KmrA multidrug efflux pump from Klebsiella pneumoniae

We cloned a gene responsible for multidrug resistance from the chromosomal DNA of Klebsiella pneumoniae MGH78578 that showed multidrug resistance. We designated the gene kmrA. The deduced amino acid sequence of KmrA was similar to that of SmvA that is responsible for methyl viologen-resistance in Salmonella enterica sv. Typhi and Typhimurium. Introduction of the cloned kmrA gene into drug-hypersensitive Escherichia coli KAM32 cells made them resistant to acriflavine, 4',6-diamidino-2-phenylindole (DAPI), Hoechst 33342, tetraphenylphosphonium chloride (TPPCl), methyl viologen and ethidium bromide. We observed elevated energy-dependent efflux of ethidium in E. coli cells carrying the kmrA gene compared with control cells. We also cloned the smvA gene from S. enterica sv. Typhimurium LT2 and investigated the resistance pattern for several drugs. The pattern was similar between KmrA from K. pneumoniae and SmvA from S. enterica.     

Gene cloning and characterization of SdrM, a chromosomally-encoded multidrug efflux pump, from Staphylococcus aureus

There are more than 30 genes for putative multidrug efflux pumps in the chromosome of Staphylococcus aureus. Only a few of these have been analyzed so far. Here we cloned a new gene, SA1972, using a PCR method, from the chromosome of S. aureus N315. We found that the product SA1972 could lead to elevated resistance against several antimicrobial agents such as norfloxacin, acriflavine and ethidium bromide. We designated the gene as sdrM. We observed elevated energy-dependent efflux of acriflavine in S. aureus cells introduced with the sdrM gene. We conclude that SdrM is a multidrug efflux pump belonging to the major facilitator (MF) superfamily.     

Effect of glimepiride and glibenclamide on S-warfarin 7-hydroxylation by human liver microsomes, recombinant human CYP2C9.1 and CYP2C9.3

The effect of glimepiride on metabolism of S-warfarin to 7-hydroxywarfarin was studied using human liver microsomes and recombinant cytochrome P450 2C9 microsomes (CYP2C9.1 and CYP2C9.3), and was compared with the results from the experiments using glibenclamide as an inhibitor. S-Warfarin 7-hydroxylation by recombinant CYP2C9.1 and CYP2C9.3 was inhibited by glimepiride competitively. The apparent K(i) value of glimepiride was lower at CYP2C9.3 than at CYP2C9.1. Glimepiride also inhibited 7-hydroxylation of S-warfarin in a competitive manner by microsomes from human liver which showed the genotypes of CYP2C9, as CYP2C9*1/*1 or CYP2C9*1/*3. The apparent K(i) value of glimepiride was lower than that of glibenclamide. These results may provide valuable information for optimizing the anticoagulant activity of warfarin when glimepiride is co-administered to patients.     

Histological changes of muscle in a patient with pyruvate dehydrogenase deficiency

Histological changes of muscle from a 17-month-old boy with pyruvate dehydrogenase deficiency are presented. The patient had muscle hypotonia, mental retardation, seizures, lactic acidosis and hyperalaninemia. Deficient activity of the pyruvate dehydrogenase complex was found in his platelets (about 25% of normal) and of pyruvate dehydrogenase in his biopsied muscle (about 5% of normal). A muscle biopsy specimen showed an increased proportion of type IIC fibers (24%), fiber-type grouping and lipid droplet accumulation.     

Uterotropic hormones produced by ovaries of Sl/Slt mutant mice before spontaneous development of tubular adenomas

Tubular adenomas developed spontaneously in ovaries of (WB X C57BL/6)F1-Sl/Slt and -W/Wv mice after 150 days of age. The uteri of the W/Wv mice were hypoplastic before development of tubular adenomas, but the uteri of the Sl/Slt mice were not. Since oophorectomy significantly reduced the uterus weight in the Sl/Slt mice, we investigated the androgen- and estrogen-producing activity of Sl/Slt and W/Wv mice before development of tubular adenomas. Fragments of ovaries were cultured for 48 hr in serum-free medium containing either progesterone or 4-androstene-3,17-dione (androstenedione). The amount of androgens produced from progesterone by ovaries of Sl/Slt mice in the presence of human chorionic gonadotropin was comparable to that of W/Wv mice. In contrast, the amount of estrogens produced from androstenedione by ovaries of Sl/Slt mice in the presence of follicle stimulating hormone was much greater than that of W/Wv mice. Moreover, the amount of estrogens produced was much greater than the amount of androgens in the ovaries of Sl/Slt mice. Therefore, before development of tubular adenomas, estrogens rather than androgens seem to be a major uterotropic hormone produced by ovaries of Sl/Slt mice.