Interaction Of Uropathogenic Escherichia Coli With Host Urothelial Barrier In The Process Of Urinary Tract Infection

Part I Effect of doses of uropathogenic Escherichia coli and condition of bladder glycosaminoglycans on initial urinary tract infectionObjectives:To investigate relativities of doses of uropathogenic Escherichia coli (UPEC) and conditions of glycosaminoglycans (GAGs) layer on initial urinary tract infection.Methods:Female mice divided into two groups underwent bladder instillation with protamine sulfate (PS, n=40) solution or sterile saline (control, n=40) for 30 minutes. Each group consisted of four subsets, each subset contained 10 mice. Through bladder instillation, the four subsets in both group were infected by UPEC at doses of 102,103, 104,105 CFU, respectively. After Urine samples were collected, Mice were killed 12h after infections, bladder bacterial tilters were also detected.Results:There was significant difference of bladder bacterial tilters between two groups at different doses of UPEC except 10s CFU. Compared with control group, Mice in PS group, infected at 102 and 103 CFU, were more susceptible to UPEC with a higher incidence of bladder bacterial tilters (P<0.05).Whereas, all mice in both PS and control groups developed positive bladder bacterial tilter infected by UPEC at doses of 104 and 105 CFU. Mice in PS group even got bladder infected by UPEC at a low dose of 102CFU. Moreover, not all mice in both groups with positive bladder bacterial tilters were detected with bacterial urine.Conclusions:Disruptions of GAGs layer may be an essential step of initial UTIs.Part? Persistence of uropathogenic escherichia coli in bladders of female patients with sterile urine after antibiotic therapiesObjectives:To determine the persistence of uropathogenic Escherichia coli (UPEC) in female patients with recurrent urinary tract infection (UT1) caused by E.coli after antibiotic therapies, and to further provide evidence of intracellular bacterial communities (IBCs) formation in UTI.Methods:We collected biopsies of the bladder, and clean-catch urine specimens from 32 women with acute uncomplicated cystitis after antibiotic therapy. All women had histories of UTI. Urine samples and biopsies were analyzed by conventional bacteriological techniques. Phylogenetic group and 16 virulence factors (VFs) of UPEC were determined using polymerase chain reaction. The infection capability of UPEC to produce intracellular infection in bladder tissue was confirmed in a well-characterized mouse model of infection, immunofluorescence and electron microscopy was used for evidence of IBCs formation. Results:All urine specimens were detected sterile. Escherichia coli (E. coil) were cultured in 6 of 32 (18.75%) biopsies from all 32 female patients, and were identified as UPEC by PCR test. Different VFs associated with IBCs formation were identified in all six UPEC isolates. Each UPEC isolate was capable of forming IBCs within the superficial umbrella cells of the bladders of mice.Conclusions:UPEC with distinctive pathological traits and capability of IBCs was firstly found in bladders of women facing antibiotic therapy. Our finds suggests that the IBCs pathogenic pathway may occur in humans, and have important role in UTI recurrence.Part? fimH Induces apoptosis and down-regulates mRNA and protein expressions of GAGs synthetases in Bladder SV-HUC-1 CellsObjective:To investigate the effects of fimH on apoptosis of bladder SV-HUC-1 cells, and synthesis of GAGs.Methods:The apoptosis of SV-HUC-1 cells was tested by using Annexin V-FITC/PI double staining and flow cytometry. The levels of GAGs synthetases including xylosyltransferase 1(Xylt1)?xylosyltransferase 2(Xylt2)?Heparan sulfate 6-O-sulfotransferase 1 (Hs6st1)?N-deacetylase/N-sulfotransferase 1 (Ndst1)?Heparan sulfate (glucosamine)3-O-sulfotransferase 6(Hs3st6) hyaluronan synthasel(?Has1), were detected by using real time PCR and Western blotting.Results:fimH could significantly induce apoptosis of SV-HUC-1 cells in a concentration-dependent manner as compared with control group (P<0.05). After administration of 10?g/ml fimH for 24 h, apoptosis rate was significantly increased. The expression levels of Xylt1?Xylt2?Hs6st1?Ndst1?Hs3st6?Has1 in SV-HUC-1 cells treated with fimH.Conclusion:The results indicated that fimH was able to induce apoptosis of SV-HUC-1 cells, and down-regulates mRNA and protein expressions of GAGs synthetases.