Rac1 silencing, NSC23766 and EHT1864 reduce growth and actin organization of bladder smooth muscle cells.

AIMS

RacGTPase-mediated proliferation and smooth muscle contraction in the lower urinary tract has been recently suggested and may offer putative targets for treamtment of lower urinary tract symptoms. However, RacGTPase function for proliferation of detrusor smooth muscle cells is unknown and the specificity of Rac inhibitors has been questioned. Here, we examined effects of Rac1 knockdown and of the Rac inhibitors NSC23766 and EHT1864 in human bladder smooth muscle cells (hBSMCs).

MAIN METHODS

Rac1 expression was silenced by shRNA expression. Effects of silencing and Rac inhibitors were assessed by CCK-8 assay, EdU staining, RT-PCR, colony formation assay, flow cytometry, and phalloidin staining.

KEY FINDINGS

Silencing of Rac1 expression reduced the viability (up to 83% compared to scramble shRNA) and proliferation (virtually completely in proliferation assay), increased apoptosis (124%) and the number of dead cells (51%), and caused breakdown of actin organization (56% reduction of polymerized actin compared to scramble shRNA). Effects on proliferation, viability, and actin organization were mimicked by NSC23766 and EHT1864, while both compounds showed divergent effects on cell death (32-fold increase of dead cells by EHT1864, but not NSC23766). Effects of NSC23766 and EHT1864 on viability of hBSMCs were not altered by Rac1 knockdown.

SIGNIFICANCE

Rac1 promotes proliferation, viability, and cytoskeletal organization, and suppresses apoptosis in bladder smooth muscle cells, which may be relevant in overactive bladder or diabetes-related bladder dysfunction. NSC23766 and EHT1864 mimick these effects, but may act Rac1-independently, by shared and divergent effects.