Inhibition of pyroptosis through targeting the activation or pore-formation of GSDMD.

The cell envelope structure of Gram-negative bacteria is unique, composed of two lipid bilayer membranes and an aqueous periplasmic space sandwiched in between. The outer membrane constitutes an extra barrier to limit the exchange of molecules between the cells and the exterior environment. Donnan potential is a membrane potential across the outer membrane, resulted from the selective permeability of the membrane, which plays a pivotal role in the permeability of many antibiotics. In this review, we discussed factors that affect the intensity of the Donnan potential, including the osmotic strength and pH of the external media, the osmoregulated periplasmic glucans trapped in the periplasmic space, and the displacement of cell surface charges. The focus of our discussion is the impact of Donnan potential on the cellular permeability of selected antibiotics including fluoroquinolones, tetracyclines, β-lactams, and trimethoprim.

Active ingredients of medicinal plants have been used to cure several human diseases. Azadirachta indica is one of the most versatile medicinal plants having a wide spectrum of biological activity due to the presence of large number of bioactive compounds. The present study was conducted to evaluate antimicrobial and antioxidant efficiency and phytochemical screening of A. indica leaf extract using methanol as a solvent. A qualitative phytochemical screening was performed for the detection of various phytochemicals. Then, the quantitative determination of total phenols, flavonoids and proanthocyanidins was done and expressed in terms of gallic acid and rutin equivalent. Total phenolic, flavonoid and proanthocyanidin content were found to be 85.9±4.0, 104.9±5.5 and 65.4±13.9 mg/g of plant extract, respectively. Also, the antibacterial activity was performed using six different bacterial strains: Escherichia coli (ATCC 25922), Staphylococcus aureus (ATCC 25923), Salmonella typhi (ATCC 14028), Klebsiella pneumoniae (ATCC 700603), Pseudomonas aeruginosa (ATCC 27853) and Proteus vulgaris (ATCC 35659). It was found that the maximum zone of inhibition of 22±3 mm was shown against S. aureus using 700 µg plant extract. Similarly, the antioxidant activity of the methanolic extract of plant was also determined and it was found that maximum inhibition obtained was 71.23% when 500 µg plants extract was used. Key words: Azadirachta indica, methanol, antibacterial, antioxidant.

The RND family efflux pump AcrAB-TolC in E. coli and its homologs in other Gram-negative bacteria are major players in conferring multidrug resistance to the cells. While the structure of the pump complex has been elucidated with ever-increasing resolution through crystallography and Cryo-EM efforts, the dynamic assembly process remains poorly understood. Here, we tested the effect of overexpressing functionally defective pump components in wild type E. coli cells to probe the pump assembly process. Incorporation of a defective component is expected to reduce the efflux efficiency of the complex, leading to the so called “dominant negative” effect. Being one of the most intensively studied bacterial multidrug efflux pumps, many AcrA and AcrB mutations have been reported that disrupt efflux through different mechanisms. We examined five groups of AcrB and AcrA mutants, defective in different aspects of assembly and substrate efflux. We found that none of them demonstrated the expected dominant negative effect, even when expressed at concentrations many folds higher than their genomic counterpart. The assembly of the AcrAB-TolC complex appears to have a proof-read mechanism that effectively eliminated the formation of futile pump complex.

We evaluated the quality of wines produced in Nepal in terms of phenolic, flavonoid, anthocyanin and tannin content, antioxidant capacity, and color parameters using spectrophotometric methods. The total phenolic content, total flavonoid content, and total antioxidant activities in Nepali wines ranged from 85.5 to 960.0 (mean = 360.5 ± 268.7) mg/L GAE, 40.9–551.3 (mean = 188.9 ± 161.5) mg/L QE, and 66.6–905.0 (mean = 332.8 ± 296.5) mg/L AAE, respectively. These parameters were significantly higher in red wines compared to white wines. The phenolic and flavonoid content showed strong correlation with each other as well as with antioxidant activities. Additional parameters measured included various color parameters and carbohydrates. The wine color showed strong correlation with phenol, flavonoid, and antioxidant activity, whereas this correlation was not significant with anthocyanin content. Multivariate analysis was carried out to better describe and discriminate the wine samples. Finally, we compared Nepali wines with wines from other countries.

Histones are cationic nuclear proteins that are essential for the structure and functions of eukaryotic chromatin. However, extracellular histones trigger inflammatory responses and contribute to death in sepsis by unknown mechanisms. We recently reported that inflammasome activation and pyroptosis trigger coagulation activation through a tissue-factor (TF)-dependent mechanism. We used a combination of various deficient mice to elucidate the molecular mechanism of histone-induced coagulation. We showed that histones trigger coagulation activation in vivo, as evidenced by coagulation parameters and fibrin deposition in tissues. However, histone-induced coagulopathy was neither dependent on intracellular inflammasome pathways involving caspase 1/11 and gasdermin D (GSDMD), nor on cell surface receptor TLR2- and TLR4-mediated host immune response, as the deficiency of these genes in mice did not protect against histone-induced coagulopathy. The incubation of histones with macrophages induced lytic cell death and phosphatidylserine (PS) exposure, which is required for TF activity, a key initiator of coagulation. The neutralization of TF diminished the histone-induced coagulation. Our findings revealed lytic cell death as a novel mechanism of histone-induced coagulation activation and thrombosis.