Testosterone enhances susceptibility to chronic UTI. Bladders left and kidney pairs right were homogenized, serially diluted, and CFU enumerated at 2 wpi.
Associated Data
The dotted horizontal line shows the limit of detection. Mice were surgically infected 4 weeks postgonadectomy at 9 weeks of age.
Data comprise three independent experiments of four to five mice per group. The limit of detection is indicated with the dotted horizontal line. To specify whether male susceptibility to severe UTI is mediated by androgens or by another testicular factor, we implanted slow-release testosterone or placebo pellets 5 days after castration or sham operation. Consistent with the above results, castration attenuated severe UTI in placebo-treated males Figure 6C. Testosterone-complemented castrated males were statistically indistinguishable from placebo-treated sham-operated males. Collectively, these data indicate that androgens mediate male susceptibility to chronic cystitis and severe pyelonephritis.
In this study, we report the first preclinical UTI model to permit direct elucidation of sex differences in susceptibility and host response to infection, using a novel inoculation technique that bypasses anatomic differences in the lower urinary tracts of male and female mice. Further, these males developed more advanced pyelonephritis and highly prevalent abscess formation compared with females.
These phenotypes were mitigated in males who underwent gonadectomy prior to infection, but were complemented with exogenous testosterone. Our findings indicate that the influences of sex on susceptibility to multiple forms of UTI are complex and not limited to traditionally cited anatomic differences. In addition, our minimally invasive model provides a new preclinical platform for translatable studies of upper- and lower-tract UTI pathogenesis in both male and female hosts. To successfully colonize the urinary tract, UPEC must circumvent formidable host defenses, which are mechanic, biochemical, and immunologic in nature.
Males that resolved acute infection also maintained a small population of persisting UPEC within bladder tissue, likely representing the quiescent intracellular reservoirs believed to seed same-strain recurrent infection in females. The chronic cystitis phenotype has proven to be an important outcome in female models investigating the natural history of UTI and host responses to repeated UPEC inoculation.
Sex differences in immune response have been demonstrated in a number of human infections and animal models; in many of these studies, females display more robust resistance to pathogens. Consistent with this paradigm, female mice more aptly controlled both lower and upper UTI compared with males in our model. Additionally, females displayed higher local proinflammatory cytokines at 24 hpi when bladder UPEC burdens were similar between sexes , suggesting that a more pronounced acute inflammatory response may be raised in the female urinary tract to better control infection.
In addition to potential evolutionary pressure to repel urinary pathogens, some evidence has suggested that estrogen may impact immune response to UTI, although the influence of male androgens on UTI outcome has not been explored. The exact organizational or activational influences of androgens on UTI severity in both males and females represent areas of ongoing investigation. Notably, our findings suggest that clinical modulation of androgens may represent a potential therapeutic route to combat recalcitrant or recurrent UTI.
At a glance, our findings in this new model may appear to represent a stark contradiction to the female predominance in human UTI epidemiology. However, minimally invasive inoculation bypasses anatomic sex differences below the bladder. Thus, our data are in fact consistent with the long-standing presumption that UTI risk in females is potentiated by anatomic features. This paradigm is consonant with clinical data reflecting increased morbidity and mortality in males who develop pyelonephritis and complicated UTI, compared with females with these conditions. In addition to facilitating the study of sex differences, our minimally invasive model fills another substantial gap in the basic investigation of UTI.
No published approaches in experimental UTI result in more than a very small minority of wild-type animals developing an ascending renal abscess. Our work therefore opens a new avenue into modeling the development, therapy, resolution, and sequelae of severe pyelonephritis and ascending renal abscess. Beyond these niches, our method also induces visible infection of the prostate, making the model potentially useful for the study of bacterial prostatitis.
In summary, the work described here will accelerate fundamental investigation into the mechanisms of UTI initiation, progression, and persistence in male hosts, as well as expanding studies of pyelonephritis, ascending renal abscess, and the treatment of complicated UTI. These advances also represent a timely response to recent calls by the National Institutes of Health NIH 57 and in the basic and clinical literature 58 — 60 for sex-based research approaches to infectious and other diseases, including a specific focus on UTI.
Animal care and use protocols received prior approval from the Washington University Animal Studies Committee. The female murine model of cystitis with transurethral inoculation via catheter has been described in methodologic detail. A 2—3 mm, vertical, midline incision was made directly overlying the bladder, first through the abdominal skin and then the peritoneum. The bladder was exposed through the incision using gentle bilateral pressure on the abdomen, and aseptically emptied.
The inoculum was prepared in a 1-ml tuberculin syringe adapted to a gauge, 0. The bladder was allowed to expand for an additional 10 seconds before withdrawing the needle; the injection site sealed immediately with no evident leakage. The bladder was gently replaced, the peritoneum and skin were closed separately with simple, interrupted sutures, and the animal was awakened in fresh air. Infections were allowed to proceed for durations from 6 hours to 4 weeks.
Postinfection, clean-catch urine samples were collected using gentle suprapubic pressure, serially diluted and plated to Luria-Bertani agar to enumerate CFU. For organ titers, mice were euthanized by CO 2 asphyxiation, and bladders and kidney pairs were aseptically removed, homogenized in 1 ml or 0. The concentrations of 23 mouse cytokines in whole-bladder homogenates or sera were analyzed with a multiplex bead array platform Bio-Plex; Bio-Rad, Hercules, CA as described previously.
Splayed bladder halves were fixed in 2. Quantification of intracellular and extracellular bacteria in the murine bladder at 6 hpi was performed by a modified ex vivo gentamicin protection assay, as previously described. Fixed tissues were embedded in paraffin, sectioned, and stained with hematoxylin and eosin or Gomori trichrome stain. Bilateral orchiectomy castration , bilateral ovariectomy, or sham operations were performed at Harlan Laboratories Indianapolis, IN following standard procedures at 5 weeks of age.
Animals were allowed to recover 1 week before shipment. Where indicated, day continuous release pellets containing 25 mg testosterone or placebo Innovative Research of America, Sarasota, FL were implanted subcutaneously at the nape of the neck 5 days following castration or sham operation. Mice were surgically infected at 9 weeks of age i.
Androgens Enhance Male Urinary Tract Infection Severity in a New Model
To compare proportions of mice developing persistent bacteriuria and chronic cystitis or renal abscess, a two-tailed Fisher exact test was used. Beatty for technical assistance, and S. Hultgren for critical review of the manuscript. Published online ahead of print. Publication date available at www. This article contains supplemental material online at http: National Center for Biotechnology Information , U. J Am Soc Nephrol. Published online Oct 8.
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Louis, Missouri Find articles by David A. Author information Article notes Copyright and License information Disclaimer. Hunstad, Washington University, S. Received Mar 27; Accepted Aug This article has been cited by other articles in PMC. Abstract Urinary tract infections UTIs occur predominantly in females but also affect substantial male patient populations; indeed, morbidity in complicated UTI is higher in males.
Open in a separate window. Discussion In this study, we report the first preclinical UTI model to permit direct elucidation of sex differences in susceptibility and host response to infection, using a novel inoculation technique that bypasses anatomic differences in the lower urinary tracts of male and female mice. Determination of Urine and Tissue Bacterial Loads Postinfection, clean-catch urine samples were collected using gentle suprapubic pressure, serially diluted and plated to Luria-Bertani agar to enumerate CFU.
Tissue and Serum Cytokine Analysis The concentrations of 23 mouse cytokines in whole-bladder homogenates or sera were analyzed with a multiplex bead array platform Bio-Plex; Bio-Rad, Hercules, CA as described previously. Ex vivo Gentamicin Protection Assay Quantification of intracellular and extracellular bacteria in the murine bladder at 6 hpi was performed by a modified ex vivo gentamicin protection assay, as previously described.
Gonadectomy and Testosterone Implantation Bilateral orchiectomy castration , bilateral ovariectomy, or sham operations were performed at Harlan Laboratories Indianapolis, IN following standard procedures at 5 weeks of age. Supplementary Material Supplemental Data: Click here to view. Acknowledgments We thank K. Footnotes Published online ahead of print. Epidemiology of urinary tract infections: Acute pyelonephritis in US hospitals in Acute pyelonephritis in adults: Arch Intern Med Hospitalization for acute pyelonephritis in Manitoba, Canada, during the period from to ; impact of diabetes, pregnancy, and aboriginal origin.
Clin Infect Dis The epidemiology of acute pyelonephritis in South Korea, Am J Epidemiol Sickness impact of chronic nonbacterial prostatitis and its correlates. Urol Clin North Am Clin Microbiol Rev The epidemiology of urinary tract infection. Nat Rev Urol 7: Selective urological evaluation in men with febrile urinary tract infection. Recurrent urinary tract infection in women. Int J Antimicrob Agents Perineal anatomy and urine-voiding characteristics of young women with and without recurrent urinary tract infections.
A murine model of urinary tract infection. Urinary tract infection of mice to model human disease: Practicalities, implications and limitations. Crit Rev Microbiol Cell Host Microbe 1: Development of intracellular bacterial communities of uropathogenic Escherichia coli depends on type 1 pili. Bacterial invasion augments epithelial cytokine responses to Escherichia coli through a lipopolysaccharide-dependent mechanism.
Urothelial cultures support intracellular bacterial community formation by uropathogenic Escherichia coli.
Androgens Enhance Male Urinary Tract Infection Severity in a New Model
Type 1 pilus-mediated bacterial invasion of bladder epithelial cells. Intracellular bacterial biofilm-like pods in urinary tract infections. Establishment of a persistent Escherichia coli reservoir during the acute phase of a bladder infection. Integrin-mediated host cell invasion by type 1-piliated uropathogenic Escherichia coli. Bacterial penetration of bladder epithelium through lipid rafts. J Biol Chem Female cystoscopy During a cystoscopy exam, your doctor inserts a thin, flexible device called a cystoscope through the urethra into the bladder. Male cystoscopy During a cystoscopy exam, your doctor inserts a thin, flexible device called a cystoscope through the urethra into the bladder.
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