A long-term cohort study: the immune evasion and decreasing neutralization dominated the SARS-CoV-2 breakthrough infection.

Most of vaccinees and COVID-19 convalescents can build effective anti-SARS-CoV-2 humoral immunity, which helps preventing infection and alleviating symptoms. However, breakthrough viral infections caused by emerging SARS-CoV-2 variants, especially Omicron subvariants, still pose a serious threat to global health. By monitoring the viral infections and the sera neutralization ability of a long-tracked cohort, we found out that the immune evasion of emerging Omicron subvariants and the decreasing neutralization led to the mini-wave of SARS-CoV-2 breakthrough infections. Meanwhile, no significant difference had been found in the infectivity of tested SARS-CoV-2 variants, even though the affinity between human angiotensin-converting enzyme 2 (hACE2) and receptor-binding domain (RBDs) of tested variants showed an increasing trend. Notably, the immune imprinting of inactivated COVID-19 vaccine can be relieved by infections of BA.5.2 and XBB.1.5 variants sequentially. Our data reveal the rising reinfection risk of immune evasion variants like Omicron JN.1 in China, suggesting the importance of booster with updated vaccines.

Influence of abdominal fat distribution and inflammatory status on post-operative prognosis in non-small cell lung cancer patients: a retrospective cohort study.

PURPOSE: To evaluate the influence of visceral fat area (VFA), subcutaneous fat area (SFA), the systemic immune-inflammation index (SII) and total inflammation-based systemic index (AISI) on the postoperative prognosis of non-small cell lung cancers (NSCLC) patients. METHODS: 266 NSCLC patients received surgery from two academic medical centers were included. To assess the effect of abdominal fat measured by computed tomography (CT) imaging and inflammatory indicators on patients' overall survival (OS) and progression-free survival (PFS), Kaplan-Meier survival analysis and Cox proportional hazards models were used. RESULTS: Kaplan-Meier analysis showed the OS and PFS of patients in high-VFA group was better than low-VFA group (p < 0.05). AISI and SII were shown to be risk factors for OS and PFS (p < 0.05) after additional adjustment for BMI (Cox regression model II). After further adjustment for VFA (Cox regression model III), low-SFA group had longer OS (p < 0.05). Among the four subgroups based on VFA (high/low) and SFA (high/low) (p < 0.05), the high-VFA & low-SFA group had the longest median OS (108 months; 95% CI 74-117 months) and PFS (85 months; 95% CI 65-117 months), as well as the lowest SII and AISI (p < 0.05). Low-SFA was a protective factor for OS with different VFA stratification (p < 0.05). CONCLUSION: VFA, SFA, SII and AISI may be employed as significant prognostic markers of postoperative survival in NSCLC patients. Moreover, excessive SFA levels may encourage systemic inflammation decreasing the protective impact of VFA, which may help to provide targeted nutritional support and interventions for postoperative NSCLC patients with poor prognosis.

An optimized high-throughput SARS-CoV-2 dual reporter trans-complementation system for antiviral screening in vitro and in vivo.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is still epidemic around the world. The manipulation of SARS-CoV-2 is restricted to biosafety level 3 laboratories (BSL-3). In this study, we developed a SARS-CoV-2 ΔN-GFP-HiBiT replicon delivery particles (RDPs) encoding a dual reporter gene, GFP-HiBiT, capable of producing both GFP signal and luciferase activities. Through optimal selection of the reporter gene, GFP-HiBiT demonstrated superior stability and convenience for antiviral evaluation. Additionally, we established a RDP infection mouse model by delivering the N gene into K18-hACE2 KI mouse through lentivirus. This mouse model supports RDP replication and can be utilized for in vivo antiviral evaluations. In summary, the RDP system serves as a valuable tool for efficient antiviral screening and studying the gene function of SARS-CoV-2. Importantly, this system can be manipulated in BSL-2 laboratories, decreasing the threshold of experimental requirements.

Development and validation of MRI-based scoring models for predicting placental invasiveness in high-risk women for placenta accreta spectrum.

OBJECTIVES: To develop and validate MRI-based scoring models for predicting placenta accreta spectrum (PAS) invasiveness. MATERIALS AND METHODS: This retrospective study comprised a derivation cohort and a validation cohort. The derivation cohort came from a systematic review of published studies evaluating the diagnostic performance of MRI signs for PAS and/or placenta percreta in high-risk women. The significant signs were identified and used to develop prediction models for PAS and placenta percreta. Between 2016 and 2021, consecutive high-risk pregnant women for PAS who underwent placental MRI constituted the validation cohort. Two radiologists independently evaluated the MRI signs. The reference standard was intraoperative and pathologic findings. The predictive ability of MRI-based models was evaluated using the area under the curve (AUC). RESULTS: The derivation cohort included 26 studies involving 2568 women and the validation cohort consisted of 294 women with PAS diagnosed in 258 women (88%). Quantitative meta-analysis revealed that T2-dark bands, placental/uterine bulge, loss of T2 hypointense interface, bladder wall interruption, placental heterogeneity, and abnormal intraplacental vascularity were associated with both PAS and placenta percreta, and myometrial thinning and focal exophytic mass were exclusively associated with PAS. The PAS model was validated with an AUC of 0.90 (95% CI: 0.86, 0.93) for predicting PAS and 0.85 (95% CI: 0.79, 0.90) for adverse peripartum outcome; the placenta percreta model showed an AUC of 0.92 (95% CI: 0.86, 0.98) for predicting placenta percreta. CONCLUSION: MRI-based scoring models established based on quantitative meta-analysis can accurately predict PAS, placenta percreta, and adverse peripartum outcome. CLINICAL RELEVANCE STATEMENT: These proposed MRI-based scoring models could help accurately predict PAS invasiveness and provide evidence-based risk stratification in the management of high-risk pregnant women for PAS. KEY POINTS: • Accurately identifying placenta accreta spectrum (PAS) and assessing its invasiveness depending solely on individual MRI signs remained challenging. • MRI-based scoring models, established through quantitative meta-analysis of multiple MRI signs, offered the potential to predict PAS invasiveness in high-risk pregnant women. • These MRI-based models allowed for evidence-based risk stratification in the management of pregnancies suspected of having PAS.

Alerting network, cognitive flexibility in children with attention deficit hyperactivity disorder and the moderating effect of neuroticism.

Attention deficit hyperactivity disorder (ADHD) is a prevalent neurodevelopmental disorder, and cognitive flexibility is a sub-component of executive functioning. Studies have shown impairments in cognitive flexibility in ADHD, which is affected by attentional processes. Personality, as a long-standing trait, has a profound effect on ADHD. However, previous studies have not assessed the relationship between attentional function, personality traits, and cognitive flexibility in children with ADHD. This study explored the association between attention networks, personality, and cognitive flexibility in ADHD, filling a gap in the related field. We expect our findings will provide insights into and clues for the prevention and interventional treatment of ADHD. This study primarily aimed to analyze differences in cognitive flexibility between individuals with ADHD and those without and further examine associations between attention networks, personality, and cognitive flexibility in children with ADHD. Overall, 55 children aged 7 to 11 years diagnosed with ADHD and 40 children without ADHD participated in this study. Cognitive flexibility, personality traits, and attentional networks were assessed using the Wisconsin Card Sorting Test, Eysenck Personality Questionnaire, and Attention Network Test, respectively. Additionally, the association between personality traits and strong attentional functioning and cognitive flexibility was investigated using multiple regression analysis. Children with ADHD had significant deficits in cognitive flexibility. A multiple regression analysis revealed that the alerting effect was highly associated with cognitive flexibility at high levels of neuroticism. This association was not salient in individuals with low levels of neuroticism. This study demonstrated that the ADHD group experienced lower cognitive flexibility than the control group. In addition, we showed the effect of neuroticism and alerting networks on cognitive flexibility. These findings may help psychiatrists provide intervention strategies to mitigate the impairment of social functioning in ADHD with cognitive spirituality deficits.

Field observations of C2 and C3 organosulfates and insights into their formation mechanisms at a suburban site in Hong Kong.

Organosulfates (OSs) are formed from volatile organic compounds (VOCs) and their oxidation products in the presence of sulfate particles. While OSs represent an important component in secondary organic aerosol, the knowledge of their formation driving force, mechanisms, and environmental impact remain inadequately understood. In this study, we report ambient observations of C2-3 oxygenated VOCs derived OSs (C2-3 OSs) at a suburban location of Hong Kong during autumn 2016. The C2-3 OSs, including glycolaldehyde sulfate (GS), hydroxyacetone sulfate (HAS), glycolic acid sulfate (GAS), and lactic acid sulfate (LAS), were quantified/semi-quantified using offline liquid chromatography-mass spectrometry analysis of aerosol filter samples. The average sum concentration of C2-3 OSs was 36 ng/m3. Correlation analysis revealed that sulfate, surface area, and liquid water content were important factors influencing C2-3 OS formation. Online measurement with an iodide High-Resolution Time-of-Flight Chemical-Ionization Mass Spectrometer (HR-ToF-CIMS) coupled with the Filter Inlet for Gases and AEROsols (FIGAERO) was also conducted to monitor C2-3 OSs, and their potential oxygenated VOC precursors in both gas- and particle-phase, and aerosol acidity tracer simultaneously. Our measurements support that glycolaldehyde/glyoxal, hydroxyacetone, glycolic acid/glyoxal, and lactic acid/methylglyoxal are likely precursors for GS, HAS, GAS, and LAS, respectively. Additionally, we found strong correlation between C2-3 OSs and H3S2O8-, a marker for aerosol acidity, providing field observational evidence for acid-catalyzed formation of small OSs. Based on both online and offline measurements, acid-catalyzed formation mechanisms in particle/aqueous phase are proposed. Specifically, the unique structure of adjacent carbonyl and hydroxyl groups in the C2-3 oxygenated VOC precursors can facilitate the formation of (1) a five-member ring intermediate via intramolecular hydrogen bond to react with sulfur trioxide through heterogenous reaction or (2) cyclic sulfate intermediate via particle-phase reaction with sulfuric acid to generate C2-3 OSs. These proposed mechanisms provide an alternative pathway for the liquid-phase production of C2-3 OSs.

Broadly neutralizing antibodies derived from the earliest COVID-19 convalescents protect mice from SARS-CoV-2 variants challenge.

Coronavirus disease 2019 (COVID-19) was first reported three years ago, when a group of individuals were infected with the original SARS-CoV-2 strain, based on which vaccines were developed. Here, we develop six human monoclonal antibodies (mAbs) from two elite convalescents in Wuhan and show that these mAbs recognize diverse epitopes on the receptor binding domain (RBD) and can inhibit the infection of SARS-CoV-2 original strain and variants of concern (VOCs) to varying degrees, including Omicron strains XBB and XBB.1.5. Of these mAbs, the two most broadly and potently neutralizing mAbs (7B3 and 14B1) exhibit prophylactic activity against SARS-CoV-2 WT infection and therapeutic effects against SARS-CoV-2 Delta variant challenge in K18-hACE2 KI mice. Furthermore, post-exposure treatment with 7B3 protects mice from lethal Omicron variants infection. Cryo-EM analysis of the spike trimer complexed with 14B1 or 7B3 reveals that these two mAbs bind partially overlapped epitopes onto the RBD of the spike, and sterically disrupt the binding of human angiotensin-converting enzyme 2 (hACE2) to RBD. Our results suggest that mAbs with broadly neutralizing activity against different SARS-CoV-2 variants are present in COVID-19 convalescents infected by the ancestral SARS-CoV-2 strain, indicating that people can benefit from former infections or vaccines despite the extensive immune escape of SARS-CoV-2.

NETosis promotes chronic inflammation and fibrosis in systemic lupus erythematosus and COVID-19.

Pulmonary fibrosis, a serious complication of systemic lupus erythematosus (SLE) and coronavirus disease 2019 (COVID-19), leads to irreversible lung damage. However, the underlying mechanism of this condition remains unclear. In this study, we revealed the landscape of transcriptional changes in lung biopsies from individuals with SLE, COVID-19-induced pulmonary fibrosis, and idiopathic pulmonary fibrosis (IPF) using histopathology and RNA sequencing, respectively. Despite the diverse etiologies of these diseases, lung expression of matrix metalloproteinase genes in these diseases showed similar patterns. Particularly, the differentially expressed genes were significantly enriched in the pathway of neutrophil extracellular trap formation, showing similar enrichment signature between SLE and COVID-19. The abundance of Neutrophil extracellular traps (NETs) was much higher in the lungs of individuals with SLE and COVID-19 compared to those with IPF. In-depth transcriptome analyses revealed that NETs formation pathway promotes epithelial-mesenchymal transition (EMT). Furthermore, stimulation with NETs significantly up-regulated α-SMA, Twist, Snail protein expression, while decreasing the expression of E-cadherin protein in vitro. This indicates that NETosis promotes EMT in lung epithelial cells. Given drugs that are efficacious in degrading damaged NETs or inhibiting NETs production, we identified a few drug targets that were aberrantly expressed in both SLE and COVID-19. Among these targets, the JAK2 inhibitor Tofacitinib could effectively disrupted the process of NETs and reversed NET-induced EMT in lung epithelial cells. These findings support that the NETs/EMT axis, activated by SLE and COVID-19, contributes to the progression of pulmonary fibrosis. Our study also highlights that JAK2 as a potential target for the treatment of fibrosis in these diseases.

PM1 exposure and spatial transmission of nicotine from the simulated second-hand vapor of pod-based electronic cigarettes.

Electronic cigarettes (E-cigarettes) have gained significant popularity in recent years as a substitute for combustible cigarettes. However, there is growing concern regarding the safety of E-cigarette products for both the users and those exposed passively to second-hand emissions, which contain nicotine and other toxic substances. In particular, the characteristics of second-hand PM1 exposure and the transmission of nicotine from E-cigarettes remain unclear. In this study, the untrapped mainstream aerosols from the E-cigarette and smoke from cigarettes were exhausted by the smoking machines which were operated under standardized puffing regimes to simulate second-hand vapor or smoke exposure. The concentrations and components of PM1 released from cigarettes and E-cigarettes were compared under varying environmental conditions and regulated using a heating, ventilation, and air conditioning (HVAC) system. Additionally, the ambient nicotine concentrations and the size distribution of the generated aerosols were determined at different distances from the release source. Results showed that PM1 accounted for the highest proportion (98 %) of the released particulate matter (PM1, PM2.5, and PM10). The mass median aerodynamic diameter (MMAD) of cigarette smoke (0.5 ± 0.01 µm, geometric standard deviation (GSD) 1.97 ± 0.1) was smaller than that of E-cigarette aerosols (1.06 ± 0.14 µm, GSD 1.79 ± 0.19). The PM1 concentrations and chemical components were effectively reduced when the HVAC system was utilized. Nicotine concentrations in E-cigarette aerosols were comparable to those of combustible cigarette emissions when close to the exposure source (0 m), while they declined more rapidly than cigarette smoke emissions with increasing distance from the source. Furthermore, the maximum nicotine concentrations occurred in 1 µm and 0.5 µm particles in E-cigarette and cigarette emissions, respectively. These results provide a scientific basis for the assessment of E-cigarette and cigarette aerosol passive exposure risks, guiding the development of environmental and human health control measures for these products.

CT Radiomics to Predict Macrotrabecular-Massive Subtype and Immune Status in Hepatocellular Carcinoma.

Background Macrotrabecular-massive (MTM) subtype of hepatocellular carcinoma (HCC) is an aggressive variant associated with angiogenesis and immunosuppressive tumor microenvironment, which is expected to be noninvasively identified using radiomics approaches. Purpose To construct a CT radiomics model to predict the MTM subtype and to investigate the underlying immune infiltration patterns. Materials and Methods This study included five retrospective data sets and one prospective data set from three academic medical centers between January 2015 and December 2021. The preoperative liver contrast-enhanced CT studies of 365 adult patients with resected HCC were evaluated. The Third Xiangya Hospital of Central South University provided the training set and internal test set, while Yueyang Central Hospital and Hunan Cancer Hospital provided the external test sets. Radiomic features were extracted and used to develop a radiomics model with machine learning in the training set, and the performance was verified in the two test sets. The outcomes cohort, including 58 adult patients with advanced HCC undergoing transarterial chemoembolization and antiangiogenic therapy, was used to evaluate the predictive value of the radiomics model for progression-free survival (PFS). Bulk RNA sequencing of tumors from 41 patients in The Cancer Genome Atlas (TCGA) and single-cell RNA sequencing from seven prospectively enrolled participants were used to investigate the radiomics-related immune infiltration patterns. Area under the receiver operating characteristics curve of the radiomics model was calculated, and Cox proportional regression was performed to identify predictors of PFS. Results Among 365 patients (mean age, 55 years ± 10 [SD]; 319 men) used for radiomics modeling, 122 (33%) were confirmed to have the MTM subtype. The radiomics model included 11 radiomic features and showed good performance for predicting the MTM subtype, with AUCs of 0.84, 0.80, and 0.74 in the training set, internal test set, and external test set, respectively. A low radiomics model score relative to the median value in the outcomes cohort was independently associated with PFS (hazard ratio, 0.4; 95% CI: 0.2, 0.8; P = .01). The radiomics model was associated with dysregulated humoral immunity involving B-cell infiltration and immunoglobulin synthesis. Conclusion Accurate prediction of the macrotrabecular-massive subtype in patients with hepatocellular carcinoma was achieved using a CT radiomics model, which was also associated with defective humoral immunity. Published under a CC BY 4.0 license. Supplemental material is available for this article. See also the editorial by Yoon and Kim in this issue.

Deep white matter hyperintensity is spatially correlated to MRI-visible perivascular spaces in cerebral small vessel disease on 7 Tesla MRI.

BACKGROUND: The association between perivascular space (PVS) and white matter hyperintensity (WMH) has been unclear. Normal-appearing white matter (NAWM) around WMH is also found correlated with the development of focal WMH. This study aims to investigate the topological connections among PVS, deep WMH (dWMH) and NAWM around WMH using 7 Tesla (7T) MRI. METHODS: Thirty-two patients with non-confluent WMHs and 16 subjects without WMHs were recruited from our department and clinic. We compared the PVS burden between patients with and without WMHs using a 5-point scale. Then, the dilatation and the number of PVS within a radius of 1 cm around each dWMH were compared with those of a reference site (without WMH) in the contralateral hemisphere. In this study, we define NAWM as an area within the radius of 1 cm around each dWMH. Furthermore, we assessed the spatial relationship between dWMH and PVS. RESULTS: Higher PVS scores in the centrum semiovale were found in patients with >5 dWMHs (median 3) than subjects without dWMH (median 2, p = 0.014). We found there was a greater dilatation and a higher number of PVS in NAWM around dWMH than at the reference sites (p<0.001, p<0.001). In addition, 79.59% of the dWMHs were spatially connected with PVS. CONCLUSION: dWMH, NAWM surrounding WMH and MRI-visible PVS are spatially correlated in the early stage of cerebral small vessel disease. Future study of WMH and NAWM should not overlook MRI-visible PVS.

Association of Serum Interleukin-8 and Serum Amyloid A With Anxiety Symptoms in Patients With Cerebral Small Vessel Disease.

Objective: Cerebral small vessel disease (CSVD) is a clinical syndrome caused by pathological changes in small vessels. Anxiety is a common symptom of CSVD. Previous studies have reported the association between inflammatory factors and anxiety in other diseases, but this association in patients with CSVD remains uncovered. Our study aimed to investigate whether serum inflammatory factors correlated with anxiety in patients with CSVD. Methods: A total of 245 CSVD patients confirmed using brain magnetic resonance imaging (MRI) were recruited from December 2019 to December 2021. Hamilton Anxiety Rating Scale (HAMA) was used to assess the anxiety symptoms of CSVD patients. Patients with HAMA scores ≥7 were considered to have anxiety symptoms. The serum levels of interleukin-1ß (IL-1ß), IL-2R, IL-6, IL-8, IL-10, tumor necrosis factor-α (TNF-α), serum amyloid A (SAA), C-reactive protein (CRP), high-sensitivity C-reactive protein (hs-CRP) and erythrocyte sedimentation rate (ESR) were detected. We compared levels of inflammatory factors between the anxiety and non-anxiety groups. Logistic regression analyses examined the correlation between inflammatory factors and anxiety symptoms. We further performed a gender subgroup analysis to investigate whether this association differed by gender. Results: In the fully adjusted multivariate logistic regression analysis model, we found that lower levels of IL-8 were linked to a higher risk of anxiety symptoms. Moreover, higher levels of SAA were linked to a lower risk of anxiety symptoms. Our study identified sex-specific effects, and the correlation between IL-8 and anxiety symptoms remained significant among males, while the correlation between SAA and anxiety symptoms remained significant among females. Conclusions: In this study, we found a suggestive association between IL-8, SAA, and anxiety symptoms in CSVD participants. Furthermore, IL-8 and SAA may have a sex-specific relationship with anxiety symptoms.

Assessment of Solid Pulmonary Nodules or Masses Using Zero Echo Time MR Lung Imaging: A Prospective Head-to-Head Comparison With CT.

Objective: The aim of this study is to determine the potential of zero echo time (ZTE) MR lung imaging in the assessment of solid pulmonary nodules or masses and diagnostic consistency to CT in terms of morphologic characterization. Methods: Our Institutional Review Board approved this prospective study. Seventy-one patients with solid pulmonary nodules or masses larger than 1 cm in diameter confirmed by chest CT were enrolled and underwent further lung ZTE-MRI scans within 7 days. ZTE-MRI and CT images were compared in terms of image quality and imaging features. Unidimensional diameter and three-dimensional volume measurements on both modalities were manually measured and compared using the Wilcoxon signed-rank test, intraclass correlation coefficient (ICC), Pearson's correlation analysis, and Bland-Altman analysis. Multivariable logistic regression analysis was used to identify the factors associated with significant inter-modality variation of volume. Results: Fifty-four of 71 (76.1%) patients were diagnosed with lung cancer. Subjective image quality was superior in CT compared with ZTE-MRI (p < 0.001). Inter-modality agreement for the imaging features was moderate for emphysema (kappa = 0.50), substantial for fibrosis (kappa = 0.76), and almost perfect (kappa = 0.88-1.00) for the remaining features. The size measurements including diameter and volume between ZTE-MRI and CT showed no significant difference (p = 0.36 for diameter and 0.60 for volume) and revealed perfect inter-observer (ICC = 0.975-0.980) and inter-modality (ICC = 0.942-0.992) agreements. Multivariable analysis showed that non-smooth margin [odds ratio (OR) = 6.008, p = 0.015] was an independent predictor for the significant inter-modality variation of volume. Conclusion: ZTE lung imaging is feasible as a part of chest MRI in the assessment and surveillance for solid pulmonary nodules or masses larger than 1 cm, presenting perfect agreement with CT in terms of morphologic characterization.

B-Cell-Epitope-Based Fluorescent Quantum Dot Biosensors for SARS-CoV-2 Enable Highly Sensitive COVID-19 Antibody Detection.

A new antibody diagnostic assay with more rapid and robust properties is demanded to quantitatively evaluate anti-severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) immunity in a large population. Here, we developed a nanometer-scale fluorescent biosensor system consisting of CdSe-ZnS quantum dots (QDs) coupled with the highly sensitive B-cell epitopes of SARS-CoV-2 that could remarkably identify the corresponding antibody with a detection limit of 100 pM. Intriguingly, we found that fluorescence quenching of QDs was stimulated more obviously when coupled with peptides than the corresponding proteins, indicating that the energy transfer between QDs and peptides was more effective. Compared to the traditional enzyme-linked immunosorbent assay (ELISA), the B-cell-epitope-based QD-biosensor could robustly distinguish coronavirus disease 2019 (COVID-19) antibody-positive patients from uninfected individuals with a higher sensitivity (92.3-98.1% positive rates by QD-biosensor vs. 78.3-83.1% positive rates by ELISAs in 207 COVID-19 patients' sera) in a more rapid (5 min) and labor-saving manner. Taken together, the 'QD-peptides' biosensor provided a novel real-time, quantitative, and high-throughput method for clinical diagnosis and home-use tests.

Drastic decline in sera neutralization against SARS-CoV-2 Omicron variant in Wuhan COVID-19 convalescents.

Global concern has been raised by the emergence and rapid transmission of the heavily mutated SARS-CoV-2 Omicron variant (B.1.1.529). So far, the infection features and immune escape ability of the Omicron variant have not been extensively studied. Here, we produced the Omicron pseudovirus and compared its entry, membrane fusion, and immune escape efficiency with the original strain and the dominating Delta variant. We found the Omicron variant showed slightly higher infectivity than the Delta variant and a similar ability to compete with the Delta variant in using Angiotensin-converting enzyme 2 (ACE2) in a BHK21-ACE2 cell line. However, the Omicron showed a significantly reduced fusogenicity than the original strain and the Delta variant in both BHK21-ACE2 and Vero-E6 cells. The neutralization assay testing the Wuhan convalescents' sera one-year post-infection showed a more dramatic reduction (10.15 fold) of neutralization against the Omicron variant than the Delta variant (1.79 fold) compared with the original strain with D614G. Notably, immune-boosting through three vaccine shots significantly improved the convalescents' immunity against the Omicron variants. Our results reveal a reduced fusogenicity and a striking immune escape ability of the Omicron variant, highlighting the importance of booster shots against the challenge of the SARS-CoV-2 antigenic drift.

N7-Methylation of the Coronavirus RNA Cap Is Required for Maximal Virulence by Preventing Innate Immune Recognition.

The ongoing coronavirus (CoV) disease 2019 (COVID-19) pandemic caused by infection with severe acute respiratory syndrome CoV 2 (SARS-CoV-2) is associated with substantial morbidity and mortality. Understanding the immunological and pathological processes of coronavirus diseases is crucial for the rational design of effective vaccines and therapies for COVID-19. Previous studies showed that 2'-O-methylation of the viral RNA cap structure is required to prevent the recognition of viral RNAs by intracellular innate sensors. Here, we demonstrate that the guanine N7-methylation of the 5' cap mediated by coronavirus nonstructural protein 14 (nsp14) contributes to viral evasion of the type I interferon (IFN-I)-mediated immune response and pathogenesis in mice. A Y414A substitution in nsp14 of the coronavirus mouse hepatitis virus (MHV) significantly decreased N7-methyltransferase activity and reduced guanine N7-methylation of the 5' cap in vitro. Infection of myeloid cells with recombinant MHV harboring the nsp14-Y414A mutation (rMHVnsp14-Y414A) resulted in upregulated expression of IFN-I and ISG15 mainly via MDA5 signaling and in reduced viral replication compared to that of wild-type rMHV. rMHVnsp14-Y414A replicated to lower titers in livers and brains and exhibited an attenuated phenotype in mice. This attenuated phenotype was IFN-I dependent because the virulence of the rMHVnsp14-Y414A mutant was restored in Ifnar-/- mice. We further found that the comparable mutation (Y420A) in SARS-CoV-2 nsp14 (rSARS-CoV-2nsp14-Y420A) also significantly decreased N7-methyltransferase activity in vitro, and the mutant virus was attenuated in K18-human ACE2 transgenic mice. Moreover, infection with rSARS-CoV-2nsp14-Y420A conferred complete protection against subsequent and otherwise lethal SARS-CoV-2 infection in mice, indicating the vaccine potential of this mutant. IMPORTANCE Coronaviruses (CoVs), including SARS-CoV-2, the cause of COVID-19, use several strategies to evade the host innate immune responses. While the cap structure of RNA, including CoV RNA, is important for translation, previous studies indicate that the cap also contributes to viral evasion from the host immune response. In this study, we demonstrate that the N7-methylated cap structure of CoV RNA is pivotal for virus immunoevasion. Using recombinant MHV and SARS-CoV-2 encoding an inactive N7-methyltransferase, we demonstrate that these mutant viruses are highly attenuated in vivo and that attenuation is apparent at very early times after infection. Virulence is restored in mice lacking interferon signaling. Further, we show that infection with virus defective in N7-methylation protects mice from lethal SARS-CoV-2, suggesting that the N7-methylase might be a useful target in drug and vaccine development.

Emerging SARS-CoV-2 variants: Why, how, and what's next?

The emergence of the SARS-CoV-2 Omicron variant poses a striking threat to human society. More than 30 mutations in the Spike protein of the Omicron variant severely compromised the protective immunity elicited by either vaccination or prior infection. The persistent viral evolutionary trajectory generates Omicron-associated lineages, such as BA.1 and BA.2. Moreover, the virus recombination upon Delta and Omicron co-infections has been reported lately, although the impact remains to be assessed. This minireview summarizes the characteristics, evolution and mutation control, and immune evasion mechanisms of SARS-CoV-2 variants, which will be helpful for the in-depth understanding of the SARS-CoV-2 variants and policy-making related to COVID-19 pandemic control.

Antibody neutralization to SARS-CoV-2 and variants after 1 year in Wuhan, China.

Most COVID-19 convalescents can build effective anti-SARS-CoV-2 humoral immunity, but it remains unclear how long it can maintain and how efficiently it can prevent the reinfection of the emerging SARS-CoV-2 variants. Here, we tested the sera from 248 COVID-19 convalescents around 1 year post-infection in Wuhan, the earliest known epicenter. SARS-CoV-2 immunoglobulin G (IgG) was well maintained in most patients and potently neutralizes the infection of the original strain and the B.1.1.7 variant. However, varying degrees of immune escape was observed on the other tested variants in a patient-specific manner, with individuals showing remarkably broad neutralization potency. The immune escape can be largely attributed to several critical spike mutations. These results suggest that SARS-CoV-2 can elicit long-lasting immunity but this is escaped by the emerging variants.

High On-Treatment Platelet Reactivity as Predictor of Long-term Clinical Outcomes in Stroke Patients with Antiplatelet Agents.

The purpose was to explore the value of high on-treatment platelet reactivity (HTPR) in predicting long-term clinical outcomes for stroke patients. The platelet reactivity was assayed after being treated with either 75 mg clopidogrel or 100 mg aspirin daily with VerifyNow System in stroke patients. HTPR for clopidogrel was defined as PRU ≥ 208, and that for aspirin was defined as ARU ≥ 550. CYP2C19 genotyping was performed using the Sequenom MassARRAY iPLEX platform. The primary endpoint was a composite of recurrent ischemic stroke, transient ischemic attack, myocardial infarction, or ischemic vascular death. The safety endpoint was bleeding. In the clopidogrel group, among 345 patients recruited, 174 of them were categorized as HTPR. A total of 270 patients were followed up for 54 months. There was a significant association between HTPR and the primary endpoint (HRadj 2.13 [95% CI, 1.43-3.15], p < 0.001). Among the 314 participants genotyped for CYP2C19, 187 (59.6%) were classified as CYP2C19 loss-of-function allele carriers. Patients with at least 1 loss-of-function allele were more likely to present with HTPR (ORadj 2.61 [95%CI, 1.43-4.77], p = 0.008), and had a higher risk of the primary endpoint (HRadj 2.05 [95% CI, 1.30, 3.25], p = 0.002). In the aspirin group, among 140 patients recruited, 28 of them were categorized as HTPR. A total of 121 patients were followed up for 30 months. Similarly, there was a significant association between HTPR and the primary endpoint (HRadj 3.28 [95% CI, 1.52-7.71], p = 0.002). HTPR is an independent risk factor for ischemic events during long-term follow-up in stroke patients. Platelet function testing is helpful to evaluate the effect of antiplatelet therapy for stroke patients.

The Protective Effect of Quercetin on Endothelial Cells Injured by Hypoxia and Reoxygenation.

Background: Cerebral small vessel disease (CSVD) is a group of clinical syndromes covering all pathological processes of small vessels in the brain, which can cause stroke and serious dementia. However, as the pathogenesis of CSVD is not clear, so the treatment is limited. Endothelial cell dysfunction is earlier than clinical symptoms, such as hypertension and leukosis. Therefore, the treatment of endothelial cells is expected to be a new breakthrough. Quercetin, a flavonoid present in a variety of plants, has the function of anti-inflammation and anti-oxidation. This study aimed to investigate the protective effect of quercetin on endothelial cell injury and provide a basic theory for subsequent application in the clinic. Methods: Human brain microvascular endothelial cells (HBMECs) were cultured in vitro, and the injury model of endothelial cells was established by hypoxia and reoxygenation (H/R). The protective effects of quercetin on HBMECs were studied from the perspectives of cell viability, cell migration, angiogenesis and apoptosis. In order to further study the mechanism of quercetin, oxidative stress and endoplasmic reticulum stress were analyzed. What's more, blood-brain barrier (BBB) integrity was also studied. Results: Quercetin can promote the viability, migration and angiogenesis of HBMECs, and inhibit the apoptosis. In addition, quercetin can also activate Keap1/Nrf2 signaling pathway, reduce ATF6/GRP78 protein expression. Further study showed that quercetin could increase the expression of Claudin-5 and Zonula occludens-1. Conclusions: Our experiments show that quercetin can protect HBMECs from H/R, which contains promoting cell proliferation, cell migration and angiogenesis, reducing mitochondrial membrane potential damage and inhibiting cell apoptosis. This may be related to its antioxidation and inhibition of endoplasmic reticulum stress. At the same time, quercetin can increase the level of BBB connexin, suggesting that quercetin can maintain BBB integrity.