Hypotonic Stress Increases Cyclooxygenase-2 Expression and Prostaglandin Release from Amnion-derived WISH Cells

David W Lundgren,Robert M Moore,Patricia L Collins,John J Moore

doi:10.1074/jbc.272.32.20118

Abstract

This report examines the effect of cell volume expansion on cyclooxygenase-2 (COX-2) mRNA expression, COX-2 protein expression, and prostaglandin E2 release from human amnion-derived WISH cells. Earle's balanced salts solution (EBSS) with limited NaCl concentration was utilized as the induction medium. COX-2 mRNA was elevated 6-fold in cells incubated for 1 h in hypotonic EBSS. COX-2 mRNA expression was not increased when raffinose or sucrose were used to reconstitute low NaCl. Actinomycin D blocked COX-2 mRNA increase by hypotonic stress, while cycloheximide enhanced COX-2 mRNA expression. COX-2 mRNA and protein concentrations increased as a function of decreasing media osmolarity and incubation time in hypotonic EBSS. Hypotonic EBSS induced a 3-fold increase in prostaglandin E2 release. WISH cells transiently transfected with a luciferase expression vector driven by the human COX-2 promoter for the COX-2 gene show a 3-fold increase in luciferase activity when incubated in hypotonic EBSS. COX-2 mRNA levels in primary human amnion cells were also increased by hypotonic stress. This study suggests that amnion cell COX-2 gene expression is regulated by cell volume expansion and/or increased plasma membrane tension.

Highlights

Prostaglandins have a central role in regulating human parturition
Hypotonic Stress and COX-2 mRNA Expression—COX-2 mRNA expression was initially examined in WISH cells incubated for 1 h in hypotonic versus isotonic Earle’s balanced salts solution (EBSS)
Relative to the concentration of COX-2 mRNA from cells incubated in isotonic EBSS, COX-2 mRNA expression was markedly elevated in hypotonically stressed cells (Fig. 1, lanes 2, 5, and 8 versus lanes 3, 6, and 9, respectively)

Summary

Introduction

Prostaglandins have a central role in regulating human parturition. Prostaglandin E2 (PGE2), the primary prostaglandin produced by fetal membranes during labor, may be directly involved in the initiation and maintenance of uterine contractions (1, 2). The biochemical mechanism by which mechanical stretching increases prostaglandin release from these tissues is largely unknown, it was recently reported that mechanical stretching of cultured rat glomerular mesangial cells induces a number of immediate early response genes including the gene for COX-2 (16). Based on this knowledge, we hypothesize that an increase in the volume of amnion cells and the resulting increase in plasma membrane tension induce COX-2 gene transcription and promote PGE2 release from fetal tissue. This study provides evidence that an increase in cell volume up-regulates COX-2 mRNA expression and elevates prostaglandin biosynthesis in amnion cells

Methods

Results

Conclusion