hepatocytes and used to assemble and stabilize very low density lipoproteins (VLDL). It may have been derived through evolution from other lipid-associating proteins such as microsomal triglyceride transfer protein or vitellogenin. The correct folding of apoB requires assistance from chaperone proteins in co-translational lipidation, disulfide bond formation, and glycosylation.

Glucosamine-induced endoplasmic reticulum stress attenuates apolipoprotein B100 synthesis via PERK signaling.

Glucosamine impairs hepatic apolipoprotein B100 (apoB100) production by inducing endoplasmic reticulum (ER) stress and enhancing cotranslational and posttranslational apoB100 degradation (Qiu, W., R. K.

Mechanisms targeting apolipoprotein B100 to proteasomal degradation: evidence that degradation is initiated by BiP binding at the N terminus and the formation of a p97 complex at the C terminus.

Objective: In lipid-poor states, the ubiquitin-proteasomal pathway rapidly degrades misfolded apolipoprotein B100 (apoB) cotranslationally, although the mechanism of delivery from the ER to cytosolic proteasomes is poorly understood. Here we demonstrate key roles of BiP, an ER luminal chaperone, and p97, a cytosolic ATPase anchored to the ER membrane, in the targeting of apoB for proteasomal degradation.

Methods And Results: Using coimmunoprecipitations, we observed associations of apoB with BiP, p97, Derlin-1, VIMP, and the E3 ubiquitin ligase Hrd1 in HepG2 cells.

Fructose and the metabolic syndrome: pathophysiology and molecular mechanisms.

Emerging evidence suggests that increased dietary consumption of fructose in Western society may be a potentially important factor in the growing rates of obesity and the metabolic syndrome. This review will discuss fructose-induced perturbations in cell signaling and inflammatory cascades in insulin-sensitive tissues. In particular, the roles of cellular signaling molecules including nuclear factor kappa B (NFkB), tumor necrosis factor alpha (TNF-alpha), c-Jun amino terminal kinase 1 (JNK-1), protein tyrosine phosphatase 1B (PTP-1B), phosphatase and tensin homolog deleted on chromosome ten (PTEN), liver X receptor (LXR), farnesoid X receptor (FXR), and sterol regulatory element-binding protein-1c (SREBP-1c) will be addressed.

Mechanisms of glucosamine-induced suppression of the hepatic assembly and secretion of apolipoprotein B-100-containing lipoproteins.

Glucosamine-induced endoplasmic reticulum (ER) stress was recently shown to specifically reduce apolipoprotein B-100 (apoB-100) secretion by enhancing the proteasomal degradation of apoB-100. Here, we examined the mechanisms linking glucosamine-induced ER stress and apoB-lipoprotein biogenesis. Trypsin sensitivity studies suggested glucosamine-induced changes in apoB-100 conformation.