275 EXPRESSION OF THE ACYL-COA SYNTHETASE 4 IN THE PERI-IMPLANTATION MOUSE UTERUS

Abstract

Acyl-CoA synthetase 4 (ACS4) is an arachidonate-preferring enzyme abundant in steroidogenic tissues and postulated to modulate eicosanoid production. Arachidonate that is esterified predominantly in phospholipids is a precursor of eicosanoids. After its release by the action of cytosolic phospholipase A2 (cPLA2), arachidonate can be converted to prostaglandins, thromboxanes, and leukotrenes via the cyclo-oxygenase (COX1 and COX2) and lioxygenase pathways, respectively, depending on the cell type. It is reported that eicosanoids have influence on inflammation, vascularization, and parturition. Then again, free arachidonate released from the plasma membrane is reesterified into phospholipids to prevent constant synthesis of potent eicosanoids. In the rodent, vasoactive prostaglandins are implicated in the implantation process. To further clarify ACS4 gene expression during pregnancy, we examined developmental expression in the peri-implantation uterus of the normal mouse and regulation of ACS4 by steroid hormone in ovariectomized mice treated with estradiol-17β (E2) and/or progesterone (P4). Adult female mice (BDF1, 6 weeks old) were mated with fertile males of the same strain. The morning on which a vaginal plug was found was designated Day 0.5 of pregnancy. Mice were sacrificed between 09:00 and 10:00 h on Days 2.5 to 6.5 of pregnancy. To induce and maintain delayed implantation, mice were ovariectomized on the morning (09:00 to 10:00 h) of Day 3.5 of pregnancy and received a daily injection of P4 from Days 4.5 to 6.5. To terminate the delay and induce implantation, the P4-primed delayed mice were given an injection of E2 on Day 6.5. COX1, COX2, cPLA2, and ACS4 mRNA were analyzed by Northern blot analysis and real-time PCR. The expression of COX1 mRNA did not show much variation on the various days in the normal pregnancy and ovariectomized mice. The levels of COX2 mRNA were high on Day 4.5 of pregnancy and dramatically decreased on Day 5.5 of pregnancy. In the ovariectomized mouse uterus, COX2 mRNA levels declined rapidly at Day 4.5 of pregnancy. The expression of cPLA2 mRNA was not altered in the normal pregnancy. However, the expression was increased in the ovariectomized mouse as compared with that in the normal pregnancy mice. An ACS4 transcript was already present in the uterus at low level on Day 3.5 until the initiation of attachment reaction after which the expression was up-regulated. In the ovariectomized mouse uterus, ACS4 mRNA was increased at Day 4.5 of pregnancy as compared with the normal pregnancy mice. To determine whether ovarian steroids influenced the induction of ACS4 gene, ovariectomized mice were treated with (E2) and/or (P4). Treatment with P4 maintained the expression of ACS4 mRNA in the ovariectomized mouse uterus. In contrast, combined treatment with P4 and E2 modestly decreased the levels of ACS4 mRNA as compared with Day 7.5 of normal pregnancy. Overall, these results suggest that the ACS4 gene is regulated in the implantation process and influenced by ovarian steroids.