Expression and immunolocalisation of neutral endopeptidase in prostate cancer.
Albrecht M., Mittler A., Wilhelm B., Lundwall A., Lilja H., Aumüller G., Bjartell A.
BACKGROUND: Neutral Endopeptidase (NEP) is a cell surface enzyme that cleaves and inactivates neuropeptides. When present on androgen-dependent prostate cancer (PC) cells, NEP inactivates growth stimulatory neuropeptides. After androgen ablation NEP expression decreases and neuropeptides can enhance cell growth, leading to the development of androgen-independent, neuropeptide stimulated PC. Aim of the study was to analyse the expression, localisation and distribution of NEP in benign and malignant prostatic tissues and its relation to the cytoskeleton. METHODS: Immunohistochemistry (IHC) was performed to localise NEP in fixed specimens from normal prostatic tissue, benign prostate hyperplasia (BPH) and PC of Gleason grade 2-5. In situ hybridisation and Western blotting experiments were carried out to confirm NEP gene expression and translation to mature protein in BPH and PC tissue. Confocal laser scanning microscopy was utilised to investigate whether development of high grade prostate tumours was accompanied by changes in intracellular actin/NEP colocalisation patterns. Finally, the proliferative activity in relation to loss of NEP expression was investigated by dual staining of NEP and Ki-67 in prostatic tumours. RESULTS: In situ hybridisation studies revealed preserved expression of NEP mRNA in epithelial cells of PC. NEP was by IHC shown to be located in the apical plasma membrane of normal epithelial cells and BPH tissue. In PC a Gleason grade dependent shift of the NEP distribution pattern towards a heterogeneous, partly cytoplasmic allocation of the protein was found. Compared to BPH tissue, specimens derived from PC showed very low IHC-staining intensity for NEP protein. In high grade PC the typical apical colocalisation of actin and NEP was lost and a strong granular cytoplasmic NEP staining was found. PC areas with a high expression of NEP displayed diminished proliferative activity i.e. low staining intensity for Ki-67. CONCLUSIONS: NEP is differentially expressed in the normal and the pathologically altered prostate with a clear shift from a membrane bound to a cytoplasmic distribution pattern in high-grade tumours and loss of NEP expression in areas of high proliferative activity. The data presented support an active involvement of NEP in the progression of androgen-independent PC. Further studies are needed to unravel the mechanisms underlying the cytoplasmic NEP distribution in PC.