title: Leiomyoma cells in 3-dimensional cultures demonstrate an attenuated response to fasudil, a rho-kinase inhibitor, when compared to 2-dimensional cultures.
abstract: Uterine leiomyomata are common benign tumors in women of reproductive age and demonstrate an attenuated response to mechanical signaling that involves Rho and integrins. To further characterize the impairment in Rho signaling, we studied the effect of Rho-kinase inhibitor, fasudil, on extracellular matrix production, in 2-dimensional (2D) and 3-dimensional (3D) cultures of leiomyoma and myometrial cells. Leiomyoma 2D cultures demonstrated a rapid decrease in gene transcripts and protein for fibronectin, procollagen 1A, and versican. In 3D cultures, fibronectin and procollagen 1A proteins demonstrated increased levels at lower concentrations of fasudil, followed by a concentration-dependent decrease. Versican protein increased up to 3-fold, whereas fibromodulin demonstrated a significant decrease of 1.92-fold. Myometrial 2D or 3D cultures demonstrated a decrease in all proteins after 72 hours of treatment. The 3D leiomyoma cultures demonstrated a significant increase in active RhoA, followed by a concentration-dependent decrease at higher concentrations. A concentration-dependent increase in phospho-extracellular regulated signal kinase and proapoptotic protein Bax was observed in 3D leiomyoma cultures. Fasudil relaxed the contraction of the 3D collagen gels caused by myometrium and leiomyoma cell growth. These findings indicate that the altered state of Rho signaling in leiomyoma was more clearly observed in 3D cultures. The results also suggest that fasudil may have clinical applicability for treatment of uterine leiomyoma.
authors.affiliation: Department of Obstetrics and Gynecology, Uniformed Services, University of the Health Sciences, Bethesda, MD, USA.
authors.forename: Minnie
authors.initials: M
authors.lastname: Malik
authors.affiliation: Program in Reproductive and Adult Endocrinology, Eunice Kennedy Shriver National Institute of Child Health and Human Development, NIH, Bethesda, MD, USA.
authors.forename: Joy
authors.initials: J
authors.lastname: Britten
authors.affiliation: Department of Obstetrics and Gynecology, Uniformed Services, University of the Health Sciences, Bethesda, MD, USA Program in Reproductive and Adult Endocrinology, Eunice Kennedy Shriver National Institute of Child Health and Human Development, NIH, Bethesda, MD, USA.
authors.forename: James
authors.initials: J
authors.lastname: Segars
authors.affiliation: Department of Obstetrics and Gynecology, Uniformed Services, University of the Health Sciences, Bethesda, MD, USA Program in Reproductive and Adult Endocrinology, Eunice Kennedy Shriver National Institute of Child Health and Human Development, NIH, Bethesda, MD, USA william.catherino@usuhs.edu.
authors.forename: William H
authors.initials: WH
authors.lastname: Catherino
mesh_terms: D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine / Q000031:analogs & derivatives* / Q000494:pharmacology / Q000627:therapeutic use; D000818:Animals; D018929:Cell Culture Techniques / Q000379:methods*; D002461:Cell Line, Transformed; D005260:Female; D006801:Humans; D007889:Leiomyoma / Q000188:drug therapy / Q000201:enzymology*; D047428:Protein Kinase Inhibitors / Q000494:pharmacology* / Q000627:therapeutic use; D051381:Rats; D016896:Treatment Outcome; D014407:Tumor Cells, Cultured; D014594:Uterine Neoplasms / Q000188:drug therapy / Q000201:enzymology*; D054460:rho-Associated Kinases / Q000037:antagonists & inhibitors* / Q000378:metabolism
chemical_list: D047428:Protein Kinase Inhibitors; D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D054460:rho-Associated Kinases; C049347:fasudil
keywords: 2D and 3D cultures; RhoA; extracellular matrix; fasudil; gel contraction; uterine leiomyoma
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