P-303 Supplementing IVM medium with mesenchymal stem cell-conditioned medium, L-carnitine, and Repaglinide to improve maturation and developmental competence of oocytes derived from normal and endometriosis mice

Abstract

Abstract Study question Does supplementing in vitro maturation (IVM) medium with bone marrow mesenchymal stem cell-conditioned medium (BMSC-CM), L-carnitine (LC), and Repaglinide (RG) enhance the developmental competence of oocytes derived from endometriosis-induced mice? Summary answer L-carnitine and BMSC-CM supplementation improved maturation and fertilization, and developmental competence of oocytes following IVF by modulating nitro-oxidative stress and accelerating nuclear maturation of oocytes. What is known already Endometriosis (EMS) is a detrimental condition of the female reproductive system affecting up to 15% of women of reproductive age. Oocytes retrieved from EMS-affected ovaries are more likely to fail IVM and to show altered morphology and lower cytoplasmic mitochondrial content. Mesenchymal stem cells (MSCs) secrete various types of cytokines, growth factors, bioactive factors, and tissue regenerative components into mesenchymal stem cell-conditioned medium. L-carnitine is an antioxidant playing an important role in cell metabolism and is crucial for fatty acid metabolism. Repaglinide (RG) is an anti-hyperglycemic medication that induces insulin secretion by attaching to β cells of the pancreas. Study design, size, duration To induce EMS, two groups of mice were used. The first group, as donor mice, were intraperitoneally injected with estradiol-17β depot for one week. Then, they were sacrificed on day 14, and their uterine horns were removed. The suspension including tissue fragments from uterine horns was injected intraperitoneally to the recipient mice group. To ensure the induction of EMS, ovaries of some mice in both normal and EMS groups were analyzed by histological examination. Participants/materials, setting, methods Adult female NMRI mice (6-8 weeks old) were divided into two normal and EMS-induced mice. Oocytes obtained from normal and EMS-induced mice were cultured in the IVM medium supplemented with RG, LC, and BMSC-CM for 24 hours. Subsequently, IVM, IVF, embryo development rates, blastocyst cell numbers were assessed. Moreover, the levels of reactive oxygen species (ROS), nitric oxide (NO), and total antioxidant capacity (TAC) of the IVM medium were evaluated. Main results and the role of chance Microscopic studies of ovarian tissue samples showed that folliculogenesis, quality of follicles, and oocyte quality reduced in the EMS-induced group. The IVM results represent that 0.3 mg/ml and 0.6 mg/ml of LC, and 25% and 50% of BMSC-CM enhanced the percentage of MII oocytes significantly. Indeed, the highest IVM rate in both the normal and EMS-induced mice was observed in the BMSC-CM 50% group. More importantly, 0.3 and 0.6 mg/ml of LC, and 25% and 50% of BMSC-CM treatments showed a dramatic decrease in ROS and NO levels and a significant enhancement in TAC levels compared to the control group. Our results also indicated that there was a substantial improvement in terms of IVF, cleavage, and blastocyst rates in all treatment groups compared to the control group. The highest blastocyst formation rate was obtained in 50% of the BMSC-CM group. In normal groups, there was a dramatic increase in the mean of total cell number and TE cells in 1µM RG, 0.3 and 0.6 mg/ml LC, 25% and 50% BMSC-CM. In EMS-induced groups, except for the 1M RG group, all treatment groups exhibited a significant difference in the mean total cell population and TE cells compared to the control group. Limitations, reasons for caution Due to limited laboratory facilities, the levels of growth factors, pro-inflammatory cytokines, anti-apoptotic, and antioxidants were not measured in the IVM medium. We, therefore, do not know which of the constituents of the BMSC-CM, in particular, had promoter effects on the oocyte maturation, fertilization, and developmental competence of IVF embryos. Wider implications of the findings These approaches may outline the clinical applications of LC and BMSC-CM in the assisted reproductive technology (ART) setting and pave the way to improve the maturation culture. Also, these findings can advance the efficiency of fertility outcomes in endometriosis-related infertile couples. Trial registration number N/A