Functional Significance of Gap Junctional Coupling in Preimplantation Development1

Houghton, Franchesca D.; Barr, Kevin; Walter, Glenn A.; Gabriel, Heinz-Dieter; Grümmer, Ruth; Traub, Otto; Leese, Henry J.; Winterhager, Elke; Kidder, Gerald M.

doi:10.1095/biolreprod66.5.1403

Cited by 58 publications

(62 citation statements)

References 57 publications

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“…The three connexin proteins (Cx43, Cx45, and Cx31) expressed in ES cells were detected throughout early embryonic development at the morula stage, as well as the blastocyst stage, in mouse [35] and rat [36]. Furthermore, we found expression of these three connexins during differentiation of HM1-mouse ES cells within the embryoid body (data not shown).…”

Section: Discussionmentioning

confidence: 76%

Connexin Expression and Functional Analysis of Gap Junctional Communication in Mouse Embryonic Stem Cells

Wörsdörfer¹,

Maxeiner²,

Markopoulos³

et al. 2007

Stem Cells

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Gap junctional intercellular communication (GJIC) has been suggested to be necessary for cellular proliferation and differentiation. We wanted to investigate the function of GJIC in mouse embryonic stem (ES) cells using pharmacological inhibitors or a genetic approach to inhibit the expression of connexins, that is, the subunit proteins of gap junction channels. For this purpose, we have analyzed all known connexin genes in mouse ES cells but found only three of them, Cx31, Cx43, and Cx45, to be expressed as proteins. We have demonstrated by coimmunoprecipitation that Cx31 and Cx43, as well as Cx43 and Cx45, probably form heteromeric gap junction channels, whereas Cx31 and Cx45 do not. The pharmacological inhibitors reduced GJIC between ES cells to approximately 3% and initiated apoptosis, suggesting an antiapoptotic effect of GJIC. In contrast to these results, reduction of GJIC to approximately 5% by decreased expression of Cx31 or Cx45 via RNA interference in homozygous Cx43-deficient ES cells did not lead to apoptosis. Additional studies suggested that apoptotic death of ES cells and adult stem cells reported in the literature is likely due to a cytotoxic side effect of the inhibitors and not due to a decrease of GJIC. Using the connexin expression pattern in mouse ES cells, as determined in this study, multiple connexin-deficient ES cells can now be genetically engineered in which the level of GJIC is further decreased, to clarify whether the differentiation of ES cells is qualitatively or quantitatively compromised. STEM CELLS 2008;26: 431-439 Disclosure of potential conflicts of interest is found at the end of this article.

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Section: Discussionmentioning

confidence: 76%

Connexin Expression and Functional Analysis of Gap Junctional Communication in Mouse Embryonic Stem Cells

Wörsdörfer¹,

Maxeiner²,

Markopoulos³

et al. 2007

Stem Cells

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“…In our study, levels of CDH1 did not differ between blastocysts and parthenotes, so the CDH1 contribution to compaction and blastulation is not altered in parthenotes. However, GJA1 levels are diminished in parthenotes, although coexpressed connexins may compensate for one another (Houghton et al 2002).…”

Section: Development and Gene Expression In Parthenotesmentioning

confidence: 99%

Biological differences between in vitro produced bovine embryos and parthenotes

Gómez¹,

Gutiérrez‐Adán²,

Díez³

et al. 2009

Reproduction

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Parthenotes may represent an alternate ethical source of stem cells, once biological differences between parthenotes and embryos can be understood. In this study, we analyzed development, trophectoderm (TE) differentiation, apoptosis/necrosis, and ploidy in parthenotes and in vitro produced bovine embryos. Subsequently, using real-time PCR, we analyzed the expression of genes expected to underlie the observed differences at the blastocyst stage. In vitro matured oocytes were either fertilized or activated with ionomycin C6-DMAP and cultured in simple medium. Parthenotes showed enhanced blastocyst development and diploidy and reduced TE cell counts. Apoptotic and necrotic indexes did not vary, but parthenotes evidenced a higher relative proportion of apoptotic cells between inner cell mass and TE. The pluripotence-related POU5F1 and the methylation DNMT3A genes were downregulated in parthenotes. Among pregnancy recognition genes, TP-1 was upregulated in parthenotes, while PGRMC1 and PLAC8 did not change. Expression of p66 shc and BAX/BCL2 ratio were higher, and p53 lower, in parthenotes. Among metabolism genes, SLC2A1 was downregulated, while AKR1B1, PTGS2, H6PD, and TXN were upregulated in parthenotes, and SLC2A5 did not differ. Among genes involved in compaction/blastulation, GJA1 was downregulated in parthenotes, but no differences were detected within ATP1A1 and CDH1. Within parthenotes, the expression levels of SLC2A1, TP-1, and H6PD, and possibly AKR1B1, resemble patterns described in female embryos. The pro-apoptotic profile is more pronounced in parthenotes than in embryos, which may differ in their way to channel apoptotic stimuli, through p66 shc and p53respectively, and in their mechanisms to control pluripotency and de novo methylation.Reproduction (2009) 137 285-295

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“…In our study, the levels of E-CAD-HERIN did not differ between the IVF blastocysts and parthenotes, suggesting that the E-CADHERIN contribution to the compaction process and blastocyst formation is not altered in parthenotes. However, the CX43 levels are diminished in parthenotes; however, even if the level of CX43 is altered, its function could be compensated by coexpressed connexins [52].The composition and accumulation of fluid in the blastocoele is regulated by Na/K-ATPase. Embryonic Na/K-ATPase activity increases when a nascent blastocoele cavity forms and then decreases to the baseline level at full expansion [53].…”

mentioning

confidence: 99%

Gene Expression in Early Expanded Parthenogenetic and In Vitro Fertilized Bovine Blastocysts

Gómez

Caamaño

Bermejo-Álvarez³

et al. 2009

Journal of Reproduction and Development

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Abstract. Mammalian oocytes can undergo artificial parthenogenesis in vitro and develop to the blastocyst stage. In this study, using real-time PCR, we analyzed the expression of genes representative of essential events in development. In vitro matured oocytes were either fertilized or activated with ionomycin + 6-DMAP and cultured in simple medium. The pluripotency-related gene Oct3/4 was downregulated in parthenotes, while the de novo methylation DNMT3A gene was unchanged. Among the pregnancy recognition genes, IFN-t was upregulated, PGRMC1 was downregulated and PLAC8 was unchanged in parthenotes. Among the metabolism genes, SLC2A1 was downregulated, while AKR1B1, COX2, H6PD and TXN were upregulated in parthenotes; there was no difference in SLC2A5. Among the genes involved in compaction/blastulation, GJA1 expression increased in parthenotes, but no differences were detected within ATP1A1 and CDH1. Expression of p66 shc and the Bax/Bcl2 ratio were higher in parthenotes, and there was no difference in p53. Parthenotes and embryos may differ in the way they stimulate apoptosis, with a preponderant role for p66 shc within parthenotes. Differentially affected functions may also include pluripotency, de novo methylation and early embryonic signalling. Key words: Bovine, Embryo, Gene expression, Parthenote (J. Reprod. Dev. 55: [607][608][609][610][611][612][613][614] 2009) arthenogenesis, a form of reproduction not spontaneous in mammals, is a process by which the oocyte develops without the male gamete. Mammalian parthenotes are unable to develop to term in utero (developing for a maximum of 48 days in the cow; [1]), and this is thought to be due to alterations in their genomic imprinting, a mechanism that provides complementary, but not equivalent, specific expression between maternal and paternal genomes [2]. During fertilization, the spermatozoon triggers multiple and rhythmic oscillations of intracellular free calcium that release the oocyte from this blocked stage [3]. These intracellular calcium patterns can be mimicked by a number of chemicals that activate the oocyte without sperm, yielding parthenogenetic embryos [4]. In cattle, production of reconstructed embryos by somatic cell nuclear transfer (SCNT) [5] and intracytoplasmic sperm injection (ICSI) [6] depends on successful oocyte activation. Therefore, procedures involving SCNT and ICSI could benefit from a better knowledge of mechanisms that differ between embryos and parthenotes.Contrary to the research in mice and humans, most studies have failed to detect putatively imprinted genes in bovine early development [7,8], suggesting that monoallelic expression may not develop until after Day 21 of development, as occurs in ovine species [9]. In fact, a recent study found imprinted expression of bovine Mest-1 in Day-21 but not Day-14 embryos, while 7 other putatively imprinted genes were shown to be not imprinted at the two stages analyzed [10]. However, gene expression does differ between blastocysts from embryos and parthenotes in domestic species [11...

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Functional Significance of Gap Junctional Coupling in Preimplantation Development1

Cited by 58 publications

References 57 publications

Connexin Expression and Functional Analysis of Gap Junctional Communication in Mouse Embryonic Stem Cells

Connexin Expression and Functional Analysis of Gap Junctional Communication in Mouse Embryonic Stem Cells

Biological differences between in vitro produced bovine embryos and parthenotes

Gene Expression in Early Expanded Parthenogenetic and In Vitro Fertilized Bovine Blastocysts

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