Middle East respiratory syndrome coronavirus (MERS-CoV) is a lineage C betacoronavirus that since its emergence in 2012 has caused outbreaks in human populations with case-fatality rates of ∼36%. As ...in other coronaviruses, the spike (S) glycoprotein of MERS-CoV mediates receptor recognition and membrane fusion and is the primary target of the humoral immune response during infection. Here we use structure-based design to develop a generalizable strategy for retaining coronavirus S proteins in the antigenically optimal prefusion conformation and demonstrate that our engineered immunogen is able to elicit high neutralizing antibody titers against MERS-CoV. We also determined high-resolution structures of the trimeric MERS-CoV S ectodomain in complex with G4, a stem-directed neutralizing antibody. The structures reveal that G4 recognizes a glycosylated loop that is variable among coronaviruses and they define four conformational states of the trimer wherein each receptor-binding domain is either tightly packed at the membrane-distal apex or rotated into a receptor-accessible conformation. Our studies suggest a potential mechanism for fusion initiation through sequential receptor-binding events and provide a foundation for the structure-based design of coronavirus vaccines.
Traditionally, the emergence of coronaviruses (CoVs) has been attributed to a gain in receptor binding in a new host. Our previous work with severe acute respiratory syndrome (SARS)-like viruses ...argued that bats already harbor CoVs with the ability to infect humans without adaptation. These results suggested that additional barriers limit the emergence of zoonotic CoV. In this work, we describe overcoming host restriction of two Middle East respiratory syndrome (MERS)-like bat CoVs using exogenous protease treatment. We found that the spike protein of PDF2180-CoV, a MERS-like virus found in a Ugandan bat, could mediate infection of Vero and human cells in the presence of exogenous trypsin. We subsequently show that the bat virus spike can mediate the infection of human gut cells but is unable to infect human lung cells. Using receptor-blocking antibodies, we show that infection with the PDF2180 spike does not require MERS-CoV receptor DPP4 and antibodies developed against the MERS spike receptor-binding domain and S2 portion are ineffective in neutralizing the PDF2180 chimera. Finally, we found that the addition of exogenous trypsin also rescues HKU5-CoV, a second bat group 2c CoV. Together, these results indicate that proteolytic cleavage of the spike, not receptor binding, is the primary infection barrier for these two group 2c CoVs. Coupled with receptor binding, proteolytic activation offers a new parameter to evaluate the emergence potential of bat CoVs and offers a means to recover previously unrecoverable zoonotic CoV strains.
Overall, our studies demonstrate that proteolytic cleavage is the primary barrier to infection for a subset of zoonotic coronaviruses. Moving forward, the results argue that both receptor binding and proteolytic cleavage of the spike are critical factors that must be considered for evaluating the emergence potential and risk posed by zoonotic coronaviruses. In addition, the findings also offer a novel means to recover previously uncultivable zoonotic coronavirus strains and argue that other tissues, including the digestive tract, could be a site for future coronavirus emergence events in humans.
The rapid spread of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) B.1.1.529 (Omicron) variant and its resistance to neutralization by vaccinee and convalescent sera are driving a ...search for monoclonal antibodies with potent neutralization. To provide insight into effective neutralization, we determined cryo-electron microscopy structures and evaluated receptor binding domain (RBD) antibodies for their ability to bind and neutralize B.1.1.529. Mutations altered 16% of the B.1.1.529 RBD surface, clustered on an RBD ridge overlapping the angiotensin-converting enzyme 2 (ACE2)-binding surface and reduced binding of most antibodies. Substantial inhibitory activity was retained by select monoclonal antibodies-including A23-58.1, B1-182.1, COV2-2196, S2E12, A19-46.1, S309, and LY-CoV1404-that accommodated these changes and neutralized B.1.1.529. We identified combinations of antibodies with synergistic neutralization. The analysis revealed structural mechanisms for maintenance of potent neutralization against emerging variants.
Abstract The triglyceride glucose-body mass index (TyG-BMI) is a convenient and clinically significant indicator of insulin resistance. This study aims to investigate the correlation between TyG-BMI ...and the onset of new-onset diabetes and determine an optimal reflection point for TyG-BMI. An analysis was conducted on 1917 participants from the risk evaluation of cancers in Chinese diabetic individuals: a lONgitudinal (REACTION) study. Participants were categorized based on their TyG-BMI, and the relationship between TyG-BMI and the incidence of new-onset diabetes was explored through logistic regression models, smoothed curve fitting with restricted cubic spline, and a two-piecewise logistic regression model. The mean age of the participants was 57.60 ± 8.89 years, with 66.5% being females. The mean TyG-BMI was 223.3 ± 32.8. Ultimately, 137 individuals (7.1%) progressed to diabetes after three years. After adjusting for covariates, TyG-BMI exhibited a positive correlation with new-onset diabetes (odd ratios (OR) for each standard deviation increase = 1.330, 95% CI 1.110–1.595). The relationship between TyG-BMI and new-onset diabetes was non-linear, with a inflcetion point at 202.9. This study reveals a positive non-linear relationship between TyG-BMI and the risk of new-onset diabetes in Chinese middle-aged and elderly individuals. When TyG-BMI exceeds 202.9, there is a significantly heightened risk of new-onset diabetes. These findings offer valuable insights for preventing new-onset diabetes.
The potential for future coronavirus outbreaks highlights the need to broadly target this group of pathogens. We used an epitope-agnostic approach to identify six monoclonal antibodies that bind to ...spike proteins from all seven human-infecting coronaviruses. All six antibodies target the conserved fusion peptide region adjacent to the S2' cleavage site. COV44-62 and COV44-79 broadly neutralize alpha- and betacoronaviruses, including severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Omicron subvariants BA.2 and BA.4/5, albeit with lower potency than receptor binding domain-specific antibodies. In crystal structures of COV44-62 and COV44-79 antigen-binding fragments with the SARS-CoV-2 fusion peptide, the fusion peptide epitope adopts a helical structure and includes the arginine residue at the S2' cleavage site. COV44-79 limited disease caused by SARS-CoV-2 in a Syrian hamster model. These findings highlight the fusion peptide as a candidate epitope for next-generation coronavirus vaccine development.
The MERS-CoV is an emerging virus, which already infected more than 1,300 humans with high (∼36%) mortality. Here, we show that m336, an exceptionally potent human anti-MERS-CoV antibody, is almost ...germline with only one somatic mutation in the heavy chain. The structure of Fab m336 in complex with the MERS-CoV receptor-binding domain reveals that its IGHV1-69-derived heavy chain provides more than 85% binding surface and that its epitope almost completely overlaps with the receptor-binding site. Analysis of antibodies from 69 healthy humans suggests an important role of the V(D)J recombination-generated junctional and allele-specific residues for achieving high affinity of binding at such low levels of somatic hypermutation. Our results also have important implications for development of vaccine immunogens based on the newly identified m336 epitope as well as for elucidation of mechanisms of neutralization by m336-like antibodies and their elicitation in vivo.
Background Mesenchymal stem cell (MSC)-based therapy is currently considered to be an effective treatment strategy for diabetes and hepatic disorders, such as liver cirrhosis and non-alcoholic fatty ...liver disease. Exosomes are important mediators of cellular connections, and increasing evidence has suggested that exosomes derived from MSCs may be used as direct therapeutic agents; their mechanisms of action, however, remain largely unclear. Here, we evaluated the efficacy and molecular mechanisms of human umbilical cord MSC-derived exosomes (HucMDEs) on hepatic glucose and lipid metabolism in type 2 diabetes mellitus (T2DM). Methods HucMDEs were used to treat T2DM rats, as well as palmitic acid (PA)-treated L-O2 cells, in order to determine the effects of HucMDEs on hepatic glucose and lipid metabolism. To evaluate the changes in autophagy and potential signaling pathways, autophagy-related proteins (BECN1, microtubule-associated protein 1 light chain 3 beta MAP 1LC3B), autophagy-related genes (ATGs, ATG5, and ATG7), AMP-activated protein kinase (AMPK), and phosphorylated AMPK (p-AMPK) were assessed by Western blotting. Results HucMDEs promoted hepatic glycolysis, glycogen storage, and lipolysis, and reduced gluconeogenesis. Additionally, autophagy potentially contributed to the effects of HucMDE treatment. Transmission electron microscopy revealed an increased formation of autophagosomes in HucMDE-treated groups, and the autophagy marker proteins, BECN1 and MAP 1LC3B, were also increased. Moreover, autophagy inhibitor 3-methyladenine significantly reduced the effects of HucMDEs on glucose and lipid metabolism in T2DM rats. Based on its phosphorylation status, we found that the AMPK signaling pathway was activated and induced autophagy in T2DM rats and PA-treated L-O2 cells. Meanwhile, the transfection of AMPK siRNA or application of the AMPK inhibitor, Comp C, weakened the therapeutic effects of HucMDEs on glucose and lipid metabolism. Conclusions These findings demonstrate that HucMDEs improved hepatic glucose and lipid metabolism in T2DM rats by activating autophagy via the AMPK pathway, which provides novel evidence suggesting the potential for HucMDEs in clinically treating T2DM patients. Keywords: Exosome, Mesenchymal stem cell, Glucose metabolism, Type 2 diabetes mellitus, Autophagy
The Fokas–Lenells equation and its multi-component coupled forms have attracted the attention of many mathematical physicists. The Fokas–Lenells equation and two coupled Fokas–Lenells equations are ...investigated from the perspective of Lie symmetries and conservation laws. The three systems have been turned into real multi-component coupled systems by appropriate transformations. By procedures of symmetry analysis, Lie symmetries of the three real systems are obtained. Explicit conservation laws are constructed using the symmetry/adjoint symmetry pair method, which depends on Lie symmetries and adjoint symmetries. The relationships between the multiplier and the adjoint symmetry are investigated.
Abstract Background The use of stem cell-derived exosomes (Exos) as therapeutic vehicles is receiving increasing attention. Exosome administration has several advantages over cell transplantation, ...thus making exosomes promising candidates for large-scale clinical implementation and commercialization. However, exosome extraction and purification efficiencies are relatively low, and therapeutic heterogeneity is high due to differences in culture conditions and cell viability. Therefore, in this study, we investigated a priming procedure to enhance the production and therapeutic effects of exosomes from human umbilical cord mesenchymal stem cells (hucMSCs). After preconditioning hucMSCs with agonists/inhibitors that target the Wnt/β-catenin pathway, we assessed both the production of exosomes and the therapeutic efficacy of the optimized exosomes in the context of diabetic wound healing, hoping to provide a safer, more stable and more effective option for clinical application. Results The Wnt signalling pathway agonist CHIR99021 increased exosome production by 1.5-fold without causing obvious changes in the characteristics of the hucMSCs or the size of the exosome particles. Further studies showed that CHIR99021 promoted the production of exosomes by facilitating exocytosis. This process was partly mediated by SNAP25. To further explore whether CHIR99021 changed the cargo that was loaded into the exosomes and its therapeutic effects, we performed proteomic and transcriptomic analyses of exosomes from primed and control hucMSCs. The results showed that CHIR99021 significantly upregulated the expression of proteins that are associated with cell migration and wound healing. Animal experiments confirmed that, compared to control hucMSC-derived exosomes, CHIR99021-pretreated hucMSC-derived exosomes (CHIR-Exos) significantly accelerated wound healing in diabetic mice, enhanced local collagen deposition, promoted angiogenesis, and reduced chronic inflammation. Subsequent in vitro experiments confirmed that the CHIR-Exos promoted wound healing by facilitating cell migration, inhibiting oxidative stress-induced apoptosis, and preventing cell cycle arrest. Conclusions The Wnt agonist CHIR99021 significantly increased exosome secretion by hucMSCs, which was partly mediated by SNAP25. Notably, CHIR99021 treatment also significantly increased the exosomal levels of proteins that are associated with wound healing and cell migration, resulting in enhanced acceleration of wound healing. All of these results suggested that pretreatment of hucMSCs with CHIR99021 not only promoted exosome production but also improved the exosome therapeutic efficacy, thus providing a promising option for large-scale clinical implementation and commercialization. Graphical Abstract
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