Contraction of Cultured Human Uterine Smooth Muscle Cells after Stimulation with Endothelin-1

Abstract

To our knowledge, the problem of how to maintain isolated smooth cells in a "contractile" phenotypic state without deviation after subculturing has yet to be resolved. The present study characterized the in vitro contractile response of human uterine smooth muscle cell to endothelin-1, which induces contractions in isolated uterine strips. Contractile effects were qualitatively investigated using silicone rubber substrata. Endothelin-1 was able to distort and reduce the wrinkles in the silicone surface. Contractions were also quantified by measuring the resulting change in the collagen lattice area. Endothelin-1 significantly increased the contractile response in a dose-dependent manner by selectively activating endothelin A receptors. When myometrial cells were cultured within collagen lattices, a microfilament-disrupting agent, cytochalasin B, abolished contractions, and no change was observed in smooth muscle alpha-actin immunostaining. Taken together, these observations show that the uterine smooth muscle cells are contractile and respond appropriately to a potent uterotonic agent. Based on these findings, a cultured uterine smooth muscle cell model, which could be used to elucidate the mechanisms controlling uterine activity, is proposed.

Novel in vitro system for functional assessment of oxytocin action

“…We were next interested to determine to what extent these cells also exhibited the prototypical physiological response of myometrial cells to OT stimulation, i.e., cellular contraction. To address this question, we developed an in vitro assay system consisting of a modified collagen lattice contraction assay that was based on a published procedure (6). In contrast to the published procedure, in which cells were imbedded in a collagen matrix, in the present system cells are layered on top of a preformed collagen matrix.…”

“…As an integral component of the assay system, we have characterized two novel nontransformed human myometrial cell lines with respect to their biochemical and physiological OT response characteristics. In the present in vitro contraction assay, cells were layered on top of a preestablished collagen lattice, whereas in the original approach cells were embedded within the lattice (6). With the present approach, less cells are needed per lattice (i.e., 25,000 vs. 150,000 cells).…”

“…The present in vitro system used to measure myometrial cell contraction in response to OT was developed on the basis of a technique originally described by Dallot et al (6). Collagen type 1 from rat tail (Sigma) was resuspended overnight in 0.01 N HCl to prepare a 5 mg/ml stock solution and kept at 4°C until use.…”

Secreted surfactant protein A from fetal membranes induces stress fibers in cultured human myometrial cells

“…After collection, the myometrial biopsies were placed in DMEM supplemented with 100 U/ml penicillin and 100 g/ml streptomycin. Human uterine smooth muscle cells were obtained by the explants method, as described previously (6). Cells were cultured in DMEM supplemented with antibiotic solution and 10% FCS and routinely passaged when 90 -95% of cells were confluent.…”

Effect of low-intensity focused ultrasound on endothelin-1, nitrogen monoxide and oxytocin receptor in the uterine tissues of Sprague-Dawley rats following abortion

“…ET is the strongest and most persistent vasoconstrictor peptide known, and has four isoenzyme isomers: ET-1, ET-2, ET-3 and vasoactive intestinal peptide contraction. Uterine tissue mainly generates ET-1 (7), which promotes muscle contraction by Gq-phospholipase C-inositol 1,4,5-trisphosphate signal transduction pathways, protein kinase Cθ (PKCθ) and phospholipase A2 to activate the synthesis of arachidonic acid, PKC and sphingosine kinase, which in turn activate RhoA/Rho kinase (7)(8)(9). NO is an endothelium-derived relaxing factor, which activates the guanylyl cyclase (GC) to inhibit spontaneous uterine smooth muscle contraction (10,11).…”