John R. Pepperell

Dissociation between intracellular calcium elevation and development of human oocytes treated with calcium ionophore

OBJECTIVE To develop an acceptable model system to study calcium activation of human oocytes. DESIGN Study of oocyte development and intracellular calcium [Ca]i dynamics of activated oocytes. SETTING Research center affiliated with infertility service. MAIN OUTCOME MEASURE Morphologic evidence of meiotic maturation and cell division under high-power Hoffman optics with an inverted microscope. Meiotic maturation was determined by the number of polar bodies or the presence of a pronucleus, and cell division was determined by evidence of a cleavage furrow or presence of blastomeres. To monitor the effect of calcium ionophore on [Ca]i levels, oocytes were incubated with fura-2 (2 microM) for 30 minutes and [Ca]i was determined by rationing the emission fluorescence (510-nm long-pass filter) during simultaneous excitation at 340 and 380 nm with a microspectrofluorimeter. RESULT(S) All oocytes loaded with fura-2 and then exposed to ionophore exhibited an isolated elevation of [Ca]i, followed by prompt return to baseline levels. None of the oocytes showed signs of cleavage or of meiotic maturation after treatment with calcium ionophore. CONCLUSION(S) Human oocytes activated with calcium ionophore A23187 or ionomycin exhibited elevated [Ca]i but remained resistant to subsequent meiotic maturation and cleavage. Our results differ from some reports of parthenogenetic activation of human oocytes. These differences may result from different activation protocols or culture conditions. Because none of the 126 oocytes cleaved after the activation protocols used in these experiments, this approach should provide an ethically acceptable model system to study calcium dynamics in human oocytes.

Intraovarian Regulation by the Ovarian Renin-Angiotensin System

Pituitary and placental gonadotrophins may be considered the major regulators of ovarian function, however, current developments in ovarian endocrinology show that other peptides also have important regulatory actions in the ovary. These latter peptides include such factors as insulin-like growth factor 1 (IGF-l), transforming growth factor-b (TGF-b), tumor necrosis factor-a (TNF-a), TNF-b, and other cytokines. Also included with these ovarian regulatory peptides are the angiotensin peptides. In this review we will discuss evidence that the ovary contains an intrinsic renin-angiotensin system which produces a variety of bioactive angiotensin peptides that can have important implications for the ovary, both in health and disease. The renal renin-angiotensin system is classically considered to produce the bioactive products, angiotensin I1 (Ang 11) and Ang 111 by a cascade reaction. Thus, activation of inactive prorenin to the active enzyme by cleavage of a 43 amino acid prosegment initiates the cascade. Active renin is an aspartyl proteinase that degrades angiotensinogen to produce Ang I by the cleavage of 4 amino acids from the carboxy terminus of angiotensinogen. Subsequently, activity of the dipeptidyl peptidase, angiotensin converting enzyme (ACE), clips a further 2 amino acids from the carboxy terminus of Ang I, to produce the octapeptide Ang 11. Further proteolytic removal of the amino terminal aspartyl residue by angiotensinase results in the formation of the septapeptide, Ang 111 (reviewed in 1).

Potential Role of the Renin-Angiotensin System in Polycystic Ovaries

Developments over the last ten years have identified that cyclical regulation of the ovary is due not only to complex interactions between gonadotrophins and steroids, but also to other factors that effect ovarian function. Our work has focused on a new ovarian autocrine system, the ovarian reninangiotensin system (OVRAS) that accumulating evidence points to having effects in regulation of ovulation, steroidogenesis, follicular atresia, and polycystic ovarian syndrome.