RESUMEN
Severe COVID-19 is characterized by lung abnormalities, including the presence of syncytial pneumocytes. Syncytia form when SARS-CoV-2 spike protein expressed on the surface of infected cells interacts with the ACE2 receptor on neighboring cells. The syncytia forming potential of spike variant proteins remain poorly characterized. Here, we first assessed Alpha (B.1.1.7) and Beta (B.1.351) spread and fusion in cell cultures, compared with the ancestral D614G strain. Alpha and Beta replicated similarly to D614G strain in Vero, Caco-2, Calu-3, and primary airway cells. However, Alpha and Beta formed larger and more numerous syncytia. Variant spike proteins displayed higher ACE2 affinity compared with D614G. Alpha, Beta, and D614G fusion was similarly inhibited by interferon-induced transmembrane proteins (IFITMs). Individual mutations present in Alpha and Beta spikes modified fusogenicity, binding to ACE2 or recognition by monoclonal antibodies. We further show that Delta spike also triggers faster fusion relative to D614G. Thus, SARS-CoV-2 emerging variants display enhanced syncytia formation.
Asunto(s)
Enzima Convertidora de Angiotensina 2/metabolismo , Anticuerpos Monoclonales/farmacología , Células Gigantes/virología , Mutación , SARS-CoV-2/fisiología , Glicoproteína de la Espiga del Coronavirus/genética , Animales , Células CACO-2 , Línea Celular , Chlorocebus aethiops , Células Gigantes/efectos de los fármacos , Células Gigantes/metabolismo , Células HEK293 , Humanos , SARS-CoV-2/efectos de los fármacos , SARS-CoV-2/genética , Células Vero , Replicación Viral/efectos de los fármacosRESUMEN
BACKGROUND: Sleep apnea is a chronic, widely underdiagnosed condition characterized by disruption of sleep architecture and intermittent hypoxia due to short cessations of breathing. It is a major independent risk factor for myocardial infarction, congestive heart failure and stroke as well as one of the rare modifiable risk factors for Alzheimer's Dementia. Reliable animal disease models are needed to understand the link between sleep apnea and the various clinically linked disorders. NEW METHOD: An automated system for inducing hypoxia was developed, in which the major improvement was the possibility to efficiently adjust the length and intensity of hypoxia in two different periods. The chamber used a small volume of gas allowing for fast exchanges of different oxygen levels. The mice were kept in their cages adapted with the system on the cage lid. As a proof of principle, they were exposed to a three week period of intermittent hypoxia for 8 hours a day, with 90 s intervals of 5, 7% and 21% oxygen to validate the model. Treated (n = 8) and control mice (no hypoxia, n = 7) were handled in the same manner and their hippocampal brain regions compared by histology. RESULTS: The chamber provided a fast, reliable and precise intermittent hypoxia, without inducing noticeable side effects to the animals. The validation experiment showed that apoptotic neurons in the hippocampus were more numerous in the mice exposed to intermittent hypoxia than in the control group, in all tested hippocampal regions (cornu ammonis 1 (CA1) P <0.001; cornu ammonis 3 (CA3) P <0.001; and dentate gyrus (DG) P = 0.023). In both, control and hypoxic conditions, there was a significantly higher number of apoptotic neurons in the DG compared to the CA1 and CA3 subfields (P <0.001). CONCLUSION: The new design of a hypoxic chamber provides a fast, adjustable and reliable model of obstructive sleep apnea, which was validated by apoptosis of hippocampal neurons.