Angiotensin II‐mediated calcium signals and mitogenesis in human prostate stromal cell line hPCPs

Abstract

Western blots and immunocytochemistry were used to detect angiotensin 1 (AT(1)) and angiotensin 2 (AT(2)) receptors in human primary cultures of the prostate stromal compartment (hPCPs). Immunohistochemistry was performed on human prostate tissue-embedded paraffin. In addition, pharmacological tools were applied in combination with photometry experiments to characterize the physiological activity of AT(1) and AT(2) receptors in hPCPs cell culture. A proliferation assay was used to describe the mitogenic activity of angiotensin II (Ang II) on hPCPs cells. Only the AT(1) receptor was detected in Western blot analysis. Immunocytochemistry of hPCPs cells showed that the AT(1) receptor is present in both the smooth muscle type and the fibroblastic type. In the stromal compartment of human prostate tissue, immunoreaction with antibodies against the AT(1) receptor was detectable.Fura-2-loaded hPCPs cells showed an instantaneous and linear rise in free intracellular calcium ion concentration ([Ca(2+)](i)) after local perfusion with Ang II in concentrations of 10 nM. Removing of external calcium or emptying intracellular calcium stores before Ang II application diminished or abolished this [Ca(2+)](i) response. The response to Ang II was also diminished when hPCPs cells were perfused with the AT(1) receptor inhibitor losartan prior to Ang II application. No inhibition of the [Ca(2+)](i) increase was detectable after perfusion with PD 123319, a specific inhibitor of the AT(2) receptor.hPCPs cells were stimulated with Ang II in various concentrations over a period of 2 days. The subsequently performed proliferation assay revealed a mitogenic effect of Ang II on hPCPs in concentrations starting at 10 nM. This effect could be inhibited by losartan.