A uropathogenic E. coli UTI89 model of prostatic inflammation and collagen accumulation for use in studying aberrant collagen production in the prostate.
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ABSTRACT:
SUBMITTER: Ruetten H
PROVIDER: S-EPMC7847049 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:We genetically engineered expression of an activated form of P110 alpha, the catalytic subunit of PI3K, in mouse prostate epithelium to create a mouse model of direct PI3K activation (Pbsn-cre4Prb;PI3KGOF/+ ). We hypothesized that direct activation would cause rapid neoplasia and cancer progression. Pbsn-cre4Prb;PI3KGOF/+ mice developed widespread prostate intraepithelial hyperplasia, but stromal invasion was limited and overall progression was slower than anticipated. However, the model produced profound and progressive stromal remodeling prior to explicit epithelial neoplasia. Increased stromal cellularity and inflammatory infiltrate were evident as early as 4 months of age and progressively increased through 12 months of age, the terminal endpoint of this study. Prostatic collagen density and phosphorylated SMAD2-positive prostatic stromal cells were expansive and accumulated with age, consistent with pro-fibrotic TGF-β pathway activation. Few reported mouse models accumulate prostate-specific collagen to the degree observed in Pbsn-cre4Prb;PI3KGOF/+ . Our results indicate a signaling process beginning with prostatic epithelial PI3K and TGF-β signaling that drives prostatic stromal hypertrophy and collagen accumulation. These mice afford a unique opportunity to explore molecular mechanisms of prostatic collagen accumulation that is relevant to cancer progression, metastasis, inflammation and urinary dysfunction. © 2019 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.
| S-EPMC7071816 | biostudies-literature
Project description:Uropathogenic Escherichia coli (UPEC) is the major cause of urinary tract infections. Nearly half of all UPEC strains secrete hemolysin, a cytotoxic pore-forming toxin. Here, we show that the prevalence of the hemolysin toxin gene (hlyA) is highly variable among the most common 83 E. coli sequence types (STs) represented on the EnteroBase genome database. To explore this diversity in the context of a defined monophyletic lineage, we contextualized sequence variation of the hlyCABD operon within the genealogy of the globally disseminated multidrug-resistant ST131 clone. We show that sequence changes in hlyCABD and its newly defined 1.616-kb-long leader sequence correspond to phylogenetic designation, and that ST131 strains with the strongest hemolytic activity belong to the most extensive multidrug-resistant sublineage (clade C2). To define the set of genes involved in hemolysin production, the clade C2 strain S65EC was completely sequenced and subjected to a genome-wide screen by combining saturated transposon mutagenesis and transposon-directed insertion site sequencing with the capacity to lyse red blood cells. Using this approach, and subsequent targeted mutagenesis and complementation, 13 genes were confirmed to be specifically required for production of active hemolysin. New hemolysin-controlling elements included discrete sets of genes involved in lipopolysaccharide (LPS) inner core biosynthesis (waaC, waaF, waaG, and rfaE) and cytoplasmic chaperone activity (dnaK and dnaJ), and we show these are required for hemolysin secretion. Overall, this work provides a unique description of hemolysin sequence diversity in a single clonal lineage and describes a complex multilevel system of regulatory control for this important toxin.IMPORTANCE Uropathogenic E. coli (UPEC) is the major cause of urinary tract infections and a frequent cause of sepsis. Nearly half of all UPEC strains produce the potent cytotoxin hemolysin, and its expression is associated with enhanced virulence. In this study, we explored hemolysin variation within the globally dominant UPEC ST131 clone, finding that strains from the ST131 sublineage with the greatest multidrug resistance also possess the strongest hemolytic activity. We also employed an innovative forward genetic screen to define the set of genes required for hemolysin production. Using this approach, and subsequent targeted mutagenesis and complementation, we identified new hemolysin-controlling elements involved in LPS inner core biosynthesis and cytoplasmic chaperone activity, and we show that mechanistically they are required for hemolysin secretion. These original discoveries substantially enhance our understanding of hemolysin regulation, secretion and function.
| S-EPMC6775461 | biostudies-literature
Project description:Benign prostatic hyperplasia (BPH) is an age-related debilitating prostatic disease that is frequently associated with prostatic inflammation and bothersome lower urinary tract symptoms (LUTS). Animal models have shown that formalin- and bacterial-induced prostatic inflammation can induce bladder dysfunction; however, the underlying mechanisms contributing to prostatic inflammation in BPH and bladder dysfunction are not clear. We previously reported that E-cadherin expression in BPH is downregulated in hyperplastic nodules compared with expression in adjacent normal tissues. Here, we explored the potential consequences of prostatic E-cadherin downregulation on the prostate and bladder in vivo using an inducible murine model of prostate luminal epithelial-specific deletion of Cdh1. The prostate-specific antigen (PSA)-CreERT2 transgenic mouse strain expressing tamoxifen-inducible CreERT2 recombinase driven by a 6-kb human PSA promoter/enhancer was crossed with the B6.129-Cdh1tm2Kem/J mouse to generate bigenic PSA-CreERT2/Cdh1-/- mice. Deletion of E-cadherin was induced by transient administration of tamoxifen when mice reached sexual maturity (7 weeks of age). At 21 to 23 weeks of age, the prostate, bladder, and prostatic urethra were examined histologically, and bladder function was assessed using void spot assays and cystometry. Mice with Cdh1 deletion had increased prostatic inflammation, prostatic epithelial hyperplasia, and stromal changes at 21 to 23 weeks of age, as well as changes in bladder voiding function compared with age-matched controls. Thus, loss of E-cadherin in the murine prostate could result in prostatic defects that are characteristic of BPH and LUTS, suggesting that E-cadherin downregulation could be a driving force in human BPH development and progression.
| S-EPMC7745638 | biostudies-literature
Project description:Uropathogenic Escherichia coli (UPEC) is a major cause of urinary tract and bloodstream infections and possesses an array of virulence factors for colonization, survival, and persistence. One such factor is the polysaccharide K capsule. Among the different K capsule types, the K1 serotype is strongly associated with UPEC infection. In this study, we completely sequenced the K1 UPEC urosepsis strain PA45B and employed a novel combination of a lytic K1 capsule-specific phage, saturated Tn5 transposon mutagenesis, and high-throughput transposon-directed insertion site sequencing (TraDIS) to identify the complement of genes required for capsule production. Our analysis identified known genes involved in capsule biosynthesis, as well as two additional regulatory genes (mprA and lrhA) that we characterized at the molecular level. Mutation of mprA resulted in protection against K1 phage-mediated killing, a phenotype restored by complementation. We also identified a significantly increased unidirectional Tn5 insertion frequency upstream of the lrhA gene and showed that strong expression of LrhA induced by a constitutive Pcl promoter led to loss of capsule production. Further analysis revealed loss of MprA or overexpression of LrhA affected the transcription of capsule biosynthesis genes in PA45B and increased sensitivity to killing in whole blood. Similar phenotypes were also observed in UPEC strains UTI89 (K1) and CFT073 (K2), demonstrating that the effects were neither strain nor capsule type specific. Overall, this study defined the genome of a UPEC urosepsis isolate and identified and characterized two new regulatory factors that affect UPEC capsule production.IMPORTANCE Urinary tract infections (UTIs) are among the most common bacterial infections in humans and are primarily caused by uropathogenic Escherichia coli (UPEC). Many UPEC strains express a polysaccharide K capsule that provides protection against host innate immune factors and contributes to survival and persistence during infection. The K1 serotype is one example of a polysaccharide capsule type and is strongly associated with UPEC strains that cause UTIs, bloodstream infections, and meningitis. The number of UTIs caused by antibiotic-resistant UPEC is steadily increasing, highlighting the need to better understand factors (e.g., the capsule) that contribute to UPEC pathogenesis. This study describes the original and novel application of lytic capsule-specific phage killing, saturated Tn5 transposon mutagenesis, and high-throughput transposon-directed insertion site sequencing to define the entire complement of genes required for capsule production in UPEC. Our comprehensive approach uncovered new genes involved in the regulation of this key virulence determinant.
| S-EPMC5654933 | biostudies-literature
Project description:In Escherichia coli, the small regulatory noncoding RNA (sRNA) RyhB and the global ferric uptake regulator (Fur) mediate iron acquisition and storage control. Iron is both essential and potentially toxic for most living organisms, making the precise maintenance of iron homeostasis necessary for survival. While the roles of these regulators in iron homeostasis have been well studied in a nonpathogenic E. coli strain, their impact on the production of virulence-associated factors is still unknown for a pathogenic E. coli strain. We thus investigated the roles of RyhB and Fur in iron homeostasis and virulence of the uropathogenic E. coli (UPEC) strain CFT073. In a murine model of urinary tract infection (UTI), deletion of fur alone did not attenuate virulence, whereas a ΔryhB mutant and a Δfur ΔryhB double mutant showed significantly reduced bladder colonization. The Δfur mutant was more sensitive to oxidative stress and produced more of the siderophores enterobactin, salmochelins, and aerobactin than the wild-type strain. In contrast, while RyhB was not implicated in oxidative stress resistance, the ΔryhB mutant produced lower levels of siderophores. This decrease was correlated with the downregulation of shiA (encoding a transporter of shikimate, a precursor of enterobactin and salmochelin biosynthesis) and iucD (involved in aerobactin biosynthesis) in this mutant grown in minimal medium or in human urine. iucD was also downregulated in bladders infected with the ΔryhB mutant compared to those infected with the wild-type strain. Our results thus demonstrate that the sRNA RyhB is involved in production of iron acquisition systems and colonization of the urinary tract by pathogenic E. coli.
| S-EPMC4249264 | biostudies-literature
Project description:In addition to their pivotal role in thrombosis and wound repair, platelets participate in inflammatory responses. We investigated the role of platelets in the autoimmune disease rheumatoid arthritis. We identified platelet microparticles--submicrometer vesicles elaborated by activated platelets--in joint fluid from patients with rheumatoid arthritis and other forms of inflammatory arthritis, but not in joint fluid from patients with osteoarthritis. Platelet microparticles were proinflammatory, eliciting cytokine responses from synovial fibroblasts via interleukin-1. Consistent with these findings, depletion of platelets attenuated murine inflammatory arthritis. Using both pharmacologic and genetic approaches, we identified the collagen receptor glycoprotein VI as a key trigger for platelet microparticle generation in arthritis pathophysiology. Thus, these findings demonstrate a previously unappreciated role for platelets and their activation-induced microparticles in inflammatory joint diseases.
| S-EPMC2927861 | biostudies-literature
Project description:ObjectiveTo evaluate predictors of prostatic chronic inflammation (PCI) and prostate cancer (PCa) in patients undergoing transperineal baseline random prostatic needle biopsies (BNB).Patient and methodsAccording to BNB outcomes, patients were divided into four groups: cases without PCI or PCa (Control group), cases with PCI only (PCI group), cases with PCa and PCI (PCa+PCI group) and cases with PCa only (PCa group). A multinomial logistic regression model was used to evaluate the association of clinical factors with BNB outcomes. Additionally, clinical factors associated with the risk of PCa in the overall population were investigated using a multivariable logistic regression model (univariate and multivariate analysis).ResultsOverall, 945 patients were evaluated and grouped as follows: Control group, 308 patients (32.6%); PCI group, 160 (16.9%); PCa+PCI group, 45 (4.8%); and PCa group, 432 (45.7%). Amongst these, PCa was independently predicted by age (odds ratio [OR] 1.081), prostate specific-antigen level (PSA; OR 1.159), transition zone volume (TZV; OR 0.916), and abnormal digital rectal examination (DRE; OR 1.962). PCa and PCI (4.8%) were independently predicted by age (OR 1.081), PSA level (OR 1.122) and TZV (OR 0.954). In the group without PCa, the PSA level was the only factor associated with the risk of PCI when compared to the control group (OR 1.051, P = 0.042). Among patients with PCa, independent factors associated with the risk of only PCa compared to cases with PCA+PCI were TZV (OR 0.972) and number of positive cores (OR 1.149). In the overall population, PCI was the strongest predictor of a decreased risk of PCa (multivariate model, OR 0.212; P < 0.001).ConclusionsAt BNB, PCI was associated with both a decreased risk of PCa and less aggressive tumour biology amongst patients with PCa. The presence of PCI on biopsy cores should be reported because of its implications in clinical practice.AbbreviationsBGG: biopsy Gleason Group; BPC: biopsy positive (cancer) cores; BMI: body mass index; FGF-2: fibroblast growth factor 2; IL: interleukin; ISUP: International Society of Urologic Pathology; NIH: National Institutes of Health; OR: odds ratio; PCa: prostate cancer; PCI: prostatic chronic inflammation; TGF: transforming growth factor; TPV: total prostate volume; TZV: transition zone volume.
| S-EPMC7473292 | biostudies-literature
Project description:Adrenal androgens dehydroepiandrosterone (DHEA) and DHEA-sulfate (DHEAS) are potential substrates for intracrine production of testosterone (T) and dihydrotestosterone (DHT), or directly to DHT, by prostate cancer (PCa) cells. Production of DHT from DHEAS and DHEA, and the role of steroid sulfatase (STS), were evaluated ex vivo using fresh human prostate tissue and in vitro using human PCa cell lines. STS was expressed in benign prostate tissue and PCa tissue. DHEAS at a physiological concentration was converted to DHT in prostate tissue and PCa cell lines, which was STS-dependent. DHEAS activation of androgen receptor (AR) and stimulation of PCa cell growth were STS-dependent. DHEA at a physiological concentration was not converted to DHT ex vivo and in vitro, but stimulated in vivo tumor growth of the human PCa cell line, VCaP, in castrated mice. The findings suggest that targeting metabolism of DHEAS and DHEA may enhance androgen deprivation therapy.
| S-EPMC6438375 | biostudies-literature
Project description:Uropathogenic E. coli (UPEC) is the dominant cause of urinary tract infections, clinically described as cystitis. UPEC express CUP pili, which are extracellular fibers tipped with adhesins that bind mucosal surfaces of the urinary tract. Here we identify the role of the F9/Yde/Fml pilus for UPEC persistence in the inflamed urothelium. The Fml adhesin FmlH binds galactose β1-3 N-acetylgalactosamine found in core-1 and -2 O-glycans. Deletion of fmlH had no effect on UPEC virulence in an acute mouse model of cystitis. However, FmlH provided a fitness advantage during chronic cystitis, which is manifested as persistent bacteriuria, high bladder bacterial burdens, and chronic inflammation. In situ binding confirmed that FmlH bound avidly to the inflamed, but not the naive bladder. In accordance with its pathogenic profile, vaccination with FmlH significantly protected mice from chronic cystitis. Thus, UPEC employ separate CUP pili to adapt to the rapidly changing niche during bladder infection.
| S-EPMC5294914 | biostudies-literature
Project description:BACKGROUND:Prostate stiffness and increased collagen content both associate with the presence of urinary symptoms in men but mechanisms responsible, including impact of age and androgens, are unknown. Dogs develop prostate-related urinary dysfunction similar to humans, but mechanisms are also unknown. Mice have been used to examine how prostatic collagen accumulation affects voiding but whether mouse prostatic collagen organization resembles human or dog has not been evaluated. Here, we have constructed the first comprehensive, comparative maps of collagen architecture in canine, human, and mouse prostate and test whether canine prostatic collagen content is increased by aging and reduced by castration. METHODS:Complete transverse prostate sections were stained with picrosirius red and imaged with confocal microscopy to reveal and compare collagen architecture across species. Canine prostatic collagen fiber length, diameter, and density in prostatic urethral, periurethral, peripheral, and capsular regions were quantified and compared among four experimental groups: young intact, young neutered, old intact, and old neutered dogs. RESULTS:Surprisingly, the majority of collagen was localized to the prostatic urethra in canine, human, and mouse. In canine and human, capsular regions also featured a dense collagen network but it appeared less dense than around prostatic urethra. Older, intact male canines exhibited overall denser prostate collagen fibers and had thicker capsular fibers than young, intact males. Prostatic glandular regions undergo dramatic atrophy and regression following castration, and our finding of neutered animals having increased collagen fiber density in both periurethral and peripheral regions is consistent with glandular contraction and increased proportion of stroma. CONCLUSIONS:Collagen architecture in dog appears similar to that in humans when cross sections are compared side-by-side. Canine collagen organization is affected by both age and androgen status, suggesting these factors may contribute to collagen accumulation in some males.
| S-EPMC6356104 | biostudies-literature