157 LEUKEMIA INHIBITORY FACTOR IMPROVES OOCYTE MATURATION AND DEVELOPMENTAL COMPETENCE IN PIGS

2014 ◽  
Vol 26 (1) ◽  
pp. 192 ◽  
Author(s):  
S. Haraguchi ◽  
T. Q. Dang-Nguyen ◽  
K. Kikuchi ◽  
F. Tanihara ◽  
S. Bodo ◽  
...  
Keyword(s):  
Embryo Development ◽  
Oocyte Maturation ◽  
Leukemia Inhibitory Factor ◽  
Es Cells ◽  
Embryonic Stem ◽  
Developmental Competence ◽  
Inhibitory Factor ◽  
Protein Levels ◽  
Underlying Mechanisms

A variety of growth factors and cytokines that are present in follicular fluid provide oocytes with a suitable environment for their maturation. One such cytokine is leukemia inhibitory factor (LIF). Although LIF-supplemented medium enhances embryo development in human, mouse, and bovine, studies investigating the effects of LIF on in vitro maturation (IVM) and subsequent embryo development are inconclusive. Additionally, the underlying mechanisms of LIF in oocyte maturation and embryo development after IVF have not been studied yet. In the present study, we examined the effect of recombinant porcine LIF (pLIF), produced in our laboratory, on porcine oocyte maturation and the mechanism of how LIF involves in oocyte maturation process at molecular level. The biological activity of pLIF was evaluated by sustenance of mouse embryonic stem (ES) cells with an undifferentiating state in ES medium supplemented with pLIF, and the final concentration (1 : 200, equivalent to 1000 U mL–1 of mouse LIF) was determined by serial dilution. Porcine cumulus–oocyte complexes (COC) were cultured in modified NCSU-37 medium supplemented with pLIF during the first 22 h [pLIF (+, –)], the latter 22 h [pLIF (–, +)], or whole 44 h [pLIF (+, +)] of IVM and the proportion of metaphase II (M-II) stage oocytes was observed. Oocyte maturation was enhanced in each group by supplementation with pLIF [pLIF (+, –): 76.1%, n = 138; pLIF (–, +): 82.1%, n = 140; pLIF (+, +): 86.6%, n = 127], when compared with control [pLIF (–, –): 69.6%, n = 112], in which a significant increase of M-II rate (P < 0.05 by ANOVA) and cumulus expansion were observed in the pLIF (+, +) group. The effect of pLIF was only seen for COC but not for denuded oocytes. When oocytes were subjected to IVF (Kikuchi et al. 2002), those matured in pLIF (+, +)-supplemented medium demonstrated higher blastocyst developmental rates (21.1% v. 16.2%; P = 0.07) with increased cell numbers (50.2 cells v. 45.0 cells; P = 0.12) compared with pLIF (–, –) on Day 6 of embryo culture (IVF = 0). Examination of transcripts and proteins of the LIF signalling pathway revealed that mRNA and protein levels of LIF, LIF receptors, and signal transducer and activator of transcription 3 (STAT3) were similar in both pLIF (–, –) and pLIF (+, +) samples. However, notable phosphorylation of STAT3 was observed in the pLIF (+, +) sample. These results suggest that the LIF/STAT3-pathway is functional during oocyte maturation in pigs. Therefore, supplementation of maturation medium with pLIF could improve the developmental competence of oocytes by activation of this pathway. This project was supported by JSPS and HAS under the Japan-Hungary Research Cooperative Program.

scholarly journals Differences in Gene Expression between Wild Type and Hoxa1 Knockout Embryonic Stem Cells after Retinoic Acid Treatment or Leukemia Inhibitory Factor (LIF) Removal

2005 ◽  
Vol 280 (16) ◽  
pp. 16484-16498 ◽  
Author(s):  
Eduardo Martinez-Ceballos ◽  
Pierre Chambon ◽  
Lorraine J. Gudas
Keyword(s):  
Gene Expression ◽  
Cell Proliferation ◽  
Retinoic Acid ◽  
Leukemia Inhibitory Factor ◽  
Target Genes ◽  
Hox Genes ◽  
Es Cells ◽  
Embryonic Stem ◽  
Inhibitory Factor ◽  
Hoxa Cluster

Homeobox (Hox) genes encode a family of transcription factors that regulate embryonic patterning and organogenesis. In embryos, alterations of the normal pattern of Hox gene expression result in homeotic transformations and malformations. Disruption of theHoxa1gene, the most 3′ member of the Hoxa cluster and a retinoic acid (RA) direct target gene, results in abnormal ossification of the skull, hindbrain, and inner ear deficiencies, and neonatal death. We have generated Hoxa1-/-embryonic stem (ES) cells (named Hoxa1-15) from Hoxa1-/-mutant blastocysts to study the Hoxa1 signaling pathway. We have characterized in detail these Hoxa1-/-ES cells by performing microarray analyses, and by this technique we have identified a number of putative Hoxa-1 target genes, including genes involved in bone development (e.g. Col1a1,Postn/Osf2, and the bone sialoprotein gene orBSP), genes that are expressed in the developing brain (e.g. Nnat,Wnt3a,BDNF,RhoB, andGbx2), and genes involved in various cellular processes (e.g. M-RAS,Sox17,Cdkn2b,LamA1,Col4a1,Foxa2,Foxq1,Klf5, andIgf2). Cell proliferation assays and Northern blot analyses of a number of ES cell markers (e.g. Rex1,Oct3/4,Fgf4, andBmp4) suggest that the Hoxa1 protein plays a role in the inhibition of cell proliferation by RA in ES cells. Additionally, Hoxa1-/-ES cells express high levels of various endodermal markers, includingGata4andDab2, and express much lessFgf5after leukemia inhibitory factor (LIF) withdrawal. Finally, we propose a model in which the Hoxa1 protein mediates repression of endodermal differentiation while promoting expression of ectodermal and mesodermal characteristics.

scholarly journals Maturation of human oocytes in vitro and their developmental competence

Reproduction ◽  
2001 ◽  
pp. 51-75 ◽  
Author(s):  
A Trounson ◽  
C Anderiesz ◽  
G Jones
Keyword(s):  
Embryo Development ◽  
Oocyte Maturation ◽  
Growth Phase ◽  
Developmental Competence ◽  
Minimum Size ◽  
Success Rates ◽  
Human Oocytes ◽  
Maturation In Vitro

Complete maturation of oocytes is essential for the developmental competence of embryos. Any interventions in the growth phase of the oocyte and the follicle in the ovary will affect oocyte maturation, fertilization and subsequent embryo development. Oocyte size is associated with maturation and embryo development in most species examined and this may indicate that a certain size is necessary to initiate the molecular cascade of normal nuclear and cytoplasmic maturation. The minimum size of follicle required for developmental competence in humans is 5-7 mm in diameter. Maturation in vitro can be accomplished in humans, but is associated with a loss of developmental competence unless the oocyte is near completion of its preovulatory growth phase. This loss of developmental competence is associated with the absence of specific proteins in oocytes cultured to metaphase II in vitro. The composition of culture medium used successfully for maturation of human oocytes is surprisingly similar to that originally developed for maturation of oocytes in follicle culture in vitro. The presence of follicle support cells in culture is necessary for the gonadotrophin-mediated response required to mature oocytes in vitro. Gonadotrophin concentration and the sequence of FSH and FSH-LH exposure may be important for human oocytes, particularly those not exposed to the gonadotrophin surge in vivo. More research is needed to describe the molecular and cellular events, the presence of checkpoints and the role of gene expression, translation and protein uptake on completing oocyte maturation in vitro and in vivo. In the meantime, there are very clear applications for maturing oocytes in human reproductive medicine and the success rates achieved in some of these special applications are clinically valuable.

Isolation of embryonic stem (ES) cells in media supplemented with recombinant leukemia inhibitory factor (LIF)

1990 ◽  
Vol 141 (2) ◽  
pp. 344-352 ◽  
Author(s):  
Shirley Pease ◽  
Paola Braghetta ◽  
David Gearing ◽  
Dianne Grail ◽  
R.Lindsay Williams
Keyword(s):  
Leukemia Inhibitory Factor ◽  
Es Cells ◽  
Embryonic Stem ◽  
Inhibitory Factor

Enhancement of In Vitro Murine Embryo Development by Recombinant Leukemia Inhibitory Factor

1994 ◽  
Vol 1 (3) ◽  
pp. 215-219 ◽  
Author(s):  
Michael H. Mitchell ◽  
Robert J. Swanson ◽  
Gary D. Hodgen ◽  
Sergio Oehninger
Keyword(s):  
Embryo Development ◽  
Leukemia Inhibitory Factor ◽  
Inhibitory Factor ◽  
Murine Embryo

Leukemia inhibitory factor enhances bovine oocyte maturation and early embryo development

2014 ◽  
Vol 81 (7) ◽  
pp. 608-618 ◽  
Author(s):  
Xianhong Mo ◽  
Guoquan Wu ◽  
Dianshuai Yuan ◽  
Baoyu Jia ◽  
Cong Liu ◽  
...  
Keyword(s):  
Embryo Development ◽  
Oocyte Maturation ◽  
Leukemia Inhibitory Factor ◽  
Early Embryo ◽  
Inhibitory Factor ◽  
Bovine Oocyte ◽  
Early Embryo Development

scholarly journals Lipids and oocyte developmental competence: the role of fatty acids and β-oxidation

Reproduction ◽  
2014 ◽  
Vol 148 (1) ◽  
pp. R15-R27 ◽  
Author(s):  
Kylie R Dunning ◽  
Darryl L Russell ◽  
Rebecca L Robker
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
Embryo Development ◽  
Oocyte Maturation ◽  
Unsaturated Fatty Acids ◽  
Cumulus Cells ◽  
Developmental Competence ◽  
Oocyte Developmental Competence

Metabolism and ATP levels within the oocyte and adjacent cumulus cells are associated with quality of oocyte and optimal development of a healthy embryo. Lipid metabolism provides a potent source of energy and its importance during oocyte maturation is being increasingly recognised. The triglyceride and fatty acid composition of ovarian follicular fluid has been characterised for many species and is influenced by nutritional status (i.e. dietary fat, fasting, obesity and season) as well as lactation in cows. Lipid in oocytes is a primarily triglyceride of specific fatty acids which differ by species, stored in distinct droplet organelles that re-localise during oocyte maturation. The presence of lipids, particularly saturated vs unsaturated fatty acids, in in vitro maturation systems affects oocyte lipid content as well as developmental competence. Triglycerides are metabolised by lipases that have been localised to cumulus cells as well as oocytes. Fatty acids generated by lipolysis are further metabolised by β-oxidation in mitochondria for the production of ATP. β-oxidation is induced in cumulus–oocyte complexes (COCs) by the LH surge, and pharmacological inhibition of β-oxidation impairs oocyte maturation and embryo development. Promoting β-oxidation with l-carnitine improves embryo development in many species. Thus, fatty acid metabolism in the mammalian COC is regulated by maternal physiological and in vitro environmental conditions; and is important for oocyte developmental competence.

scholarly journals 296 LEUKEMIA INHIBITORY FACTOR INFLUENCES SHEEP OOCYTE PARTHENOGENETIC DEVELOPMENT DURING THE TRANSITION FROM GERMINAL VESICLE TO EARLY PRONUCLEAR STAGE

2005 ◽  
Vol 17 (2) ◽  
pp. 298
Author(s):  
G. Ptak ◽  
F. Lopes ◽  
P. Loi
Keyword(s):  
Embryo Development ◽  
Leukemia Inhibitory Factor ◽  
Calf Serum ◽  
Blastocyst Stage ◽  
Inhibitory Factor ◽  
Control Group ◽  
Blastocyst Development ◽  
Human Follicular Fluid ◽  
Oocyte Competence

Leukemia inhibitory factor (LIF) is an indispensable cytokine for female fertility. The influence of LIF on embryo development and particularly implantation has been recently confirmed; however, the effect of this cytokine on the oocyte has not been studied. The presence of LIF in human follicular fluid implies its possible role in the acquisition of oocyte competence. Furthermore, the up-regulation of LIF by steroid hormones in sheep makes entirely feasible the hypothesis that ovulatory estradiol peak plays a role in the preparation of female gamete for fertilization. With this in mind, we studied the effect of LIF during in vitro development of sheep oocytes mimicking the physiological expression of LIF induced by the ovulatory peak of estradiol in mice. GV stage oocytes matured and chemically activated in the presence of LIF and anti-LIF antibody were cultured to the blastocyst stage in our standard media. To eliminate the effect of the putative presence of LIF in heat inactivated fetal calf serum used for oocyte maturation, aliquots of LIF were treated at 56°C for 30 min and added to the maturation medium. The proportion of embryos that reached the blastocyst stage in vitro was significantly higher (P < 0.001) for oocytes matured and activated with LIF (36/93; 39%) than for the group incubated with antibody against LIF (6/68; 9%). The significant effect of anti-LIF antibody (P < 0.001) was also observed when compared with blastocysts developed from the control group of oocytes matured without LIF addition (31/106; 29%). Although the beneficial influence of LIF treatment on embryo development demonstrated with those preliminary data was not confirmed statistically, due to low number of oocytes involved, the proportion of embryos reaching the blastocyst stage in vitro was about 10% higher for those incubated with LIF than for either those cultured without the cytokine or those, matured in the presence of heat-treated LIF (15/55; 27%); however, the rate of blastocyst development appeared very similar to that of the control group. This study revealed for the first time a role of LIF in determining oocyte competence. Further investigation to determine how LIF achieves its effects on the oocyte are ongoing in our laboratory. This work was supported by FIRB RBNE01HPMX, COFIN 2002074357, COFIN 2003073943 002, and British Council 2004.

scholarly journals CD9 Is Associated with Leukemia Inhibitory Factor-mediated Maintenance of Embryonic Stem Cells

2002 ◽  
Vol 13 (4) ◽  
pp. 1274-1281 ◽  
Author(s):  
Masahiro Oka ◽  
Kenichi Tagoku ◽  
Thomas L. Russell ◽  
Yuka Nakano ◽  
Takashi Hamazaki ◽  
...  
Keyword(s):  
Mrna Expression ◽  
Leukemia Inhibitory Factor ◽  
Transmembrane Protein ◽  
Es Cells ◽  
Embryonic Stem ◽  
Immunohistochemical Analysis ◽  
Inhibitory Factor ◽  
Cdna Subtraction ◽  
Differentiated Cells ◽  
Undifferentiated State

Mouse embryonic stem (ES) cells can proliferate indefinitely in an undifferentiated state in the presence of leukemia inhibitory factor (LIF), or differentiate into all three germ layers upon removal of this factor. To determine cellular factors associated with self-renewal of undifferentiated ES cells, we used polymerase chain reaction-assisted cDNA subtraction to screen genes that are expressed in undifferentiated ES cells and down-regulated after incubating these cells in a differentiation medium without LIF for 48 h. The mRNA expression of a tetraspanin transmembrane protein, CD9, was high in undifferentiated ES cells and decreased shortly after cell differentiation. An immunohistochemical analysis confirmed that plasma membrane-associated CD9 was expressed in undifferentiated ES cells but low in the differentiated cells. Addition of LIF to differentiating ES cells reinduced mRNA expression of CD9, and CD9 expression was accompanied with a reappearance of undifferentiated ES cells. Furthermore, activation of STAT3 induced the expression of CD9, indicating the LIF/STAT3 pathway is critical for maintaining CD9 expression. Finally, addition of anti-CD9 antibody blocked ES cell colony formation and reduced cell viability. These results indicate that CD9 may play a role in LIF-mediated maintenance of undifferentiated ES cells.

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