Cytochrome c Oxidase (CcO) is a multisubunit bigenomic protein complex which catalyzes the last step of the mitochondrial electron transport chain. The nuclear encoded subunits are thought to have roles either in regulation or in the structural stability of the enzyme. Subunit Vb is a peripheral nuclear-encoded subunit of mammalian CcO that is dramatically reduced under hypoxia. Although it has been shown to contain different ligand binding sites and undergo modifications, its precise function is not known. In the present study we generated a cell line from RAW 264.7 murine macrophages, that has more than 80% reduced level of Vb. Functional analysis of these cells showed a loss of CcO activity, membrane potential and lower ability to generate ATP. Resolution of complexes on Blue native gel and two dimensional electrophoretic analysis showed an accumulation of subcomplexes of CcO and also reduced association with super complexes of the electron transfer chain. Furthermore, the mitochondria from CcO Vb knock down cells generated increased ROS and the cells were unable to grow on galactose containing medium. Pulse chase experiments suggest the role of CcO Vb subunit in the assembly of the complex. We show for the first time the role of a peripheral, non-transmembrane subunit in the formation as well as function of the terminal CcO complex.

Infection by SARS-CoV-2 is associated with a high risk of thrombosis. The laboratory documentation of hypercoagulability and impaired fibrinolysis remains a challenge. Our aim was to assess the potential usefulness of viscoelastometric testing (VET) to predict thrombotic events in COVID-19 patients according to the literature. We also (i) analyzed the impact of anticoagulation and the methods used to neutralize heparin, (ii) analyzed whether maximal clot mechanical strength brings more information than Clauss fibrinogen, and (iii) critically scrutinized the diagnosis of hypofibrinolysis. We performed a systematic search in PubMed and Scopus databases until December 31st, 2020. VET methods and parameters, and patients’ features and outcomes were extracted. VET was performed for 1063 patients (893 intensive care unit (ICU) and 170 non-ICU, 44 studies). There was extensive heterogeneity concerning study design, VET device used (ROTEM, TEG, Quantra and ClotPro) and reagents (with non-systematic use of heparin neutralization), timing of assay, and definition of hypercoagulable state. Notably, only 4 out of 25 studies using ROTEM reported data with heparinase (HEPTEM). The common findings were increased clot mechanical strength mainly due to excessive fibrinogen component and impaired to absent fibrinolysis, more conspicuous in the presence of an added plasminogen activator. Only 4 studies out of the 16 that addressed the point found an association of VETs with thrombotic events. So-called functional fibrinogen assessed by VETs showed a variable correlation with Clauss fibrinogen. Abnormal VET pattern, often evidenced despite standard prophylactic anticoagulation, tended to normalize after increased dosing. VET studies reported heterogeneity, and small sample sizes do not support an association between the poorly defined prothrombotic phenotype of COVID-19 and thrombotic events.

Coronavirus disease 2019 (COVID-19) is associated with extreme inflammatory response, disordered hemostasis and high thrombotic risk. A high incidence of thromboembolic events has been reported despite thromboprophylaxis, raising the question of a more effective anticoagulation. First-line hemostasis tests such as activated partial thromboplastin time, prothrombin time, fibrinogen and D-dimers are proposed for assessing thrombotic risk and monitoring hemostasis, but are vulnerable to many drawbacks affecting their reliability and clinical relevance. Specialized hemostasis-related tests (soluble fibrin complexes, tests assessing fibrinolytic capacity, viscoelastic tests, thrombin generation) may have an interest to assess the thrombotic risk associated with COVID-19. Another challenge for the hemostasis laboratory is the monitoring of heparin treatment, especially unfractionated heparin in the setting of an extreme inflammatory response. This review aimed at evaluating the role of hemostasis tests in the management of COVID-19 and discussing their main limitations.

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Low tissue factor pathway inhibitor (TFPI) together with low antithrombin allows sufficient thrombin generation in neonates.

Pituitary-thyroid function was assessed in 40 patients with chronic renal failure undergoing regular maintenance hemodialysis and in 35 normal subjects. Serum thyroid hormone levels were significantly lower in the dialysis patients than in the normal subjects (P less than 0.001) and were in the hypothyroid range in a high proportion of dialysis patients (total T3, 25%; free T3, 45%; total T4, 55%; free T4, 45%; and free T4 index, 38%). The reduced free thyroid hormone levels could not be explained by currently recognized assay artefacts. Serum TSH levels were higher than in the normal subjects (P less than 0.01), but still within the normal range for most (35 of 40) dialysis patients and did not correlate significantly with total or free thyroid hormone concentrations in either group. These results suggest some impairment in the thyroidal response to TSH and impaired pituitary response to low serum thyroid hormone levels, the latter implying resetting of the normal feedback mechanism such that diminished thyroid hormone production evokes a smaller than normal increase in TSH secretion. This diminished thyrotroph sensitivity to reduced serum thyroid hormone levels may be beneficial in severe nonthyroidal illness.

Platelet count, indices (mean volume, young—immature platelet fraction) and aggregation are widely used laboratory parameters to investigate primary hemostasis. We performed a systematic, thorough evaluation of the influence of the time-interval since blood draw from 20 healthy individuals and of the anticoagulation of collected blood on such parameters. Blood was anticoagulated with citrate, K2-ethylenediaminetetraacetic acid (EDTA) and hirudin and analyzed 5, 30, 60, 120 and 180 min after blood draw. Multiple electrode aggregometry (MEA) was performed with either hirudin (half-diluted with NaCl) or citrate samples (half-diluted with NaCl or CaCl2 3 mM). Platelet count and indices (Sysmex XN-20) were rather stable over time with EDTA blood. MEA results were lower with citrate blood than with hirudin blood; supplementation with calcium was partially compensatory. MEA results were also lower when performed less than 30 or more than 120 min after blood draw. Platelet clumping, quantitatively estimated with microscope examination of blood smears, was more important in hirudin blood than citrate or EDTA blood and could explain some of the differences observed between preanalytical variables. The results stress once more the importance of preanalytical variables in hemostasis laboratory testing. Decision thresholds based on those tests are only applicable within specific preanalytical conditions.

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