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Biomedical subjects

J W Warren

Publications and source records attributed to J W Warren.

At least 55 records · Page 3Linked to original sources
PubMed1995

Cytolethality of hemolytic Escherichia coli to primary human renal proximal tubular cell cultures obtained from different donors.

OBJECTIVES: In earlier experiments, we confirmed epidemiologic studies demonstrating the prominence in acute pyelonephritis of Escherichia coli expressing P fimbriae and hemolysin, produced the disease with pyelonephritogenic strains in an animal model, and developed in vitro assays using human renal proximal tubular cells that demonstrated bacterial adherence by P fimbriae and killing of the renal cells by hemolysin. In the present series of experiments, we sought to determine whether P-fimbriated hemolytic E coli killed human renal proximal tubular epithelial cells obtained from different human donors. METHODS: Human renal proximal tubular cells, putative target cells for bacteria causing acute pyelonephritis, were cultured from 9 donors and cell death was measured by two methods. RESULTS: We showed that the E coli strain was significantly more cytolethal for renal cells of all donors than its hemolysin-negative mutant. CONCLUSIONS: This work suggests that the pathogenesis of acute pyelonephritis by P-fimbriated hemolytic E coli, characteristics of the causative organism in about 50% of human cases, may be at least in part through killing of human renal epithelial cells by hemolysin.

Cell Survival↗
PubMed1995

Culture of bladder epithelium from cystoscopic biopsies of patients with interstitial cystitis.

Interstitial cystitis is a chronic disease of unknown etiology characterized by bladder pain and urinary frequency and urgency. The epithelium may be critical in its pathogenesis; the hallmarks of the disease are visible epithelial defects (Hunner's ulcers and epithelial ruptures). Areas denuded of epithelium are commonly seen, and defects in epithelial permeability are characteristic. We report here the culture and characterization of epithelial cells from cystoscopic bladder biopsies obtained from 7 female patients with interstitial cystitis. Within 4 to 14 days cellular outgrowths appeared from explants incubated in cell medium. Monolayers reached confluence after 6 weeks. Cells of the monolayer were cytokeratin-positive and smooth muscle actin-negative, confirming their epithelial origin. They exhibited epithelial cell ultrastructure including intermediate filaments and junctional complexes. Vesicles bounded by a trilaminar plasma membrane and lateral interdigitations were also present. This is the first report of the culture of bladder epithelium from interstitial cystitis patients. Epithelial cells may be targets for initiating agents and inflammatory effects of interstitial cystitis and should be useful for studies of the pathogenesis of this disease.

Adult↗
PubMed1995

Organ-limited autopsies. Obtaining permission for postmortem examination of the urinary tract.

OBJECTIVE: To develop a systematic pre- and postmortem program for obtaining permission for autopsies limited to the urinary tract. DESIGN: Comparison versus historical control of autopsy permission using a new process comprising premortem education, modification of the permission process, and autopsy limited to the urinary tract. PARTICIPANTS AND SETTING: Patients, family members, and healthcare providers associated with a nursing home. INTERVENTION: Education of healthcare providers and family members and modification of autopsy consent and performance. MAIN OUTCOME MEASURE: Permission rate and demographic characteristics of patients and family members giving permission. RESULTS: Of the 361 patients who died during the study, we received permission for postmortem examination of the urinary tract for 129 (36%). This was substantially better than the autopsy rate in the year prior to the study, 0.6%. A significantly higher permission rate was associated with patients and contact persons of white race, a higher level of education of the contact person, a nonrelative contact person, and two or fewer individuals making the decision. CONCLUSION: Permission for organ-limited autopsies can be obtained with a systematic approach.

Aged↗
PubMed1994

Is interstitial cystitis an infectious disease?

Interstitial cystitis (IC) is manifest by years of urinary frequency, urgency, and bladder pain and on cystoscopy, is diagnosed by petechial hemorrhages or ulcers. The etiology is unknown; the prominent theories are that IC is an autoimmune disease or is linked to increased permeability of the bladder mucosa. Although sought, no infectious agent has ever been identified. The disease has many characteristics of a chronic infection and the author's opinion is that an infectious disease has not been properly ruled out. To do so would require culture of bladder epithelium (not just urine) using special culture and non-culture techniques such as polymerase chain reaction. Infection can easily be integrated into the autoimmune and permeability theories of IC pathogenesis. A possible analogue for this disease is chronic gastritis in which Helicobacter pylori has been identified as an etiological agent.

Bacteria↗
PubMed1994

Colonization of urinary catheters by Escherichia coli and Providencia stuartii in a laboratory model system.

A laboratory model system was developed to investigate the progressive colonization of urinary catheters by Escherichia coli Providencia stuartii, prominent organisms in the polymicrobial bacteriuria of the long-term catheterized urinary tract. We hypothesized that colonization of the catheter and artificial urine by E. coli is influenced by the presence of P. stuartii. E. coli or P. stuartii in pure culture both rapidly colonized the artificial urine and catheters, and both persisted throughout all experiments. In systems containing both organisms, P. stuartii occurred in significantly higher numbers in the artificial urine and on the catheters than E. coli (p < 0.05). To obtain similar numbers of E. coli and P. stuartii in the artificial urine, citrate was eliminated; however, P. stuartii still dominated on the catheter surfaces. The presence of P. stuartii appeared to facilitate growth of E. coli in the artificial urine, yet reduce numbers of E. coli on the catheter. In a separate experiment using different strains of E. coli and P. stuartii, the latter was dominant in the artificial urine and on the catheter surfaces. However, this strain of P. stuartii (which was urease positive) did not facilitate growth of E. coli. The interaction between these strains may have been considerably affected by urea hydrolysis, which resulted in an increase in pH (6.5 to > 8.5) and considerable precipitate formation in the model system. The paradox of P. stuartii enhancing colonization by E. coli in the artificial urine, yet inhibiting its colonization on the catheter surface, illustrates the complexity of polymicrobial interactions in colonization of the catheterized urinary tract.

Bacteriuria↗
PubMed1994

Binding to and killing of human renal epithelial cells by hemolytic P-fimbriated E. coli.

Acute pyelonephritis is a common invasive infection frequently caused by E. coli that possess P-fimbriae and secrete hemolysin. We have examined the role of P fimbriae and hemolysin in the killing of putative target cells of acute pyelonephritis, that is, human renal epithelial cells (HRPTEC). Cultures of HRPTEC were overlaid with (1) a prototypic pyelonephritogenic E. coli (CFT073) which expresses both P fimbriae and hemolysin; (2) its hemolysin-negative isogenic mutant (CFT073hlyD::TnphoA); or (3) a prototypic nonpyelonephritogenic fecal E. coli (FN414) which is negative for both P fimbriae and hemolysin. CFT073 and CFT073hlyD::TnphoA but not FN414 adhered to HRPTEC, as demonstrated by electron microscopy and direct counting. Adherence was diminished by antisera directed against P fimbriae and by a monoclonal antibody recognizing the epithelial receptor for P fimbriae. CFT073 was significantly more cytolethal for HRPTEC than its hemolysin-negative mutant. The bacteria-free filtrate of CFT073 was both hemolytic and cytolethal whereas that of CFT073hyD::TnphoA was not hemolytic and was significantly less cytolethal. Finally, we demonstrated that CFT073 passed through monolayers of HRPTEC at a higher rate than CFT073hlyD::TnphoA, indicating that hemolysin damages HRPTEC, facilitating passage of bacteria through the epithelial barrier. With HRPTEC and a pyelonephritogenic strain of E. coli we have reproduced in vitro bacterial attachment and toxin delivery by P fimbriae and hemolysin, factors epidemiologically associated with acute pyelonephritis in patients.

Bacterial Adhesion↗
PubMed1994

Catheter-associated bacteriuria in long-term care facilities.

Catheter-associated bacteriuria is the most common infection acquired in long-term care facilities. Complications include fever, acute pyelonephritis, bacteremia, catheter obstruction, urinary stones, chronic interstitial nephritis, renal failure, and death. The closed-catheter system has been the only innovation in this traditional method of care that has led to prevention of bacteriuria. Antimicrobial agents rarely are indicated to prevent or treat bacteriuria in long-term catheterized patients, except for those with symptomatic infection. Alternative devices are available and often may be preferable to the indwelling urethral catheter. These patients and their reservoirs of bacteriuric organisms are sources of nosocomial outbreaks. Such outbreaks can be prevented and controlled with attention to catheter hygiene, prevention of patient-to-patient transmission on the hands of caregivers, and possibly use of antimicrobials to diminish bacterial concentrations in the urine.

Anti-Bacterial Agents↗
PubMed1994

Internalization of Escherichia coli into human kidney epithelial cells: comparison of fecal and pyelonephritis-associated strains.

A gentamicin survival assay, using primary human renal epithelial cells and Escherichia coli strains isolated from the feces of asymptomatic individuals and from the urine or blood of patients with acute pyelonephritis, was used to investigate bacterial internalization as a model for renal parenchymal invasion in pyelonephritis. E. coli strains, regardless of their origin, efficiently entered into human renal epithelial cells, a process inhibited by cytochalasin D. While the percentage of survival of nonhemolytic pyelonephritis isolates did not differ from that of fecal isolates, survival of hemolytic pyelonephritis strains was lower than that of nonhemolytic strains, perhaps as a consequence of the greater cytotoxicity of hemolytic strains. There was no evidence of intracellular multiplication of E. coli. These results demonstrate that human renal epithelial cells are capable of efficient uptake of E. coli regardless of the source of the bacteria.

Cells, Cultured↗
PubMed1994

Long-term urethral catheterization increases risk of chronic pyelonephritis and renal inflammation.

OBJECTIVE: To determine the prevalences of chronic pyelonephritis and chronic renal inflammation in elderly nursing home patients at the time of death and to assess correlation with urethral catheterization and other putative risk factors. DESIGN: Prospective assessment of risk factors with the prevalences of chronic pyelonephritis and renal inflammation at autopsy. SETTING: A 240-bed long-term care facility. PARTICIPANTS: All residents > or = 65 years old who died and were autopsied during a 2-year period. MEASUREMENTS: Antemortem assessment of risk factors for renal inflammation, including a search for any urethral catheterization in the person's life. Prospective assessment of urethral catheterization, catheter obstruction, and use of anti-inflammatory medications and urine cultures. Urinary tract pathology was assessed for gross and microscopic evidence of inflammation and urinary tract stones. RESULTS: The duration of catheterization was significantly associated with increasing prevalence of bacteriuria, polymicrobial bacteriuria, chronic pyelonephritis, and chronic renal inflammation. The prevalence of chronic pyelonephritis at death was 10 percent (5/52) for patients catheterized > 90 days during their last year of life and zero (0/65) when catheterized < or = 90 days (P < 0.02; Fisher's exact test). Chronic pyelonephritis was significantly associated with renal stones and hydronephrosis. The prevalence of chronic renal inflammation without chronic pyelonephritis was significantly greater than that of chronic pyelonephritis: the prevalence was 43 percent (20/47) when catheterized > 90 days and 18 percent (12/65) when < or = 90 days (P < 0.05). Chronic renal inflammation was associated with hydronephrosis, ureteral dilatation, acute pyelonephritis and diastolic hypertension. CONCLUSION: Chronic pyelonephritis and chronic renal inflammation are associated with long-term catheterization.

Age Factors↗
PubMed1994

Internalization of Proteus mirabilis by human renal epithelial cells.

Proteus mirabilis, a common agent of bacteriuria in humans, causes acute pyelonephritis and bacteremia. Renal epithelium provides a barrier between luminal organisms and the renal interstitium. We have hypothesized that P. mirabilis may be internalized into renal epithelium. To test this hypothesis, we added suspensions of three P. mirabilis strains (10(8) CFU) to confluent monolayers of primary cultures of human renal proximal tubular epithelial cells (HRPTEC) and, after 3 h, found the bacteria internalized within membrane-bound vacuoles by light and electron microscopy. Internalization of bacteria by HRPTEC was corroborated by using the gentamicin protection assay. Cytolysis of HRPTEC by the HpmA hemolysin, however, was a confounding factor in this assay, and therefore a hemolysin-negative hpmA mutant was used in subsequent experiments. The nonhemolytic mutant WPM111 did not disrupt the monolayer and was recovered in numbers that were 10- to 100-fold higher than those of the hemolytic parent BA6163. Cytochalasin D (20 micrograms/ml) inhibited internalization of Salmonella typhimurium but not that of P. mirabilis, suggesting that the latter species enters HRPTEC by a mechanism that is not dependent on actin polymerization. We suggest that HpmA hemolysin-mediated cytotoxicity and internalization of bacteria by HRPTEC may play a role in the development of Proteus pyelonephritis.

Cells, Cultured↗
PubMed1994

Construction of an MR/P fimbrial mutant of Proteus mirabilis: role in virulence in a mouse model of ascending urinary tract infection.

Proteus mirabilis, a cause of acute pyelonephritis, produces at least four types of fimbriae, including MR/P (mannose-resistant/Proteus-like) fimbriae. To investigate the contribution of MR/P fimbriae to colonization of the urinary tract, we constructed an MR/P fimbrial mutant by allelic exchange. A 4.2-kb BamHI fragment carrying the mrpA gene was subcloned into a mobilizable plasmid, pSUP202. A 1.3-kb Kanr cassette was inserted into the mrpA open reading frame, and the construct was transferred to the parent P. mirabilis strain by conjugation. Following passage on nonselective medium, 1 of 500 transconjugants screened was found to have undergone allelic exchange as demonstrated by Southern blot. Colony immunoblot, Western immunoblot, and immunogold labeling with a monoclonal antibody to MR/P fimbriae revealed that MrpA was not expressed. Complementation with cloned mrpA restored MR/P expression as shown by hemagglutination, Western blot, and immunogold electron microscopy. To assess virulence, we challenged 40 CBA mice transurethrally with 10(7) CFU of wild-type or mutant strains. After 1 week, geometric means of log10 CFU per milliliter of urine or per gram of bladder or kidney for the wild-type and mutant strains were as follows: urine, 7.79 (wild type) versus 7.02 (mutant) (P = 0.035); bladder, 6.22 versus 4.78 (P = 0.019); left kidney, 5.02 versus 3.31 (P = 0.009); and right kidney, 5.28 versus 4.46 (P = 0.039). Mice challenged with the wild-type strain showed significantly more severe renal damage than did mice challenged with the MR/P-negative mutant (P = 0.007). We conclude that MR/P fimbriae contribute significantly to colonization of the urinary tract and increase the risk of development of acute pyelonephritis.

Animals↗
PubMed1994

Interstitial cystitis as an infectious disease.

In the author's opinion, infection has not been properly ruled out as a factor in the development of interstitial cystitis. Dismissing infection as a cause would require special techniques to search for microorganisms not only in urine but also in bladder tissue. Infection is not necessarily exclusive of other theories concerning the pathogenesis of interstitial cystitis, such as autoimmunity or increased bladder permeability.

Autoimmune Diseases↗
PubMed1993

Isogenic P-fimbrial deletion mutants of pyelonephritogenic Escherichia coli: the role of alpha Gal(1-4) beta Gal binding in virulence of a wild-type strain.

Escherichia coli strains causing acute pyelonephritis often express multiple fimbrial types and haemolysin, which may contribute to their ability to adhere to, and interact with, kidney epithelial cells. Strain CFT073, a pap+, sfa+, pil+, hly+ pyelonephritis strain, previously established as virulent in the CBA mouse model of ascending urinary tract infection and cytotoxic for cultured human renal epithelial cells, was selected for construction of isogenic strains. From a gene bank of this strain, two distinct copies of the pap operon were isolated. The two P-fimbrial determinants were subcloned into pCVD442, a positive selection suicide vector containing the sacB gene of Bacillus subtilis. Deletion mutations were introduced into each of the two constructs, within papEFG of one operon and papDEFG of the other. Suicide vectors carrying pap deletions were mobilized from E. coli SM10 lambda pir into CFT073 (NalR) and cointegrates were passaged on non-selective medium. The first pap mutation was identified by screening a Southern blot of DNA from sucrose-resistant colonies using a papEFG probe. This mutant retained the MRHA+ phenotype since a second functional copy of pap was still present. A double pap-deletion mutant, UPEC76, confirmed by Southern blotting, was unable to agglutinate human type O erythrocytes or alpha Gal(1-4)beta Gal-coated latex beads. CBA mice (N = 100) were challenged transurethrally with 10(5), 10(6), 10(7), or 10(9) cfu of strains CFT073 or UPEC76. After one week, quantitative cultures of urine, bladder, and kidney were done and histologic changes were examined. No substantive differences in organism concentration or histological findings between parent and mutant were detected in urine, bladder, or kidney at any challenge concentration. We conclude that adherence by P fimbriae of uropathogenic E. coli strain CFT073 plays only a subtle role in the development of acute pyelonephritis in the CBA mouse model.

Acute Disease↗
PubMed1993

Contribution of Proteus mirabilis urease to persistence, urolithiasis, and acute pyelonephritis in a mouse model of ascending urinary tract infection.

Proteus mirabilis, a significant cause of bacteriuria and acute pyelonephritis in humans, produces urease. This high-molecular-weight, multimeric, cytoplasmic enzyme hydrolyzes urea to ammonia and carbon dioxide. To assess the role of urease in colonization, urolithiasis, and acute pyelonephritis in an animal model of ascending urinary tract infection, we compared a uropathogenic strain of P. mirabilis with its isogenic urease-negative mutant, containing an insertion mutation within ureC, the gene encoding the large subunit of the enzyme. Mice challenged transurethrally with the parent strain developed significant bacteriuria and urinary stones. The urease-negative mutant had a 50% infective dose of 2.7 x 10(9) CFU, a value more than 1,000-fold greater than that of the parent strain (2.2 x 10(6) CFU). The urease-positive parent strain reached significantly higher concentrations and persisted significantly longer in the bladder and kidney than did the mutant. Indeed, in the kidney, the parent strain increased in concentration while the mutant concentration fell so that, by 1 week, the parent strain concentration was 10(6) times that of the mutant. Similarly, the urease-positive parent produced significantly more severe renal pathology than the mutant. The initial abnormalities were in and around the pelvis and consisted of acute inflammation and epithelial necrosis. By 1 week, pyelitis was more severe, crystals were seen in the pelvis, and acute pyelonephritis, with acute interstitial inflammation, tubular epithelial cell necrosis, and in some cases abscesses, had developed. By 2 weeks, more animals had renal abscesses and radial bands of fibrosis. We conclude that the urease of P. mirabilis is a critical virulence determinant for colonization, urolithiasis, and severe acute pyelonephritis.

Acute Disease↗
PubMed1993

Urethral obstruction of 6 hours or less causes bacteriuria, bacteremia, and pyelonephritis in mice challenged with "nonuropathogenic" Escherichia coli.

Urethral obstruction may be caused by prostatic hypertrophy, urethral stricture, or encrustation of a urethral-catheter lumen. Bacteriuria often complicates these obstructions. The sequelae include fever, acute pyelonephritis, chronic renal inflammation, and death. We hypothesized that even brief obstruction of the urinary tract containing a nonvirulent bacterium would result in these complications. Mice challenged transurethrally with Escherichia coli FN414, which is rapidly eliminated from normal mice without causing bacteriuria, bacteremia, or renal pathology, were subjected to reversible urethral obstruction by coating the urethral meatus with collodion for 1, 3, or 6 h. The majority of mice obstructed for 1 h demonstrated parenchymal renal inflammation 48 h later. At the end of 3 h of obstruction, 9 of 10 mice were bacteremic; some bacteremias were present at 48 h after removal of the obstruction. At that time, more severe renal inflammation was seen in these mice. As little as 6 h of obstruction resulted not only in the acute changes described above but also in chronic renal inflammation and fibrosis in the majority of animals sacrificed 3 and 6 weeks later. Additional studies demonstrated that urethral obstruction enhanced the uropathogenicity of another nonpathogenic E. coli strain (K-12 strain HB101) and caused more severe renal lesions in mice challenged with E. coli CFT073, isolated from a patient with symptoms of pyelonephritis. These findings demonstrate that brief urethral obstruction may (i) induce organisms which are cleared rapidly from the normal urinary tract to cause bacteriuria, bacteremia, and pyelonephritis and (ii) intensify the renal lesions caused by a uropathogen.

Animals↗
PubMed1992

Visual and profilometric wear measurements.

Wear of composites can be estimated by the degree of marginal discrepancy between the prepared cavity wall and the occlusal margins of composites. Such evaluations are done on casts by comparing and rating the marginal discrepancy with those on standard casts. We analyzed the reliability of this technique on metal and stone specimens. These specimens contained grooves of different width and depth. For the visual comparison we used stone casts of machined standards of known groove depth. We measured the depths of the metal specimens with a profilometer and made stone casts of these original specimens. Using the stone casts of the standards, five dentists estimated the unknown groove depths on the remaining stone casts. These estimates were done under standardized conditions and repeated by each dentist on five different occasions. The results showed that visual depth evaluations of die stone specimens underestimated the depths when compared with the values measured with a profilometer on the original metal models. One investigator gave significantly different (p less than 0.05) groove depth estimates at different occasions.

Calcium Sulfate↗
PubMed1992

Catheter-associated bacteriuria.

Catheter-associated bacteriuria is the most common nosocomial infection in both acute and long-term care facilities. Complications include fever, bacteremia, acute pyelonephritis, urinary stones, chronic renal inflammation, and death. Catheter-associated bacteriuria can be postponed by maintaining the closed nature of the catheter system. Postponement may become prevention of bacteriuria if the catheter is removed as soon as possible.

Aged↗