Intermediate cells in human prostate epithelium are enriched in proliferative inflammatory atrophy

Abstract

Within the human prostate epithelium four cell populations can be discriminated based on their expression of keratins (K). Basal cells express high levels of K5 and K14, as well as p63, whereas they have very low levels of androgen receptor, prostate-specific antigen (PSA), K8, and K18. Luminal secretory cells lack p63, K5, and K14 but express high levels of K8, K18, androgen receptor, and PSA. Additionally, cells have been identified with a keratin phenotype intermediate between basal and luminal cells that co-express high levels of K5 and K18 (K5/18) as well as hepatocyte growth factor receptor c-MET. Although intermediate cells have been proposed as precursor cells of prostate cancer, their biology is ill defined. Epithelial cells in proliferative inflammatory atrophy (PIA) appear to be cycling rapidly as indicated by expression of Ki-67, and morphological transitions have been identified between PIA and high-grade prostate intraepithelial neoplasia. Many of the atrophic epithelial luminal cells in PIA are candidates for intermediate cells based in part on weak expression of PSA and androgen receptor, high levels of K8/18, and lack of p63. The objective of this study was to further clarify the phenotype of the proposed intermediate cells in PIA and to quantitatively determine the level in which these intermediate cells preferentially occur in PIA lesions. Intermediate cells were immunohistochemically demonstrated using antibodies to K5, K14, K18, and c-MET. Using radical prostatectomy specimens (n = 15) the area fraction of intermediate cells in normally differentiated prostate epithelium and PIA were quantified by a grid point counting method. Atrophic luminal cells of PIA lesions expressed K5 in 39.2 +/- 7.4% of cells compared to 2.4 +/- 2.3% in normal epithelium (P < 0.00001). By contrast, K14 was only expressed in 3.0 +/- 3.2% of the luminal cells. Previous studies have shown that virtually 100% of these atrophic luminal cells are strongly positive for K8/18. c-MET was present in 44.1 +/- 14.1% of luminal cells in PIA but only in 2.1 +/- 2.8% of luminal cells in normal epithelium (P < 0.00001). To unambiguously determine whether intermediate luminal cells in PIA show increased proliferative activity and decreased p27(kip1) expression, double-staining immunofluorescence of Ki-67 and K5, as well as p27(Kip1) and K5 was performed. Luminal cells in PIA often co-expressed K5 and Ki-67. Although p27(Kip1) was strongly expressed in K5-negative differentiated cells in normal epithelium, p27(Kip1) staining was absent in many of the K5-positive cells in the luminal compartment of PIA. We conclude that cells phenotypically intermediate between basal and secretory cells are enriched in PIA lesions. The finding of a large number of highly proliferating intermediate cells in PIA provides further support that these cells may serve as preferred target cells in prostate carcinogenesis.

Figure 1.

Morphological and K5 phenotype of PIA. A: Low-power image showing region of simple atrophy. Arrows indicate normal appearing epithelium with abundant clear cytoplasm. Note basophilic appearance of the majority of the acini. H&E stain. B: High-power image of area indicated in boxed region in A. Note two cell layers consisting of cuboidal and basophilic luminal cells (arrowheads) and darker underlying basal cells (arrows). H&E. C: Medium-power view of normal appearing epithelium (n) and PIA stained with anti-K5 antibodies. Note strong staining of basal layer in all epithelia, lack of staining of luminal cells in normal acini, and weak but positive staining in many of the luminal cells in atrophic acini. D: Higher power view of an atrophic acinus from C. Arrowheads indicate luminal secretory cells and arrows indicate basal cells. Original magnifications: ×40 (A); ×400 (B, D); ×100 (C).

Figure 2.

c-MET expression in PIA. A: Low-power image showing region of simple atrophy (bottom) and normal appearing epithelium (top). Note strong staining in atrophic epithelium for c-MET. B: High-power image of area indicated in boxed region in A. n, Normal acinus. Arrowhead indicates atrophic luminal epithelial cell. Original magnifications: ×50 (A); ×400 (B).

Figure 3.

Box plot of stereological grid point counting data from immunohistochemical staining results for K5, K14, and c-MET in PIA versus normal prostate luminal cells. Box extends from first to third quartiles, median is indicated by the line within the box. Whiskers include all remaining data points. A, K5; B, c-MET; C, K14. Note complete lack of overlap for both K5 and c-MET and lack of difference for K14.

Figure 4.

Immunofluorescence double labeling of PIA. A: PIA acinus stained for K5 (green), Ki-67 (red), and nuclei (blue). L, lumen; S, stroma. Note near continuous basal layer showing strong K5 staining and weaker but positive K5 staining in the majority of luminal cells. Arrow indicates a single Ki-67-positive cell that is clearly localized in the luminal compartment and is co-expressing K5. B: High-power view from another area of PIA showing Ki-67-positive luminal cell. C: Normal appearing prostate acinus stained for p27^Kip1 (red), K5 (green), and nuclei (blue). Note strong staining for p27^Kip1 in the vast majority of columnar luminal secretory cells. D: Area of PIA from same specimen as in C. Note absence of p27^Kip1 staining in a group of K5-positive luminal cells (arrows). Original magnifications: ×400 (A, C, D); ×1000 (B).