We characterized the tension response of clathrin-mediated endocytosis by using various cell manipulation methodologies. Elevated tension in a cell hinders clathrin-mediated endocytosis through inhibition of coat initiation, elongation of clathrin coat lifetimes and reduction of high-magnitude growth rates. Actin machinery supplies an inward pulling force necessary for internalization of clathrin coats under high tension. These findings suggest that the physical cues cells receive from their microenvironment are major determinants of clathrin-mediated endocytic activity.
Positively charged polymeric materials have been an alternative to combat bacteria as they exhibit inherently antibacterial potency via bacteria membrane disruption. In this study, we report facile preparation of positively charged core‐crosslinked polymeric micelles with inherent antibacterial properties. Spherical micelles were prepared by self‐assembling of poly(4‐vinylpyridine)‐b‐(oligoethylene glycol methyl ether methacrylate) copolymer in aqueous solution. Herein, quaternization reaction was utilized for the first time to core crosslink the micelles through the pyridine rings utilizing their hydrophobic core and thus resulting positively charged nanostructures. Dynamic light scattering (DLS) results identified that the micelles have an average hydrodynamic diameter of 114 nm with a polydispersity index ranging between 0.105 and 0.114. The electrophoretic light scattering (ELS) measurements demonstrated that the micelles have zeta potential values ranging from +38 to +63 mV. It was evident from both ELS and DLS results that the micelles in solution exhibit long‐term stability as the samples were able to maintain their size and charge even after a year of storage. Further, the micelles exhibited inherently antibacterial activity against Escherichia coli and furthermore, this antibacterial efficacy was sustained over a period of 1 year. These stable core‐crosslinked micelles are proposed to have great potential as antibacterial materials for long‐term applications. © 2019 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2020, 137, 48393.
Background: Brucellosis is a zoonotic disease that causes serious economic losses due to factors such as miscarriages and decreased milk yield in animals. Existing live vaccines have some disadvantages, so effective vaccines need to be developed with new technological approaches. Objectives: The primary objectives of this study were the expression and purification of recombinant Omp25 fusion protein from B. abortus and the evaluation of the effect of the Omp25 protein on cell viability and inflammatory response. Methods: The omp25 gene region was amplified by a polymerase chain reaction and cloned into a Pet102/D-TOPO expression vector. The protein expression was carried out using the procaryotic expression system. The recombinant Omp25 protein was purified with affinity chromatography followed by GPC (Gel Permeation Chromatography). The MTS assay and cytokine-release measurements were carried out to evaluate cell viability and inflammatory response, respectively Results: It was determined that doses of the recombinant Omp25 protein doses greater than 0.1 μg/mL are toxic to RAW cells. Doses of 1 µg/mL and lower significantly increased inflammation due to nitric oxide (NO) levels. ELISA results show that IFN-γ was produced in stimulated RAW 264.7 cells at a dose that did not affect the viability (0.05 µg/mL). However, IL-12, which is known to have a dual role in the activation of macrophages, did not show a statistically significant difference at the same dose. Conclusion: Studies of cell viability and Th1‐related cytokine release suggest that Omp25 protein is a promising candidate molecule for vaccine development.
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