Abstracts Figures will be available only online

Downloaded from rsx.sagepub.com at UNIV OF PITTSBURGH on June 19, 2015

Downloaded from rsx.sagepub.com at UNIV OF PITTSBURGH on June 19, 2015

Scientific Abstracts

Reproductive Sciences Vol. 22, Supplement 1, March 2015


O-002 Hsa-miR-30d Is Transported By Exosomes From Human Endometrial Epithelium To the Trophectoderm of Preimplantation Embryos and Modulates Embryo Adhesion. Nuria Balaguer,1 Felipe Vilella,1 Juan M Moreno,1 Maria Herrero,1 Carlos Simon.1,2 1Fundación IVI, Fundación Instituto Valenciano de Infertilidad (FIVI), Valencia, Spain; 2Department of Ob/Gyn (C.S), Standford University School of Medecine, Standford, CA, USA. INTRODUCTION: The knowledge about the molecular regulation of the maternal-embryo communication during embryonic development is gaining momentum. Our previous work described how endometrial miRNAs were secreted as exosome-associated molecules and then taken up into the trophectoderm. The present research aims to trace hsa-miR-30d in the dialogue between human endometrium and blastocyst. METHODS: Proteomic assay was performed to identify the proteins responsible for the loading of hsa-miR-30d into exosomes. To identify the hsa-miR-30d production, human epithelial cells (hEEC) were incubated 20 h. with a SmartFlare probe™ (miR-30d or scramble) (Millipore). hEEC were co-culture with day-2 mouse embryos and visualized, after 10 h., with a confocal microscope (Olympus). Embryo transcriptomic modifications were assessed by expression microarrays and adhesive modifications by embryo adhesion assays. Student’s t test was used. RESULTS: Proteomic analysis showed 15 proteins that bind to miR30d. Functional analysis of these proteins revealed a predominant association with biological functions related to RNA-post transcriptional modifications, cell adhesion and embryonic morphogenesis. Among them, hnRNPC1 constitutes the main candidate for conducting the load of miR-30d into exosomes, since its role in the stability and transport of mRNAs from the nucleus to the cytoplasm is already established. The transfer of miR-30d from the hEEC to the trophoectoderm of mouse embryos was demonstrated by using a Smartflare probe. However, this internalization only occurred when a complete cell to cell contact was established. Transcriptomic analysis revealed that miR-30d triggered the up-regulation of embryonic cadherin and integrin-mediated signaling

pathways. To confirm this data, functional embryo adhesion was tested comparing control, scrambled miRNA, miR-30d mimic, and miR-30d inhibitor (n=360). After 32h, miR-30d mimic increased adhesion vs scramble and this was abolished with miR-30d inhibitor (35.22±7.4% vs 53.44±6.4% vs 18.55±3.72%, respectively p=0.001). CONCLUSIONS: These findings represent an advance in our understanding of how functional miRNAs secreted from the endometrium are transported to the embryo and modifies their transcriptomic signature. NB & FV contributed equally

O-003 A Targeted Drug Delivery System for the Uterus. Jonathan Paul,1,3 Susan Hua,2 Roger Smith.1,3,4 1School of Medicine and Public Health, University of Newcastle, Newcastle, NSW, Australia; 2School of Biomedical Sciences and Pharmacy, University of Newcastle, Newcastle, NSW, Australia; 3Mothers and Babies Research Centre, Hunter Medical Research Institute, Newcastle, NSW, Australia; 4Endocrinology Unit, Hunter New England Area Health, Newcastle, NSW, Australia. INTRODUCTION: Current approaches to tocolysis are hampered by off-target effects of therapeutic agents. Use of liposomes as targeted nanocarriers has been explored previously, but not in the context of targeting the uterus. We have designed liposomes to specifically recognise the oxytocin receptor (OTR), and here we report a promising targeted drug delivery system for the human uterus. METHODS: In Vitro: Strips of non-labouring human myometrial tissue, or uterine tissue from pregnant mice, were suspended in organ baths connected to force transducers. Upon onset of contractions, strips were treated with non-targeted (IgG coated) or targeted (anti-OTR coated) liposomes loaded with nifedipine, salbutamol, rolipram or dofetilide. The effect on contractions was recorded. In Vivo: To assess their localization, fluorescently-tagged liposomes loaded with salbutamol were administered to pregnant mice daily from fetal gestation day 17 until labor. Following labor, internal organs were harvested and liposome localization was imaged using an IVIS-100. RESULTS: In Vitro: Non-targeted liposomes loaded with therapeutic agents had no effect on human uterine contractility in vitro. However, OTR-targeted liposomes loaded with nifedipine, salbutamol or rolipram abolished contractions, whereas targeted liposomes loaded with dofetilide increased contraction duration. The same results were observed for mouse uterine tissue. In Vivo: Non-targeted liposomes administered to pregnant mice localized only to the liver. OTR-targeted liposomes localised strongly to the uterus as well as liver, along with low detection in mammary tissue, but not in the brain or neonate. *Figure(s) will be available online. CONCLUSIONS: Our targeted liposomes demonstrate specificity for the OTR, and are capable of delivering drugs to uterine tissue that block or promote contractions. In a mouse model the liposomes successfully localize to the uterus, and therefore represent a promising drug delivery system for this tissue.

O-004 Molecular Evidence of Functional Progesterone Withdrawal in Human Labor. Lubna Nadeem,1 Oksana Shynlova,1 Xuesen Dong,3 Stephen Lye.1,2 1Lunenfeld-Tanenbaum Res. Inst., Mount Sinai Hosp., Toronto, ON, Canada; 2Dept Physiol&Ob/Gyn, Univ. of Toronto, Toronto, ON, Canada; 3Dept Urologic Sci., Univ of British Columbia, Vancouver, BC, Canada. INTRODUCTION: Progesterone (P4) withdrawal is essential for labor initiation. In animals this is achieved by reduced P4 levels, but in humans P4 levels remain elevated. While it has been suggested that there is a functional withdrawal of P4 the causal mechanisms remain unknown. We have shown that P4 suppresses the labor gene, connexin43 (Cx43), through a P4 receptor (PRA/B) mediated pathway in which: 1) PR binds AP-1 dimers and recruits transcriptional repressors to the AP-1 site of Cx43 promoter; 2) AP-1 factors have differential affinity for PR isoforms ie, PRA has stronger affinity for Fos members than PRB and 3) unliganded

Downloaded from rsx.sagepub.com at UNIV OF PITTSBURGH on June 19, 2015

Thursday Orals

Impaired Development of Early Endometriotic Lesions in CD 44 Knockout Mice. Jennifer F Knudtson, Marlen I Tellez Santos, Peter A Binkley, Rajeshwar R Tekmal, Robert S Schenken. Obstetrics and Gynecology, University of Texas Health Science Center, San Antonio, TX, USA. INTRODUCTION: Endometrial stromal and epithelial cells from women with endometriosis over express several CD44 specific splice variants compared to cells of women without endometriosis. CD44 functions as the principal receptor for hyaluronic acid. Previous studies have shown mesothelial cell-associated hyaluronic acid (HA) is involved in the attachment of endometrial stroma and epithelial cells to peritoneal mesothelial cells. Here, we assessed the role of CD44 in the formation of the early endometriotic lesion using a CD44 knockout mouse model. METHODS: Using an established murine model, endometrial tissue from donor mice [wild type (WT) and CD44 knockout (CD44-KO)] was used to induce endometriosis in recipient mice (WT and CD44-KO). Four groups of donor/recipients were created: 1) WT/WT, 2)CD44-KO/CD44-KO, 3) CD44-KO/WT, 4) WT/CD44-KO. Endometriotic lesions were visualized by fluorescent stereo microscopy and confirmed by hematoxylin and eosin stain. Immunohistochemistry, RNA expression and protein expression was performed to confirm decreased CD44 expression in knockout mice compared to wild type. RESULTS: Significantly (p=0.002) fewer lesions developed in Groups 2, 3, and 4 when compared to control Group 1. The greatest impairment of lesion development occurred when there was decreased CD44 expression in both donor endometrial and recipient peritoneal tissues (p20 were more likely to have a 5 minute APGAR 20mm Hg

≤20mm Hg


RR(95% CI)

aRR(95% CI)

APGAR 1 min

Abstracts of the 62nd Annual Scientific Meeting, March 26-28, 2015, San Francisco, California.

Abstracts of the 62nd Annual Scientific Meeting, March 26-28, 2015, San Francisco, California. - PDF Download Free
13MB Sizes 4 Downloads 51 Views