Identification of VEGFR2 as the Histatin-1 receptor in endothelial cells.

Histatin-1 is a salivary peptide with antimicrobial and wound healing promoting activities, which was previously shown to stimulate angiogenesis in vitro and in vivo via inducing endothelial cell migration. The mechanisms underlying the proangiogenic effects of Histatin-1 remain poorly understood and specifically, the endothelial receptor for this peptide, is unknown. Based on the similarities between Histatin-1-dependent responses and those induced by the prototypical angiogenic receptor, vascular endothelial growth factor receptor 2 (VEGFR2), we hypothesized that VEGFR2 is the Histatin-1 receptor in endothelial cells. First, we observed that VEGFR2 is necessary for Histatin-1-induced endothelial cell migration, as shown by both pharmacological inhibition studies and siRNA-mediated ablation of VEGFR2. Moreover, Histatin-1 co-immunoprecipitated and co-localized with VEGFR2, associating spatial proximity between these proteins with receptor activation. Indeed, pulldown assays with pure, tagged and non-tagged proteins showed that Histatin-1 and VEGFR2 directly interact in vitro. Optical tweezers experiments permitted estimating kinetic parameters and rupture forces, indicating that the Histatin-1-VEGFR2 interaction is transient, but specific and direct. Sequence alignment and molecular modeling identified residues Phe26, Tyr30 and Tyr34 within the C-terminal domain of Histatin-1 as relevant for VEGFR2 binding and activation. This was corroborated by mutation and molecular dynamics analyses, as well as in direct binding assays. Importantly, these residues were required for Histatin-1 to induce endothelial cell migration and angiogenesis in vitro. Taken together, our findings reveal that VEGFR2 is the endothelial cell receptor of Histatin-1 and provide insights to the mechanism by which this peptide promotes endothelial cell migration and angiogenesis.

Analysis by real-time PCR of five transport and conservation mediums of nasopharyngeal swab samples to COVID-19 diagnosis in Santiago of Chile.

Due to the COVID-19 pandemic, many transport kits have been manufactured to preserve and transport nasopharyngeal swab samples (NPSs) from patients. However, there is no information on the performance of the different virus transport media (VTM) used in COVID-19 diagnosis in the population of Santiago de Chile. We compared the RT-qPCR amplification profile of five different viral transport kit mediums, including DNA/RNA Shield™, NAT, VTM-N, Ezmedlab™, and phosphate-buffered saline (PBS), for NPSs from Central Metropolitan Health Service, Santiago, Chile. The DNA/RNA Shield™ medium showed a better performance in terms of Cq and RFU values for the internal reference RNase P and viral ORF1ab probes. By contrast, the PBS transport medium registered higher Cq values for the viral and reference gene, compared to the other VTM. DNA/RNA Shield™ shows higher relative fluorescence units (RFUs) and lower Cq values for the reference gene. Collectively, our results suggest that the PBS medium could compromise the sample diagnosis because of its lower RT-qPCR performance. The NAT, Ezmedlab and VTM-N, and DNA/RNA Shield™ media show acceptable RT-qPCR parameters and, consequently, seem suitable for use in COVID-19 diagnosis.

Histatin-1 is a novel osteogenic factor that promotes bone cell adhesion, migration, and differentiation.

Histatin-1 is a salivary antimicrobial peptide involved in the maintenance of enamel and oral mucosal homeostasis. Moreover, Histatin-1 has been shown to promote re-epithelialization in soft tissues, by stimulating cell adhesion and migration in oral and dermal keratinocytes, gingival and skin fibroblasts, endothelial cells and corneal epithelial cells. The broad-spectrum activity of Histatin-1 suggests that it behaves as a universal wound healing promoter, although this is far from being clear yet. Here, we report that Histatin-1 is a novel osteogenic factor that promotes bone cell adhesion, migration, and differentiation. Specifically, Histatin-1 promoted cell adhesion, spreading, and migration of SAOS-2 cells and MC3T3-E1 preosteoblasts in vitro, when placed on a fibronectin matrix. Besides, Histatin-1 induced the expression of osteogenic genes, including osteocalcin, osteopontin, and Runx2, and increased both activity and protein levels of alkaline phosphatase. Furthermore, Histatin-1 promoted mineralization in vitro, as it augmented the formation of calcium deposits in both SAOS-2 and MC3T3-E1 cells. Mechanistically, although Histatin-1 failed to activate ERK1/2, FAK, and Akt, which are signaling proteins associated with osteogenic differentiation or cell migration, it triggered nuclear relocalization of ß-catenin. Strikingly, the effects of Histatin-1 were recapitulated in cells that are nonosteogenically committed, since it promoted surface adhesion, migration, and the acquisition of osteogenic markers in primary mesenchymal cells derived from the apical papilla and dental pulp. Collectively, these observations indicate that Histatin-1 is a novel osteogenic factor that promotes bone cell differentiation, surface adhesion and migration, as crucial events required for bone tissue regeneration.