Expression of a cytosolic phospholipase A(2) by ovine endometrium on days 11-14 of a simulated oestrous cycle

Oxytocin stimulates the synthesis and secretion of PGP(2 alpha) from uterin e tissues in vivo and in vitro late in the ovine oestrous cycle. The synthe sis of eicosanoids is dependent upon the availability of free arachidonic a cid which is released through the activity of arachidonate releasing phosph olipases. In the present study, the following hypothesis was tested: the ov ine endometrium expresses a cytosolic phospholipase A(2) (cPLA(2)) and expr ession or activity of cPLA(2) increases as uterine secretory responsiveness to oxytocin develops late in the oestrous cycle. Endometrial tissue was co llected from cyclic ewes on day 15 of the oestrous cycle for the preparatio n of tissue homogenates and isolation of mRNA to determine whether ovine en dometrium expressed a cPLA(2). A 110 kDa band was detected by western blott ing, indicating the presence of a putative ovine cPLA(2). A 834 bp fragment of the ovine cPLA(2) shared 87% homology with human and mouse cDNA, and no rthern blot hybridization analysis indicated a single 3.4 kb transcript. A total of 20 ewes were ovariectomized and treated with progesterone and oest rogen to simulate the oestrous cycle to determine whether the expression or activity of ovine cPLA(2) changed during the onset of uterine secretory re sponsiveness to oxytocin in vivo. On days 11-14 (n = 5 per day) of a simula ted oestrous cycle, caruncular endometrium was evaluated for expression of ovine cPLA(2) mRNA. and protein and the synthesis of PGF(2 alpha) in respon se to melittin (a potent stimulator of PLA, activity). Immunoreactive cPLA( 2) and cPLA(2) mRNA were observed on all days and did not increase during t he development of uterine responsiveness to oxytocin in vivo. Similarly, me littin increased the synthesis of PGF(2 alpha) irrespective of day, indicat ing the presence of a functional cPLA(2) on all days examined. These data i ndicate that the ovine endometrium expresses a functional cPLA(2), and that ample concentrations of cPLA(2) are present by day 11 of a simulated oestr ous cycle.