Host-pathogen checkpoints and population bottlenecks in persistent and intracellular uropathogenic Escherichia coli bladder infection

Bladder infections affect millions of people yearly, and recurrent symptomatic infections (cystitis) are very common. The rapid increase in infections caused by multidrug-resistant uropathogens threatens to make recurrent cystitis an increasingly troubling public health concern. Uropathogenic Escherichia coli (UPEC) cause the vast majority of bladder infections. Upon entry into the lower urinary tract, UPEC face obstacles to colonization that constitute population bottlenecks, reducing diversity, and selecting for fit clones. A critical mucosal barrier to bladder infection is the epithelium (urothelium). UPEC bypass this barrier when they invade urothelial cells and form intracellular bacterial communities (IBCs), a process which requires type 1 pili. IBCs are transient in nature, occurring primarily during acute infection. Chronic bladder infection is common and can be either latent, in the form of the quiescent intracellular reservoir (QIR), or active, in the form of asymptomatic bacteriuria (ASB/ABU) or chronic cystitis. In mice, the fate of bladder infection, QIR, ASB, or chronic cystitis, is determined within the first 24 h of infection and constitutes a putative host–pathogen mucosal checkpoint that contributes to susceptibility to recurrent cystitis. Knowledge of these checkpoints and bottlenecks is critical for our understanding of bladder infection and efforts to devise novel therapeutic strategies.

Escherichia coli isolates causing asymptomatic bacteriuria in catheterized and noncatheterized individuals possess similar virulence properties

Urinary tract infections (UTIs) are among the most common infectious diseases of humans, with Escherichia coli being responsible for >80% of all cases. Asymptomatic bacteriuria (ABU) occurs when bacteria colonize the urinary tract without causing clinical symptoms and can affect both catheterized patients (catheter-associated ABU [CA-ABU]) and noncatheterized patients. Here, we compared the virulence properties of a collection of ABU and CA-ABU nosocomial E. coli isolates in terms of antibiotic resistance, phylogenetic grouping, specific UTI-associated virulence genes, hemagglutination characteristics, and biofilm formation. CA-ABU isolates were similar to ABU isolates with regard to the majority of these characteristics; exceptions were that CA-ABU isolates had a higher prevalence of the polysaccharide capsule marker genes kpsMT II and kpsMT K1, while more ABU strains were capable of mannose-resistant hemagglutination. To examine biofilm growth in detail, we performed a global gene expression analysis with two CA-ABU strains that formed a strong biofilm and that possessed a limited adhesin repertoire. The gene expression profile of the CA-ABU strains during biofilm growth showed considerable overlap with that previously described for the prototype ABU E. coli strain, 83972. This is the first global gene expression analysis of E. coli CA-ABU strains. Overall, our data suggest that nosocomial ABU and CA-ABU E. coli isolates possess similar virulence profiles.

Relevance of the deletion polymorphisms of the glutathione S-transferases GSTT1 and GSTM1 in pharmacology and toxicology

Although cytosolic glutathione S-transferase (GST) enzymes occupy a key position in biological detoxification processes, two of the most relevant human isoenzymes, GSTT1-1 and GSTM1-1, are genetically deleted (non-functional alleles GSTT1*0 and GSTM1*0) in a high percentage of the human population, with major ethnic differences. The structures of the GSTT and GSTM gene areas explain the underlying genetic processes. GSTT1-1 is highly conserved during evolution and plays a major role in phase-II biotransformation of a number of drugs and industrial chemicals, e.g. cytostatic drugs, hydrocarbons and halogenated hydrocarbons. GSTM1-1 is particularly relevant in the deactivation of carcinogenic intermediates of polycyclic aromatic hydrocarbons. Several lines of evidence suggest that hGSTT1-1 and/or hGSTM1-1 play a role in the deactivation of reactive oxygen species that are likely to be involved in cellular processes of inflammation, ageing and degenerative diseases. There is cumulating evidence that combinations of the GSTM1*0 state with other genetic traits affecting the metabolism of carcinogens (CYP1A1, GSTP1) may predispose the aero-digestive tract and lung, especially in smokers, to a higher risk of cancer. The GSTM1*0 status appears also associated with a modest increase in the risk of bladder cancer, consistent with a GSTM1 interaction with carcinogenic tobacco smoke constituents. Both human GST deletions, although largely counterbalanced by overlapping substrate affinities within the GST superfamily, have consequences when the organism comes into contact with distinct man-made chemicals. This appears relevant in industrial toxicology and in drug metabolism.

Association of CYP1B1 codon 432 mutant allele in head and neck squamous cell cancer is reflected by somatic mutations of p53 in tumor tissue

Tobacco use is causally associated with head and neck squamous cell cancer (HNSCC). Here, we present the results of a case-control study that investigated the effects that the genetic variants of the cytochrome (CYP)1A1, CYP1B1, glutathione-S-transferase (GST)M1, GSTT1, and GSTP1 genes have on modifying the risk of smoking-related HNSCC. Allelisms of the CYP1A1, GSTT1, GSTM1, and GSTT1 genes alone were not associated with an increased risk. CYP1B1 codon 432 polymorphism was found to be a putative susceptibility factor in smoking-related HNSCC. The frequency of CYP1B1 polymorphism was significantly higher (P < 0.001) in the group of smoking cases when compared with smoking controls. Additionally, an odds ratio (OR) of 4.53 (2.62-7.98) was discovered when investigating smoking and nonsmoking cases for the susceptible genotype CYP1B1*2/*2, when compared with the presence of the genotype wild type. In combination with polymorphic variants of the GST genes, a synergistic-effect OR was observed. The calculated OR for the combined genotype CYP1B1*2/*2 and GSTM1*2/*2 was 12.8 (4.09-49.7). The calculated OR for the combined genotype was 13.4 (2.92-97.7) for CYP1B1*2/*2 and GSTT1*2/*2, and 24.1 (9.36-70.5) for the combination of CYP1B1*2/*2 and GSTT1-expressors. The impact of the polymorphic variants of the CYP1B1 gene on HNSCC risk is reflected by the strong association with the frequency of somatic mutations of the p53 gene. Smokers with susceptible genotype CYP1B1*2/*2 were 20 times more likely to show evidence of p53 mutations than were those with CYP1B1 wild type. Combined genotype analysis of CYP1B1 and GSTM1 or GSTT1 revealed interactive effects on the occurrence of p53 gene mutations. The results of the present study indicate that polymorphic variants of CYP1B1 relate significantly to the individual susceptibility of smokers to HNSCC.

DNA binding study of isophorone in rats and mice

Following isophorone exposure, in a 2-year study with F344 rats and B6C3F1 mice performed under the National Toxicology Program (NTP), an elevated incidence of tumors was observed in male rats (kidney tumors) and male mice (liver tumors). Female rats and mice showed no elevation of tumor rates by isophorone (NTP 1986).

Species differences in the glutathione transferase GSTT1-1 activity towards the model substrates methyl chloride and dichloromethane in liver and kidney

Glutathione transferase (GST) GSTT1-1 is involved in the biotransformation of several chemicals widely used in industry, such as butadiene and dichloro methane DCM. The polymorphic hGSTT1-1 may well play a role in the development of kidney tumours after high and long-term occupational exposure against trichloroethylene. Although several studies have investigated the association of this polymorphism with malignant diseases little is known about its enzyme activity in potential extrahepatic target tissues. The known theta-specific substrates methyl chloride (MC) dichloromethane and 1,2-epoxy-3-(p-nitrophenoxy)propane (EPNP) were used to assay GSTT1-1 activity in liver and kidney of rats, mice, hamsters and humans differentiating the three phenotypes (non-conjugators, low conjugators, high conjugators) seen in humans. In addition GSTT1-1 activity towards MC and DCM was determined in human erythrocytes. No GSTT1-1 activity was found in any tissue of non-conjugators (NC). In all organs high conjugators (HC) showed twofold higher activity towards MC and DCM than low conjugators (LC). The activity in human samples towards EPNP was too close to the detection limit to differentiate between the three conjugator phenotypes. GSTT1-1 activity towards MC was two to seven-times higher in liver cytosol than in kidney cytosol. The relation for MC between species was identical in both organs: mouse > HC > rat > LC > hamster > NC. In rats, mice and hamsters GSTT1-1 activity in liver cytosol towards DCM was also two to seven-times higher than in the kidney cytosol. In humans this activity was twice as high in kidney cytosol than in liver cytosol. The relation between species was mouse > rat > HC > LC > hamster > NC for liver, but mouse > HC > LC/rat > hamster/NC for kidney cytosol. The importance to heed the specific environment at potential target sites in risk assessment is emphasized by these results.

Comparison of GST theta activity in liver and kidney of four species

Glutathione transferases (GSTs) catalyzing the conjugation of glutathione with electrophilic substrates are important enzymes in the metabolism of xenobiotics. Several isozymes exhibit polymorphisms in humans. The two deletion polymorphisms of hGSTM1 and hGSTT1 result in total loss of enzyme activity in homozygous null genotype (GSTM1*0 and GSTT1*0 respectively) individuals (Seidegård et al. 1988; Pemble et al. 1994). Individuals that are heterozygous for hGSTT1 show distinctly lower enzyme activities than individuals carrying two functional alleles of hGSTT1 (Wiebel et al. 1996). A similar effect is conceivable for the hGSTM1 polymorphism but has not been verified so far.

Determination of the ethylene oxide adduct S-(2-hydroxyethyl)cysteine by a fluorometric HPLC method in albumin and globin from human blood

A new method has been developed for the quantification of 2-hydroxyethylated cysteine resulting as adduct in blood proteins after human exposure to ethylene oxide, by reversed-phase HPLC with fluorometric detection. The specific adduct is analysed in albumin and in globin. After isolation of albumin and globin from blood, acid hydrolysis of the protein and precolumn derivatisation of the digest with 9-fluorenylmethoxycarbonylchloride, the levels of derivatised S-hydroxyethylcysteine are analysed by RP-HPLC and fluorescence detection, with a detection limit of 8 nmol/g protein. Background levels of S-hydroxyethylcysteine were quantified in both albumin and globin, under special consideration of the glutathione transferase GSTT1 and GSTM1 polymorphisms. GSTT1 polymorphism had a marked influence on the physiological background alkylation of cysteine. While S-hydroxyethylcysteine levels in "non-conjugators" were between 15 and 50 nmol/g albumin, "low conjugators" displayed levels between 8 and 21 nmol/g albumin, and "high conjugators" did not show levels above the detection limit. The human GSTM1 polymorphism had no apparent effect on background levels of blood protein 2-hydroxyethylation.

Cerebellar grey-matter deficits, cannabis use and first-episode schizophrenia in adolescents and young adults

Epidemiological data link adolescent cannabis use to psychosis and schizophrenia, but its contribution to schizophrenia neuropathology remains controversial. First-episode schizophrenia (FES) patients show regional cerebral grey- and white-matter changes as well as a distinct pattern of regional grey-matter loss in the vermis of the cerebellum. The cerebellum possesses a high density of cannabinoid type 1 receptors involved in the neuronal diversification of the developing brain. Cannabis abuse may interfere with this process during adolescent brain maturation leading to ‘schizophrenia-like’ cerebellar pathology. Magnetic resonance imaging and cortical pattern matching techniques were used to investigate cerebellar grey and white matter in FES patients with and without a history of cannabis use and non-psychiatric cannabis users. In the latter group we found lifetime dose-dependent regional reduction of grey matter in the right cerebellar lobules and a tendency for more profound grey-matter reduction in lobule III with younger age at onset of cannabis use. The overall regional grey-matter differences in cannabis users were within the normal variability of grey-matter distribution. By contrast, FES subjects had lower total cerebellar grey-matter : total cerebellar volume ratio and marked grey-matter loss in the vermis, pedunculi, flocculi and lobules compared to pair-wise matched healthy control subjects. This pattern and degree of grey-matter loss did not differ from age-matched FES subjects with comorbid cannabis use. Our findings indicate small dose-dependent effects of juvenile cannabis use on cerebellar neuropathology but no evidence of an additional effect of cannabis use on FES cerebellar grey-matter pathology.