Central residues of the amphipathic ß-hairpin loop control the properties of Clostridium perfringens epsilon-toxin channel.

Clostridium perfringens epsilon toxin (ETX) is a heptameric pore-forming toxin of the aerolysin toxin family. ETX is the most potent toxin of this toxin family and the third most potent bacterial toxin with high cytotoxic and lethal activities in animals. In addition, ETX shows a demyelinating activity in nervous tissue leading to devastating multifocal central nervous system white matter disease in ruminant animals. Pore formation in target cell membrane is most likely the initial critical step in ETX biological activity. Eight single to quadruple ETX mutants were generated by replacement of polar residues (serine, threonine, glutamine) in middle positions of the ß-strands forming the ß-barrel and facing the channel lumen with charged glutamic residues. Channel activity and ion selectivity were monitored in artificial lipid monolayer membranes and cytotoxicity was investigated in MDCK cells by the viability MTT test and propidium iodide entry. All the mutants formed channels with similar conductance in artificial lipid membranes and increasing cation selectivity for increasing number of mutations. Here, we show that residues in the central position of each ß-strand of the amphipathic ß-hairpin loop that forms the transmembrane pore, control the size and ion selectivity of the channel. While the highest cationic ETX mutants were not cytotoxic, no strict correlation was observed between ion selectivity and cytotoxicity.

The protective role of erythropoietin induced by hypoxic preconditioning on cerebral injury and cognition after global cerebral ischemia / 中国中西医结合急救杂志

Objective To observe the protective effects of hypoxic preconditioning(HPC) on the improvement of the cognitive dysfunction(learning and memory) and the damage in hippocampus induced by global cerebral ischemia/reperfusion(I/R) in CA1 and CA3 for 5 days in rats,and on the regulation of expression of erythropoietin(EPO) protein to approach the mechanism of the protection.Methods One hundred and twenty adult male Sprague-Dawley(SD) rats were divided into four groups randomly: sham group,I/R group,HPC24 group(hypoxia for 24 hours before I/R) and HPC48 group(hypoxia for 48 hours before I/R).Hang(motor function),passive avoidance and Morris water maze tests were carried out on the 5th day after I/R to measure the motor and cognition functions;hematoxylin-eosin(HE) staining was used to detected histopathological changes in hippocampus tissues;and the contents of EPO were tested by immunohistochemistry at 1 hour and 4 hours after I/R from hippocampus CA1 and CA3 regions.Results Hang,passive avoidance and Morris water maze tests showed that I/R can injure rat cognition;the improvement of cognition was marked in HPC groups, and it was shown that the effects were more significant in HPC48 group than those in the HPC24 group(P