Epigallocatechin Gallate-Mediated Cell Death Is Triggered by Accumulation of Reactive Oxygen Species Induced via the Cpx Two-Component System in Escherichia coli

The high antimicrobial activity of epigallocatechin gallate (EGCG), the most bioactive component of tea polyphenol with a number of health benefits, is well-known. However, little is known about the mechanism involved. Here, we discovered the relationship between reactive oxygen species (ROS), the Cpx system, and EGCG-mediated cell death. We first found an increase in ampicillin resistance as well as the transcription level of a LD-transpeptidase (LD-TPase) involved in cell wall synthesis; ycbB transcription was upregulated whereas that of another LD-TPase, ynhG, appeared to be constant after a short exposure of Escherichia coli to sub-inhibitory doses of EGCG. Additionally, the transcription level of cpxP, a downstream gene belonging to the Cpx regulon, was positively correlated with the concentration of EGCG, and significant upregulation was detected when cells were treated with high doses of EGCG. Through analysis of a cpxR deletion strain (ΔcpxR), we identified a constant ROS level and a notable increase in the survival rate of ΔcpxR, while the ROS level increased and the survival rate decreased remarkably in the wild-type strain. Furthermore, thiourea, which is an antioxidant, reduced the ROS level and antimicrobial activity of EGCG. Taken together, these results suggest that EGCG induces ROS formation by activating the Cpx system and mediates cell death

Porcine β-defensin 2 inhibits proliferation of pseudorabies virus in vitro and in transgenic mice

BACKGROUND: Porcine β-defensin 2 (PBD-2), produced by host cells, is an antimicrobial cysteine-rich cationic peptide with multi-functions. Previous studies have demonstrated that PBD-2 can kill various bacteria, regulate host immune responses and promote growth of piglets. However, the antiviral role of PBD-2 is rarely investigated. This study aimed to reveal the antiviral ability of PBD-2 against pseudorabies virus (PRV), the causative pathogen of Aujeszky’s disease, in PK-15 cells and in a PBD-2 expressing transgenic (TG) mouse model. METHODS: In this study, the cytotoxicity of PBD-2 on PK-15 cells was measured by CCK-8 assay. PK-15 cells were incubated with PRV pre-treated with different concentrations of PBD-2 and PRV titers in cell culture supernatants were determined by real-time quantitative PCR (RT-qPCR). TG mice and wild-type (WT) mice were intraperitoneally injected with PRV and the survival rate was recorded for 10 days. Meanwhile, tissue lesions in brain, spleen and liver of infected mice were observed and the viral loads of PRV in brain, liver and lung were analyzed by RT-qPCR. RESULTS: PBD-2 at a maximum concentration of 80 μg/mL displayed no significant cytotoxicity on PK-15 cells. A threshold concentration of PBD-2 at 40 μg/mL was required to inhibit PRV proliferation in PK-15 cells. The survival rate in PBD-2 TG mice was 50% higher than that of WT mice. In addition, TG mice showed alleviated tissue lesions in brain, spleen and liver compared with their WT littermates after PRV challenge, while viral loads of PRV in brain, liver and lung of TG mice were significantly lower than that of WT mice. CONCLUSIONS: PBD-2 could inhibit PRV proliferation in PK-15 cells and protect mice from PRV infection, which confirmed the antiviral ability of PBD-2 both in vitro and in vivo. The application of PBD-2 in developing anti-viral drugs or disease-resistant animals can be further investigated

Demonstrating Biological Fate of Nanoparticle-Loaded Dissolving Microneedles with Aggregation-Caused Quenching Probes: Influence of Application Sites

Integrating dissolving microneedles (DMNs) and nanocarriers (NC) holds great potential in transdermal drug delivery because it can simultaneously overcome the stratum corneum barrier and achieve efficient and controlled drug delivery. However, different skin sites with different thicknesses and compositions can affect the transdermal diffusion of NC-loaded DMNs. There are few reports on the biological fate (especially transdermal diffusion) of NC-loaded DMNs, and inaccurate bioimaging information of intact NC limits the accurate understanding of the in vivo fate of NC-loaded DMNs. The aggregation-caused quenching (ACQ) probes P4 emitted intense fluorescence signals in intact NC while quenched after the degradation of NC, had been demonstrated the feasibility of label intact NC. In this study, P4 was loaded in solid lipid nanoparticles (SLNs), and further encapsulated into DMNs, to track the transdermal diffusion of SLNs delivered at different skin sites. The results showed that SLNs had excellent stability after being loaded into DMNs with no significant changes in morphology and fluorescence properties. The in vivo live and ex vivo imaging showed that the transdermal diffusion rate of NC-loaded DMNs was positively correlated with skin thickness, with the order ear > abdomen > back. In conclusion, this study confirmed the site-dependency of transdermal diffusion in NC-loaded DMNs

Multifunctional Nanostructure RAP‐RL Rescues Alzheimer's Cognitive Deficits through Remodeling the Neurovascular Unit

Abstract Cerebrovascular dysfunction characterized by the neurovascular unit (NVU) impairment contributes to the pathogenesis of Alzheimer's disease (AD). In this study, a cerebrovascular‐targeting multifunctional lipoprotein‐biomimetic nanostructure (RAP‐RL) constituted with an antagonist peptide (RAP) of receptor for advanced glycation end‐products (RAGE), monosialotetrahexosyl ganglioside, and apolipoprotein E3 is developed to recover the functional NVU and normalize the cerebral vasculature. RAP‐RL accumulates along the cerebral microvasculature through the specific binding of RAP to RAGE, which is overexpressed on cerebral endothelial cells in AD. It effectively accelerates the clearance of perivascular Aβ, normalizes the morphology and functions of cerebrovasculature, and restores the structural integrity and functions of NVU. RAP‐RL markedly rescues the spatial learning and memory in APP/PS1 mice. Collectively, this study demonstrates the potential of the multifunctional nanostructure RAP‐RL as a disease‐modifying modality for AD treatment and provides the proof of concept that remodeling the functional NVU may represent a promising therapeutic approach toward effective intervention of AD