Osmotic stress response is altered in leiomyoma cells

Abstract

OBJECTIVE: Clinical treatment of leiomyomas with GnRH analogue produces a rapid reduction in leiomyoma volume. Rapid leiomyoma regrowth when GnRH treatment is withdrawn suggests the reduction in volume is due to transport of water, yet little is known about the response of leiomyoma cells to osmotic stress. Nuclear factor of activated T cells 5 (NFAT5) is the key osmo-sensing transcription factor in human cells. Our goal was to characterize osmosensing via NFAT5 in leiomyoma cells. DESIGN: Prospective, human model experiments MATERIALS AND METHODS: Osmotic stress was induced by culturing immortalized human leiomyoma and myometrial cells in DMEM-10% FBS with additional osmolytes of NaCl at 25, 50 or 100 mM for 8 hours. Proliferation assays were performed by the sulforhodamine B method. Apoptosis was assessed by caspase 3/7 activity assay. Expression of NFAT5 in tissue and cells was determined with qRT-PCR and western blot analysis. RESULTS: Leiomyoma cell growth was inhibited 18% at 50 mM NaCl compared to 48% in myometrial cells (p<0.05), consistent with altered osmosensing in leiomyoma cells. This finding was not due to apoptosis, as evidenced by no change in caspase activity. NFAT5 transcripts were increased in leiomyoma 1.5 fold ± 0.19 compared to myometrium in 4 of 7 patient samples (p<0.05). Concentration dependent upregulation of NFAT5 mRNA was observed in osmotically stressed leiomyoma cells. At 50mM NaCl, NFAT5 transcripts were increased 2.5-fold compared to control. CONCLUSIONS: Leiomyoma cells were more resistant to changes in osmotic stress than myometrial cells, suggesting dysregulation in osmotic signaling in leiomyomas. This change was accompanied by differential expression of NFAT5 in leiomyoma tissue. Altered osmotic signaling in leiomyoma cells suggests a central role for water regulation in the rapid change of leiomyoma volume with GnRH analogue treatment.