Inhibition of human prostate smooth muscle contraction by the integrin α2β1-selective inhibitor BTT3033 and the integrin-linked kinase inhibitor Cpd22

Introduction: Mirabegron is available for treatment of storage symptoms in overactive bladder, which may be improved by β3-adrenoceptor-induced bladder smooth muscle relaxation. In addition to storage symptoms, lower urinary tract symptoms in men include obstructive symptoms attributed to benign prostatic hyperplasia, caused by increased prostate smooth muscle tone and prostate enlargement. In contrast to the bladder and storage symptoms, effects of mirabegron on prostate smooth muscle contraction and obstructive symptoms are poorly understood. Evidence from non-human smooth muscle suggested antagonism of α1-adrenoceptors as an important off-target effect of mirabegron. As α1-adrenergic contraction is crucial in pathophysiology and medical treatment of obstructive symptoms, we here examined effects of mirabegron on contractions of human prostate tissues and on proliferation of prostate stromal cells.Methods: Contractions were induced in an organ bath. Effects of mirabegron on proliferation, viability, and cAMP levels in cultured stromal cells were examined by EdU assays, CCK-8 assays and enzyme-linked immunosorbent assay.Results: Mirabegron in concentrations of 5 and 10 μM, but not 1 µM inhibited electric field stimulation-induced contractions of human prostate tissues. Mirabegron in concentrations of 5 and 10 µM shifted concentration response curves for noradrenaline-, methoxamine- and phenylephrine-induced contractions to the right, including recovery of contractions at high concentrations of α1-adrenergic agonists, increased EC50 values, but unchanged Emax values. Rightshifts of noradrenaline concentration response curves and inhibition of EFS-induced contractions were resistant to L-748,337, l-NAME, and BPIPP. 1 µM mirabegron was without effect on α1-adrenergic contractions. Endothelin-1- and U46619-induced contractions were not affected or only inhibited to neglectable extent. Effects of mirabegron (0.5–10 µM) on proliferation and viability of stromal cells were neglectable or small, reaching maximum decreases of 8% in proliferation assays and 17% in viability assays. Mirabegron did not induce detectable increases of cAMP levels in cultured stromal cells.Conclusion: Mirabegron inhibits neurogenic and α1-adrenergic human prostate smooth muscle contractions. This inhibition may be based on antagonism of α1-adrenoceptors by mirabegron, and does not include activation of β3-adrenoceptors and requires concentrations ranging 50-100fold higher than plasma concentrations reported from normal dosing. Non-adrenergic contractions and proliferation of prostate stromal cells are not inhibited by mirabegron.

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Ghrelin Aggravates Prostate Enlargement in Rats with Testosterone-Induced Benign Prostatic Hyperplasia, Stromal Cell Proliferation, and Smooth Muscle Contraction in Human Prostate Tissues

Oxidative Medicine and Cellular Longevity ◽

10.1155/2019/4748312 ◽

2019 ◽

Vol 2019 ◽

pp. 1-14 ◽

Cited By ~ 1

Author(s):

Xiaolong Wang ◽

Yiming Wang ◽

Christian Gratzke ◽

Christian Sterr ◽

Qingfeng Yu ◽

...

Keyword(s):

Smooth Muscle ◽

Benign Prostatic Hyperplasia ◽

Muscle Contraction ◽

Stromal Cell ◽

Molecular Mechanisms ◽

Smooth Muscle Contraction ◽

Prostatic Hyperplasia ◽

Human Prostate ◽

Mrna Levels ◽

Prostate Enlargement

Epidemiologic studies revealed a context between lower urinary tract symptoms (LUTS) suggestive of benign prostatic hyperplasia (BPH) and metabolic syndrome. However, molecular mechanisms underlying this relationship are largely unknown. Prostate enlargement and increased prostate smooth muscle tone are important factors in the pathophysiology of LUTS suggestive of BPH. In the present study, we studied effects of the metabolic hormone ghrelin on prostate enlargement in rats with experimentally induced BPH, growth of cultured stromal cells from human prostate (WPMY-1), and smooth muscle contraction of human prostate tissues. Ghrelin (20 nmol/kg daily, p.o., 2 weeks) increased prostate size in rats with testosterone-induced BPH. Microarray identified 114 ghrelin-upregulated genes (2-fold or more) in these prostates, with possible roles in growth, smooth muscle contraction, or metabolism. 12 genes were selected for further analyses. In human prostate tissues, mRNA levels of 11 of them correlated positively with ghrelin receptor (GHSR) expression, but only two with the degree of BPH. Accordingly, no correlation was evident between GHSR expression level and BPH in human prostate tissues. In WPMY-1 cells, the GHRS agonist MK0677 upregulated 11 of the selected genes. MK0677 induced proliferation of WPMY-1 cells, shown by EdU assay, colony formation, proliferation markers, flow cytometry, and viability. In myographic measurements, GHSR agonists enhanced contractions of human prostate strips. Together, ghrelin may aggravate prostate enlargement, stromal cell growth, and prostate smooth muscle contraction in BPH. Ghrelin may deteriorate urethral obstruction independently from BPH, qualifying the ghrelin system as an attractive new target to be tested for LUTS treatment in BPH.

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MP19-09 NSC23766, A RAC GTPASE INHIBITOR, INHIBITS SMOOTH MUSCLE CONTRACTION IN THE HYPERPLASTIC HUMAN PROSTATE

The Journal of Urology ◽

10.1016/j.juro.2014.02.710 ◽

2014 ◽

Vol 191 (4S) ◽

Author(s):

Thomas Kunit ◽

Andrea Schreiber ◽

Christian Gratzke ◽

Beata Rutz ◽

Raphaela Waidelich ◽

...

Keyword(s):

Smooth Muscle ◽

Muscle Contraction ◽

Smooth Muscle Contraction ◽

Human Prostate ◽

Rac Gtpase

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Inhibition of smooth muscle force generation by focal adhesion kinase inhibitors in the hyperplastic human prostate

AJP Renal Physiology ◽

10.1152/ajprenal.00011.2014 ◽

2014 ◽

Vol 307 (7) ◽

pp. F823-F832 ◽

Cited By ~ 14

Author(s):

Thomas Kunit ◽

Christian Gratzke ◽

Andrea Schreiber ◽

Frank Strittmatter ◽

Raphaela Waidelich ◽

...

Keyword(s):

Smooth Muscle ◽

Western Blot ◽

Muscle Contraction ◽

Focal Adhesion Kinase ◽

Focal Adhesion ◽

Blot Analysis ◽

Western Blot Analysis ◽

Smooth Muscle Contraction ◽

Human Prostate ◽

Focal Adhesion Proteins

Smooth muscle contraction may be critical for lower urinary tract symptoms (LUTS) in patients with benign prostate hyperplasia and requires stable anchorage of the cytoskeleton to the cell membrane. These connections are regulated by focal adhesion kinase (FAK). Here, we addressed the involvement of FAK in the regulation of smooth muscle contraction in hyperplastic human prostate tissues. Prostate tissues were obtained from radical prostatectomy. Expression of FAK and focal adhesion proteins was assessed by Western blot analysis and immunohistochemical stainings. Effects of the FAK inhibitors PF-573228 and Y-11 on contraction of prostate strips were examined in the organ bath. Expression of FAK and focal adhesion proteins (integrin-5α, paxilin, and c-Src) was detected by Western blot analysis in prostate samples. By double immunofluorescence staining with calponin and pan-cytokeratin, expression of FAK was observed in stromal and epithelial cells. Immunoreactivity for FAK colocalized with integrin-5α, paxilin, talin, and c-Src. Stimulation of prostate tissues with the α1-adrenergic agonist phenylephrine increased the phosphorylation state of FAK at Tyr397 and Tyr925 with different kinetics, which was blocked by the α1-adrenoceptor antagonist tamsulosin. Norepinephrine and phenylephrine induced concentration-dependent contractions of prostate strips. Both FAK inhibitors PF-573228 and Y-11 significantly inhibited norepinephrine- and phenylephrine-induced contractions. Finally, PF-573228 and Y-11 inhibited contractions induced by electric field stimulation, which was significant at the highest frequency. In conclusion, α1-adrenergic smooth muscle contraction or its regulation involves FAK in the human prostate. Consequently, FAK may be involved in the pathophysiology of LUTS and in current or future LUTS therapies.

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