P-649 Unconventional action of Luteinizing Hormone

Abstract

Abstract Study question Has Luteinizing Hormone (LH) a role in the activation/survival of follicles of the primary ovarian reserve? Summary answer LH promotes survival and perhaps activation/growth of primordial and primary follicles, proliferation of granulosa cells of secondary follicles, reduces follicle atresia at later growing stages. What is known already In the mammalian ovary, Primordial Follicles (PMFs) constitutes the primary ovarian reserve, which declines over time due to physiological events such as atresia, recruitment, and ovulation. While in the adult, the role of LH in steroidogenesis and ovulation is well established, recent work from our laboratory has shown that under chemotherapeutic treatments, LH supports PMFs survival in prepuberal mouse ovary. Study design, size, duration LH effects on follicle dynamics and some activities of pre-granulosa cells were investigated in vitro and in vivo. In vitroexperiments were performed on PD4 GFP/Kit mice ovarian fragments and on PD8 CD1 mice ovarian somatic cells cultured up to six days in continuous presence of 0.2, 0.5, 1IU LH. In vivo experiments were performed on 9 PD4 and 8 PD28 CD1 mice receiving an intra-peritoneal injection for eight consecutive days of 1-5IU LH. Participants/materials, setting, methods Number and diameters of the GFP stained oocyte were counted in the ovarian fragments. Kit-Ligand and LH-R mRNA expression levels were analyzed by qPCR in pre-granulosa cells. For in vivo experiments, ovarian volume was measured by Micro-Ct and follicle counts were performed on histological sections. Estrous cycle phases were monitored by vaginal smear during the 8 days of treatment. Statistics was performed using Student’s t-test. Main results and the role of chance In fragments of PD4 mouse ovaries cultured in vitro for six days, 1IU LH significantly increased the number of oocytes with a diameter >40 mm (CTRL 17.7% vs LH 36.07%, p < 0,05), suggesting activation of PMFs. Moreover, the hormone increased the expression of mRNA for LHR and KL in the somatic ovarian cells and promoted their proliferation in vitro. The effects of 1IU LH on PD4 females can be summarized as follows: 1. Ovary volume increase (CTRL 0.3 mm3 vs LH 0.36 mm3, p = 0.07), 2. higher percent of PMFs (CTRL 68.227% vs LH 78.29%, p < 0,05) associated with lower percent of primary (CTRL 7.2% vs LH 4.05%, p < 0.04) and growing follicles (CTRL 14.18% vs LH 8.56%, p = 0.1), 3. higher number of granulosa cell layers (> three layers) in secondary follicles (CTRL 42.8% vs LH 54.6%), and 4. lower percent of atretic follicles (CTRL 2.64% vs LH 1.72%, p = 0.1). Similar results were obtained on 28 dpp mice. Morever, we observed that corpora lutea were present only in ovaries of LH-treated females and that all such females were in estrus. Limitations, reasons for caution The study was conducted only in mice. Wider implications of the findings These studies have highlighted new possible roles of LH on follicle classes (primordial/primary) whose development is considered gonadotropin independent. The results, if confirmed in humans, imply that LH might be used as a treatment for patients with ovarian disorders involved the survival/dynamics of the ovarian reserve. Trial registration number NA