Cloaking the ACE2 receptor with salivary cationic proteins inhibits SARS-CoV-2 entry.

Saliva contributes to the innate immune system, which suggests that it can prevent SARS-CoV-2 entry. We studied the ability of healthy salivary proteins to bind to angiotensin-converting enzyme 2 (ACE2) using biolayer interferometry and pull-down assays. Their effects on binding between the receptor-binding domain of the SARS-CoV-2 spike protein S1 (S1) and ACE2 were determined using an enzyme-linked immunosorbent assay. Saliva bound to ACE2 and disrupted the binding of S1 to ACE2 and four ACE2-binding salivary proteins were identified, including cationic histone H2A and neutrophil elastase, which inhibited the S1-ACE2 interaction. Calf thymus histone (ct-histone) also inhibited binding as effectively as histone H2A. The results of a cell-based infection assay indicated that ct-histone suppressed SARS-CoV-2 pseudoviral invasion into ACE2-expressing host cells. Manufactured polypeptides, such as ε-poly-L-lysine, also disrupted S1-ACE2 binding, indicating the importance of the cationic properties of salivary proteins in ACE2 binding. Overall, we demonstrated that positively charged salivary proteins are a barrier against SARS-CoV-2 entry by cloaking the negatively charged surface of ACE2 and provided a view that the cationic polypeptides represent a preventative and therapeutic treatment against COVID-19.

An upstream estrogen response element linked to exogenous p53 tumor suppressor gene expression differentiates effects of the codon 72 polymorphism.

The objective of this study was to assess the effects of an upstream estrogen response element (ERE) on exogenous p53 tumor suppressor gene with a codon 72 polymorphism about which there have been controversial reports in relation to cancer risk. The p53 gene (bases 166-1143 from start codon) with the codon 72 polymorphism, inserted into the pIRES-hrGFP II plasmid with or without upstream ERE, were transfected into HHUA endometrial cancer cells expressing the estrogen receptor. The ERE-linked p53 gene with the proline variant at codon 72 showed lower transfection rates than the gene without ERE or with the arginine variant at codon 72. p21 expression was significantly higher in HHUA cells transfected with the proline variant gene than in those transfected with the arginine variant gene. We consider that the presence of an upstream ERE promotes the transcriptional effects of the exogenous p53 gene with the proline variant, which strengthens the expression of p21, and results in lower transfection rates through cell cycle inhibition.

Prevention of venous thrombosis by preoperative glycyrrhizin infusion in a rat model.

BACKGROUND: Glycyrrhizin is an agent with the capacity to bind to selectin molecules expressed on vascular endothelial cells and potentially prevent the adherence of neutrophils to the vascular endothelial surface. It has been found to prevent intravenous thrombus formation. METHODS: Venous thrombosis was induced in male rats by ligation of the inferior vena cava (IVC) for 6 h. Before the ligation, the study rats were given intravenous injections of glycyrrhizin through the IVC. After 6 h of venous ligation, the rats were sacrificed and the IVC segments were harvested. Thrombus within the IVC was collected to measure the wet weight. Gene expression of P-, L-, and E-selectin was detected by reverse transcriptase polymerase chain reaction using extracts of mRNA from the IVC vein wall. As baseline controls, IVC samples without ligation were harvested immediately after laparotomy. Neutrophil adhesion to the luminal surface of IVC was assessed on histological sections stained with hematoxylin and eosin. Blood samples were collected through the IVC proximal to the ligation after 6 h to estimate activated partial thromboplastin time (APTT) and prothrombin time (PT). To investigate the effect of glycyrrhizin on binding capacity of P-selectin to human neutrophils, real-time biospecific interaction analysis was performed with the Biacore 2000 system. RESULTS: The mean weight of thrombus in the glycyrrhizintreated group was 12.9 +/- 11.1 mg, which is significantly lower than that of the saline-treated control group (21.3 +/- 12.5 mg). The expression level of P-and L-selectin mRNA in both saline-and glycyrrhizin-treated groups was significantly higher than that of the baseline control. Histological studies of cross sections of IVC showed significantly fewer neutrophils adhering to the luminal surface with glycyrrhizin treatment than in the saline-treated controls. There was no significant difference in the values of coagulation parameters with or without glycyrrhizin treatment. In vitro analysis showed that glycyrrhizin caused a dose-dependent reduction of neutrophils binding to immobilized recombinant P-selectin. CONCLUSIONS: Preoperative treatment with glycyrrhizin is potentially useful for preventing venous thrombosis by suppressing the adherence of neutrophils to the venous endothelium during the initial phase of thrombus formation without reducing coagulation capacity and the subsequent risk for increased bleeding.

Identification and characterization of endoplasmic reticulum-associated protein, ERp43.

Disposal of misfolded proteins from the lumen of the endoplasmic reticulum (ER) is one of the quality control mechanisms present in the protein secretory pathway. Through ER-associated degradation, misfolded substrates are targeted to the cytosol where they are degraded by proteasomes. Here we describe the identification of a human ER-associated 43-kD protein (ERp43) by sequencing of the subtraction suppression hybridization cDNA library from ER stress-treated cells. The ERp43 gene encodes a protein of 383 amino acid residues that contains a potential transmembrane domain. Analysis revealed that ERp43 is primarily located in the ER. Quantitative reverse transcriptase-polymerase chain reaction demonstrated that gene expression was relatively high in the neuronal tissues and in the kidney, with ERp43 protein highly expressed in the spinal cord and in the kidney. In cultured cells, overexpression of ERp43 accelerated cell growth and inhibited ER stress-induced cell death, while down-regulation of ERp43 expression decreased proliferation rate and enhanced this type of cell death. These findings indicate that ERp43 plays important roles in cell growth and ER stress-induced cell death.