OBJECTIVES Overexpression of efflux pumps in Acinetobacter baumannii is a common mechanism of multidrug resistance in this nosocomial pathogen. Increased efflux pump expression is often assumed from MICs of antibiotics and dyes, without measurement of efflux levels. This study describes a safe, rapid and simple 96-well plate assay that measures the accumulation of a fluorescent dye, Hoechst (H)...

Sugars such as glucose are transported into Escherichia coli by a coupled phosphorylation mechanism (the phosphoenolpyruvate:sugar phosphotransferase system, PTS). Transport of sugars through the PTS results in inhibition of adenylate cyclase [ATP pyrophosphate-lyase (cyclizing), EC 4.6.1.1] activity by a mechanism involving a change in the state of phosphorylation of PTS proteins. Other sugars...

Assay conditions for the measurement of 25-hydroxyvitamin D3-24-hydroxylase activity in rat kidney mitochondria have been worked out. The product, 24,25-dihydroxyvitamin D3 was quantitated either by high pressure liquid chromatography or by isotope dilution-mass spectrometry. By these procedures, the enzyme activity could be measured with saturating concentration (greater than 2.5 X 10(-6) M) o...

BACKGROUNDS Autophagy is an important process in the pathogenesis of diabetes and plays a critical role in maintaining cellular homeostasis. However, the autophagic response and its mechanism in diabetic vascular endothelium remain unclear. METHODS AND RESULTS We studied high-glucose-induced renin-angiotensin system (RAS)-mitochondrial damage and its effect on endothelial cells. With regard t...

Differentiation-inducing factor-1 [1-(3,5-dichloro-2,6-dihydroxy-4-methoxyphenyl)hexan-1-one (DIF-1)] is an important regulator of cell differentiation and chemotaxis in the development of the cellular slime mold Dictyostelium discoideum However, the entire signaling pathways downstream of DIF-1 remain to be elucidated. To characterize DIF-1 and its potential receptor(s), we synthesized two flu...

We examined the effects of metabolic inhibition on intracellular Ca(2+) release in single pulmonary arterial smooth muscle cells (PASMCs). Severe metabolic inhibition with cyanide (CN, 10 mM) increased intracellular calcium concentration ([Ca(2+)](i)) and activated Ca(2+)-activated Cl(-) currents [I(Cl(Ca))] in PASMCs, responses that were greatly inhibited by BAPTA-AM or caffeine. Mild metaboli...

Multidrug-resistant strain Acinetobacter baumannii BM4454 was isolated from a patient with a urinary tract infection. The adeB gene, which encodes a resistance-nodulation-cell division (RND) protein, was detected in this strain by PCR with two degenerate oligodeoxynucleotides. Insertional inactivation of adeB in BM4454, which generated BM4454-1, showed that the corresponding protein was respons...

Salivary histatin 5 (Hst 5) is a cationic salivary protein with high fungicidal activity against Candida albicans. Binding to the cell wall followed by intracellular translocation is required for killing; however, specific binding components and critical toxic events are not understood. In this study, laminarin (beta-1,3-glucan) but not sialic acid, mannan or pustulan mediated Hst 5 binding to ...

Rotaviruses are nonenveloped viruses that infect enterocytes of the small intestine and cause severe infantile gastroenteritis. It was previously thought that rotavirus exits cells by lysis, but this behavior does not match the local pathogenesis of the virus. In this study, we have investigated the release of the simian rotavirus strain (RRV) from the polarized intestinal Caco-2 cells. We foun...

myo-Inositol hexakisphosphate (InsP6), which is found in soil and most, if not all, plant and animal cells, has been estimated to have an affinity for Fe3+ in the range of 10(25) to 10(30) M-1. In this report, we demonstrate that the Fe-InsP6 complex has siderophore activity and is able to reverse the iron-restricted growth inhibition of Pseudomonas aeruginosa by ethylene diamine di(o-hydroxyph...