Inducible prostate luminal epithelial cell-specific deletion of E-cadherin (CDH1) induces prostatic inflammation, proliferation and bladder overactivity – Characterization of a potential animal model for benign prostatic hyperplasia

Mizoguchi S1, Pascal L1, Dhir R1, Rigatti L1, Chen W1, Wang K1, Nelson J1, Chambon P2, Metzger D2, Wang Z1, Yoshimura N1

Research Type

Pure and Applied Science / Translational

Abstract Category

Male Lower Urinary Tract Symptoms (LUTS) / Voiding Dysfunction

Abstract 294
E-Poster 2
Scientific Open Discussion Session 18
Thursday 5th September 2019
13:25 - 13:30 (ePoster Station 1)
Exhibition Hall
Benign Prostatic Hyperplasia (BPH) Overactive Bladder Animal Study
1.University of Pittsburgh, 2.Université de Strasbourg
Presenter
S

Shinsuke Mizoguchi

Links

Abstract

Hypothesis / aims of study
There is increasing evidence indicating a positive correlation between chronic prostatic inflammation and development of benign prostatic hyperplasia (BPH) and male lower urinary tract symptoms (LUTS). Recent studies have shown that prostate specific antigen (PSA), which is exclusively expressed in the luminal epithelial cells of the prostate, can be found in the stromal compartment of prostatic specimens from BPH patients in association with downregulation of E-cadherin, which is an important adherens junction protein required for the epithelial barrier in mucous membrane [1]. These studies suggest that junctional dysfunction by E-cadherin downregulation allows substances in prostatic secretions including PSA to leak into the stromal compartment, which could lead to chronic prostatic inflammation and promote BPH pathogenesis. Thus, in the present study, we examined the impact of reduced barrier function induced by a conditional loss of E-cadherin in the mice prostatic epithelium on bladder function and inflammation-related changes in the prostate tissue.
Study design, materials and methods
The PSA-CreERT2 transgenic mouse strain expressing tamoxifen-inducible Cre-ERT2 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-/- (Cdh1-/-) mice. Deletion of E-cadherin was performed by transient administration of tamoxifen when mice reached sexual maturity (7 weeks of age). At 150 days of age, prostate tissues were harvested and separated to ventrolateral lobes, dorsal lobes and anterior lobes for histological examination and inflammatory related gene expression analyses. Loss of E-cadherin was confirmed by immunostaining.   Epithelial proliferation and inflammatory cell infiltration were evaluated by immunohistostaining for Ki-67 and CD19, respectively. Furthermore, mice with the second tamoxifen administration at 150 days of age were used (WT; n=5, cdh-/-; n=5) to investigate changes in voiding function by voiding spot assay and awake cystometry.  In voiding spot assay, each group of mice were placed individually in a metabolic cage with filter papers for four hours after subcutaneous injection of saline at volume of 20 μl/g (Body weight) followed by free access to regular foods, but not to water. Thereafter, filter papers were collected, dried and evaluated for urine spots under ultraviolet light.  Each urine spot area was analysed with the Fiji version of ImageJ software to determine the number of voiding events and voided volume by using a standard formula generated by the correlation between the amount of urine and stained area.  One day after voiding spot assay, awake cystometry was performed to evaluate intercontraction intervals (ICI) and the number of non-voiding contractions (NVCs) during each voiding cycle.
Results
A mosaic pattern of E-cadherin loss was displayed in all lobes of the prostate, while the bladder showed no changes in E-cadherin, confirming the prostate-specific deletion of E-cadherin. Compared to WT, all lobes of the prostate from mice with Cdh1 deletion demonstrated an increased number of Ki-67 positive cells in epithelium and CD19 positive cells as well as collagen fibres in the stromal compartment.  In qPCR analysis of inflammation-related genes, IL6, IL8, TGFβ, and COX2 mRNA levels were significantly increased in the ventrolateral lobes in the Cdh1-/- group compared to WT.  Voiding spot assay showed significantly decreased single voided volume and higher voiding frequency compared to WT although there was no significant change in total urine volume. In cystometric investigation, there was no significant change in ICI between WT and Cdh1-/- groups while mice with cdh1 deletion demonstrated a significant increase in NVCs/min compared to WT.
Interpretation of results
Mice with conditional knockout of E-cadherin in the prostatic epithelium exhibited significant epithelial proliferative activity, chronic inflammation and stromal fibrosis as evidenced by increases in the number of Ki-67 positive cells in the prostatic epithelium, a B cell marker (CD19 positive cells), as well as collagen fibers in the stroma. These findings were correlated with gene upregulation of IL6, IL8, TGFβ and COX2, which are reportedly to be implicated in prostatic epithelial proliferation and stromal fibrosis. A recent study has shown that downregulated E-cadherin can induce increased prostatic epithelial permeability [2]; therefore, in this study, it is likely that leakage of intraluminal substances into the stromal compartment through potentially increased permeability by E-cadherin knockout induced chronic prostatic inflammation followed by increased proliferation and fibrosis. Additionally, mice with double injections of tamoxifen showed a bladder overactive condition as evidenced by significantly smaller single voiding volume, higher frequency in voiding spot assay and NVCs/min in cystometry. Therefore, prostatic inflammation could be involved in the development of bladder overactivity in this model, possibly through prostate-to-bladder cross-talk, as recently reported in a rat model of chemically induced prostatic inflammation [3].
Concluding message
Conditional loss of E-cadherin in the prostatic epithelium induced chronic prostatic inflammation and increased epithelial proliferation as well as bladder overactivity, which are characteristics of BPH and male LUTS. Thus, a mouse model of conditional Cdh1 knockout in the prostatic luminal epithelial cells could be useful for further investigation of BPH pathophysiology and new approaches for the treatment of male LUTS associated with prostatic inflammation.
Figure 1
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References
  1. O'Malley KJ, et al. Proteomic analysis of patient tissue reveals PSA protein in the stroma of benign prostatic hyperplasia. Prostate. 2014;74(8):892-900.
  2. Li F, et al. AB012. E-cadherin is down-regulated in benign prostate hyperplasia and required for tight junction formation and permeability barrier in prostatic epithelial cell monolayer. Transl Androl Urol. 2018;7(Suppl 5):AB012.
  3. Funahashi, Y. , et al. Bladder overactivity and afferent hyperexcitability induced by prostate-to-bladder cross-sensitization in rats with prostatic inflammation. J Physiol. 2019 doi:10.1113/JP277452
Disclosures
Funding NIH U54 DK112079 Clinical Trial No Subjects Animal Species Rat Ethics Committee University of Pittsburgh Institutional Animal Care and Use Committees
28/03/2024 07:45:28