Beta-catenin is elevated in human benign prostatic hyperplasia specimens compared to histologically normal prostate tissue

Abstract

Benign prostatic hyperplasia (BPH) is linked to lower urinary tract symptoms (LUTS) such as incomplete bladder emptying, urinary frequency and urgency. Mechanisms responsible for BPH are not fully known. Here, we tested whether beta-catenin (CTNNB1) immunostaining intensity and distribution differ in human glandular BPH tissue specimens compared to normal prostate tissue. Multiplex immunostaining of CTNNB1, its putative transcriptional target gene lymphoid enhancer binding factor 1 (LEF1), and the epithelial marker E-cadherin were examined in clinical human prostate specimens with or without histological BPH (pure epithelial or mixed stromal-epithelial nodules). BPH specimens were obtained from 24 men who experienced LUTS and underwent transurethral resection of the prostate surgery. Control specimens were tumor-adjacent histologically normal prostate tissue from 48 patients who underwent radical prostatectomy. The resulting multispectral images were unmixed and optical densities recorded to quantify staining abundance, cellular (membranous, cytoplasmic, and nuclear) and tissue localization (stromal versus epithelial), and determination of percentage of CTNNB1-positive cells. The following CTNNB1 indices were significantly higher in BPH compared to normal prostate tissue: overall staining intensity, staining intensity in prostate stromal cell membranes, cytoplasm and nuclei, and prostate epithelial cell nuclei. The following LEF1 indices were significantly lower in BPH compared to tumor-adjacent normal prostate tissue: stromal LEF1 staining intensity, percentage of LEF1-positive stromal cells, and intensity of LEF1 staining in stromal cell membranes, cytoplasm, and nuclei. The percentage of stromal cells with CTNNB1(+)/LEF1(-) nuclei was higher and percentage of stromal cells with CTNNB1(-)/LEF1(+) nuclei was lower in BPH compared to tumor-adjacent normal prostate tissues. These results support the hypothesis that CTNNB1 expression increases in specific BPH tissue compartments. Further, since nuclear LEF1 staining does not coincide with cytoplasmic or nuclear CTNNB1 staining, it does not appear to be a reliable index of CTNNB1 activity in adult human prostate.

Keywords: LEF1; Prostate; beta-catenin; cellular localization; multispectral; stromal-epithelial; tissue microarray.

Figure 1

LEF1 and CTNNB1 localization and staining abundance in normal prostate and glandular benign prostatic hyperplasia (BPH). Nuance® software was used to build a spectral library to unmix multiplexed immunohistochemical staining, resulting in separation of hematoxylin (left column), Bajoran purple (middle column), and 3,3’-Diaminobenzidine (DAB; right column) chromogens (A). CTNNB1 and LEF1 staining intensity was quantified using inForm® software (B). Total CTNNB1 staining was significantly higher in BPH (mean = 0.093, SEM = 0.002) compared to benign prostatic tissue (BPT; 0.085 ± 0.002; p = 0.02) (C). Total LEF1 staining was not significantly different between BPT (0.058 ± 0.006) and BPH (0.046 ± 0.004; p = 0.17).

Figure 2

Quantitative analysis of tissue-specific CTNNB1 or LEF1 staining abundance by mean optical density (OD) and percentage of immunopositive cells within tissue. No differences in epithelial CTNNB1 staining were observed as assessed by mean OD (p = 0.15) or positivity (p = 0.54) analysis (A). Stromal CTNNB1 staining was significantly higher in BPH (mean = 0.028, SEM = 0.001) than BPT (0.021 ± 0.001; p<0.0001) in mean OD analysis. Stromal CTNNB1 positivity was higher in BPH than BPT but failed to reach significance (p = 0.06). Epithelial LEF1 staining was similar between BPT and BPH in both mean OD (p = 0.99) and positivity (p = 0.90) analysis (B). Analysis of stromal LEF1 staining by mean OD displayed significantly lower expression in BPH (0.042 ± 0.005) compared to BPT (0.070 ± 0.008; p = 0.02). LEF1 positivity in the stroma also showed a significantly lower amount of positive cells in BPH (mean = 7.84%, SEM = 1.89) compared to BPT (20.01 ± 3.28; p = 0.02).