Coronaviruses (CoV) are enveloped positive-stranded RNA viruses and, historically, there are seven known human-infecting CoVs with varying degrees of virulence. CoV attachment to the host is the first step of viral pathogenesis and mainly relies on the spike glycoprotein located on the viral surface. Among the human-infecting CoVs, only the infection of SARS CoV 2 (SARS2) among humans resulted to a pandemic which would suggest that the protein structural conformation of SARS2 spike protein is distinct as compared to other human-infecting CoVs. Surprisingly, the possible differences and similarities in the protein structural conformation between the various human-infecting CoV spike proteins have not been fully elucidated. In this study, we utilized a computational approach to generate models and analyze the seven human-infecting CoV spike proteins, namely: HCoV 229E, HCoV OC43, HCoV NL63, HCoV HKU1, SARS CoV, MERS CoV, and SARS2. Model quality assessment of all CoV models generated, structural superimposition of the whole protein model and selected S1 domains (S1-CTD and S1-NTD), and structural comparison based on RMSD values, Tm scores, and contact mapping were all performed. We found that the structural orientation of S1-CTD is a potential structural feature associated to both the CoV phylogenetic cluster and lineage. Moreover, we observed that spike models in the same phylogenetic cluster or lineage could potentially have similar protein structure. Additionally, we established that there are potentially three distinct S1-CTD orientation (Pattern I, Pattern II, Pattern III) among the human-infecting CoVs. Furthermore, we postulate that human-infecting CoVs in the same phylogenetic cluster may have similar S1-CTD and S1-NTD structural orientation. Taken together, we propose that the SARS2 spike S1-CTD follows a Pattern III orientation which has a higher degree of similarity with SARS1 and some degree of similarity with both OC43 and HKU1 which coincidentally are in the same phylogenetic cluster and lineage, whereas, the SARS2 spike S1-NTD has some degree of similarity among human-infecting CoVs that are either in the same phylogenetic cluster or lineage.

Butyric acid (BA) is produced by periodontopathic bacterial pathogens and contributes to periodontal disease (PD) induction. Moreover, PD has been associated with detrimental effects which subsequently may lead to systemic disease (SD) development affecting certain organs. Surprisingly, the potential systemic manifestations and organ-localized effects of BA have never been elucidated. Here, we simulated BA-based oral infection among young (20-week-old) rats and isolated blood cytosol to determine BA effects on stress network-related signals [total heme, hydrogen peroxide (H₂O₂), catalase (CAT), glutathione reductase (GR), free fatty acid (FFA), NADP/NADPH], inflammation-associated signals [caspases (CASP12 and CASP1), IL-1β, TNF-α, metallomatrix proteinase-9 (MMP-9), and toll-like receptor-2 (TLR2)], and neurological blood biomarkers [presenilin (PS1 and PS2) and amyloid precursor protein (APP)]. Similarly, we extracted the brain from both control and BA-treated rats, isolated the major regions (hippocampus, pineal gland, hypothalamus, cerebrum, and cerebellum), and, subsequently, measured stress network-related signals [oxidative stress: total heme, NADPH, H₂O₂, GR, and FFA; ER stress: GADD153, calcium, CASP1, and CASP3] and a brain neurodegenerative biomarker (Tau). In the blood, we found that BA was no longer detectable. Nevertheless, oxidative stress and inflammation were induced. Interestingly, amounts of representative inflammatory signals (CASP12, CASP1, IL-1β, and TNF-α) decreased while MMP-9 levels increased which we believe would suggest that inflammation was MMP-9-modulated and would serve as an alternative inflammatory mechanism. Similarly, TLR2 activity was increased which would insinuate that neurological blood biomarkers (APP, PS1, and PS2) were likewise affected. In the brain, BA was not detected, however, we found that both oxidative and ER stresses were likewise altered in all brain regions. Interestingly, tau protein amounts were significantly affected in the cerebellar and hippocampal regions which coincidentally are the major brain regions affected in several neurological disorders. Taken together, we propose that gingival BA can potentially cause systemic inflammation ascribable to prolonged systemic manifestations in the blood and localized detrimental effects within the brain organ.

Butyric acid (BA) is a common secondary metabolite by-product produced by oral pathogenic bacteria and is detected in high amounts in the gingival tissue of patients with periodontal disease. Previous works have demonstrated that BA can cause oxidative stress in various cell types; however, this was never explored using neuronal cells. Here, we exposed nerve growth factor (NGF)-treated PC12 cells to varying BA concentrations (0.5, 1.0, 5.0 mM). We measured total heme, H₂O₂, catalase, and calcium levels through biochemical assays and visualized the neurite outgrowth after BA treatment. Similarly, we determined the effects of other common periodontal short-chain fatty acids (SCFAs) on neurite outgrowth for comparison. We found that high (1.0 and 5.0 mM) BA concentrations induced oxidative stress and altered calcium homeostasis, whereas low (0.5 mM) BA concentration had no significant effect. Moreover, compared to other SCFAs, we established that only BA was able to induce neurite retraction.

Butyric acid (BA) induces jugular blood mitochondrial oxidative stress, whereas heme-induced oxidative stress was previously reported to inhibit SIRT1 in vitro. This would imply that BA-induced oxidative stress may similarly affect SIRT1. Here, we elucidated the BA effects on jugular blood cytosolic oxidative stress and SIRT1. Jugular blood cytosol was collected 0, 60, and 180 min after BA injection into rat gingival tissues and used throughout the study. Blood cytosolic oxidative stress induction, heme accumulation, NADPH oxidase (NOX) activation, nicotinamide adenine dinucleotide (NAD⁺) and NADP pool levels, NAD kinase (NADK), and SIRT1 amounts were determined. We found that BA retention in the gingival tissue induces blood cytosolic oxidative stress and heme accumulation which we correlated to both NOX activation and NADP pool increase. Moreover, we showed that BA-related NADP pool build-up is associated with NADK increase which we suspect decreased NAD⁺ levels and consequentially lowered SIRT1 amounts in the rat blood cytosol.

Fusobacterium nucleatum (Fn) is generally an opportunistic oral pathogen that adheres to mammalian mucosal sites, triggering a host inflammatory response. In general, Fn is normally found within the human oral cavity; however, it was previously reported that Fn is a risk factor for certain respiratory diseases. Surprisingly, this was never fully elucidated. Here, we investigated the virulence potential of heat-killed Fn on primary human tracheal, bronchial, and alveolar epithelial cells. In this study, we measured the secretion of inflammatory- (IL-8 and IL-6), stress- (total heme and hydrogen peroxide), and cell death-related (caspase-1 and caspase-3) signals. We established that the inflammatory response mechanism varies in each epithelial cell type: (1) along tracheal cells, possible Fn adherence would trigger increased heme secretion and regulated inflammatory response; (2) along bronchial cells, potential Fn adherence would simultaneously initiate an increase in secreted H₂O₂ and inflammatory response (ascribable to decreased secreted heme amounts); and (3) along alveolar cells, putative Fn adherence would instigate the increased secretion of inflammatory responses attributable to a decrease in secreted heme levels. Moreover, regardless of the epithelial cell-specific inflammatory mechanism, we believe these are putative, not harmful. Taken together, we propose that any potential Fn-driven inflammation along the respiratory tract would be initiated by differing epithelial cell-specific inflammatory mechanisms that are collectively dependent on secreted heme.