Recent evidence has shown that nanoparticles that have been used to improve or create new functional properties for common products may pose potential risks to human health. Silicon dioxide (SiO₂) has emerged as a promising therapy vector for the heart. However, its potential toxicity and mechanisms of damage remain poorly understood. This study provides the first exploration of SiO₂-induced toxicity in cultured cardiomyocytes exposed to 7- or 670-nm SiO₂ particles. We evaluated the mechanism of cell death in isolated adult cardiomyocytes exposed to 24-h incubation. The SiO₂ cell membrane association and internalization were analyzed. SiO₂ showed a dose-dependent cytotoxic effect with a half-maximal inhibitory concentration for the 7 nm (99.5 ± 12.4 µg/ml) and 670 nm (>1,500 µg/ml) particles, which indicates size-dependent toxicity. We evaluated cardiomyocyte shortening and intracellular Ca²⁺ handling, which showed impaired contractility and intracellular Ca²⁺ transient amplitude during β-adrenergic stimulation in SiO₂ treatment. The time to 50% Ca²⁺ decay increased 39%, and the Ca²⁺ spark frequency and amplitude decreased by 35 and 21%, respectively, which suggest a reduction in sarcoplasmic reticulum Ca²⁺-ATPase (SERCA) activity. Moreover, SiO₂ treatment depolarized the mitochondrial membrane potential and decreased ATP production by 55%. Notable glutathione depletion and H₂O₂ generation were also observed. These data indicate that SiO₂ increases oxidative stress, which leads to mitochondrial dysfunction and low energy status; these underlie reduced SERCA activity, shortened Ca²⁺ release, and reduced cell shortening. This mechanism of SiO₂ cardiotoxicity potentially plays an important role in the pathophysiology mechanism of heart failure, arrhythmias, and sudden death. NEW & NOTEWORTHY Silica particles are used as novel nanotechnology-based vehicles for diagnostics and therapeutics for the heart. However, their potential hazardous effects remain unknown. Here, the cardiotoxicity of silica nanoparticles in rat myocytes has been described for the first time, showing an impairment of mitochondrial function that interfered directly with Ca²⁺ handling.

The hemagglutinin (HA) surface glycoprotein promotes influenza virus entry and is the key protective antigen in natural immunity and vaccines. The HA protein is a trimeric envelope glycoprotein consisting of a globular receptor-binding domain (HA-RBD) that is inserted into a membrane fusion-mediating stalk domain. Similar to other class I viral fusion proteins, the fusogenic stalk domain spontaneously refolds into its postfusion conformation when expressed in isolation, consistent with this domain being trapped in a metastable conformation. Using X-ray crystallography, we show that the influenza virus HA-RBD refolds spontaneously into its native, immunogenic structure even when expressed in an unglycosylated form in Escherichia coli . In the 2.10-Å structure of the HA-RBD, the receptor-binding pocket is intact and its conformational epitopes are preserved. Recombinant HA-RBD is immunogenic and protective in ferrets, and the protein also binds with specificity to sera from influenza virus-infected humans. Overall, the data provide a structural basis for the rapid production of influenza vaccines in E. coli . From an evolutionary standpoint, the ability of the HA-RBD to refold spontaneously into its native conformation suggests that influenza virus acquired this domain as an insertion into an ancestral membrane-fusion domain. The insertion of independently folding domains into fusogenic stalk domains may be a common feature of class I viral fusion proteins.

The A/H1N1/2009 influenza pandemic made evident the need for faster and higher-yield methods for the production of influenza vaccines. Platforms based on virus culture in mammalian or insect cells are currently under investigation. Alternatively, expression of fragments of the hemagglutinin (HA) protein in prokaryotic systems can potentially be the most efficacious strategy for the manufacture of large quantities of influenza vaccine in a short period of time. Despite experimental evidence on the immunogenic potential of HA protein constructs expressed in bacteria, it is still generally accepted that glycosylation should be a requirement for vaccine efficacy. We expressed the globular HA receptor binding domain, referred to here as HA_63–286-RBD, of the influenza A/H1N1/2009 virus in Escherichia coli using a simple, robust and scalable process. The recombinant protein was refolded and purified from the insoluble fraction of the cellular lysate as a single species. Recombinant HA_63–286-RBD appears to be properly folded, as shown by analytical ultracentrifugation and bio-recognition assays. It binds specifically to serum antibodies from influenza A/H1N1/2009 patients and was found to be immunogenic, to be capable of triggering the production of neutralizing antibodies, and to have protective activity in the ferret model. Projections based on our production/purification data indicate that this strategy could yield up to half a billion doses of vaccine per month in a medium-scale pharmaceutical production facility equipped for bacterial culture. Also, our findings demonstrate that glycosylation is not a mandatory requirement for influenza vaccine efficacy.

Intracellular Ca²⁺ mishandling is an underlying mechanism in hypoxia/reoxygenation (H/R) injury that results in mitochondrial dysfunction and cardiomyocytes death. These events are mediated by mitochondrial Ca²⁺ (mCa²⁺) overload that is facilitated by the mitochondrial calcium uniporter (MCU) channel. Along this line, we evaluated the effect of siRNA‐targeting MCU in cardiomyocytes subjected to H/R injury. First, cardiomyocytes treated with siRNA demonstrated a reduction of MCU expression by 67%, which resulted in significant decrease in mitochondrial Ca²⁺ transport. siRNA treated cardiomyocytes showed decreased mitochondrial permeability pore opening and oxidative stress trigger by Ca²⁺ overload. Furthermore, after H/R injury MCU silencing decreased necrosis and apoptosis levels by 30% and 50%, respectively, and resulted in reduction in caspases 3/7, 9, and 8 activity. Our findings are consistent with previous conclusions that demonstrate that MCU activity is partly responsible for cellular injury induced by H/R and support the concept of utilizing siRNA‐targeting MCU as a potential therapeutic strategy.