Ruth Kaplan‐Kraicer

Tel Aviv University

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Pigment Epithelium Derived Factor (PEDF) a Crucial Regulator of Ovarian Angiogenesis.

Biology of Reproduction (2012)

Dana Chuderland Ido Ben‐Ami Ruth Kaplan‐Kraicer Hadas Grossman Ronit Satchi Fainaro

Angiogenesis is critical for the development of ovarian follicles. Blood vessels are abrogated from the follicle until ovulation, when they invade the follicle to support the developing corpus luteum. Granulosa cells secrete anti-angiogenic factors, of a yet undefined molecular identity, that shield against premature vascularization. Abnormalities in ovarian angiogenesis contribute to several pathologies such as ovarian hyper-stimulation syndrome (OHSS).Our aim was to illustrate the role of PEDF, a potent anti-angiogenic factor, in ovarian angiogenesis and to evaluate its potential quality as a therapy agent for OHSS. To study the role of PEDF in the ovary we employed a broad range of biochemical and molecular methodologies on ovarian histological sections, as well as on oocytes and granulosa cells. We further employed a variety of established angiogenesis assays to examine the effect of PEDF on the well-known properties of human umbilical vein endothelial cells. OHSS was induced in mice by administration of excess doses of PMSG followed by hCG. The therapeutic properties of PEDF were tested by injecting it along with the hCG. OHSS was assessed by monitoring two classical parameters (body weight and vascular permeability). We are the first to demonstrate a cardinal role for PEDF in the ovary. Immunostaining of PEDF has localized it within the follicle with significantly strong staining in the oocyte. Interestingly PEDF was found to be synthesized and secreted only by granulosa cells in a dynamically regulated fashion, mediated by gonadotropins, estradiol and progesterone. We found that the levels of PEDF and VEGF are reciprocally regulated; PEDF level decreases towards ovulation while VEGF level increases, thereby regulating endothelial cells activity, and by that, angiogenesis. We showed that the PEDF-VEGF balance changes when OHSS is induced and that administration of PEDF to mice treated with access gonadotropins, significantly reduced the measured OHSS parameters (10 mice/treatment*3 groups; p<0.05; t-test). To summarize, our findings have considerable implications in both physiological and clinical settings. The finding that PEDF is highly expressed in the ovary in reciprocity to VEGF suggests its function as an ovarian angiogenesis regulator, and offers a rational for a potential use of PEDF as a therapeutic agent.

PEDF

10.1093/biolreprod/87.s1.575

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Citations (1)

Viscosity and refractive index of follicular fluid in relation to in vitro fertilization

Journal of Assisted Reproduction and Genetics (1996)

Benjamin Fisch Linda Harel S. Amit Ruth Kaplan‐Kraicer Naomi Mor

Follicular fluid

10.1007/bf02066526

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Citations (5)

Practical and informative charts for in vitro fertilization and embryo transfer

Journal of In Vitro Fertilization and Embryo Transfer (1987)

Yona Tadir Ruth Kaplan‐Kraicer Richard D. Kelly R. Tepper S. Amit

10.1007/bf01555385

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Citations (2)

A novel regulatory pathway in granulosa cells, the LH/human chorionic gonadotropin-microRNA-125a-3p-Fyn pathway, is required for ovulation

The FASEB Journal (2015)

Hadas Grossman Dana Chuderland Lihi Ninio-Many Noa Hasky Ruth Kaplan‐Kraicer

Granulosa cells support the developing oocytes and serve as transducers of the ovulatory stimulus induced by LH surge. Fyn kinase is expressed in granulosa cells, though its role in these cells has not been studied. In human embryonic kidney 293T cells, microRNA (miR)-125a-3p down-regulates Fyn expression, causing a decrease in cells' migratory ability. Our aim was to explore the role of miR-125a-3p and Fyn in granulosa cells toward ovulation, focusing on migration as a possible mechanism. We demonstrate expression of miR-125a-3p and Fyn in mouse mural granulosa cells of preovulatory follicles and miR-125a-3p-induced down-regulation of Fyn expression in a granulosa cell line (rat). Administration of human chorionic gonadotropin (hCG; LH analog) caused a 75% decrease in the in vivo miR-125a-3p:Fyn mRNA ratio, followed by a 2-fold increased migratory ability of mural granulosa cells. In the hCG-treated granulosa cell line, miR-125a-3p expression was decreased, followed by Fyn up-regulation and phosphorylation of focal adhesion kinase and paxillin, enabling cell migration. An in vivo interference with miR-125a-3p:Fyn mRNA ratio in granulosa cells by intrabursal injections of Fyn small interfering RNA or miR-125a-3p mimic caused a 33 or 55% decrease in the number of ovulated oocytes, respectively. These observations reveal a new regulatory pathway in mural granulosa cells under the regulation of LH/hCG. Modulation of cell migration may account for the significance of the LH/hCG-miR-125a-3p-Fyn pathway to ovulation.

FYN

Human chorionic gonadotropin

Granulosa cell

10.1096/fj.14-269449

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Citations (25)

De novo synthesis of protein phosphatase 1A, magnesium dependent, alpha isoform (PPM1A) during oocyte maturation

Cellular & Molecular Biology Letters (2012)

Dana Chuderland Zeev Dvashi Ruth Kaplan‐Kraicer Daniella Ben‐Meir Ruth Shalgi

Abstract Oocyte maturation in mammals is a multiple-stage process that generates fertilizable oocytes. Ovarian oocytes are arrested at prophase of the first meiotic division characterized by the presence of a germinal vesicle. Towards ovulation, the oocytes resume meiosis and proceed to the second metaphase in a process known as maturation; they undergo nuclear and cytoplasmic changes that are accompanied by translation and degradation of mRNA. Protein phosphatase 1A, magnesium dependent, alpha isoform (PPM1A), which belongs to the metal-dependent serine/threonine protein phosphatase family, is highly conserved during evolution. PPM1A plays a significant role in many cellular functions such as cell cycle progression, apoptosis and cellular differentiation. It works through diverse signaling pathways, including p38 MAP kinase JNK and transforming growth factor beta (TGF-β). Herein we report that PPM1A is expressed in mouse oocytes and that its mRNA level rises during oocyte maturation. Using quantitative real-time polymerase chain reaction (qPCR) and western blot analysis, we found that PPM1A mRNA is synthesized at the beginning of the maturation process and remains elevated in the mature oocytes, promoting the accumulation of PPM1A protein. Since PPM1A function is mainly affected by its level, we propose that it might have an important role in oocyte maturation.

Germinal vesicle

10.2478/s11658-012-0022-7

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Citations (8)

The Role of Pigment Epithelium-Derived Factor in the Pathophysiology and Treatment of Ovarian Hyperstimulation Syndrome in Mice

The Journal of Clinical Endocrinology & Metabolism (2013)

Dana Chuderland Ido Ben‐Ami Ruth Kaplan‐Kraicer Hadas Grossman Raphaël Ron-El

Ovarian hyperstimulation syndrome (OHSS) is a potentially life-threatening complication of assisted reproduction. OHSS is induced by an ovarian release of vasoactive, angiogenic substances that results in vascular hyperpermeability, leakage, and shift of fluids from blood vessels into the extravascular space with consequent ascites and edema that are attributed to vascular endothelial growth factor (VEGF). Our objective was to examine a physiological approach for preventing and treating OHSS, based on negating the VEGF network. We used a mouse OHSS model and cultured granulosa cells. Changes in pigment epithelium-derived factor (PEDF) and VEGF were measured by quantitative PCR and Western blot analysis. OHSS was recorded by changes in body weight and in peritoneal vascular leakage, quantified by the modified Miles vascular permeability assay. Granulosa cells produced and secreted the anti-angiogenic factor, PEDF, in an inverse fashion to VEGF. The physiological PEDF-VEGF counterbalance was found to be impaired in the mouse OHSS model. Treatment of OHSS-induced mice with low doses of recombinant PEDF (rPEDF) alleviated OHSS signs including edema (P < .001) and vascular leakage (P < .001) and reduced the level of ovarian VEGF mRNA. Low doses of rPEDF also reduced VEGF mRNA levels in granulosa cells in vitro. However, these effects were not seen at higher doses of rPEDF, suggesting a hormetic mechanism of rPEDF action. These observations provide a new perspective into the pathophysiology of OHSS, namely, high expression level of VEGF together with a nearly undetectable level of PEDF. A replacement therapy with rPEDF is suggested as an innovative physiological treatment for OHSS. Finally, control of the PEDF-VEGF reciprocal relationship could open new therapeutic avenues for other angiogenic-related fertility pathologies.

PEDF

Ovarian hyperstimulation syndrome

Vascular permeability

Pathophysiology

10.1210/jc.2012-3037

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Citations (39)

Individualized Approach to Selection of Ovum Pick-up Method for In Vitro Fertilization

Journal of Gynecologic Surgery (1989)

Yona Tadir Benjamin Fisch Haim Pinkas Arik Kahane Ruth Kaplan‐Kraicer

Treatment guidelines are given defining individualized criteria for patient selection for various modes of follicular aspiration as part of a comprehensive approach to the investigation and treatment of the infertile patient. The controversies concerning the treatment of choice and comparative results are reviewed. (J GYNECOL SURG 5:133, 1989)

10.1089/gyn.1989.5.133

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Citations (3)

Myristoylated alanine-rich C kinase substrate, but not Ca2+/calmodulin-dependent protein kinase II, is the mediator in cortical granules exocytosis

Reproduction (2008)

Lina Tsaadon Ruth Kaplan‐Kraicer Ruth Shalgi

Sperm–egg fusion induces cortical granules exocytosis (CGE), a process that ensures the block to polyspermy. CGE can be induced independently by either a rise in intracellular calcium concentration or protein kinase C (PKC) activation. We have previously shown that myristoylated alanine-rich C kinase substrate (MARCKS) cross-links filamentous actin (F-actin) and regulates its reorganization. This activity is reduced either by PKC-induced MARCKS phosphorylation (PKC pathway) or by its direct binding to calmodulin (CaM; CaM pathway), both inducing MARCKS translocation, F-actin reorganization, and CGE. Currently, we examine the involvement of Ca 2 + /CaM-dependent protein kinase II (CaMKII) and MARCKS in promoting CGE and show that PKC pathway can compensate for lack of Ca 2 + /CaM pathway. Microinjecting eggs with either overexpressed protein or complementary RNA of constitutively active αCaMKII triggered resumption of second meiotic division, but induced CGE of an insignificant magnitude compared with CGE induced by wt αCaMKII. Microinjecting eggs with mutant-unphosphorylatable MARCKS reduced the intensity of 12- O -tetradecanoylphorbol 13-acetate or ionomycin-induced CGE by 50%, indicating that phosphorylation of MARCKS by novel and/or conventional PKCs (n/cPKCs) is a pivotal event associated with CGE. Moreover, we were able to demonstrate cPKCs involvement in ionomycin-induced MARCKS translocation and CGE. These results led us to propose that MARCKS, rather than CaMKII, as a key mediator of CGE.

MARCKS

10.1530/rep-07-0554

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Citations (21)

Molecular cloning of rat sperm galactosyl receptor, a C-type lectin with in vitro egg binding activity

Molecular Reproduction and Development (2000)

Eugene Rivkin Laura L. Tres Ruth Kaplan‐Kraicer Ruth Shalgi Abraham L. Kierszenbaum

Rat sperm galactosyl receptor is a member of the C-type animal lectin family showing preferential binding to N-acetylgalactosamine compared to galactose. Binding is mediated by a Ca(2+)-dependent carbohydrate-recognition domain (CRD) identical to that of the minor variant of rat hepatic lectin receptor 2/3 (RHL-2/3). The molecular organization of the genomic DNA, cDNA, and derived amino acid sequence of rat testis galactosyl receptor have been determined and in vitro fertilization studies were conducted to ascertain its role. We have determined that the rat testis galactosyl receptor gene generates two mRNA species: one species, designated liver-type, is identical to RHL-2/3; the other, designated testis-type, contains one unspliced intron (86 nt) which alters the reading frame and changes the amino acid sequence of the carboxyl terminus. As a result, the CRD (glutamine-proline-aspartic acid/QPD) and flanked Ca(2+)-binding amino acid sequences were not present in the testis-type protein. Northern and Southern blots demonstrated presence of transcripts with unspliced intron in rat sperm but not liver. Similarly, antibody, raised against a synthetic 12-amino acid peptide (p12) encoded by the unspliced intron, recognized in immunoblots a 54 kDa receptor protein in protein extracts from testis but not from liver. Immunofluorescence and immunogold electron microscopy studies demonstrated that both protein species localized on the plasma membrane surface of the head and tail of rat sperm. Furthermore, capacitated rat sperm preincubated with polyclonal antisera to RHL-2/3 or to the CRD of the liver-type galactosyl receptor showed a statistically significant decrease in the in vitro fertilization rate. We conclude that rat sperm galactosyl receptor may play a role in egg binding and that an undetermined molecular mechanism operates to generate two proteins with identical intracellular amino terminal domain but only one of them displays a CRD and associated Ca(2+)-binding sites at the carboxyl terminal extracellular domain.

C-type lectin

10.1002/1098-2795(200007)56:3<401::aid-mrd11>3.0.co;2-7

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Citations (15)

The role of Src family kinases in egg activation

Developmental Biology (2007)

Tomashov-Matar Reut Mattan Levi Dafna Tchetchik Ruth Kaplan‐Kraicer Ruth Shalgi

FYN

Src family kinase

10.1016/j.ydbio.2007.09.010

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Citations (19)