Differential pathogenic and molecular features in neurological infection of SARS-CoV-2 Omicron BA.5.2 and BA.2.75 and Delta.

The Coronavirus disease 2019 (COVID-19) pandemic caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) remains a global threat, exacerbated by the emergence of viral variants. Two variants of SARS-CoV-2, Omicron BA.2.75 and BA.5, led to global infection peaks between May 2022 and May 2023, yet their precise characteristics in pathogenesis are not well understood. In this study, we compared these two Omicron sublineages with the previously dominant Delta variant using a human angiotensin-converting enzyme 2 knock-in mouse model. As expected, Delta exhibited higher viral replication in the lung and brain than both Omicron sublineages which induced less severe lung damage and immune activation. In contrast, the Omicron variants especially BA.5.2 showed a propensity for cellular proliferation and developmental pathways. Both Delta and BA.5.2 variants, but not BA.2.75, led to decreased pulmonary lymphocytes, indicating differential adaptive immune response. Neuroinvasiveness was shared with all strains, accompanied by vascular abnormalities, synaptic injury, and loss of astrocytes. However, Immunostaining assays and transcriptomic analysis showed that BA.5.2 displayed stronger immune suppression and neurodegeneration, while BA.2.75 exhibited more similar characteristics to Delta in the cortex. Such differentially infectious features could be partially attributed to the weakened interaction between Omicron Spike protein and host proteomes decoded via co-immunoprecipitation followed by mass spectrometry in neuronal cells. Our present study supports attenuated replication and pathogenicity of Omicron variants but also highlights their newly infectious characteristics in the lung and brain, especially with BA.5.2 demonstrating enhanced immune evasion and neural damage that could exacerbate neurological sequelae.

Primate-specific BTN3A2 protects against SARS-CoV-2 infection by interacting with and reducing ACE2.

BACKGROUND: Coronavirus disease 2019 (COVID-19) is an immune-related disorder caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The complete pathogenesis of the virus remains to be determined. Unraveling the molecular mechanisms governing SARS-CoV-2 interactions with host cells is crucial for the formulation of effective prophylactic measures and the advancement of COVID-19 therapeutics. METHODS: We analyzed human lung single-cell RNA sequencing dataset to discern the association of butyrophilin subfamily 3 member A2 (BTN3A2) expression with COVID-19. The BTN3A2 gene edited cell lines and transgenic mice were infected by live SARS-CoV-2 in a biosafety level 3 (BSL-3) laboratory. Immunoprecipitation, flow cytometry, biolayer interferometry and competition ELISA assays were performed in BTN3A2 gene edited cells. We performed quantitative real-time PCR, histological and/or immunohistochemical analyses for tissue samples from mice with or without SARS-CoV-2 infection. FINDINGS: The BTN3A2 mRNA level was correlated with COVID-19 severity. BTN3A2 expression was predominantly identified in epithelial cells, elevated in pathological epithelial cells from COVID-19 patients and co-occurred with ACE2 expression in the same lung cell subtypes. BTN3A2 targeted the early stage of the viral life cycle by inhibiting SARS-CoV-2 attachment through interactions with the receptor-binding domain (RBD) of the Spike protein and ACE2. BTN3A2 inhibited ACE2-mediated SARS-CoV-2 infection by reducing ACE2 in vitro and in vivo. INTERPRETATION: These results reveal a key role of BTN3A2 in the fight against COVID-19. Identifying potential monoclonal antibodies which mimic BTN3A2 may facilitate disruption of SARS-CoV-2 infection, providing a therapeutic avenue for COVID-19. FUNDING: This study was supported by the National Natural Science Foundation of China (32070569, U1902215, and 32371017), the CAS "Light of West China" Program, and Yunnan Province (202305AH340006).

Specific inhibition of the NLRP3 inflammasome suppresses immune overactivation and alleviates COVID-19 like pathology in mice.

BACKGROUND: The Coronavirus Disease 2019 (COVID-19) pandemic has been a great threat to global public health since 2020. Although the advance on vaccine development has been largely achieved, a strategy to alleviate immune overactivation in severe COVID-19 patients is still needed. The NLRP3 inflammasome is activated upon SARS-CoV-2 infection and associated with COVID-19 severity. However, the processes by which the NLRP3 inflammasome is involved in COVID-19 disease remain unclear. METHODS: We infected THP-1 derived macrophages, NLRP3 knockout mice, and human ACE2 transgenic mice with live SARS-CoV-2 in Biosafety Level 3 (BSL-3) laboratory. We performed quantitative real-time PCR for targeted viral or host genes from SARS-CoV-2 infected mouse tissues, conducted histological or immunofluorescence analysis in SARS-CoV-2 infected mouse tissues. We also injected intranasally AAV-hACE2 or intraperitoneally NLRP3 inflammasome inhibitor MCC950 before SARS-CoV-2 infection in mice as indicated. FINDINGS: We have provided multiple lines of evidence that the NLRP3 inflammasome plays an important role in the host immune response to SARS-CoV-2 invasion of the lungs. Inhibition of the NLRP3 inflammasome attenuated the release of COVID-19 related pro-inflammatory cytokines in cell cultures and mice. The severe pathology induced by SARS-CoV-2 in lung tissues was reduced in Nlrp3-/- mice compared to wild-type C57BL/6 mice. Finally, specific inhibition of the NLRP3 inflammasome by MCC950 alleviated excessive lung inflammation and thus COVID-19 like pathology in human ACE2 transgenic mice. INTERPRETATION: Inflammatory activation induced by SARS-CoV-2 is an important stimulator of COVID-19 related immunopathology. Targeting the NLRP3 inflammasome is a promising immune intervention against severe COVID-19 disease. FUNDING: This work was supported by grants from the Bureau of Frontier Sciences and Education, CAS (grant no. QYZDJ-SSW-SMC005 to Y.G.Y.), the key project of the CAS "Light of West China" Program (to D.Y.) and Yunnan Province (202001AS070023 to D.Y.).

Corrigendum: brain microvascular endothelial cell-derived HMGB1 facilitates monocyte adhesion and transmigration to promote JEV neuroinvasion.

[This corrects the article DOI: 10.3389/fcimb.2021.701820.].

Characteristics of replication and pathogenicity of SARS-CoV-2 Alpha and Delta isolates.

The continuously arising of SARS-CoV-2 variants has been posting a great threat to public health safety globally, from B.1.17 (Alpha), B.1.351 (Beta), P.1 (Gamma), B.1.617.2 (Delta) to B.1.1.529 (Omicron). The emerging or re-emerging of the SARS-CoV-2 variants of concern is calling for the constant monitoring of their epidemics, pathogenicity and immune escape. In this study, we aimed to characterize replication and pathogenicity of the Alpha and Delta variant strains isolated from patients infected in Laos. The amino acid mutations within the spike fragment of the isolates were determined via sequencing. The more efficient replication of the Alpha and Delta isolates was documented than the prototyped SARS-CoV-2 in Calu-3 and Caco-2 â€‹cells, while such features were not observed in Huh-7, Vero E6 and HPA-3 â€‹cells. We utilized both animal models of human ACE2 (hACE2) transgenic mice and hamsters to evaluate the pathogenesis of the isolates. The Alpha and Delta can replicate well in multiple organs and cause moderate to severe lung pathology in these animals. In conclusion, the spike protein of the isolated Alpha and Delta variant strains was characterized, and the replication and pathogenicity of the strains in the cells and animal models were also evaluated.

Brain Microvascular Endothelial Cell-Derived HMGB1 Facilitates Monocyte Adhesion and Transmigration to Promote JEV Neuroinvasion.

Infection with Japanese encephalitis virus (JEV) induces high morbidity and mortality, including potentially permanent neurological sequelae. However, the mechanisms by which viruses cross the blood-brain barrier (BBB) and invade into the central nervous system (CNS) remain unclear. Here, we show that extracellular HMGB1 facilitates immune cell transmigration. Furthermore, the migration of immune cells into the CNS dramatically increases during JEV infection which may enhance viral clearance, but paradoxically expedite the onset of Japanese encephalitis (JE). In this study, brain microvascular endothelial cells (BMECs) were utilized for the detection of HMGB1 release, and leucocyte, adhesion, and the integrity of the BBB in vitro. Genetically modified JEV-expressing EGFP (EGFP-JEV) and the BBB model were established to trace JEV-infected immune cell transmigration, which mimics the process of viral neuroinfection. We find that JEV causes HMGB1 release from BMECs while increasing adhesion molecules. Recombinant HMGB1 enhances leukocyte-endothelium adhesion, facilitating JEV-infected monocyte transmigration across endothelia. Thus, JEV successfully utilizes infected monocytes to spread into the brain, expanding inside of the brain, and leading to the acceleration of JE onset, which was facilitated by HMGB1. HMGB1-promoted monocyte transmigration may represent the mechanism of JEV neuroinvasion, revealing potential therapeutic targets.

Corrigendum: Brain Microvascular Endothelial Cell-Derived HMGB1 Facilitates Monocyte Adhesion and Transmigration to Promote JEV Neuroinvasion.

[This corrects the article DOI: 10.3389/fcimb.2021.701820.].

SARS-CoV-2 Mpro inhibitors with antiviral activity in a transgenic mouse model.

The COVID-19 pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) continually poses serious threats to global public health. The main protease (Mpro) of SARS-CoV-2 plays a central role in viral replication. We designed and synthesized 32 new bicycloproline-containing Mpro inhibitors derived from either boceprevir or telaprevir, both of which are approved antivirals. All compounds inhibited SARS-CoV-2 Mpro activity in vitro, with 50% inhibitory concentration values ranging from 7.6 to 748.5 nM. The cocrystal structure of Mpro in complex with MI-23, one of the most potent compounds, revealed its interaction mode. Two compounds (MI-09 and MI-30) showed excellent antiviral activity in cell-based assays. In a transgenic mouse model of SARS-CoV-2 infection, oral or intraperitoneal treatment with MI-09 or MI-30 significantly reduced lung viral loads and lung lesions. Both also displayed good pharmacokinetic properties and safety in rats.