Fig. 5

IF and IHC analysis of the distribution of long-lived LRCs and expression of cell lineage and SC markers in the adult mouse prostate. a A transversal HTX stained section of an adult prostate; (b) shows KRT-8 expression (green) in a consecutive section to (a), with dotted lines marking the border between distal and proximal/ductal epithelium; the white line demarks the proximal VP, blue line demarks the proximal AP, and the DLP is demarked in red. A yellow dotted line demarks the sphincter muscle. In (c), the abrupt decrease in TROP-2 (green) at the proximal-distal border is indicated by arrows, and a representational magnification of BrdU (green) and TROP-2 (red) from the proximal AP is highlighted in the inset (arrowhead, c). In contrast, CD133 (DAB) was absent from the proximal lobes and ducts (arrow, d), but strongly expressed by luminal cells of the distal lobes (d). CD133 (green) and TROP-2 (red) expression showed some overlap at the very proximal-distal border of the AP lobes (green arrow points distally), but was mutually excluding at the cellular level (e). f, g LRCs (green) positive for AR expression (red) (arrows, f–h). The arrow in (i) highlights a basal LRC (green) that is negative for AR. Basal LRCs (green) expressed p63 (red) (arrow, j), and both basal (arrow) and luminal (arrowhead) LRCs (green) in ducts were found to express CD44 (red). l KRT-7 (green) and TROP-2 (red) were co-expressed in prostate ducts. However, KRT-7 (green) had a more restricted expression pattern in distal epithelium (m) as compared to TROP-2 (red). Basal (arrowhead) and luminal (arrow) LRCs (green) co-expressed KRT-7 (red) in prostate ducts (n). o Co-expression of KRT-7 (red) and AR (green) is seen in the dorso-lateral prostate (DLP) (arrows); distal DLP epithelium is indicated by white dotted lines, and the sphincter muscle (Sp) is demarked by a yellow dotted line. Note a rare KRT-7 and AR co-expressing cell in the distal DLP and the high frequency of co-expressing cells in the ducts and proximal DLP (o). Rare KRT-7-expressing cells (red) in the distal AP further co-expressed high levels of AR (green) (arrow, p). The expression level of c-kit was lower in adult prostates and the harsh BrdU protocol hindered proper IF detection of c-kit; however, using double sequential chromogenic detection, c-kit (NBT/BCIP)-positive LRCs (DAB) could be detected (arrow, q). r A single LRC (green) co-expressing Sca-1 is indicated in the distal VP (arrow). No LRCs (green) (arrow, s) were found positive for the proliferative marker Ki67 (red) (arrowhead, s). The majority of proliferating (Ki67; green) cells were found in the luminal cell layer (arrow, t) and were negative for p63 (red) (arrowheads, t) expression. Counterstaining when applied was either with HTX (IHC) or DAPI (IF). a–d are consecutive composite images of several individual photomicrographs. Scale bar = 500 μm (a–d), 25 μm (i, o, p, r), 50 μm (h, j, l–n, q), and 100 μm (e–g, k, s-t)