SARS-CoV-2 infection causes periodontal fibrotic pathogenesis through deregulating mitochondrial beta-oxidation.

The global high prevalence of COVID-19 is a major challenge for health professionals and patients. SARS-CoV-2 virus has four structural protein components: the spike protein, envelope protein, membrane protein, and nucleocapsid protein. The SARS-CoV-2 virus mutates predominantly in the spike proteins, whilst the other key viral components usually remain stable. Essentially the pathological functions of the SARS-CoV-2 virus on different cell types are still largely unknown. Previous studies have shown that the human oral cavity can potentially act as reservoir of the SARS-CoV-2 virus. However, the consequence of SARS-CoV-2 viral infection on human oral health has not been systematically examined. COVID-19 can cause severe oral mucosa lesions and is likely to be connected with poor periodontal conditions. Fibroblasts are the major cell type inside periodontal ligament (PDL) and express the SARS-CoV-2 receptor: Angiotensin-converting enzyme 2 (ACE2), whose expression level can increase upon bacterial infection hence potentially provide a direct route of SARS-CoV-2 infection to PDL fibroblasts. In this research, we aimed to study the pathogenicity of SARS-CoV-2 viral components on human fibroblasts. We found that by exposing to SARS-CoV-2, especially to the viral envelope and membrane proteins, the human periodontal fibroblasts could develop fibrotic pathogenic phenotypes, including hyperproliferation that was simultaneously induced with increased apoptosis and senescence. The fibrotic degeneration was mediated by a down-regulation of mitochondrial ß-oxidation in the fibroblasts. Fatty acid ß-oxidation inhibitor, etomoxir treatment could mirror the same pathological consequence on the cells, similar to SARS-CoV-2 infection. Our results therefore provide novel mechanistic insights into how SARS-CoV-2 infection can affect human periodontal health at the cell and molecular level with potential new therapeutic targets for COVID-19 induced fibrosis.

Associations between brain gene expression perturbations implicated by COVID-19 and psychiatric disorders.

BACKGROUND: Currently, there is increasing evidence from clinic, epidemiology, as well as neuroimaging, demonstrating neuropsychiatric abnormalities in COVID-19, however, whether there were associations between brain changes caused by COVID-19 and genetic susceptibility of psychiatric disorders was still unknown. METHODS: In this study, we performed a meta-analysis to investigate these associations by combing single-cell RNA sequencing datasets of brain tissues of COVID-19 and genome-wide association study summary statistics of psychiatric disorders. RESULTS: The analysis demonstrated that among ten psychiatric disorders, gene expression perturbations implicated by COVID-19 in excitatory neurons of choroid plexus were significantly associated with schizophrenia. CONCLUSIONS: Our analysis might provide insights for the underlying mechanism of the psychiatric consequence of COVID-19.

Mechanism of an RBM-targeted rabbit monoclonal antibody 9H1 neutralizing SARS-CoV-2.

The COVID-19 pandemic, caused by SARS-CoV-2, has led to over 750 million infections and 6.8 million deaths worldwide since late 2019. Due to the continuous evolution of SARS-CoV-2, many significant variants have emerged, creating ongoing challenges to the prevention and treatment of the pandemic. Therefore, the study of antibody responses against SARS-CoV-2 is essential for the development of vaccines and therapeutics. Here we perform single particle cryo-electron microscopy (cryo-EM) structure determination of a rabbit monoclonal antibody (RmAb) 9H1 in complex with the SARS-CoV-2 wild-type (WT) spike trimer. Our structural analysis shows that 9H1 interacts with the receptor-binding motif (RBM) region of the receptor-binding domain (RBD) on the spike protein and by directly competing with angiotensin-converting enzyme 2 (ACE2), it blocks the binding of the virus to the receptor and achieves neutralization. Our findings suggest that utilizing rabbit-derived mAbs provides valuable insights into the molecular interactions between neutralizing antibodies and spike proteins and may also facilitate the development of therapeutic antibodies and expand the antibody library.

Associations between brain gene expression perturbations implicated by COVID-19 and psychiatric disorders

Background Currently, there is increasing evidence from clinic, epidemiology, as well as neuroimaging, demonstrating neuropsychiatric abnormalities in COVID-19, however, whether there were associations between brain changes caused by COVID-19 and genetic susceptibility of psychiatric disorders was still unknown. Methods In this study, we performed a meta-analysis to investigate these associations by combing single-cell RNA sequencing datasets of brain tissues of COVID-19 and genome-wide association study summary statistics of psychiatric disorders. Results The analysis demonstrated that among ten psychiatric disorders, gene expression perturbations implicated by COVID-19 in excitatory neurons of choroid plexus were significantly associated with schizophrenia. Conclusions Our analysis might provide insights for the underlying mechanism of the psychiatric consequence of COVID-19.

Mechanism of a rabbit monoclonal antibody broadly neutralizing SARS-CoV-2 variants.

Due to the continuous evolution of SARS-CoV-2, the Omicron variant has emerged and exhibits severe immune evasion. The high number of mutations at key antigenic sites on the spike protein has made a large number of existing antibodies and vaccines ineffective against this variant. Therefore, it is urgent to develop efficient broad-spectrum neutralizing therapeutic drugs. Here we characterize a rabbit monoclonal antibody (RmAb) 1H1 with broad-spectrum neutralizing potency against Omicron sublineages including BA.1, BA.1.1, BA.2, BA.2.12.1, BA.2.75, BA.3 and BA.4/5. Cryo-electron microscopy (cryo-EM) structure determination of the BA.1 spike-1H1 Fab complexes shows that 1H1 targets a highly conserved region of RBD and avoids most of the circulating Omicron mutations, explaining its broad-spectrum neutralization potency. Our findings indicate 1H1 as a promising RmAb model for designing broad-spectrum neutralizing antibodies and shed light on the development of therapeutic agents as well as effective vaccines against newly emerging variants in the future.

How Servant Leadership Influence Employee Service Quality of Fitness Centers During COVID-19: The Interacting Effects of Self-Efficacy.

Purpose: Drawing on conservation of resource theory, social cognitive theory and person-environment fit theory, this study aims to investigate the impact of servant leadership and self-efficacy on service quality in fitness centers. Patients and Methods: Cross-sectional data came from 771 employees of fitness center. All participants completed the perceived servant leadership scale, self-efficacy scale and service quality scale. The polynomial regression and response surface analysis techniques were used to investigate the mediating influence and boundary conditions of self-efficacy. Results: Self-efficacy acts as a mediator between servant leadership and employee service quality, with servant leadership having a positive, substantial impact on both. Meanwhile, regarding the relationship, the level of service quality is inversely correlated with the degree to which servant leadership and employee self-efficacy are aligned. The smaller the degree of alignment, the lower the level of employee service quality. Conclusion: The findings of this study can help fitness centre practitioners better improve the service quality of employees through service-oriented servant leadership during the COVID-19 pandemic, thereby contributing to the development of the sports and service industries.

Time-course RNA-Seq profiling reveals isoform-level gene expression dynamics of the cGAS-STING pathway.

The cGAS-STING pathway, orchestrating complicated transcriptome-wide immune responses, is essential for host antiviral defense but can also drive immunopathology in severe COVID-19. Here, we performed time-course RNA-Seq experiments to dissect the transcriptome expression dynamics at the gene-isoform level after cGAS-STING pathway activation. The in-depth time-course transcriptome after cGAS-STING pathway activation within 12 h enabled quantification of 48,685 gene isoforms. By employing regression models, we obtained 13,232 gene isoforms with expression patterns significantly associated with the process of cGAS-STING pathway activation, which were named activation-associated isoforms. The combination of hierarchical and k-means clustering algorithms revealed four major expression patterns of activation-associated isoforms, including two clusters with increased expression patterns enriched in cell cycle, autophagy, antiviral innate-immune functions, and COVID-19 coronavirus disease pathway, and two clusters showing decreased expression pattern that mainly involved in ncRNA metabolism, translation process, and mRNA processing. Importantly, by merging four clusters of activation-associated isoforms, we identified three types of genes that underwent isoform usage alteration during the cGAS-STING pathway activation. We further found that genes exhibiting protein-coding and non-protein-coding gene isoform usage alteration were strongly enriched for the factors involved in innate immunity and RNA splicing. Notably, overexpression of an enriched splicing factor, EFTUD2, shifted transcriptome towards the cGAS-STING pathway activated status and promoted protein-coding isoform abundance of several key regulators of the cGAS-STING pathway. Taken together, our results revealed the isoform-level gene expression dynamics of the cGAS-STING pathway and uncovered novel roles of splicing factors in regulating cGAS-STING pathway mediated immune responses.

A mechanism for SARS-CoV-2 RNA capping and its inhibition by nucleotide analog inhibitors.

Decoration of cap on viral RNA plays essential roles in SARS-CoV-2 proliferation. Here, we report a mechanism for SARS-CoV-2 RNA capping and document structural details at atomic resolution. The NiRAN domain in polymerase catalyzes the covalent link of RNA 5' end to the first residue of nsp9 (termed as RNAylation), thus being an intermediate to form cap core (GpppA) with GTP catalyzed again by NiRAN. We also reveal that triphosphorylated nucleotide analog inhibitors can be bonded to nsp9 and fit into a previously unknown "Nuc-pocket" in NiRAN, thus inhibiting nsp9 RNAylation and formation of GpppA. S-loop (residues 50-KTN-52) in NiRAN presents a remarkable conformational shift observed in RTC bound with sofosbuvir monophosphate, reasoning an "induce-and-lock" mechanism to design inhibitors. These findings not only improve the understanding of SARS-CoV-2 RNA capping and the mode of action of NAIs but also provide a strategy to design antiviral drugs.

A mechanism for SARS-CoV-2 RNA capping and its inhibition by nucleotide analogue inhibitors

Decoration of cap on viral RNA plays essential roles in SARS-CoV-2 proliferation. Here we report a mechanism for SARS-CoV-2 RNA capping and document structural details at atomic resolution. The NiRAN domain in polymerase catalyzes the covalent link of RNA 5’ end to the first residue of nsp9 (termed as RNAylation), thus being an intermediate to form cap core (GpppA) with GTP catalyzed again by NiRAN. We also reveal that triphosphorylated nucleotide analogue inhibitors can be bonded to nsp9 and fit into a previously unknown ‘Nuc-pocket’ in NiRAN, thus inhibiting nsp9 RNAylation and formation of GpppA. S-loop (residues 50-KTN-52) in NiRAN presents a remarkable conformational shift observed in RTC bound with sofosbuvir monophosphate, reasoning an ‘induce-and-lock’ mechanism to design inhibitors. These findings not only improve the understanding of SARS-CoV-2 RNA capping and the mode of action of NAIs, but also provide a strategy to design antiviral drugs. Graphical Structural analyses reveal how proteins from SARS-CoV-2 cooperate and use GTP to form the cap on viral mRNA, and how this process is interrupted by nucleotide analogues that serve as antiviral drugs.

Immunogenicity and protective efficacy of a DNA vaccine inducing optimal expression of the SARS-CoV-2 S gene in hACE2 mice.

The wide spread of coronavirus disease 2019 (COVID-19) has significantly threatened public health. Human herd immunity induced by vaccination is essential to fight the epidemic. Therefore, highly immunogenic and safe vaccines are necessary to control SARS-CoV-2, whose S protein is the antigenic determinant responsible for eliciting antibodies that prevent viral entry and fusion. In this study, we developed a SARS-CoV-2 DNA vaccine expressing the S protein, named pVAX-S-OP, which was optimized according to the human-origin codon preference and using polyinosinic-polycytidylic acid as an adjuvant. pVAX-S-OP induced specific antibodies and neutralizing antibodies in BALB/c and hACE2 transgenic mice. Furthermore, we observed 1.43-fold higher antibody titers in mice receiving pVAX-S-OP plus adjuvant than in those receiving pVAX-S-OP alone. Interferon gamma production in the pVAX-S-OP-immunized group was 1.58 times (CD3+CD4+IFN-gamma+) and 2.29 times (CD3+CD8+IFN-gamma+) lower than that in the pVAX-S-OP plus adjuvant group but higher than that in the control group. The pVAX-S-OP vaccine was also observed to stimulate a Th1-type immune response. When, hACE2 transgenic mice were challenged with SARS-CoV-2, qPCR detection of N and E genes showed that the viral RNA loads in pVAX-S-OP-immunized mice lung tissues were 104 times and 106 times lower than those of the PBS control group, which shows that the vaccine could reduce the amount of live virus in the lungs of hACE2 mice. In addition, pathological sections showed less lung damage in the pVAX-S-OP-immunized group. Taken together, our results demonstrated that pVAX-S-OP has significant immunogenicity, which provides support for developing SARS-CoV-2 DNA candidate vaccines.

Advances in application of single-cell RNA sequencing in cardiovascular research.

Single-cell RNA sequencing (scRNA-seq) provides high-resolution information on transcriptomic changes at the single-cell level, which is of great significance for distinguishing cell subtypes, identifying stem cell differentiation processes, and identifying targets for disease treatment. In recent years, emerging single-cell RNA sequencing technologies have been used to make breakthroughs regarding decoding developmental trajectories, phenotypic transitions, and cellular interactions in the cardiovascular system, providing new insights into cardiovascular disease. This paper reviews the technical processes of single-cell RNA sequencing and the latest progress based on single-cell RNA sequencing in the field of cardiovascular system research, compares single-cell RNA sequencing with other single-cell technologies, and summarizes the extended applications and advantages and disadvantages of single-cell RNA sequencing. Finally, the prospects for applying single-cell RNA sequencing in the field of cardiovascular research are discussed.

Cylindrical Tube Stethoscopes: The Value of Practical Equipment in the Management of Patients with Infectious Diseases.

Background: The COVID-19 pandemic has continued for more than two years since its outbreak. Due to the clinical auscultation needs of doctors when wearing airtight protective clothing, a cylindrical tube stethoscope was proposed to address this problem. However, the idea has been questioned by some experts. Methods: To address these questions, we performed three-part experiments using cylindrical tube stethoscopes. First, we performed laboratory tests to detect the sound intensity from a cylindrical tube stethoscope. Second, we improved the cylindrical tube stethoscope to achieve better results. Third, we revealed the difference in the auscultation effects of the cylindrical tube stethoscope and a conventional professional 3 M stethoscope. Results: From these experiments, we found that a narrow cylindrical tube with a diameter of 4.2 cm and a length of 20 cm equipped with a silicone gasket better auscultation of heart sounds. A cylindrical tube stethoscope and a 3 M stethoscope were used to perform stethoscope tests on 10 volunteers. The alveolar lung sounds were 44.478 decibels vs 49.529 decibels, the heart sounds were 46.631 decibels vs 41.109 decibels, and the intestinal sounds were 40.132 decibels vs 43.787 decibels, respectively. Conclusion: This improved cylindrical tube stethoscope can meet the auscultation requirements for cardiorespiratory and abdominal diagnosis during infectious disease pandemics.

Structures of Omicron spike complexes and implications for neutralizing antibody development.

The emergence of the SARS-CoV-2 Omicron variant is dominant in many countries worldwide. The high number of spike mutations is responsible for the broad immune evasion from existing vaccines and antibody drugs. To understand this, we first present the cryo-electron microscopy structure of ACE2-bound SARS-CoV-2 Omicron spike. Comparison to previous spike antibody structures explains how Omicron escapes these therapeutics. Secondly, we report structures of Omicron, Delta, and wild-type spikes bound to a patient-derived Fab antibody fragment (510A5), which provides direct evidence where antibody binding is greatly attenuated by the Omicron mutations, freeing spike to bind ACE2. Together with biochemical binding and 510A5 neutralization assays, our work establishes principles of binding required for neutralization and clearly illustrates how the mutations lead to antibody evasion yet retain strong ACE2 interactions. Structural information on spike with both bound and unbound antibodies collectively elucidates potential strategies for generation of therapeutic antibodies.

Continuous Renal Replacement Therapy With oXiris Filter May Not be an Effective Resolution to Alleviate Cytokine Release Syndrome in Non-AKI Patients With Severe and Critical COVID-19.

In this study, we aimed to determine whether continuous renal replacement therapy (CRRT) with oXiris filter may alleviate cytokine release syndrome (CRS) in non-AKI patients with severe and critical coronavirus disease 2019 (COVID-19). A total of 17 non-AKI patients with severe and critical COVID-19 treated between February 14 and March 26, 2020 were included and randomly divided into intervention group and control group according to the random number table. Patients in the intervention group immediately received CRRT with oXiris filter plus conventional treatment, while those in the control group only received conventional treatment. Demographic data were collected and collated at admission. During ICU hospitalization, the concentrations of circulating cytokines and inflammatory chemokines, including IL-2, IL-4, IL-6, IL-10, TNF-α, and IFN-γ, were quantitatively measured daily to reflect the degree of CRS induced by SARS-CoV-2 infection. Clinical data, including the severity of COVID-19 white blood cell count (WBC), neutrophil proportion (NEUT%), lymphocyte count (LYMPH), lymphocyte percentage (LYM%), platelet (PLT), C-reaction protein (CRP), high sensitivity C-reactive protein (hs-CRP), alanine aminotransferase (ALT), aspartate aminotransferase (AST), total bilirubin (TB), albumin (ALB), serum creatinine (SCr), D-Dimer, fibrinogen (FIB), IL-2, IL-4, IL-6, IL-10, TNF-α, IFN-γ, number of hospital days and sequential organ failure assessment (SOFA) score were obtained and collated from medical records, and then compared between the two groups. Age, and SCr significantly differed between the two groups. Besides the IL-2 concentration that was significantly lower on day 2 than that on day 1 in the intervention group, and the IL-6 concentrations that were significantly higher on day 1, and day 2 in the intervention group compared to the control group, similar to the IL-10 concentration on day 5, there were no significant differences between the two groups. To sum up, CRRT with oXiris filter may not effectively alleviate CRS in non-AKI patients with severe and critical COVID-19. Thus, its application in these patients should be considered with caution to avoid increasing the unnecessary burden on society and individuals and making the already overwhelmed medical system even more strained (IRB number: IRB-AF/SC-04).

Cytokine levels in sputum, not serum, may be more helpful for indicating the damage in the lung and the prognosis of severe COVID-19 - A case series.

PURPOSE: To describe the relationship between the severity of lung damage and cytokine levels in sputum, bronchoalveolar lavage fluid (BALF), serum. METHOD: Eight severe patients infected with coronavirus disease 2019 (COVID-19) were admitted and their cytokines and chest computed tomography (CT) were analyzed. RESULTS: Compared with in serum, IL-6 and TNF-α in sputum and in BALF show more directly reflect the severity of COVID-19 critical patients. The gradient ratio of IL-6 levels may predict the prognosis of severe patients. CONCLUSION: Cytokine levels in the sputum may be more helpful for indicating lung damage. Local intervention through the respiratory tract is expected to benefit patients with severe COVID-19.

Remdesivir overcomes the S861 roadblock in SARS-CoV-2 polymerase elongation complex.

Remdesivir (RDV), a nucleotide analog with broad-spectrum features, has exhibited effectiveness in COVID-19 treatment. However, the precise working mechanism of RDV when targeting the viral RNA-dependent RNA polymerase (RdRP) has not been fully elucidated. Here, we solve a 3.0-Å structure of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RdRP elongation complex (EC) and assess RDV intervention in polymerase elongation phase. Although RDV could induce an "i+3" delayed termination in meta-stable complexes, only pausing and subsequent elongation are observed in the EC. A comparative investigation using an enterovirus RdRP further confirms similar delayed intervention and demonstrates that steric hindrance of the RDV-characteristic 1'-cyano at the -4 position is responsible for the "i+3" intervention, although two representative Flaviviridae RdRPs do not exhibit similar behavior. A comparison of representative viral RdRP catalytic complex structures indicates that the product RNA backbone encounters highly conserved structural elements, highlighting the broad-spectrum intervention potential of 1'-modified nucleotide analogs in anti-RNA virus drug development.

Potent SARS-CoV-2 neutralizing antibodies with protective efficacy against newly emerged mutational variants.

Accumulating mutations in the SARS-CoV-2 Spike (S) protein can increase the possibility of immune escape, challenging the present COVID-19 prophylaxis and clinical interventions. Here, 3 receptor binding domain (RBD) specific monoclonal antibodies (mAbs), 58G6, 510A5 and 13G9, with high neutralizing potency blocking authentic SARS-CoV-2 virus display remarkable efficacy against authentic B.1.351 virus. Surprisingly, structural analysis has revealed that 58G6 and 13G9 both recognize the steric region S470-495 on the RBD, overlapping the E484K mutation presented in B.1.351. Also, 58G6 directly binds to another region S450-458 in the RBD. Significantly, 58G6 and 510A5 both demonstrate prophylactic efficacy against authentic SARS-CoV-2 and B.1.351 viruses in the transgenic mice expressing human ACE2 (hACE2), protecting weight loss and reducing virus loads. Together, we have evidenced 2 potent neutralizing Abs with unique mechanism targeting authentic SARS-CoV-2 mutants, which can be promising candidates to fulfill the urgent needs for the prolonged COVID-19 pandemic.

Guidelines on the treatment with integrated traditional Chinese medicine and western medicine for severe coronavirus disease 2019.

Severe Coronavirus Disease 2019 (COVID-19) is characterized by numerous complications, complex disease, and high mortality, making its treatment a top priority in the treatment of COVID-19. Integrated traditional Chinese medicine (TCM) and western medicine played an important role in the prevention, treatment, and rehabilitation of COVID-19 during the epidemic. However, currently there are no evidence-based guidelines for the integrated treatment of severe COVID-19 with TCM and western medicine. Therefore, it is important to develop an evidence-based guideline on the treatment of severe COVID-19 with integrated TCM and western medicine, in order to provide clinical guidance and decision basis for healthcare professionals, public health personnel, and scientific researchers involved in the diagnosis, treatment, and care of COVID-19 patients. We developed and completed the guideline by referring to the standardization process of the "WHO handbook for guideline development", the Grading of Recommendations Assessment, Development and Evaluation (GRADE) system, and the Reporting Items for Practice Guidelines in Healthcare (RIGHT).

Cryo-EM Structure of an Extended SARS-CoV-2 Replication and Transcription Complex Reveals an Intermediate State in Cap Synthesis.

Transcription of SARS-CoV-2 mRNA requires sequential reactions facilitated by the replication and transcription complex (RTC). Here, we present a structural snapshot of SARS-CoV-2 RTC as it transitions toward cap structure synthesis. We determine the atomic cryo-EM structure of an extended RTC assembled by nsp7-nsp82-nsp12-nsp132-RNA and a single RNA-binding protein, nsp9. Nsp9 binds tightly to nsp12 (RdRp) NiRAN, allowing nsp9 N terminus inserting into the catalytic center of nsp12 NiRAN, which then inhibits activity. We also show that nsp12 NiRAN possesses guanylyltransferase activity, catalyzing the formation of cap core structure (GpppA). The orientation of nsp13 that anchors the 5' extension of template RNA shows a remarkable conformational shift, resulting in zinc finger 3 of its ZBD inserting into a minor groove of paired template-primer RNA. These results reason an intermediate state of RTC toward mRNA synthesis, pave a way to understand the RTC architecture, and provide a target for antiviral development.

Role of Renin-Angiotensin System in Acute Lung Injury Caused by Viral Infection.

The renin-angiotensin system (RAS) is the most important regulatory system of electrolyte homeostasis and blood pressure and acts through angiotensin-converting enzyme (ACE)/angiotensin II (Ang II)/Ang II type 1 (AT1) receptor axis and angiotensin-converting enzyme 2 (ACE2)/angiotensin (1-7)/MAS receptor axis. RAS dysfunction is related to the occurrence and development of acute lung injury (ALI) and acute respiratory distress syndrome (ARDS) and causes a serious prognosis and even death. ALI/ARDS can be induced by various ways, one of which is viral infections, such as SARS-CoV, SARS-CoV-2, H5N1, H7N9, and EV71. This article reviews the specific mechanism on how RAS dysfunction affects ALI/ARDs caused by viral infections. SARS-CoV and SARS-CoV-2 enter the host cells by binding with ACE2. H5N1 and H7N9 avian influenza viruses reduce the ACE2 level in the body, and EV71 increases Ang II concentration. Treatment with angiotensin-converting enzyme inhibitor and angiotensin AT1 receptor blocker can alleviate ALI/ARDS symptoms. This review provides suggestions for the treatment of lung injury caused by viral infections.