25-Hydroxycholesterol and 27-hydroxycholesterol inhibit human rotavirus infection by sequestering viral particles into late endosomes

A novel innate immune strategy, involving specific cholesterol oxidation products as effectors, has begun to reveal connections between cholesterol metabolism and immune response against viral infections. Indeed, 25-hydroxycholesterol (25HC) and 27-hydroxycholesterol (27HC), physiologically produced by enzymatic oxidation of cholesterol, act as inhibitors of a wide spectrum of enveloped and non-enveloped human viruses. However, the mechanisms underlying their protective effects against non-enveloped viruses are almost completely unexplored. To get insight into this field, we investigated the antiviral activity of 25HC and 27HC against a non-enveloped virus causing acute gastroenteritis in children, the human rotavirus (HRV). We found that 25HC and 27HC block the infectivity of several HRV strains at 50% inhibitory concentrations in the low micromolar range in the absence of cell toxicity. Both molecules affect the final step of virus penetration into cells by preventing the association of two cellular proteins: the oxysterol binding protein (OSBP) and the vesicle-associated membrane protein-associated protein-A (VAP-A). By altering the activity of these cellular mediators, 25HC and 27HC disturb the recycling of cholesterol between the endoplasmic reticulum and the late endosomes which are exploited by HRV to penetrate into the cell. The substantial accumulation of cholesterol in the late endosomal compartment results in sequestering viral particles inside these vesicles thereby preventing cytoplasmic virus replication. These findings suggest that cholesterol oxidation products of enzymatic origin might be primary effectors of host restriction strategies to counteract HRV infection and point to redox active lipids involvement in viral infections as a research area of focus to better focus in order to identify novel antiviral agents targets

New chitosan nanobubbles for ultrasound-mediated gene delivery: preparation and in vitro characterization

BACKGROUND: The development of nonviral gene delivery systems is one of the most intriguing topics in nanomedicine. However, despite the advances made in recent years, several key issues remain unsettled. One of the main problems relates to the difficulty in designing nanodevices for targeted delivery of genes and other drugs to specific anatomic sites. In this study, we describe the development of a novel chitosan nanobubble-based gene delivery system for ultrasound-triggered release. METHODS AND RESULTS: Chitosan was selected for the nanobubble shell because of its low toxicity, low immunogenicity, and excellent biocompatibility, while the core consisted of perfluoropentane. DNA-loaded chitosan nanobubbles were formed with a mean diameter of less than 300 nm and a positive surface charge. Transmission electron microscopic analysis confirmed composition of the core-shell structure. The ability of the chitosan nanobubbles to complex with and protect DNA was confirmed by agarose gel assay. Chitosan nanobubbles were found to be stable following insonation (2.5 MHz) for up to 3 minutes at 37°C. DNA release was evaluated in vitro in both the presence and absence of ultrasound. The release of chitosan nanobubble-bound plasmid DNA occurred after just one minute of insonation. In vitro transfection experiments were performed by exposing adherent COS7 cells to ultrasound in the presence of different concentrations of plasmid DNA-loaded nanobubbles. In the absence of ultrasound, nanobubbles failed to trigger transfection at all concentrations tested. In contrast, 30 seconds of ultrasound promoted a moderate degree of transfection. Cell viability experiments demonstrated that neither ultrasound nor the nanobubbles affected cell viability under these experimental conditions. CONCLUSION: Based on these results, chitosan nanobubbles have the potential to be promising tools for ultrasound-mediated DNA delivery

Inhibition of pathogenic non-enveloped viruses by 25-hydroxycholesterol and 27-hydroxycholesterol

Recent studies reported a broad but selective antiviral activity of 25-hydroxycholesterol (25HC) against enveloped viruses, being apparently inactive against non-enveloped viruses. Here we show that 25HC is endowed with a marked antiviral activity against three pathogenic non-enveloped viruses, i.e. human papillomavirus-16 (HPV-16), human rotavirus (HRoV), and human rhinovirus (HRhV), thus significantly expanding its broad antiviral spectrum, so far recognized to be limited to viruses with envelope. Moreover, here we disclose the remarkable antiviral activity of another oxysterol of physiological origin, i.e. 27-hydroxycholesterol (27HC), against HPV-16, HRoV and HRhV. We have also identified a much weaker antiviral activity of other oxysterols of pathophysiological relevance, i.e 7α-hydroxycholesterol, 7β-hydroxycholesterol, and 7-ketocholesterol. These findings suggest that appropriate modulation of endogenous production of oxysterols might be a primary host strategy to counteract a broad panel of viral infections. Moreover, 25HC and 27HC could be considered for new therapeutic strategies against HPV-16, HRoV and HRhV