Receptor Angiotensin-converting Enzyme Research Articles

In vitro and in silico study of biosynthesized silver nanoparticles using Nigella sativa extract against SARS-CoV-2 and Candida albicans

Pathogens that are resistant to multiple drugs, commonly known as superbugs, including viruses and fungi, are posing a significant danger to human health. These superbugs are responsible for causing high rates of morbidity and mortality each year, and the numbers are continuing to rise. Therefore, the search for potent, safe, and viable alternative antimicrobial agents is imperative. In the current study, a rapid and eco-friendly method for synthesizing silver nanoparticles (AgNPs) using Nigella sativa seed extract was developed. Various characterization techniques, including ultraviolet–visible (UV–vis) spectroscopy, Fourier transform infrared (FTIR) spectroscopy, X-ray diffraction (XRD), scanning electron microscopy associated with microanalysis by dispersive energy of X-rays (SEM-EDX), and Zeta-potential, were employed to confirm the formation of AgNPs. Agglomerated particles with a spherical shape and a size of approximately 50 nm were identified in the SEM images of AgNPs. The optical bandgap energy of the AgNPs was determined to be 2.43 eV for direct bandgap and 1.87 eV for allowable indirect band gap. The study also investigated the antiviral activity of AgNPs against the XBB.1.5 SARS-CoV-2 variant and assessed their Anti-Candida Assay against Candida albicans. Furthermore, we conducted molecular docking studies to explore the interactions between AgNPs and the Omicron receptor-binding motif (RBM). The spike protein RBD binds to the receptor angiotensin-converting enzyme 2 (ACE2) human cellular receptor (hACE2) that helps the virus enter the host cell. The results indicated that AgNPs can bind to the spike protein RBD, potentially elucidating their antiviral effects against SARS-CoV-2. The findings of this study suggest that silver nanoparticles (AgNPs) hold promise as potent agents for therapeutic applications, particularly in addressing multidrug-resistant strains of viruses and Candida. However, further research is essential to thoroughly elucidate the mode of action and assess the biosafety of AgNPs.

SARS-CoV-2 spike protein expression as an identification in quality control testing for Adenovector based COVID-19 vaccine

AimQuality control testing of the vaccine for lot release is of paramount importance in public health. A recent pandemic caused by the SARS-CoV-2 virus brought together all spheres of vaccine to combat the virus. The scientific advancement in the development of vaccines facilitated the scientists to develop the vaccine against SARS-CoV-2 in a record time. Thus, these vaccines should be stringently monitored for their safety and efficacy as per the latest WHO and national regulatory guidelines, and quality control evaluation of the product should be done at national control laboratories before releasing the product into the market as it assures the quality and safety of the vaccine. MethodsThe SARS-CoV-2 exploited the ACE2 (Angiotensin Converting Enzyme 2) receptor, a surface protein on mammalian cells to gain entry into the host cells. The viral surface protein that interacted with the ACE2 receptor is the Spike protein of SARS-CoV-2. Thus, in the development of the vaccine and assessing its quality, the Spike protein of SARS-CoV-2 became an attractive immunodominant antigen. In National Institute of Biologicals, an apex body in the testing of biologicals in India, received the Adenovector (Adenovirus + vector) based COVID-19 vaccine, a finished product for quality evaluation. Due to the lack of a pharmacopeial monograph, the testing of the vaccine was done as per the manufacturer's specifications and methods. The routine assays of identification employed by the manufacturer do not reflect the expression of Spike protein which is required for the immune system to get activated. In this report, we showed the determination of Spike protein expression by immunoblotting and immunofluorescence for identification parameters in the quality testing of the COVID-19 vaccine. We determined the translation of the SARS-CoV-2 Spike gene cloned into an Adenovector. ResultsThe results from these experiments indicated the expression of Spike protein upon infection of mammalian cells with viral particles suggested that the expression of immunodominant Spike protein of SARS-CoV-2 may be employed by quality control laboratories as a parameter for identification. ConclusionThe study suggested that the determination of the expression of Spike protein is pertinent to identifying the Adenovector based vaccines against COVID-19.

Ligand- and Structure-Based Virtual Screening Identifies New Inhibitors of the Interaction of the SARS-CoV-2 Spike Protein with the ACE2 Host Receptor.

The Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) is a fast-spreading viral pathogen and poses a serious threat to human health. New SARS-CoV-2 variants have been arising worldwide; therefore, is necessary to explore more therapeutic options. The interaction of the viral spike (S) protein with the angiotensin-converting enzyme 2 (ACE2) host receptor is an attractive drug target to prevent the infection via the inhibition of virus cell entry. In this study, Ligand- and Structure-Based Virtual Screening (LBVS and SBVS) was performed to propose potential inhibitors capable of blocking the S receptor-binding domain (RBD) and ACE2 interaction. The best five lead compounds were confirmed as inhibitors through ELISA-based enzyme assays. The docking studies and molecular dynamic (MD) simulations of the selected compounds maintained the molecular interaction and stability (RMSD fluctuations less than 5 Å) with key residues of the S protein. The compounds DRI-1, DRI-2, DRI-3, DRI-4, and DRI-5 efficiently block the interaction between the SARS-CoV-2 spike protein and receptor ACE2 (from 69.90 to 99.65% of inhibition) at 50 µM. The most potent inhibitors were DRI-2 (IC50 = 8.8 µM) and DRI-3 (IC50 = 2.1 µM) and have an acceptable profile of cytotoxicity (CC50 > 90 µM). Therefore, these compounds could be good candidates for further SARS-CoV-2 preclinical experiments.

Bioinformatics Study of Flavonoids From Genus Erythrina As Ace2 inhibitor Candidates For Covid-19 Treatment.

This study aimed to screen potential drug candidates from the flavonoids of the genus Erythrina for the Corona Virus Disease 2019 (COVID-19) treatment. A comprehensive screening was conducted on the structures of 473 flavonoids derived from the genus Erythrina, focusing on their potential toxicity and pharmacokinetic profiles. Subsequently, flavonoids that were non-toxic and possessed favorable pharmacokinetic properties underwent further analysis to explore their interactions with the angiotensin-converting enzyme 2 (ACE2) receptor, employing molecular docking and molecular dynamics simulations. Among 473 flavonoids, 104 were predicted to be safe from being mutagenic, hepatotoxic, and inhibitors of the human ether-a-go-go-related gene (hERG). Among these 104 flavonoids, 18 compounds were predicted not to be substrates of P-glycoprotein (P-gp). Among these 18 flavonoids, gangetinin (471) and erybraedin D (310) exhibit low binding affinities and root mean square deviation (RMSD) values, indicating stable binding to the ACE2 receptor. The physicochemical attributes of compounds 310 and 471 suggest that they possess drug-like properties. Gangetinin (471) and erybraedin D (310) may serve as promising candidates for COVID-19 treatment due to their potential to inhibit the ACE2-RBD interaction. This warrants further investigation into their inhibitory effects on ACE2-RBD binding through in vitro experiments.

Glycosylation Modulation Dictates Trafficking and Interaction of SARS-CoV-2 S1 Subunit and ACE2 in Intestinal Epithelial Caco-2 Cells.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) mainly targets the upper respiratory tract. It gains entry by interacting with the host cell receptor angiotensin-converting enzyme 2 (ACE2) via its heavily glycosylated spike glycoprotein. SARS-CoV-2 can also affect the gastrointestinal tract. Given the significant role of glycosylation in the life cycle of proteins and the multisystem target of SARS-CoV-2, the role of glycosylation in the interaction of S1 with ACE2 in Caco-2 cells was investigated after modulation of their glycosylation patterns using N-butyldeoxynojirimycin (NB-DNJ) and 1-deoxymannojirimycin (dMM), in addition to mutant CHO cells harboring mutations at different stages of glycosylation. The data show a substantial reduction in the interactions between the altered glycosylation forms of S1 and ACE2 in the presence of NB-DNJ, while varied outcomes resulted from dMM treatment. These results highlight the promising effects of NB-DNJ and its potential use as an off-label drug to treat SARS-CoV-2 infections.

Susceptibility and transmissibility of SARS-CoV-2 variants in transgenic mice expressing the cat angiotensin-converting enzyme 2 (ACE-2) receptor

The emergence of SARS-CoV-2 in 2019 and its rapid spread throughout the world has caused the largest pandemic of our modern era. The zoonotic origin of this pathogen highlights the importance of the One Health concept and the need for a coordinated response to this kind of threats. Since its emergence, the virus has caused >7 million deaths worldwide. However, the animal source for human outbreaks remains unknown. The ability of the virus to jump between hosts is facilitated by the presence of the virus receptor, the highly conserved angiotensin-converting enzyme 2 (ACE2), found in various mammals. Positivity for SARS-CoV-2 has been reported in various species, including domestic animals and livestock, but their potential role in bridging viral transmission to humans is still unknown. Additionally, the virus has evolved over the pandemic, resulting in variants with different impacts on human health. Therefore, suitable animal models are crucial to evaluate the susceptibility of different mammalian species to this pathogen and the adaptability of different variants. In this work, we established a transgenic mouse model that expresses the feline ACE2 protein receptor (cACE2) under the human cytokeratin 18 (K18) gene promoter's control, enabling high expression in epithelial cells, which the virus targets. Using this model, we assessed the susceptibility, pathogenicity, and transmission of SARS-CoV-2 variants. Our results show that the sole expression of the cACE2 receptor in these mice makes them susceptible to SARS-CoV-2 variants from the initial pandemic wave but does not enhance susceptibility to omicron variants. Furthermore, we demonstrated efficient contact transmission of SARS-CoV-2 between transgenic mice that express either the feline or the human ACE2 receptor.

Long COVID: Epidemiology, post-COVID-19 manifestations, possible mechanisms, treatment, and prevention strategies – A review

Background and objectives: The respiratory disease COVID-19 began in 2019 and quickly became a pandemic infecting millions of individuals across the globe. Many patients show lingering effects of the infection several days after testing negative for the disease. This has become known as “long COVID” and is defined by various sources as lasting anywhere from 4 weeks to periods. This is a review of the existing literature on long COVID which offersextensive insights into its clinical features, diagnosis, and treatment. Materials and method: Information on clinical features, mechanisms, treatment options, preventive measures, and epidemiology of long COVID is derived from an extensive review of scientific journals and pertinent authoritative sources. Results: The virus enters the cells via angiotensin-converting enzyme 2(ACE2) receptors. ACE2 receptors are present on numerous cell types throughout the body and thus the virus can affect several organs resulting in avariety of different symptoms. Long COVID symptoms include fatigue, dyspnea, headache, brain fog, and symptoms related to cardiovascular and pulmonary systems. Fatigue can affect upwards of 93% of patients suffering from long COVID. Failure of the body to clear the virus could initiate this chronic effect. Studies indicate that the use of antiviral drugs at the early phase of COVID-19 could prevent long COVID symptoms. Vaccines against SARS-CoV-2 also might help prevent long COVID. Conclusion: Diagnosing and managing long COVID is challenging due to diverse symptoms, including mental health issues like anxiety and depression. Longitudinal studies and patient-oriented approaches are crucial for treatment, supported by policies and educational campaigns. Understanding the pathophysiology remains a top priority. July 2024; Vol. 18(2):003. DOI: https://doi.org/10.55010/imcjms.18.015 *Correspondence: M. S. Zaman, Department of Biological Sciences, Alcorn State University, Lorman, MS 39096, USA; Department of Biology, South Texas College, McAllen, TX 78501, USA. Emails: zaman@alcorn.edu; mzaman@southtexascollege.edu

Cell type-specific adaptation of the SARS-CoV-2 spike.

Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) can infect various human tissues and cell types, principally via interaction with its cognate receptor angiotensin-converting enzyme-2 (ACE2). However, how the virus evolves in different cellular environments is poorly understood. Here, we used experimental evolution to study the adaptation of the SARS-CoV-2 spike to four human cell lines expressing different levels of key entry factors. After twenty passages of a spike-expressing recombinant vesicular stomatitis virus (VSV), cell-type-specific phenotypic changes were observed and sequencing allowed the identification of sixteen adaptive spike mutations. We used VSV pseudotyping to measure the entry efficiency, ACE2 affinity, spike processing, TMPRSS2 usage, and entry pathway usage of all the mutants, alone or in combination. The fusogenicity of the mutant spikes was assessed with a cell-cell fusion assay. Finally, mutant recombinant VSVs were used to measure the fitness advantage associated with selected mutations. We found that the effects of these mutations varied across cell types, both in terms of viral entry and replicative fitness. Interestingly, two spike mutations (L48S and A372T) that emerged in cells expressing low ACE2 levels increased receptor affinity, syncytia induction, and entry efficiency under low-ACE2 conditions. Our results demonstrate specific adaptation of the SARS-CoV-2 spike to different cell types and have implications for understanding SARS-CoV-2 tissue tropism and evolution.

A Rationally Designed Synthetic Antiviral Peptide Binder Targeting the Receptor-Binding Domain of SARS-CoV-2.

The severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), a novel coronavirus, is the causative agent responsible for the spread of the COVID19 pandemic across the globe. The global impact of the COVID19 pandemic, the successful approval of vaccines for controlling the pandemic, and the further resurgence of COVID19 necessitate the exploration and validation of alternative therapeutic avenues targeting SARS-CoV-2. The initial entry and further invasion by SARS-CoV-2 require strong protein-protein interactions (PPIs) between the receptor-binding domain (RBD) of the SARS-CoV-2 spike protein and the human angiotensin-converting enzyme 2 (ACE2) receptors expressed on the cell surfaces of various tissues. In principle, disruption of the PPIs between the RBD of SARS-CoV-2 and the ACE2 receptor by designer peptides with optimized pharmacology appears to be an ideal choice for potentially preventing viral entry with minimal immunogenicity. In this context, the current study describes a short, synthetic designer peptide (codenamed SR16, ≤18 aa, molecular weight ≤2.5 kDa), which has a few noncoded amino acids, demonstrates a helical conformation in solution, and also engages the RBD of SARS-CoV-2 through a high-affinity interaction, as judged from a battery of biophysical studies. Further, the designer peptide demonstrates resistance to trypsin degradation, appears to be nontoxic to mammalian cells, and also does not induce hemolysis in freshly isolated human erythrocytes. In summary, SR16 appears to be an ideal peptide binder targeting the RBD of SARS-CoV-2, which has the potential for further optimization and development as an antiviral agent targeting SARS-CoV-2.

SARS-CoV-2 superinfection in CD14+ monocytes with latent human cytomegalovirus (HCMV) promotes inflammatory cascade

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the etiologic agent of coronavirus disease 2019 (COVID-19), has posed significant challenges to global health. While much attention has been directed towards understanding the primary mechanisms of SARS-CoV-2 infection, emerging evidence suggests co-infections or superinfections with other viruses may contribute to increased morbidity and mortality, particularly in severe cases of COVID-19. Among viruses that have been reported in patients with SARS-CoV-2, seropositivity for Human cytomegalovirus (HCMV) is associated with increased COVID-19 risk and hospitalization. HCMV is a ubiquitous beta-herpesvirus with a seroprevalence of 60–90 % worldwide and one of the leading causes of mortality in immunocompromised individuals. The primary sites of latency for HCMV include CD14+ monocytes and CD34+ hematopoietic cells. In this study, we sought to investigate SARS-CoV-2 infection of CD14+ monocytes latently infected with HCMV. We demonstrate that CD14+ cells are susceptible and permissive to SARS-CoV-2 infection and detect subgenomic transcripts indicative of replication. To further investigate the molecular changes triggered by SARS-CoV-2 infection in HCMV-latent CD14+ monocytes, we conducted RNA sequencing coupled with bioinformatic differential gene analysis. The results revealed significant differences in cytokine-cytokine receptor interactions and inflammatory pathways in cells superinfected with replication-competent SARS-CoV-2 compared to the heat-inactivated and mock controls. Notably, there was a significant upregulation in transcripts associated with pro-inflammatory response factors and a decrease in anti-inflammatory factors. Taken together, these findings provide a basis for the heightened inflammatory response, offering potential avenues for targeted therapeutic interventions among HCMV-infected severe cases of COVID-19. SummaryCOVID-19 patients infected with secondary viruses have been associated with a higher prevalence of severe symptoms. Individuals seropositive for human cytomegalovirus (HCMV) infection are at an increased risk for severe COVID-19 disease and hospitalization. HCMV reactivation has been reported in severe COVID-19 cases with respiratory failure and could be the result of co-infection with SARS-CoV-2 and HCMV. In a cell culture model of superinfection, HCMV has previously been shown to increase infection of SARS-CoV-2 of epithelial cells by upregulating the human angiotensin-converting enzyme-2 (ACE2) receptor. In this study, we utilize CD14+ monocytes, a major cell type that harbors latent HCMV, to investigate co-infection of SARS-CoV-2 and HCMV. This study is a first step toward understanding the mechanism that may facilitate increased COVID-19 disease severity in patients infected with SARS-CoV-2 and HCMV.

Analysis of Humoral Immunity SARS-CoV-2 Infection Model with ACE2 Receptor and Latent Phase

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is RNA virus which causes the coronavirus disease 2019 (COVID-19). SARS-CoV-2 infects the epithelial (target) cells by binding its spike protein, S, to the Angiotensin-Converting Enzyme 2 (ACE2) receptor on the surface of epithelial cells. ACE2 is an essential mediating factor in the SARS-CoV-2 infection pathway. In this study, we build a mathematical model for characterizing the dynamics of SARS-CoV-2 within the host while taking into account the impact of humoral immunity and the function of the ACE2 receptor. We incorporate the cells that are latently infected into the model. We consider three distributed delays: (i) delay in development of latently infected epithelial cells, (ii) delay in the latently infected epithelium cells’ activation, and (iii) delay in the maturation of recently released SARS-CoV-2 virions. We first address the fundamental properties of the delayed system, then find all possible equilibria. We demonstrate the existence and stability of the equilibria on the basis of two threshold parameters, namely basic reproduction number (ℜ₀) and humoral immune activation number (ℜ1). By building appropriate Lyapunov functions and applying LaSalle’s invariance principle, we prove the global asymptotic stability for all equilibria. We do numerical simulations to demonstrate the theoretical conclusions. We do sensitivity analysis and determine the most vulnerable parameters. Discussion is had on how the dynamics of the SARS-CoV-2 are affected by ACE2 receptors, humoral immunity, latent phase and time delays. It is shown that vigorous activation of humoral immunity can suppress viral multiplication. We found that, ℜ₀ is influenced by the rates of ACE2 receptor growth and degradation, and this may offer valuable guidance for the creation of potential receptor-targeted vaccinations and medications. Further, it is shown that, increasing time delays can effectively decrease ℜ₀ and then inhibit the SARS-CoV-2 replication. Finally, we showed that, excluding the latently infected cells in the model would result in an overestimation of ℜ₀. Our findings may be useful in understanding the dynamics of SARS-CoV-2 infection in the host as well as in the development of novel therapies.

Sites and Zones of Maximum Reactivity of the most Stable Structure of the Receptor-binding Domain of Wild-type SARS-CoV-2 Spike Protein: A Quantum Density Functional Theory Study

Today, it is well known that Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) has four types of proteins within its structure, between them the spike protein (S). The infection mechanism is carried out by the entry of the virus into the human host cell through the S protein, which strongly interacts with the human cell receptor angiotensin-converting enzyme 2 (ACE2). In this work, we propose an atomic model of the Receptor Binding Domain (RBD) of the S spike protein of the wild-type SARS-CoV-2 virus. The molecular structure of the model was composed of 50 amino acids that were chemically bonded, starting with Leucine and ending with one amino acid Tyrosine. The novelty of our work lies in the importance of knowing the sites and zones of maximum reactivity of the RBD from the fundamental levels of quantum mechanics considering the atomic structure of matter. For this, the local and global reactivity indices of the RBD were calculated, such as frontier orbitals, Highest Occupied Molecular Orbital (HOMO) and Lowest Unoccupied Molecular Orbital (LUMO), Fukui indices, chemical potential, chemical hardness, electrophilicity index; with this, it will be possible to know what type of molecules are more likely to interact with the RBD structure, and in this way, new knowledge will be generated at the quantum, atomic and molecular level to inhibit the virulent effects of wild-type SARS-CoV-2. Finally, in order to identify the functional groups within the most stable structure and thereby verify the future reactions that can be carried out between the RBD structure and biomolecules, the Infrared (IR) absorption spectrum was calculated. For this work, we used Material Studio v6.0 which uses the density functional theory (DFT) implemented in its DMol3 computational code. The IR spectrum was obtained using the Spartan ‘94 computer code. One novelty would be that we found nine amino acids more that could make the RBD and ACE2 binding further the already known. Thus, the Mulliken charge distribution indicates that the highest concentrations of positive and negative charge are found in the zones 477S, 478T, 484E, and 501N amino acids letting ionic or Van der Waals possible interactions with other structures.

A dual-target SPR screening system for simultaneous ligand discovery of SARS-CoV-2 spike protein and its receptor ACE2 from Chinese herbs

Traditional Chinese Medicine (TCM) is a supremely valuable resource for the development of drug discovery. Few methods are capable of hunting for potential molecule ligands from TCM towards more than one single protein target. In this study, a novel dual-target surface plasmon resonance (SPR) biosensor was developed to perform targeted compound screening of two key proteins involved in the cellular invasion process of the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2): the spike (S) protein receptor binding domain (RBD) and the angiotensin-converting enzyme 2 (ACE2). The screening and identification of active compounds from six Chinese herbs were conducted taking into consideration the multi-component and multi-target nature of Traditional Chinese Medicine (TCM). Puerarin from Radix Puerariae Lobatae was discovered to exhibit specific binding affinity to both S protein RBD and ACE2. The results highlight the efficiency of the dual-target SPR system in drug screening and provide a novel approach for exploring the targeted mechanisms of active components from Chinese herbs for disease treatment.

Molecular modeling of some commercially available antiviral drugs and their derivatives against SARS-CoV-2 infection.

Numerous prior studies have identified therapeutic targets that could effectively combat severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, including the angiotensin-converting enzyme 2 (ACE2) receptor, RNA-dependent RNA polymerase (RdRp), and Main protease (Mpro). In parallel, antiviral compounds like abacavir, acyclovir, adefovir, amantadine, amprenavir, darunavir, didanosine, oseltamivir, penciclovir, and tenofovir are under investigation for their potential in drug repurposing to address this infection. The aim of the study was to determine the effect of modifying the functional groups of the aforementioned antivirals in silico. Using the genetic optimization for ligand docking algorithm on software Maestro (version 11.1), the modified antivirals were docked onto ACE2 receptor, RdRp, and Mpro. Using QuickProp (Maestro v11.1), PASS (prediction of activity spectra for the substances), and altogether with SwissADME, the ADMET (absorption, distribution, metabolism, excretion, and toxicity) of the modified antivirals, as well as their bioavailability and the predicted activity spectra, were determined. Discovery studio software was used to undertake post-docking analysis. Among the 10 antivirals, N(CH3)2 derivative of darunavir, N(CH3)2 derivative of amprenavir and NCH3 derivative of darunavir exhibited best binding affinities with ACE2 receptor (docking scores: -10.333, -9.527 and -9.695 kJ/mol, respectively). Moreover, NCH3 derivative of abacavir (-6.506 kJ/mol), NO2 derivative of didanosine (-6.877 kJ/mol), NCH3 derivative of darunavir (-7.618 kJ/mol) exerted promising affinity to Mpro. In conclusion, the results of the in silico screenings can serve as a useful information for future experimental works.

Delay induced stability switch in a mathematical model of CD8 T-cell response to SARS-CoV-2 mediated by receptor ACE2.

The pathogen SARS-CoV-2 binds to the receptor angiotensin-converting enzyme 2 (ACE2) of the target cells and then replicates itself through the host, eventually releasing free virus particles. After infection, the CD8 T-cell response is triggered and appears to play a critical role in the defense against virus infections. Infected cells and their activated CD8 T-cells can cause tissue damage. Here, we established a mathematical model of within-host SARS-CoV-2 infection that incorporates the receptor ACE2, the CD8 T-cell response, and the damaged tissues. According to this model, we can get the basic reproduction number R0 and the immune reproduction number R1. We provide the theoretical proof for the stability of the disease-free equilibrium, immune-inactivated equilibrium, and immune-activated equilibrium. Finally, our numerical simulations show that the time delay in CD8 T-cell production can induce complex dynamics such as stability switching. These results provide insights into the mechanisms of SARS-CoV-2 infection and may help in the development of effective drugs against COVID-19.

378 Hydroxypropyl beta cyclodextrin barrier prevents respiratory and eye viral infections

OBJECTIVES/GOALS: Susceptible mucocutaneous membranes of the eye and nasal cavity are easily infected by viruses leading to pink eye or respiratory infections whose direct cost has been estimated as $16 billion annually in the United States. We have developed a novel and effective barrier that will be agnostic to variants enveloped viruses like coronaviruses. METHODS/STUDY POPULATION: We evaluated the efficacy of hydroxypropyl cyclodextrin barrier in preventing respiratory coronavirus infections using 25 humanized angiotensin converting enzyme-2 receptor (hACE-2) mice under a BSL3 laboratory setting. We have shown the barrier is safe and efficacious in preventing coronavirus infections in in vitro respiratory cell lines. We instilled 10 uL aliquot of the barrier into the nostril of the mouse 30 minutes before exposing them to a 10uL titer containing 10,000 plaque forming units of the SARS-CoV-2 delta variant. The control mice received the SARS-CoV-2 infection but not the barrier. The mice were observed for 5 days after which they were sacrificed. We analyzed the lungs and nasal palates for viral load using reverse transcription-polymerase chain reaction. RESULTS/ANTICIPATED RESULTS: We observed our barrier to be effective in preventing SARS-CoV-2 delta variant infection in hACE2 mice models. The lungs and nasal secretions of treated mice were less infectious with lower viral load than the control mice. The lungs of treated mice showed decrease in IFN gene expression and many cytokines and chemokines that regulate virally induced inflammatory responses such as IL-1b, IL-8, CXCL9, CXCL10, and the CCLs. We observed the plasma Angiotensin I and Angiotensin II decreased with barrier treatment, correlating with the viral load observed in the lungs. These peptides may be useful biomarkers for monitoring viral load within the lungs of virally infected individuals. DISCUSSION/SIGNIFICANCE: This supports the barrier’s efficacy to reduce transmission and prevent infections of SARS-CoV-2. This easy to use barrier can augment the mucocutaneous layers of the eye and nasal cavity. Our agnostic barrier will reduce the economic and public health burden of seasonal respiratory and eye viral infections and their related deaths amongst the public.

COVID 19- Biochemical and Other Mechanisms associated with Neuropathy and Stroke.

The Coronavirus (SARS-C0V-2) caused COVID 19 disease, commencing from China in December 2019 and soon in 2020 its neurological manifestations were noted. The viral spike (S) protein binding with host cell receptor angiotensin-converting enzyme 2 (ACE2) with assistance of membrane fusion protease (TMPRSS) caused viral entry in host cells. We searched 47 articles using PubMed from January 2021 to January 2023. The initial pooling of virus in nasal mucosa was followed by spread to respiratory and other tissues. This review highlights the neuropathy and stroke mechanisms. The role of receptors, enzymes, proteins and genes in neuropathological mechanisms are highlighted. Vascular endothelial effects, choroid plexus and Blood Brain Barrier disruptions were noted. The epigenetic neuropathological mechanisms need to be explored further to help in designing new therapeutic modalities. The post vaccination neurological manifestations as well as the post COVID effects both need further attention to promptly diagnose and start timely treatment.

SARS-CoV-2 and Angiotensin-Converting Enzyme-2 Receptor Interaction Blocker– an In-Silico Approach

The global COVID-19 pandemic, instigated by the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), has led to substantial morbidity and mortality on a worldwide scale. While COVID-19 vaccines offer hope, the emergence of mutated viral strains necessitates the development of FDA-approved drugs to address future outbreaks. Objective: To examine prospective antiviral medications through an analysis of the interaction between the spike protein of SARS-CoV-2 and Angiotensin-converting enzyme-2 (ACE-2) receptors, which play a pivotal role in facilitating viral entry into host cells. Methods: Molecular docking was employed to assess the binding affinities of various protease inhibitors with ACE-2 receptors. Natural proteases, including Furin and Transmembrane serine protease 2 (TMPRSS2), cleave viral spike proteins into S1 and S2 subunits, facilitating fusion with ACE-2 receptors. We assessed the binding energies of Indinavir, Nafamostat, Fosamprenavir, Lopinavir, and Boceprevir to inhibit this interaction with a sense of optimism for their potential therapeutic applications. Results: Our findings suggest potential treatments for COVID-19, with Indinavir, Nafamostat, Fosamprenavir, Lopinavir, and Boceprevir displaying promising binding energies of -9.6 kcal/mol, -8.4 kcal/mol, -7.7 kcal/mol, and -7.5 kcal/mol, respectively. Conclusions: While promising, further clinical trials are important to potentially evaluate the efficacy and safety of these proposed drugs in combating COVID-19 and its variants.

Agonists or positive allosteric modulators of α7 nicotinic acetylcholine receptor prevent interaction of SARS-Cov-2 receptor-binding domain with astrocytoma cells

SARS-Cov-2, the virus causing COVID-19, penetrates host target cells via the receptor of angiotensin-converting enzyme 2 (ACE2). Disrupting the virus interaction with ACE2 affords a plausible mechanism for prevention of cell penetration and inhibiting dissemination of the virus. Our studies demonstrate that ACE2 interaction with the receptor binding domain of SARS-Cov-2 spike protein (RBD) can be impaired by modulating the α7 nicotinic acetylcholine receptor (α7 nAChR) contiguous with ACE2. U373 cells of human astrocytoma origin were shown to bind both ACE2-specific antibody and recombinant RBD in Cell-ELISA. ACE2 was found to interact with α7 nAChR in U373 cell lysates studied by Sandwich ELISA. Our studies demonstrate that inhibition of RBD binding to ACE2-expressing U373 cells were defined with α7 nAChR agonists choline and PNU282987, but not a competitive antagonist methyllicaconitine (MLA). Additionally, the type 2 positive allosteric modulator (PAM2) PNU120596 and hydroxyurea (HU) also inhibited the binding. Our studies demonstrate that activation of α7 AChRs has efficacy in inhibiting the SARS-Cov-2 interaction with the ACE2 receptor and in such a way can prevent virus target cell penetration. These studies also help to clarify the consistent efficacy and positive outcomes for utilizing HU in treating COVID-19.

Effect of heart failure pharmacotherapies in patients with heart failure with mildly reduced ejection fraction.

The study sought to comprehensively investigate the effect of heart failure (HF) pharmacotherapies in patients with HF with mildly reduced ejection fraction (HFmrEF). In the absence of randomized controlled trials, guideline recommendations concerning HF-related therapies in patients with HFmrEF are limited. Consecutive patients hospitalized with HFmrEF were retrospectively included at one institution from 2016 to 2022. The prognostic value of treatment with beta-blockers (BB), angiotensin-converting enzyme inhibitors, receptor blockers, or receptor-neprilysin inhibitor (ACEi/ARB/ARNI), mineralocorticoid receptor antagonists (MRA), and sodium-glucose-linked transport protein 2 inhibitors (SGLT2i) was investigated for all-cause mortality at 30 months (a median follow-up) and HF-related rehospitalization. A total of 2109 patients with HFmrEF were included. Treatment with BB [27.0 vs. 35.0%; hazard ratio (HR) = 0.737; 95% confidence interval (CI) 0.617-0.881; P = 0.001], ACEi/ARB/ARNI (25.9 vs. 37.6%; HR = 0.612; 95% CI 0.517-0.725; P = 0.001), and SGLT2i (11.9 vs. 29.5%; HR = 0.441; 95% CI 0.236-0.824; P = 0.010) was associated with a lower risk of 30-month all-cause mortality, which was still demonstrated after multivariable adjustment and propensity score matching. In contrast, MRA treatment was not associated with long-term prognosis. The risk of HF-related rehospitalization was not affected by HF pharmacotherapies. Finally, the lowest risk of long-term all-cause mortality was observed in patients with combined use of BB, ACEi/ARB/ARNI, and SGLT2i (HR = 0.456; 95% CI 0.227-0.916; P = 0.027). Beta-blockers, ACEi/ARB/ARNI, and SGLT2i were independently associated with a lower risk of all-cause mortality in patients with HFmrEF, specifically when applied as combined 'HF triple therapy'. Randomized studies are needed to investigate the effect of HF-related pharmacotherapies in patients with HFmrEF.