Microsleep assessment enhances interpretation of the Maintenance of Wakefulness Test.

STUDY OBJECTIVES: The Maintenance of Wakefulness Test (MWT) is used to objectively evaluate an individual's ability to remain awake; however, microsleeps are not included in the assessment. We aimed to determine if microsleep data prior to sleep onset assisted in interpretation of ability to maintain wakefulness across a range of typical patient groups. METHODS: Forty-eight patients referred for overnight polysomnography and subsequent MWT were included. Patients were divided into 3 groups (treated obstructive sleep apnea [OSA], untreated OSA, or treated idiopathic hypersomnia or narcolepsy) based on prior medical diagnosis. Demographics, clinical characteristics, polysomnography, and MWT variables, including frequency, distribution, duration, and latency of microsleeps were compared between groups. RESULTS: Microsleeps were observed in MWT trials significantly more frequently in patients with treated idiopathic hypersomnia/narcolepsy over the course of the day (0.34 ± 0.06 vs 0.07 ± 0.02 microsleeps/min; P < .001) and in patients with untreated OSA toward the end of the day (0.31 ± 0.06 vs 0.05 ± 0.02 microsleeps/min; P < .001) compared to the group with treated OSA. Microsleeps were often observed in series and earlier in patients with treated idiopathic hypersomnia/narcolepsy (10.9 ± 1.6 minutes) and those with untreated OSA (16.2 ± 2.7 minutes) compared to the group with treated OSA (24.9 ± 3.0 minutes; P < .05), and, if taken into consideration, would increase the proportion of patients demonstrating inability to maintain wakefulness by 33% and 22%, respectively. CONCLUSIONS: MWT performance varies significantly across patient groups. Microsleep analysis prior to sleep onset may be a more sensitive measure of patient daytime wakefulness than sleep latency alone and should be considered in MWT assessment. CITATION: Anniss AM, Young A, O'Driscoll DM. Microsleep assessment enhances interpretation of the Maintenance of Wakefulness Test. J Clin Sleep Med. 2021;17(8):1571-1578.

Importance of Urinary Drug Screening in the Multiple Sleep Latency Test and Maintenance of Wakefulness Test.

STUDY OBJECTIVES: Multiple sleep latency testing (MSLT) and the maintenance of wakefulness test (MWT) are gold-standard objective tests of daytime sleepiness and alertness; however, there is marked variability in their interpretation and practice. This study aimed to determine the incidence of positive drug screens and their influence on MSLT, MWT, and polysomnographic variables. METHODS: All patients attending Eastern Health Sleep Laboratory for MSLT or MWT over a 21-mo period were included in the study. Urinary drug screening for amphetamines, barbiturates, benzodiazepines, cannabinoids, cocaine, methadone, and opiates was performed following overnight polysomnography (PSG). Demographics and PSG variables were compared. RESULTS: Of 69 studies, MSLT (43) and MWT (26), 16% of patients had positive urinary drug screening (7 MSLT; 4 MWT). Drugs detected included amphetamines, cannabinoids, opiates, and benzodiazepines. No patient self-reported use of these medications prior to testing. No demographic, MSLT or MWT PSG data or overnight PSG data showed any statistical differences between positive and negative drug screen groups. Of seven MSLT patients testing positive for drug use, one met criteria for the diagnosis of narcolepsy and five for idiopathic hypersomnia. On MWT, three of the four drug-positive patients had a history of a motor vehicle accident and two patients were occupational drivers. CONCLUSIONS: These findings indicate drug use is present in patients attending for daytime testing of objective sleepiness and wakefulness. These data support routine urinary drug screening in all patients undergoing MSLT or MWT studies to ensure accurate interpretation in the context of illicit and prescription drug use.

Variable adhesion of different red blood cell products to activated vascular endothelium under flow conditions.

Red blood cells (RBCs) that have been stored prior to transfusion show increased adherence to vascular endothelium in vitro, which suggests a potential for stored blood transfusion to impede blood flow in some patients. Transfusion is often required in patients with sepsis or inflammation; however, whether activation of endothelium affects stored RBC-endothelial cell (EC) interactions is unknown. We investigated whether storage time and leukocyte content of RBC products influences the adhesion of RBCs to activated ECs. RBCs from nonleukocyte-reduced (S-RBCs), buffy-coat-poor (BCP-RBCs), and leukocyte-filtered (LF-RBCs) products and cultured EC layers were pretreated with endotoxin, tumor necrosis factor-alpha (TNF-alpha), or medium alone prior to perfusion of the RBCs across the EC layer in a continuous flow microchamber. After a single day of RBC storage, the number of adherent RBCs was increased in the endotoxin and TNF-alpha pretreated groups compared to the unactivated-control group. These differences were statistically significant for S-RBCs and LF-RBC products (P < 0.05). In contrast, there was no significant difference in RBC adherence to activated and unactivated endothelium at other time-points of RBC product storage. The strength of adhesion of stored RBCs from S-RBC products to activated ECs was not altered following treatment; however, endotoxin significantly increased the adhesive strength of LF-RBCs to endothelium. These results demonstrate that while fresh RBCs show increased adhesion to activated endothelium, storage of RBCs did not promote increased adhesion to activated endothelium. However, inflammatory conditions promote stronger adhesion of stored RBCs to ECs, which may contribute to impaired tissue perfusion in some transfusion recipients.

Storage duration and white blood cell content of red blood cell (RBC) products increases adhesion of stored RBCs to endothelium under flow conditions.

BACKGROUND: Adherence of red blood cells (RBCs) to vascular endothelium impairs blood flow and decreases oxygen delivery. Although RBCs may be stored for up to 42 days before transfusion under current blood banking guidelines, little is known of how changes to RBCs during storage may affect their adherence properties. The influence of RBC product storage time and white blood cell (WBC) burden on the adherence of RBCs for transfusion to vascular endothelium under conditions of continuous flow was investigated in this study. STUDY DESIGN AND METHODS: RBC samples were collected from nonleukoreduced (S-RBC), buffy coat-poor (BCP-RBC), and leukofiltered (LF-RBC) products at fixed time points during storage. Samples were perfused, at controlled shear stress and temperature, across a confluent endothelial cell (EC) monolayer with a parallel-flow chamber mounted to an inverted microscope. RBC-EC interactions were recorded with a digital camera attached to the microscope. RESULTS: The number of RBCs adhering to the EC layer increased significantly with storage time in all RBC products; however, WBC reduction delayed this increase. LF-RBCs were also significantly less adherent than S-RBC or BCP-RBC products on Day 1 of storage (p < 0.05). The strength of RBC attachment to vascular endothelium was significantly stronger in S-RBC products compared to BCP-RBC and LF-RBC products. CONCLUSION: Our findings indicate that product storage time and WBC burden increase the number and strength of adhesion of RBCs to vascular endothelium. These results may lead to greater understanding of the interaction of transfused RBCs with recipient endothelium and the biologic consequences of this adherence.

Proteomic analysis of supernatants of stored red blood cell products.

BACKGROUND: The development of the red blood cell (RBC) storage lesion remains incompletely understood. To gain a greater insight into the mechanisms involved, a proteomics analysis was used to identify proteins that accumulate in supernatants of standard nonleukoreduced RBC products (S-RBCs) and prestorage leukofiltered RBC products (LF-RBCs) during storage. STUDY DESIGN AND METHODS: S-RBCs and LF-RBCs were collected and stored in accordance with standard blood bank procedures. Supernatant samples were collected at fortnightly intervals until product expiry (at Day 42). Maps of supernatant proteins were generated by two-dimensional (2D)-gel electrophoresis and selected proteins were identified by mass spectrometry. RESULTS: 2D-gel mapping revealed that greater numbers of proteins accumulated in supernatants of S-RBCs compared to LF-RBCs. Abundant plasma proteins were strongly represented in both products. Several potentially bioactive proteins were found to predominantly accumulate in supernatant of S-RBCs. Among these, a promoter of neutrophil adhesion and an acute-phase scavenger protein were identified. In contrast, proteins found to accumulate predominantly in supernatant of LF-RBCs were RBC-regulatory proteins. CONCLUSION: Proteomics provides a valuable approach to examine storage-related effects on RBCs. Such analytical approaches may help to elucidate the mechanisms involved in the RBC storage lesion and provide insights into the biologic consequences of transfusion of stored RBC products.

An oxysterol-binding protein family identified in the mouse.

Oxysterols are oxygenated derivatives of cholesterol. They have been shown to influence a variety of biological functions including sterol metabolism, lipid trafficking, and apoptosis. Recently, 12 human OSBP-related genes have been identified. In this study, we have identified a family of 12 oxysterol-binding protein (OSBP)-related proteins (ORPs) in the mouse. A high level of amino acid identity (88-97%) was determined between mouse and human ORPs, indicating a very high degree of evolutionary conservation. All proteins identified contained the conserved OSBP amino acid sequence signature motif "EQVSHHPP," and most contained a pleckstrin homology (PH) domain. Using RT-PCR, each mouse ORP gene was found to exhibit a unique tissue distribution with many showing high expression in testicular, brain, and heart tissues. Interestingly, the tissue distribution of ORP-4 and ORP-10 were the most selective within the family. Expression of the various ORP genes was also investigated, specifically in highly purified populations of hemopoietic precursor cells defined by the lin(-) c-kit(+) Sca-1(+) (LKS(+)) and lin(-) c-kit(+) Sca-1(-) (LKS(-)) immunophenotype. Most ORP genes were expressed in both LKS(+) and LKS(-) populations, although ORP-4 appeared to be more highly expressed in the primitive, stem-cell enriched LKS(+) population, whereas ORP-10 was more highly expressed by maturing LKS(-) cells. The identification of a family of ORP proteins in the mouse, the frequently preferred animal model for in vivo studies, should further our understanding of the function of these proteins and their interactions with each other.

Expression of CD47 (integrin-associated protein) decreases on red blood cells during storage.

In light of recent studies that demonstrate that CD47 antigen is an important "self-recognition" marker involved in protecting circulating red blood cells (RBCs) from phagocytosis by macrophages, the aim of the present study was to investigate whether CD47 expression on RBCs is altered during storage. Red cell concentrates (RCCs) were prepared and stored at 4 degrees C and samples were collected on days 1, 14, 28 and 42 post donation. RBCs were labelled with anti-CD47 antibody and analysed by flow cytometry. A small but significant and progressive decrease in CD47 antigen expression was observed in non-irradiated and irradiated RBCs stored for 14 days onwards. CD47 was also detected in increasing quantities in supernatant from RCCs after 14 days of storage. It is proposed that loss of CD47 during storage, in addition to other biophysical changes, may target many transfused RBCs for clearance from the transfusion recipient's circulation.