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Individual receptor profiling as a novel tool to support diagnosis of bladder pain syndrome/interstitial cystitis (BPS/IC)

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Abstract

Purpose

Dysregulation of neurotransmitter receptors may contribute to bladder overactivity (OAB) symptoms. To address the question whether specific receptor expression patterns are associated with bladder pain syndrome/interstitial cystitis (BPS/IC), we examined the expression of muscarinic, purinergic and histamine receptors in the detrusor.

Methods

Detrusor receptor expression was investigated in bladder biopsies of female BPS/IC patients (n = 44; age 60.64 ± 13.78, mean ± SD) and carcinoma patients (n = 11; age 58.91 ± 12.72) undergoing cystectomy. Protein expression of muscarinic (M2, M3), purinergic (P2X1–3) and histamine receptors (H1, H2) was analysed by confocal immunofluorescence, and gene expression was quantified by real-time polymerase chain reaction (qPCR).

Results

M2, P2X1, P2X2 and H1 receptor immunoreactivity (-IR) was significantly enhanced in BPS/IC compared to the control group, while there was no difference for M3-, P2X3- and H2-IR. We calculated a score, which separated BPS/IC from control patients with an AUC of 89.46%, showing 84.09% sensitivity and 90.91% specificity. Patients had a 9.25 times enhanced calculated risk for BPS/IC. In addition, two patient subgroups (M2 > M3 and M3 > M2) were observed, which differed in associated purinergic and histamine receptor expression.

Conclusions

M2, P2X1, P2X2 and H1 were significantly upregulated in BPS/IC patients, and H2 was occasionally highly overexpressed. There was no significant correlation between receptor protein and gene expression, implying posttranslational mechanisms being responsible for the altered receptor expressions. On the basis of individual receptor profiles, upregulated receptors could be targeted by monotherapy or combination therapy with already approved receptor inhibitors, thereby promoting tailored therapy for patients suffering from BPS/IC-like symptoms.

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Acknowledgment

We thank Mrs. A. Weimann, Mrs. M. Berndt and Mrs. K. Büttner for their excellent technical assistance.

Conflict of interest

The authors declare that they have no conflict of interest.

Author information

Authors and Affiliations

  1. Department of Urology, University Hospital Leipzig, Liebigstr. 20, 04103, Leipzig, Germany

    Jochen Neuhaus, Jens-Uwe Stolzenburg, Siegurd Hartenstein & Thilo Schwalenberg

  2. Department of Urology, St. Hedwig Hospital, Medical School of Charité Universitätsmedizin Berlin, Berlin, Germany

    Heinrich Schulte-Baukloh

  3. CISUC, Department of Informatics Engineering, University of Coimbra, Coimbra, Portugal

    Pietro Speroni di Fenizio

  4. Department of Pathology, University of Leipzig, Leipzig, Germany

    Lars-Christian Horn

  5. Department of Anesthesiology, Intensive Care, Pain Therapy and Palliative Medicine, HELIOS Hospital Borna, Borna, Germany

    Henrik Rüffert

  6. Department of Urology, University of Regensburg, Caritas Hospital St. Josef, Regensburg, Germany

    Maximilian Burger

  7. Urological Practice, Markkleeberg, Germany

    Matthias Schulze

Authors
  1. Jochen Neuhaus
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  2. Heinrich Schulte-Baukloh
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  3. Jens-Uwe Stolzenburg
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  10. Thilo Schwalenberg
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Corresponding author

Correspondence to Jochen Neuhaus.

Electronic supplementary material

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345_2011_774_MOESM1_ESM.doc

Table S1: Confocal immunofluorescence analysis results. Numbers represent fluorescence intensities of the receptor stainings normalized to background fluorescence (FIrec/FIbkgd). (DOC 41 kb)

345_2011_774_MOESM2_ESM.doc

Table S2: Correlation analysis. Nonparametric correlation coefficient (Spearman r) is reported. Except for P2RX1 receptor, there was no significant correlation between receptor gene expression and protein expression. (DOC 32 kb)

345_2011_774_MOESM3_ESM.doc

Table S3: Correlation analysis. Nonparametric correlation coefficient (Spearman r) is reported. There was no relevant correlation of receptor expression with age in either group. Note the high Spearman r for CHRM2 and P2RX1 with R_Score, indicating major contribution of those receptors. * p < 0.05; ** p < 0.01; *** p < 0.001; na = not applicable. (DOC 59 kb)

345_2011_774_MOESM4_ESM.doc

Table S4: Correlation between receptor immunofluorescence in SM2 and SM3 patients. 82% of the patients showed M2 > M3 expression (SM2). Note the high number of significant correlations between receptors (Spearman r, *p < 0.05). 18% of the patients showed M3 > M2 expression (SM3) with only few significant correlations of receptor expression. * p < 0.05; ** p < 0.01; *** p < 0.001. (DOC 53 kb)

345_2011_774_MOESM5_ESM.tif

Figure S1: Workflow of confocal immunofluorescence quantitative analysis. (a) Confocal image of staining control (StainCo); (b) M2 receptor staining; regions of interest (ROIs) marked. (c-d) Pixel intensity distribution is measured in 35 ROIs, defined as alpha-smooth muscle cell actin-positive areas. Nuclei were excluded from analysis. (e) Pixel distribution normalized to StainCo (FIRec/FIStainCo). Note the excellent signal to noise ratio as demonstrated by clear separation of the M2 receptor staining (clear area) and staining control (dark grey). (f) Receptor profile including statistical analysis by ANOVA with Tukey’s post hoc test; bars: p < 0.05 was considered significant; § indicates no significant receptor expression. (TIFF 3212 kb)

345_2011_774_MOESM6_ESM.tif

Figure S2: Assessment of antibody specificity. Antibody specificity was tested by comparison of confocal immunofluorescence intensities of sections stained with specific receptor antibodies and corresponding sections where the peptide, to which the antibody was raised, was used to block the antibody-antigen detection. The peptides were purchased from the supplier of the corresponding antibody. The samples were processed and analysed according to the standard protocol. (a) Quantitative analysis revealed significant diminution of the fluorescence intensity by blocking peptide (dilution 1:100–1:2000). (b) Representative receptor immunofluorescence confocal images. (TIFF 5101 kb)

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Neuhaus, J., Schulte-Baukloh, H., Stolzenburg, JU. et al. Individual receptor profiling as a novel tool to support diagnosis of bladder pain syndrome/interstitial cystitis (BPS/IC). World J Urol 30, 693–700 (2012). https://doi.org/10.1007/s00345-011-0774-0

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  • DOI: https://doi.org/10.1007/s00345-011-0774-0

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