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OBJECTIVE: Severe Acute Respiratory Syndrome Coronavirus (SARS-CoV-2) infection may yield a hypercoagulable state with fibrinolysis impairment. We conducted a single-center observational study with the aim of analyzing the coagulation patterns of intensive care unit (ICU) COVID-19 patients with both standard laboratory and viscoelastic tests. The presence of coagulopathy at the onset of the infection and after seven days of systemic anticoagulant therapy was investigated. PATIENTS AND METHODS: Forty consecutive SARS-CoV-2 patients, admitted to the ICU of a University hospital in Italy between 29th February and 30th March 2020 were enrolled in the study, providing they fulfilled the acute respiratory distress syndrome criteria. They received full-dose anticoagulation, including Enoxaparin 0.5 mg·kg-1 subcutaneously twice a day, unfractionated Heparin 7500 units subcutaneously three times daily, or low-intensity Heparin infusion. Thromboelastographic (TEG) and laboratory parameters were measured at admission and after seven days. RESULTS: At baseline, patients showed elevated fibrinogen activity [rTEG-Ang 80.5° (78.7 to 81.5); TEG-ACT 78.5 sec (69.2 to 87.9)] and an increase in the maximum amplitude of clot strength [FF-MA 42.2 mm (30.9 to 49.2)]. No alterations in time of the enzymatic phase of coagulation [CKH-K and CKH-R, 1.1 min (0.85 to 1.3) and 6.6 min (5.2 to 7.5), respectively] were observed. Absent lysis of the clot at 30 minutes (LY30) was observed in all the studied population. Standard coagulation parameters were within the physiological range: [INR 1.09 (1.01 to 1.20), aPTT 34.5 sec (29.7 to 42.2), antithrombin 97.5% (89.5 to 115)]. However, plasma fibrinogen [512.5 mg·dl-1 (303.5 to 605)], and D-dimer levels [1752.5 ng·ml-1 (698.5 to 4434.5)], were persistently increased above the reference range. After seven days of full-dose anticoagulation, average TEG parameters were not different from baseline (rTEG-Ang p = 0.13, TEG-ACT p = 0.58, FF-MA p = 0.24, CK-R p = 0.19, CKH-R p = 0.35), and a persistent increase in white blood cell count, platelet count and D-dimer was observed (white blood cell count p < 0.01, neutrophil count p = 0.02, lymphocyte count p < 0.01, platelet count p = 0.13 < 0.01, D-dimer levels p= 0.02). CONCLUSIONS: SARS-CoV-2 patients with acute respiratory distress syndrome show elevated fibrinogen activity, high D-dimer levels and maximum amplitude of clot strength. Platelet count, fibrinogen, and standard coagulation tests do not indicate a disseminated intravascular coagulation. At seven days, thromboelastographic abnormalities persist despite full-dose anticoagulation.
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Anticoagulantes/uso terapéutico , Trastornos de la Coagulación Sanguínea/sangre , COVID-19/sangre , Síndrome de Dificultad Respiratoria/sangre , Tromboelastografía , Anciano , Anciano de 80 o más Años , Antitrombinas/sangre , Trastornos de la Coagulación Sanguínea/tratamiento farmacológico , Pruebas de Coagulación Sanguínea , Enoxaparina/uso terapéutico , Femenino , Productos de Degradación de Fibrina-Fibrinógeno/metabolismo , Fibrinógeno/metabolismo , Heparina/uso terapéutico , Humanos , Relación Normalizada Internacional , Recuento de Leucocitos , Recuento de Linfocitos , Masculino , Persona de Mediana Edad , Neutrófilos , Tiempo de Tromboplastina Parcial , Recuento de Plaquetas , Estudios Prospectivos , SARS-CoV-2 , Resultado del Tratamiento , Tratamiento Farmacológico de COVID-19RESUMEN
BACKGROUND: Spinal cord injury (SCI) is a debilitating condition characterized by a complex of neurological dysfunctions ranging from loss of sensation to partial or complete limb paralysis. Recently, we reported that intravenous administration of neural precursors physiologically releasing erythropoietin (namely Er-NPCs) enhances functional recovery in animals following contusive spinal cord injury through the counteraction of secondary degeneration. Er-NPCs reached and accumulated at the lesion edges, where they survived throughout the prolonged period of observation and differentiated mostly into cholinergic neuron-like cells. OBJECTIVE: The aim of this study was to investigate the potential reparative and regenerative properties of Er-NPCs in a mouse experimental model of traumatic spinal cord injury. METHODS AND RESULTS: We report that Er-NPCs favoured the preservation of axonal myelin and strongly promoted the regrowth across the lesion site of monoaminergic and chatecolaminergic fibers that reached the distal portions of the injured cord. The use of an anterograde tracer transported by the regenerating axons allowed us to assess the extent of such a process. We show that axonal fluoro-ruby labelling was practically absent in saline-treated mice, while it resulted very significant in Er-NPCs transplanted animals. CONCLUSION: Our study shows that Er-NPCs promoted recovery of function after spinal cord injury, and that this is accompanied by preservation of myelination and strong re-innervation of the distal cord. Thus, regenerated axons may have contributed to the enhanced recovery of function after SCI.
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Eritropoyetina/metabolismo , Regeneración Nerviosa/fisiología , Recuperación de la Función/fisiología , Traumatismos de la Médula Espinal/cirugía , Trasplante de Células Madre/métodos , Animales , Colina O-Acetiltransferasa/metabolismo , Dextranos/metabolismo , Modelos Animales de Enfermedad , Eritropoyetina/uso terapéutico , Colorantes Fluorescentes/administración & dosificación , Proteína GAP-43/metabolismo , Locomoción/fisiología , Masculino , Ratones , Proteínas Asociadas a Microtúbulos/metabolismo , Vaina de Mielina/efectos de los fármacos , Vaina de Mielina/patología , Regeneración Nerviosa/efectos de los fármacos , Compuestos Orgánicos/administración & dosificación , Recuperación de la Función/efectos de los fármacos , Rodaminas/metabolismo , Serotonina/metabolismo , Traumatismos de la Médula Espinal/patología , Tubulina (Proteína)/metabolismo , Tirosina 3-Monooxigenasa/metabolismoRESUMEN
Aim: Carotid endarterectomy is a widely accepted procedure for stroke prevention, and carotid clamping is a necessary surgical step. Glutathionylated haemoglobin (HbSSG) has been recently employed as a biomarker of oxidative stress, its level being increased under several conditions, including hypoxia. This study aims to evaluating whether HbSSG level in peripheral and/or jugular blood is affected during carotid surgery under normal routine operative conditions. Methods: This study enrolled 13 consecutive patients undergoing elective carotid endarterectomy under general anesthesia. At different times during surgery, blood was taken simultaneously from both a peripheral vein and the jugular vein ipsilateral to the clamped carotid. HbSSG was measured in RBC hemolysates by MALDI-ToF mass spectrometry in each sample. Results: Three patients showed a complex pattern of rise and fall of HbSSG levels in different time periods before, during and after surgery. They also showed statistically significant differences between peripheral and jugular blood, with mean HbSSG levels in jugular blood higher by approx. 30% than those of peripheral blood at the end of the period of carotid clamping. In all three patients HbSSG levels fell to pre-clamping values within 2 min from removal of carotid artery clamp. Conclusion: Although effective routine drug management allowed brain safety during carotid clamping time, a number of patients showed a fast modification over time of the HbSSG levels in jugular blood, suggesting that "resident" cerebral biochemical protection mechanisms could play some role to compensate clinically silent brain oxidative stress.
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OBJECTIVES: Assessment of maternal plasma amino acids during normal gestation and in early stages of intrauterine growth restriction (IUGR). STUDY DESIGN: Plasma amino acid concentrations were measured in: (1) non-pregnant women (n=7); (2) normal pregnant women in the first (n=13), second (n=17) and third (n=12) trimester; and (3) pregnant women in the first trimester with later development of IUGR (n=8). Amino acid levels were quantified by electrochemical detection in a reversed-phase high-performance liquid chromatography (HPLC) system. RESULTS: The levels of most essential and non-essential amino acids changed markedly in the first trimester during normal pregnancy and thereafter remained almost constant. In the first trimester of IUGR, a number of both essential and non-essential amino acids were significantly different from those observed in normal pregnancies, with values more similar to those observed in non-pregnant women. CONCLUSIONS: Levels of most maternal amino acids decrease and some increase during early gestation reflecting a metabolic adaptation that occurs in normal pregnancies. Pregnancies that later develop IUGR show a lack of these adaptations for a significant number of both essential and non-essential amino acids, suggesting a lack of adaptation.
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Aminoácidos/sangre , Retardo del Crecimiento Fetal/sangre , Adaptación Fisiológica , Cromatografía Líquida de Alta Presión , Femenino , Humanos , Embarazo , Primer Trimestre del Embarazo , Valores de ReferenciaRESUMEN
Protein folding in the endoplasmic reticulum (ER) often involves the formation of disulfide bonds. The oxidizing conditions required within this organelle were shown to be maintained through the release of small thiols, mainly cysteine and glutathione. Thiol secretion was stimulated when proteins rich in disulfide bonds were translocated into the ER, and secretion was prevented by the inhibition of protein synthesis. Endogenously generated cysteine and glutathione counteracted thiol-mediated retention in the ER and altered the extracellular redox. The secretion of thiols might link disulfide bond formation in the ER to intra- and intercellular redox signaling.
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Cisteína/metabolismo , Disulfuros/metabolismo , Retículo Endoplásmico/metabolismo , Glutatión/metabolismo , Proteínas/metabolismo , Animales , Benzopiranos/metabolismo , Brefeldino A , Cicloheximida/farmacología , Ciclopentanos/farmacología , Cistina/metabolismo , Exocitosis , Glutatión/análogos & derivados , Disulfuro de Glutatión , Aparato de Golgi/metabolismo , Glicoproteínas Hemaglutininas del Virus de la Influenza/biosíntesis , Inmunoglobulina M/biosíntesis , Cadenas lambda de Inmunoglobulina/metabolismo , Oocitos , Oxidación-Reducción , Inhibidores de la Síntesis de la Proteína/farmacología , Temperatura , Células Tumorales Cultivadas , Xenopus laevisRESUMEN
Thiol-dependent retention mechanisms involving the microsecond chain Cys575 ensure that only polymeric IgM are secreted. B lymphocytes are unable to polymerize IgM and degrade unpolymerized precursors intracellularly. Since several non-lymphoid transfectants secrete hexameric IgM, specific mechanism(s) inhibiting IgM polymerization/secretion may be active in B cells. Here, we show that Xenopus laevis oocytes are also unable to polymerize IgM and retain this isotype via Cys575 as efficiently as B cells. The mechanisms and the hierarchy of the thiol-dependent pre-Golgi retention are conserved in amphibian oocytes, as indicated by the efficient retention of secretory IgA and the slow secretion of unassembled J558 lambda chains. We also show that B cells do not lack any structural component necessary to polymerize IgM: after retention has been weakened by 2-mercaptoethanol, polymerization can occur if oxidizing conditions are restored. Since release from retention can result in polymerization, stringent retention in B cells and oocytes might be at the basis of their common inability to polymerize secretory IgM. Our findings suggest that disulfide interchange reactions in the exocytic compartment can be modulated during B cell differentiation to control IgM secretion.
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Linfocitos B/metabolismo , Inmunoglobulina M/metabolismo , Oocitos/metabolismo , Compuestos de Sulfhidrilo/metabolismo , Animales , Linfocitos B/inmunología , Línea Celular , Cisteína/fisiología , Glicósido Hidrolasas/farmacología , Humanos , Inmunoglobulina A/efectos de los fármacos , Inmunoglobulina A/metabolismo , Inmunoglobulina M/biosíntesis , Inmunoglobulina M/efectos de los fármacos , Cadenas lambda de Inmunoglobulina/metabolismo , Linfoma de Células B , Oocitos/inmunología , Células Plasmáticas/metabolismo , Polímeros , ARN/biosíntesis , Sustancias Reductoras/farmacología , Especificidad por Sustrato , Células Tumorales Cultivadas , Xenopus laevisRESUMEN
Secreted glycoproteins generally contain oligosaccharides of the complex type. However, several molecules have been described in which individual glycans are processed differently from one another. Folding, assembly and oligomerization could affect the maturation of certain glycans by hindering them to the Golgi processing machinery. We have tested this possibility by analysing a panel of engineered murine mu chains secreted as mu2L2 monomers or as polymers, and having or not the carboxy-terminal glycan (Asn563). In secreted IgM polymers, Asn563 bears high-mannose oligosaccharides, typical of endoplasmic reticulum resident proteins, while complex sugars are found at the other four sites (Brenckle and Kornfeld, 1980 Arch. Biochem. Biophys. 243, 605-618). Polymeric and monomeric IgM contain mu chains whose glycans are processed differently. We show here that this is mainly due to the differential processing at the Asn563 glycan, which undergoes Golgi-mediated processing when IgM are secreted in the monomeric form. These results indicate that the oligomerization-dependent accessibility to the sugar modifying enzymes can be one of the key features that dictate the extent of oligosaccharide processing in multimeric glycoproteins. The presence of high mannose glycans at Asn563 implies that IgM polymerization takes place before encountering mannosidase II, likely in a pre-Golgi compartment.
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Glicosilación , Inmunoglobulina M/metabolismo , Cadenas mu de Inmunoglobulina/metabolismo , Secuencia de Aminoácidos , Animales , Asparagina/química , Aparato de Golgi/metabolismo , Ratones , Datos de Secuencia Molecular , Polímeros , Polisacáridos/metabolismo , Procesamiento Proteico-PostraduccionalRESUMEN
This paper investigates the ceramide composition of the psoriatic scale compared with that of normal human SC. A method was optimalized, based on TLC separation followed by densitometry, allowing the provision of good resolution and quantification of ceramide fractions from both normal and pathological specimens. Seven ceramide fractions were isolated and submitted to compositional analysis. The obtained results suggested a revisitation of previous ceramide designation. Therefore a simple classification is suggested, based on grouping ceramides carrying structural similarities under common codes. According to these rules, ceramides were grouped into five classes designated as: (1) Cer[EOS], which contains ester-linked fatty acids, omega-OH fatty acids and sphingosines; (2) Cer[NS], which contains non-OH fatty acids and sphingosines; (3) Cer[NP], which contains non-OH fatty acids and phytosphingosines; (4) Cer[AS], which contains alpha-OH fatty acids and sphingosines; (5) Cer[AP], which contains alpha-OH fatty acids and phytosphingosines. Analysis of ceramides from the psoriatic scale, compared to those from normal human SC, resulted in an impairment of the Cer[EOS] content as well as of the ceramides containing phytosphingosine, with concurrent increase in ceramides containing sphingosine, being the total amount maintained identical. Since one of the suggested pathways for phytosphingosine biosynthesis involves the water addition to the corresponding sphingosine double bond, we can speculate that the observed alteration is due to a deranged water bioavailability, associated with psoriasis.