Inhibition of mitogenic signaling and induction of apoptosis in human bladder smooth muscle cells treated with doxazosin

Department of Cell Biology and Physiology, Washington University in St. Louis, San Luis, Missouri, United States
The Journal of Urology (Impact Factor: 4.47). 11/2004; 172(4 Pt 2):1662-5; discussion 1666. DOI: 10.1097/
Source: PubMed


The alpha1 antagonist doxazosin is used to treat lower urinary tract symptoms and is believed to function primarily as a smooth muscle relaxant. However, doxazosin has been shown to inhibit proliferation and induce apoptosis in nonbladder smooth muscle. Consequently, we examined the effects of doxazosin on human bladder smooth muscle cell (SMC) proliferation and apoptosis.
Primary human bladder SMCs were cultured in M199 with 10% fetal bovine serum (FBS) until they reached 65% confluency and then they were made quiescent by serum starvation in M199 with 0.4% FBS for 24 hours. The quiescent bladder SMCs were pretreated for 30 minutes with doxazosin or vehicle (dimethyl sulfoxide) and then stimulated with 10% FBS for 24 hours. Measurement of 5'-bromo-2'-deoxyuridine (BrdU) uptake by flow cytometry was used to determine the effect of doxazosin on cell cycle progression. Western immunoblot was used to examine cell cycle protein expression and phosphorylation of the retinoblastoma protein (Rb) and cyclin A, both of which regulate cell cycle progression.
Cellular proliferation was inhibited in a dose dependent manner by doxazosin. There was nearly a 50% decrease in BrdU uptake at 10 microM doxazosin and an approximately 90% decrease in BrdU at 25 microM doxazosin. Notably, doxazosin inhibited phosphorylation of Rb and expression of cyclin A, both of which are necessary for cell cycle progression. At concentrations of 25 microM doxazosin or greater apoptosis was induced in the bladder SMCs, as indicated by an increase in subG1 DNA content.
Our study demonstrates that doxazosin inhibits mitogen induced proliferation of human bladder SMC by blocking cell cycle progression at the of G1/S border. Doxazosin induced cell cycle inhibition appears to be at least in part due to an inhibition of mitogen induced Rb phosphorylation and cyclin A expression. At higher concentrations doxazosin induces apoptosis in human bladder SMCs.

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    • "While the induction of apoptosis by doxazosin and chemically related compounds was initially shown in prostate cells and most of the subsequent work has also been conducted in such cells, later studies demonstrated doxazosin-induced apoptosis to also occur in a range of other normal and tumor cells, indicating that this may be a global effect that is not cell type-dependent. Such other cell types include cardiac myocytes (Rodriguez-Feo et al. 2000; Eiras et al. 2006) and the H9C2 cell line derived thereof (Yang et al. 2009), vascular endothelial cells (Keledjian et al. 2005), bladder smooth muscle cells (Austin et al. 2004), urothelial cancer cells (Siddiqui et al. 2005), pituitary adenoma cells (Fernando and Heaney 2005), breast cancer cells (Hui et al. 2008), colon cancer cells, and HeLa cells (Gan et al. 2008). One of the most interesting aspects of this effect is the fact that apoptosis induction occurs independently of the α 1 antagonistic properties of these drugs (Kyprianou and Benning 2000; Anglin et al. 2002). "
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    ABSTRACT: Doxazosin and related, quinazoline-based alpha(1)-adrenoceptor antagonists can induce apoptosis in prostate and various other normal, benign, smooth muscle, endothelial and malignant cells. Such apoptosis-inducing effects occur independently of alpha(1)-adrenoceptor antagonism and typically require much high concentrations than those required for receptor occupancy. Several studies have invested efforts towards the elucidation of the molecular mechanisms underlying doxazosin-induced apoptosis. These include various tumor cells, cardiomyocytes, endothelial cells and bladder smooth muscle cells. While the high concentrations of doxazosin required to induce apoptosis challenge the use of this and related drugs for clinical optimization of apoptosis induction, such quinazoline structure may represent chemical starting points to develop more potent apoptosis-inducing agents free of alpha(1)-adrenoceptor antagonistic action and suitable for cancer treatment with minimal and well-tolerated side effects.
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    • "Furthermore, our results have confirmed that Dox induces apoptosis in epithelial cells of rat prostatic lobes, without showing interlobar variation. These results are in agreement with the Dox effect on the prostate (Kyprianou 2003), bladder smooth muscle (Austin et al. 2004), and cardiomyocytes (Eiras et al. 2006). However, Garrison and Kyprianou (2006) have shown that benign prostate cells seem to be more sensitive to the apoptotic effect of Dox than are tumor cells. "
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