Biology and Biotechnology of Reproduction

During preimplantation development, the first cell lineages differentiate while a proportion of cells retains pluripotency. Genes and mechanisms that regulate these essential events have been studied extensively in the mouse model, where reverse genetics is most advanced. In mouse embryos, loss of OCT4 causes failure of primitive endoderm development (shown by the lack of cells expressing GATA6), while cells expressing the pluripotency marker gene NANOG are not affected. In contrast, bovine OCT4 deficient embryos retain cells that express GATA6 but lose NANOG entirely (Simmet et al., PNAS 2018). A recent study on the function of OCT4 in early human development revealed the same mechanism as in bovine, suggesting bovine embryos as an interesting model for studying human development.

Biology of reproduction in livestock species is an interesting research topic since many aspects resemble human reproductive biology more closely than rodent models do. Moreover, reproductive biotechnologies are the basis for genetic modifications to perform reverse genetics studies or generate large animal models.
Maternal-to-embryonic transition is the period when maternal RNAs and proteins stored in the oocyte are gradually degraded and transcription of the embryonic genome is activated. The onset of embryonic transcription is obscured by the presence of maternal transcripts and could only be determined for genes which are not expressed in oocytes. Using RNA sequencing (collaboration with Helmut Blum, LAFUGA) of bovine germinal vesicle and metaphase II oocytes, and of four-cell, eight-cell, 16-cell and blastocyst stage embryos, we established a comprehensive transcriptome data set of bovine oocyte maturation and early development. Embryonic genome activation was analyzed by i) detection of embryonic transcripts which are not present in oocytes; ii) detection of transcripts from the paternal allele; and iii) detection of primary transcripts with intronic sequences. Using these approaches, we were able to map the onset of embryonic transcription for almost 7,400 genes.
Current studies address EGA in cloned embryos as well as developmental heterogeneity and early lineage specification events in bovine embryos at the time of major EGA. In addition, gene editing (CRISPR/Cas9) is used to dissect the roles of pluripotency networks in early bovine development.

Highlights

• OCT4/POU5F1 is necessary for NANOG expression in bovine blastocysts (Simmet et al., PNAS 2018)

• Detailed mapping of genome activation in bovine embryos (Graf et al., PNAS 2014)

• The endometrium responds differently to cloned vs. fertilized embryos (Bauersachs et al., PNAS 2009)

Publications

2022

Simmet K, Kurome M, Zakhartchenko V, Reichenbach HD, Springer C, Bähr A, Blum H, Philippou-Massier J, Wolf E. OCT4/POU5F1 is indispensable for the lineage differentiation of the inner cell mass in bovine embryos. FASEB J. 2022 Jun;36(6):e22337. PMID: 35486003

2021

Alberio R, Wolf E. 25th ANNIVERSARY OF CLONING BY SOMATIC-CELL NUCLEAR TRANSFER: Nuclear transfer and the development of genetically modified/gene edited livestock. Reproduction. 2021 Jun 11;162(1):F59-F68. PMID: 34096507

Bebbere D, Ulbrich SE, Giller K, Zakhartchenko V, Reichenbach HD, Reichenbach M, Verma PJ, Wolf E, Ledda S, Hiendleder S. Mitochondrial DNA Depletion in Granulosa Cell Derived Nuclear Transfer Tissues. Front Cell Dev Biol. 2021 May 14;9:664099. PMID: 34124044

Simmet K, Wolf E, Zakhartchenko V. Manipulating the Epigenome in Nuclear Transfer Cloning: Where, When and How. Int J Mol Sci 2020; 22:236.

Springer C, Zakhartchenko V, Wolf E, Simmet K. Cells. Hypoblast Formation in Bovine Embryos Does Not Depend on NANOG. 2021 Aug 28;10(9):2232. PMID: 34571882

Springer C, Wolf E, Simmet K. A New Toolbox in Experimental Embryology-Alternative Model Organisms for Studying Preimplantation Development. J Dev Biol. 2021 Apr 2;9(2):15. PMID: 33918361

van der Weijden VA, Schmidhauser M, Kurome M, Knubben J, Flöter VL, Wolf E, Ulbrich SE. Transcriptome dynamics in early in vivo developing and in vitro produced porcine embryos. BMC Genomics. 2021 Feb 27;22(1):139. doi: 10.1186/s12864-021-07430-7. PMID: 33639836

2020

Gegenfurtner K, Froehlich T, Flenkenthaler F, Kosters M, Fritz S, Desnoes O, Le Bourhis D, Salvetti P, Sandra O, Charpigny G, Mermillod P, Lonergan P, Wolf E, Arnold GJ. Genetic merit for fertility alters the bovine uterine luminal fluid proteome. Biology of reproduction 2020; 102:730-739

Gegenfurtner K, Flenkenthaler F, Fröhlich T, Wolf E, Arnold GJ. The impact of transcription inhibition during in vitro maturation on the proteome of bovine oocytes. Biol Reprod 2020a; 103:1000-1011

2019

Duan JE, Shi W, Jue NK, Jiang Z, Kuo L, O'Neill R, Wolf E, Dong H, Zheng X, Chen J, Tian XC. Dosage Compensation of the X Chromosomes in Bovine Germline, Early Embryos, and Somatic Tissues. Genome biology and evolution 2019; 11:242-252

Gegenfurtner K, Froehlich T, Koesters M, Mermillod P, Locatelli Y, Fritz S, Salvetti P, Forde N, Lonergan P, Wolf E, Arnold GJ. Influence of metabolic status and genetic merit for fertility on proteomic composition of bovine oviduct fluid. Biology of reproduction 2019; 101:893-905

Kurome M, Baehr A, Simmet K, Jemiller EM, Egerer S, Dahlhoff M, Zakhartchenko V, Nagashima H, Klymiuk N, Kessler B, Wolf E. Targeting alphaGal epitopes for multi-species embryo immunosurgery. Reproduction, fertility, and development 2019; 31:820-826

Locatelli Y, Forde N, Blum H, Graf A, Piegu B, Mermillod P, Wolf E, Lonergan P, Saint-Dizier M. Relative effects of location relative to the corpus luteum and lactation on the transcriptome of the bovine oviduct epithelium. BMC genomics 2019; 20:233

2018

Lavagi I, Krebs S, Simmet K, Beck A, Zakhartchenko V, Wolf E, Blum H. Single-cell RNA sequencing reveals developmental heterogeneity of blastomeres during major genome activation in bovine embryos. Scientific reports 2018; 8:4071

Simmet K, Zakhartchenko V, Philippou-Massier J, Blum H, Klymiuk N, Wolf E. OCT4/POU5F1 is required for NANOG expression in bovine blastocysts. Proceedings of the National Academy of Sciences of the United States of America 2018a; 115:2770-2775

Simmet K, Zakhartchenko V, Wolf E. Comparative aspects of early lineage specification events in mammalian embryos - insights from reverse genetics studies. Cell cycle (Georgetown, Tex) 2018b; 17:1688-1695

2017

Bauersachs S, Simintiras CA, Sturmey RG, Krebs S, Bick J, Blum H, Wolf E, Lonergan P, Forde N. Effect of metabolic status on conceptus-maternal interactions on day 19 in dairy cattle: II. Effects on the endometrial transcriptome. Biology of reproduction 2017; 97:413-425

Forde N, Simintiras CA, Sturmey RG, Graf A, Wolf E, Blum H, Lonergan P. Effect of lactation on conceptus-maternal interactions at the initiation of implantation in cattle: I. Effects on the conceptus transcriptome and amino acid composition of the uterine luminal fluid. Biology of reproduction 2017; 97:798-809

Kurome M, Leuchs S, Kessler B, Kemter E, Jemiller EM, Foerster B, Klymiuk N, Zakhartchenko V, Wolf E. Direct introduction of gene constructs into the pronucleus-like structure of cloned embryos: a new strategy for the generation of genetically modified pigs. Transgenic research 2017; 26:309-318

2016

Popken J, Schmid VJ, Strauss A, Guengoer T, Wolf E, Zakhartchenko V. Stage-dependent remodeling of the nuclear envelope and lamina during rabbit early embryonic development. The Journal of reproduction and development 2016; 62:127-135

2015

Bauersachs S, Wolf E. Uterine responses to the preattachment embryo in domestic ungulates: recognition of pregnancy and preparation for implantation. Annual review of animal biosciences 2015; 3:489-511

Demant M, Deutsch DR, Frohlich T, Wolf E, Arnold GJ. Proteome analysis of early lineage specification in bovine embryos. Proteomics 2015; 15:688-701

Flisikowski K, Boulhaeve M, Lopes FP, Wolf E, Zwierzchowski L. Expression pattern of STAT5A gene during early bovine embryogenesis. Zygote 2015; 23:307-311

Kurome M, Kessler B, Wuensch A, Nagashima H, Wolf E. Nuclear transfer and transgenesis in the pig. Methods in molecular biology 2015; 1222:37-59

Popken J, Dahlhoff M, Guengoer T, Wolf E, Zakhartchenko V. 3D structured illumination microscopy of mammalian embryos and spermatozoa. BMC developmental biology 2015a; 15:46

Popken J, Graf A, Krebs S, Blum H, Schmid VJ, Strauss A, Guengoer T, Zakhartchenko V, Wolf E, Cremer T. Remodeling of the nuclear envelope and lamina during bovine preimplantation development and its functional implications. PloS one 2015b; 10:e0124619

Simmet K, Reichenbach M, Reichenbach HD, Wolf E. Phytohemagglutinin facilitates the aggregation of blastomere pairs from Day 5 donor embryos with Day 4 host embryos for chimeric bovine embryo multiplication. Theriogenology 2015; 84:1603-1610

2014

Deutsch DR, Frohlich T, Otte KA, Beck A, Habermann FA, Wolf E, Arnold GJ. Stage-specific proteome signatures in early bovine embryo development. Journal of proteome research 2014; 13:4363-4376

Graf A, Krebs S, Heininen-Brown M, Zakhartchenko V, Blum H, Wolf E. Genome activation in bovine embryos: review of the literature and new insights from RNA sequencing experiments. Animal reproduction science 2014a; 149:46-58

Graf A, Krebs S, Zakhartchenko V, Schwalb B, Blum H, Wolf E. Fine mapping of genome activation in bovine embryos by RNA sequencing. Proceedings of the National Academy of Sciences of the United States of America 2014b; 111:4139-4144

Klymiuk N, Fezert P, Wunsch A, Kurome M, Kessler B, Wolf E. Homologous recombination contributes to the repair of zinc-finger-nuclease induced double strand breaks in pig primary cells and facilitates recombination with exogenous DNA. Journal of biotechnology 2014; 177:74-81

Li S, Flisikowska T, Kurome M, Zakhartchenko V, Kessler B, Saur D, Kind A, Wolf E, Flisikowski K, Schnieke A. Dual fluorescent reporter pig for Cre recombination: transgene placement at the ROSA26 locus. PloS one 2014; 9:e102455

Popken J, Brero A, Koehler D, Schmid VJ, Strauss A, Wuensch A, Guengoer T, Graf A, Krebs S, Blum H, Zakhartchenko V, Wolf E, Cremer T. Reprogramming of fibroblast nuclei in cloned bovine embryos involves major structural remodeling with both striking similarities and differences to nuclear phenotypes of in vitro fertilized embryos. Nucleus 2014a; 5:555-589

Popken J, Koehler D, Brero A, Wuensch A, Guengoer T, Thormeyer T, Wolf E, Cremer T, Zakhartchenko V. Positional changes of a pluripotency marker gene during structural reorganization of fibroblast nuclei in cloned early bovine embryos. Nucleus 2014b; 5:542-554

Samans B, Yang Y, Krebs S, Sarode GV, Blum H, Reichenbach M, Wolf E, Steger K, Dansranjavin T, Schagdarsurengin U. Uniformity of nucleosome preservation pattern in mammalian sperm and its connection to repetitive DNA elements. Developmental cell 2014; 30:23-35

2013

Bauersachs S, Wolf E. Immune aspects of embryo-maternal cross-talk in the bovine uterus. Journal of reproductive immunology 2013; 97:20-26

Bebbere D, Bauersachs S, Furst RW, Reichenbach HD, Reichenbach M, Medugorac I, Ulbrich SE, Wolf E, Ledda S, Hiendleder S. Tissue-specific and minor inter-individual variation in imprinting of IGF2R is a common feature of Bos taurus Concepti and not correlated with fetal weight. PloS one 2013; 8:e59564

Kurome M, Geistlinger L, Kessler B, Zakhartchenko V, Klymiuk N, Wuensch A, Richter A, Baehr A, Kraehe K, Burkhardt K, Flisikowski K, Flisikowska T, Merkl C, Landmann M, Durkovic M, Tschukes A, Kraner S, Schindelhauer D, Petri T, Kind A, Nagashima H, Schnieke A, Zimmer R, Wolf E. Factors influencing the efficiency of generating genetically engineered pigs by nuclear transfer: multi-factorial analysis of a large data set. BMC biotechnology 2013; 13:43

Samborski A, Graf A, Krebs S, Kessler B, Reichenbach M, Reichenbach HD, Ulbrich SE, Bauersachs S. Transcriptome changes in the porcine endometrium during the preattachment phase. Biol Reprod. 2013 Dec 12;89(6):134. doi: 10.1095/biolreprod.113.112177. Print 2013 Dec. PubMed PMID: 24174570.


2012

Bauersachs S, Ulbrich SE, Reichenbach HD, Reichenbach M, Buettner M, Meyer HH, Spencer TE, Minten M, Sax G, Winter G, Wolf E. Comparison of the effects of early pregnancy with human interferon, alpha 2 (IFNA2), on gene expression in bovine endometrium. Biology of reproduction 2012; 86:46

Bauersachs S, Wolf E. Transcriptome analyses of bovine, porcine and equine endometrium during the pre-implantation phase. Animal reproduction science 2012; 134:84-94

Flisikowski K, Venhoranta H, Bauersachs S, Haenninen R, Fuerst RW, Saalfrank A, Ulbrich SE, Taponen J, Lohi H, Wolf E, Kind A, Andersson M, Schnieke A. Truncation of MIMT1 gene in the PEG3 domain leads to major changes in placental gene expression and stillbirth in cattle. Biology of reproduction 2012; 87:140

Richter A, Kurome M, Kessler B, Zakhartchenko V, Klymiuk N, Nagashima H, Wolf E, Wuensch A. Potential of primary kidney cells for somatic cell nuclear transfer mediated transgenesis in pig. BMC biotechnology 2012; 12:84

Streyl D, Kenngott R, Herbach N, Wanke R, Blum H, Sinowatz F, Wolf E, Zerbe H, Bauersachs S. Gene expression profiling of bovine peripartal placentomes: detection of molecular pathways potentially involved in the release of foetal membranes. Reproduction 2012; 143:85-105

Ulbrich SE, Wolf E, Bauersachs S. Hosting the preimplantation embryo: potentials and limitations of different approaches for analysing embryo-endometrium interactions in cattle. Reproduction, fertility, and development 2012; 25:62-70

2011

Groebner AE, Rubio-Aliaga I, Schulke K, Reichenbach HD, Daniel H, Wolf E, Meyer HH, Ulbrich SE. Increase of essential amino acids in the bovine uterine lumen during preimplantation development. Reproduction 2011a; 141:685-695

Groebner AE, Schulke K, Schefold JC, Fusch G, Sinowatz F, Reichenbach HD, Wolf E, Meyer HH, Ulbrich SE. Immunological mechanisms to establish embryo tolerance in early bovine pregnancy. Reproduction, fertility, and development 2011b; 23:619-632

Groebner AE, Zakhartchenko V, Bauersachs S, Rubio-Aliaga I, Daniel H, Buettner M, Reichenbach HD, Meyer HH, Wolf E, Ulbrich SE. Reduced amino acids in the bovine uterine lumen of cloned versus in vitro fertilized pregnancies prior to implantation. Cellular reprogramming 2011c; 13:403-410

Lagutina I, Zakhartchenko V, Fulka H, Colleoni S, Wolf E, Fulka J, Jr., Lazzari G, Galli C. Formation of nucleoli in interspecies nuclear transfer embryos derived from bovine, porcine, and rabbit oocytes and nuclear donor cells of various species. Reproduction 2011; 141:453-465

Leidenfrost S, Boelhauve M, Reichenbach M, Guengoer T, Reichenbach HD, Sinowatz F, Wolf E, Habermann FA. Cell arrest and cell death in mammalian preimplantation development: lessons from the bovine model. PloS one 2011; 6:e22121

Pimentel EC, Bauersachs S, Tietze M, Simianer H, Tetens J, Thaller G, Reinhardt F, Wolf E, Koenig S. Exploration of relationships between production and fertility traits in dairy cattle via association studies of SNPs within candidate genes derived by expression profiling. Animal genetics 2011; 42:251-262

Ulbrich SE, Meyer SU, Zitta K, Hiendleder S, Sinowatz F, Bauersachs S, Buettner M, Froehlich T, Arnold GJ, Reichenbach HD, Wolf E, Meyer HH. Bovine endometrial metallopeptidases MMP14 and MMP2 and the metallopeptidase inhibitor TIMP2 participate in maternal preparation of pregnancy. Molecular and cellular endocrinology 2011; 332:48-57

Zakhartchenko V, Flisikowska T, Li S, Richter T, Wieland H, Durkovic M, Rottmann O, Kessler B, Guengoer T, Brem G, Kind A, Wolf E, Schnieke A. Cell-mediated transgenesis in rabbits: chimeric and nuclear transfer animals. Biology of reproduction 2011; 84:229-237

2010

Bauersachs S, Blum H, Krebs S, Frohlich T, Arnold GJ, Wolf E. Creating new knowledge for ruminant reproduction from rapidly expanding and evolving scientific databases. Society of Reproduction and Fertility supplement 2010; 67:29-40

de Montera B, El Zeihery D, Mueller S, Jammes H, Brem G, Reichenbach HD, Scheipl F, Chavatte-Palmer P, Zakhartchenko V, Schmitz OJ, Wolf E, Renard JP, Hiendleder S. Quantification of leukocyte genomic 5-methylcytosine levels reveals epigenetic plasticity in healthy adult cloned cattle. Cellular reprogramming 2010; 12:175-181

Groebner AE, Schulke K, Unterseer S, Reichenbach HD, Reichenbach M, Buettner M, Wolf E, Meyer HH, Ulbrich SE. Enhanced proapoptotic gene expression of XAF1, CASP8 and TNFSF10 in the bovine endometrium during early pregnancy is not correlated with augmented apoptosis. Placenta 2010; 31:168-177

Koehler D, Zakhartchenko V, Ketterl N, Wolf E, Cremer T, Brero A. FISH on 3D preserved bovine and murine preimplantation embryos. Methods in molecular biology 2010; 659:437-445

Merkl M, Ulbrich SE, Otzdorff C, Herbach N, Wanke R, Wolf E, Handler J, Bauersachs S. Microarray analysis of equine endometrium at days 8 and 12 of pregnancy. Biology of reproduction 2010; 83:874-886

Mitterhuemer S, Petzl W, Krebs S, Mehne D, Klanner A, Wolf E, Zerbe H, Blum H. Escherichia coli infection induces distinct local and systemic transcriptome responses in the mammary gland. BMC genomics 2010; 11:138

Ostrup E, Bauersachs S, Blum H, Wolf E, Hyttel P. Differential endometrial gene expression in pregnant and nonpregnant sows. Biology of reproduction 2010; 83:277-285

Reichenbach M, Lim T, Reichenbach HD, Guengoer T, Habermann FA, Matthiesen M, Hofmann A, Weber F, Zerbe H, Grupp T, Sinowatz F, Pfeifer A, Wolf E. Germ-line transmission of lentiviral PGK-EGFP integrants in transgenic cattle: new perspectives for experimental embryology. Transgenic research 2010; 19:549-556

Shimizu T, Krebs S, Bauersachs S, Blum H, Wolf E, Miyamoto A. Actions and interactions of progesterone and estrogen on transcriptome profiles of the bovine endometrium. Physiological genomics 2010; 42A:290-300

Ulbrich SE, Zitta K, Hiendleder S, Wolf E. In vitro systems for intercepting early embryo-maternal cross-talk in the bovine oviduct. Theriogenology 2010; 73:802-816

2009

Bauersachs S, Ulbrich SE, Zakhartchenko V, Minten M, Reichenbach M, Reichenbach HD, Blum H, Spencer TE, Wolf E. The endometrium responds differently to cloned versus fertilized embryos. Proceedings of the National Academy of Sciences of the United States of America 2009; 106:5681-5686

Berendt FJ, Froehlich T, Bolbrinker P, Boelhauve M, Guengoer T, Habermann FA, Wolf E, Arnold GJ. Highly sensitive saturation labeling reveals changes in abundance of cell cycle-associated proteins and redox enzyme variants during oocyte maturation in vitro. Proteomics 2009; 9:550-564

Brero A, Hao R, Schieker M, Wierer M, Wolf E, Cremer T, Zakhartchenko V. Reprogramming of active and repressive histone modifications following nuclear transfer with rabbit mesenchymal stem cells and adult fibroblasts. Cloning and stem cells 2009; 11:319-329

Koehler D, Zakhartchenko V, Froenicke L, Stone G, Stanyon R, Wolf E, Cremer T, Brero A. Changes of higher order chromatin arrangements during major genome activation in bovine preimplantation embryos. Experimental cell research 2009; 315:2053-2063

Ulbrich SE, Frohlich T, Schulke K, Englberger E, Waldschmitt N, Arnold GJ, Reichenbach HD, Reichenbach M, Wolf E, Meyer HH, Bauersachs S. Evidence for estrogen-dependent uterine serpin (SERPINA14) expression during estrus in the bovine endometrial glandular epithelium and lumen. Biology of reproduction 2009; 81:795-805

2008

Bauersachs S, Mitko K, Ulbrich SE, Blum H, Wolf E. Transcriptome studies of bovine endometrium reveal molecular profiles characteristic for specific stages of estrous cycle and early pregnancy. Experimental and clinical endocrinology & diabetes : official journal, German Society of Endocrinology [and] German Diabetes Association 2008; 116:371-384

Lepikhov K, Zakhartchenko V, Hao R, Yang F, Wrenzycki C, Niemann H, Wolf E, Walter J. Evidence for conserved DNA and histone H3 methylation reprogramming in mouse, bovine and rabbit zygotes. Epigenetics & chromatin 2008; 1:8

Mitko K, Ulbrich SE, Wenigerkind H, Sinowatz F, Blum H, Wolf E, Bauersachs S. Dynamic changes in messenger RNA profiles of bovine endometrium during the oestrous cycle. Reproduction 2008; 135:225-240

Spencer TE, Sandra O, Wolf E. Genes involved in conceptus-endometrial interactions in ruminants: insights from reductionism and thoughts on holistic approaches. Reproduction 2008; 135:165-179

2007

Bauersachs S, Mitko K, Blum H, Wolf E. Technical note: Bovine oviduct and endometrium array version 1: a tailored tool for studying bovine endometrium biology and pathophysiology. Journal of dairy science 2007; 90:4420-4423

Habermann FA, Wuensch A, Sinowatz F, Wolf E. Reporter genes for embryogenesis research in livestock species. Theriogenology 2007; 68 Suppl 1:S116-124

Paula-Lopes FF, Boelhauve M, Habermann FA, Sinowatz F, Wolf E. Leptin promotes meiotic progression and developmental capacity of bovine oocytes via cumulus cell-independent and -dependent mechanisms. Biology of reproduction 2007; 76:532-541

Wuensch A, Habermann FA, Kurosaka S, Klose R, Zakhartchenko V, Reichenbach HD, Sinowatz F, McLaughlin KJ, Wolf E. Quantitative monitoring of pluripotency gene activation after somatic cloning in cattle. Biology of reproduction 2007; 76:983-991

Yang F, Hao R, Kessler B, Brem G, Wolf E, Zakhartchenko V. Rabbit somatic cell cloning: effects of donor cell type, histone acetylation status and chimeric embryo complementation. Reproduction 2007; 133:219-230

2006

Bauersachs S, Ulbrich SE, Gross K, Schmidt SE, Meyer HH, Wenigerkind H, Vermehren M, Sinowatz F, Blum H, Wolf E. Embryo-induced transcriptome changes in bovine endometrium reveal species-specific and common molecular markers of uterine receptivity. Reproduction 2006; 132:319-331

Ewerling S, Hofmann A, Klose R, Weppert M, Brem G, Rink K, Pfeifer A, Wolf E. Evaluation of laser-assisted lentiviral transgenesis in bovine. Transgenic research 2006; 15:447-454

Hiendleder S, Wirtz M, Mund C, Klempt M, Reichenbach HD, Stojkovic M, Weppert M, Wenigerkind H, Elmlinger M, Lyko F, Schmitz OJ, Wolf E. Tissue-specific effects of in vitro fertilization procedures on genomic cytosine methylation levels in overgrown and normal sized bovine fetuses. Biology of reproduction 2006; 75:17-23

Hofmann A, Kessler B, Ewerling S, Kabermann A, Brem G, Wolf E, Pfeifer A. Epigenetic regulation of lentiviral transgene vectors in a large animal model. Molecular therapy : the journal of the American Society of Gene Therapy 2006; 13:59-66

Klein C, Bauersachs S, Ulbrich SE, Einspanier R, Meyer HH, Schmidt SE, Reichenbach HD, Vermehren M, Sinowatz F, Blum H, Wolf E. Monozygotic twin model reveals novel embryo-induced transcriptome changes of bovine endometrium in the preattachment period. Biology of reproduction 2006; 74:253-264

Machado SA, Reichenbach HD, Weppert M, Wolf E, Goncalves PB. The variability of ovum pick-up response and in vitro embryo production from monozygotic twin cows. Theriogenology 2006; 65:573-583

Rottmayer R, Ulbrich SE, Koelle S, Prelle K, Neumueller C, Sinowatz F, Meyer HH, Wolf E, Hiendleder S. A bovine oviduct epithelial cell suspension culture system suitable for studying embryo-maternal interactions: morphological and functional characterization. Reproduction 2006; 132:637-648

Ulbrich SE, Rehfeld S, Bauersachs S, Wolf E, Rottmayer R, Hiendleder S, Vermehren M, Sinowatz F, Meyer HH, Einspanier R. Region-specific expression of nitric oxide synthases in the bovine oviduct during the oestrous cycle and in vitro. The Journal of endocrinology 2006; 188:205-213

Wolf E, Hiendleder S, Bauersachs S, Froehlich T, Sinowatz F, Blum H, Arnold GJ. [Methods for transcriptome and proteome research: applications for studying the biology of reproduction in cattle]. Berliner und Munchener tierarztliche Wochenschrift 2006; 119:7-16

2005

Bauersachs S, Ulbrich SE, Gross K, Schmidt SE, Meyer HH, Einspanier R, Wenigerkind H, Vermehren M, Blum H, Sinowatz F, Wolf E. Gene expression profiling of bovine endometrium during the oestrous cycle: detection of molecular pathways involved in functional changes. Journal of molecular endocrinology 2005; 34:889-908

Berendt FJ, Frohlich T, Schmidt SE, Reichenbach HD, Wolf E, Arnold GJ. Holistic differential analysis of embryo-induced alterations in the proteome of bovine endometrium in the preattachment period. Proteomics 2005; 5:2551-2560

Boelhauve M, Sinowatz F, Wolf E, Paula-Lopes FF. Maturation of bovine oocytes in the presence of leptin improves development and reduces apoptosis of in vitro-produced blastocysts. Biology of reproduction 2005; 73:737-744

Hiendleder S, Bauersachs S, Boulesteix A, Blum H, Arnold GJ, Froehlich T, Wolf E. Functional genomics: tools for improving farm animal health and welfare. Revue scientifique et technique 2005a; 24:355-377

Hiendleder S, Zakhartchenko V, Wolf E. Mitochondria and the success of somatic cell nuclear transfer cloning: from nuclear-mitochondrial interactions to mitochondrial complementation and mitochondrial DNA recombination. Reproduction, fertility, and development 2005b; 17:69-83

Before 2005

Alberio R, Brero A, Motlik J, Cremer T, Wolf E, Zakhartchenko V. Remodeling of donor nuclei, DNA-synthesis, and ploidy of bovine cumulus cell nuclear transfer embryos: effect of activation protocol. Molecular reproduction and development 2001a; 59:371-379

Alberio R, Kubelka M, Zakhartchenko V, Hajduch M, Wolf E, Motlik J. Activation of bovine oocytes by specific inhibition of cyclin-dependent kinases. Molecular reproduction and development 2000a; 55:422-432

Alberio R, Motlik J, Stojkovic M, Wolf E, Zakhartchenko V. Behavior of M-phase synchronized blastomeres after nuclear transfer in cattle. Molecular reproduction and development 2000b; 57:37-47

Alberio R, Zakhartchenko V, Motlik J, Wolf E. Mammalian oocyte activation: lessons from the sperm and implications for nuclear transfer. The International journal of developmental biology 2001b; 45:797-809

Bauersachs S, Blum H, Mallok S, Wenigerkind H, Rief S, Prelle K, Wolf E. Regulation of ipsilateral and contralateral bovine oviduct epithelial cell function in the postovulation period: a transcriptomics approach. Biology of reproduction 2003; 68:1170-1177

Bauersachs S, Rehfeld S, Ulbrich SE, Mallok S, Prelle K, Wenigerkind H, Einspanier R, Blum H, Wolf E. Monitoring gene expression changes in bovine oviduct epithelial cells during the oestrous cycle. Journal of molecular endocrinology 2004; 32:449-466

Bruggerhoff K, Zakhartchenko V, Wenigerkind H, Reichenbach HD, Prelle K, Schernthaner W, Alberio R, Kuchenhoff H, Stojkovic M, Brem G, Hiendleder S, Wolf E. Bovine somatic cell nuclear transfer using recipient oocytes recovered by ovum pick-up: effect of maternal lineage of oocyte donors. Biology of reproduction 2002; 66:367-373

Dean W, Santos F, Stojkovic M, Zakhartchenko V, Walter J, Wolf E, Reik W. Conservation of methylation reprogramming in mammalian development: aberrant reprogramming in cloned embryos. Proceedings of the National Academy of Sciences of the United States of America 2001; 98:13734-13738

Durcova-Hills G, Prelle K, Muller S, Stojkovic M, Motlik J, Wolf E, Brem G. Primary culture of porcine PGCs requires LIF and porcine membrane-bound stem cell factor. Zygote 1998; 6:271-275

Grosse-Hovest L, Muller S, Minoia R, Wolf E, Zakhartchenko V, Wenigerkind H, Lassnig C, Besenfelder U, Muller M, Lytton SD, Jung G, Brem G. Cloned transgenic farm animals produce a bispecific antibody for T cell-mediated tumor cell killing. Proceedings of the National Academy of Sciences of the United States of America 2004; 101:6858-6863

Hiendleder S, Bebbere D, Zakhartchenko V, Reichenbach HD, Wenigerkind H, Ledda S, Wolf E. Maternal-fetal transplacental leakage of mitochondrial DNA in bovine nuclear transfer pregnancies: potential implications for offspring and recipients. Cloning and stem cells 2004a; 6:150-156

Hiendleder S, Mund C, Reichenbach HD, Wenigerkind H, Brem G, Zakhartchenko V, Lyko F, Wolf E. Tissue-specific elevated genomic cytosine methylation levels are associated with an overgrowth phenotype of bovine fetuses derived by in vitro techniques. Biology of reproduction 2004b; 71:217-223

Hiendleder S, Prelle K, Bruggerhoff K, Reichenbach HD, Wenigerkind H, Bebbere D, Stojkovic M, Muller S, Brem G, Zakhartchenko V, Wolf E. Nuclear-cytoplasmic interactions affect in utero developmental capacity, phenotype, and cellular metabolism of bovine nuclear transfer fetuses. Biology of reproduction 2004c; 70:1196-1205

Hiendleder S, Wolf E. The mitochondrial genome in embryo technologies. Reproduction in domestic animals = Zuchthygiene 2003; 38:290-304

Hiendleder S, Zakhartchenko V, Wenigerkind H, Reichenbach HD, Bruggerhoff K, Prelle K, Brem G, Stojkovic M, Wolf E. Heteroplasmy in bovine fetuses produced by intra- and inter-subspecific somatic cell nuclear transfer: neutral segregation of nuclear donor mitochondrial DNA in various tissues and evidence for recipient cow mitochondria in fetal blood. Biology of reproduction 2003; 68:159-166

Hirai M, Boersma A, Hoeflich A, Wolf E, Foll J, Aumuller TR, Braun J. Objectively measured sperm motility and sperm head morphometry in boars (Sus scrofa): relation to fertility and seminal plasma growth factors. Journal of andrology 2001; 22:104-110

Hofmann A, Kessler B, Ewerling S, Weppert M, Vogg B, Ludwig H, Stojkovic M, Boelhauve M, Brem G, Wolf E, Pfeifer A. Efficient transgenesis in farm animals by lentiviral vectors. EMBO reports 2003; 4:1054-1060

Hofmann A, Zakhartchenko V, Weppert M, Sebald H, Wenigerkind H, Brem G, Wolf E, Pfeifer A. Generation of transgenic cattle by lentiviral gene transfer into oocytes. Biology of reproduction 2004; 71:405-409

Hyttel P, Laurincik J, Viuff D, Fair T, Zakhartchenko V, Rosenkranz C, Avery B, Rath D, Niemann H, Thomsen PD, Schellander K, Callesen H, Wolf E, Ochs RL, Greve T. Activation of ribosomal RNA genes in preimplantation cattle and swine embryos. Animal reproduction science 2000; 60-61:49-60

Hyttel P, Laurincik J, Zakhartchenko V, Stojkovic M, Wolf E, Muller M, Ochs RL, Brem G. Nucleolar protein allocation and ultrastructure in bovine embryos produced by nuclear transfer from embryonic cells. Cloning 2001; 3:69-82
Kolle S, Sinowatz F, Boie G, Lincoln D, Palma G, Stojkovic M, Wolf E. Topography of growth hormone receptor expression in the bovine embryo. Histochemistry and cell biology 1998; 109:417-419

Kolle S, Stojkovic M, Boie G, Wolf E, Sinowatz F. Growth hormone inhibits apoptosis in in vitro produced bovine embryos. Molecular reproduction and development 2002; 61:180-186

Kolle S, Stojkovic M, Boie G, Wolf E, Sinowatz F. Growth hormone-related effects on apoptosis, mitosis, and expression of connexin 43 in bovine in vitro maturation cumulus-oocyte complexes. Biology of reproduction 2003; 68:1584-1589

Kolle S, Stojkovic M, Prelle K, Waters M, Wolf E, Sinowatz F. Growth hormone (GH)/GH receptor expression and GH-mediated effects during early bovine embryogenesis. Biology of reproduction 2001; 64:1826-1834

Kolle S, Stojkovic M, Reese S, Reichenbach HD, Wolf E, Sinowatz F. Effects of growth hormone on the ultrastructure of bovine preimplantation embryos. Cell and tissue research 2004; 317:101-108

Laurincik J, Schmoll F, Mahabir E, Schneider H, Stojkovic M, Zakhartchenko V, Prelle K, Hendrixen PJ, Voss PL, Moeszlacher GG, Avery B, Dieleman SJ, Besenfelder U, Muller M, Ochs RL, Wolf E, Schellander K, Maddox-Hyttel P. Nucleolar proteins and ultrastructure in bovine in vivo developed, in vitro produced, and parthenogenetic cleavage-stage embryos. Molecular reproduction and development 2003; 65:73-85

Laurincik J, Zakhartchenko V, Stojkovic M, Brem G, Wolf E, Muller M, Ochs RL, Maddox-Hyttel P. Nucleolar protein allocation and ultrastructure in bovine embryos produced by nuclear transfer from granulosa cells. Molecular reproduction and development 2002; 61:477-487

Modl J, Reichenbach HD, Wolf E, Brem G. Development of frozen-thawed porcine blastocysts in vitro and in vivo. The Veterinary record 1996; 139:208-210
Motlik J, Alberio R, Zakhartchenko V, Stojkovic M, Kubelka M, Wolf E. The effect of activation of Mammalian oocytes on remodeling of donor nuclei after nuclear transfer. Cloning and stem cells 2002; 4:245-252

Mueller S, Prelle K, Rieger N, Petznek H, Lassnig C, Luksch U, Aigner B, Baetscher M, Wolf E, Mueller M, Brem G. Chimeric pigs following blastocyst injection of transgenic porcine primordial germ cells. Molecular reproduction and development 1999; 54:244-254

Ott M, Schernthaner W, Sinowatz F, Wolf E. Effects of bovine serum albumin and estrous cow serum on development and ultrastructure of in vitro-produced porcine embryos. Anatomia, histologia, embryologia 2002; 31:151-157

Prelle K, Stojkovic M, Boxhammer K, Motlik J, Ewald D, Arnold GJ, Wolf E. Insulin-like growth factor I (IGF-I) and long R(3)IGF-I differently affect development and messenger ribonucleic acid abundance for IGF-binding proteins and type I IGF receptors in in vitro produced bovine embryos. Endocrinology 2001; 142:1309-1316

Prelle K, Zakhartchenko V, Wolf E. [Application of stem cell technology and nuclear transfer in animal models]. Zeitschrift fur arztliche Fortbildung und Qualitatssicherung 2002a; 96:434-440

Prelle K, Zink N, Wolf E. Pluripotent stem cells--model of embryonic development, tool for gene targeting, and basis of cell therapy. Anatomia, histologia, embryologia 2002b; 31:169-186

Reischl J, Prelle K, Schol H, Neumuller C, Einspanier R, Sinowatz F, Wolf E. Factors affecting proliferation and dedifferentiation of primary bovine oviduct epithelial cells in vitro. Cell and tissue research 1999; 296:371-383

Rief S, Sinowatz F, Stojkovic M, Einspanier R, Wolf E, Prelle K. Effects of a novel co-culture system on development, metabolism and gene expression of bovine embryos produced in vitro. Reproduction 2002; 124:543-556

Santl B, Wenigerkind H, Schernthaner W, Modl J, Stojkovic M, Prelle K, Holtz W, Brem G, Wolf E. Comparison of ultrasound-guided vs laparoscopic transvaginal ovum pick-up (OPU) in simmental heifers. Theriogenology 1998; 50:89-100

Santos F, Zakhartchenko V, Stojkovic M, Peters A, Jenuwein T, Wolf E, Reik W, Dean W. Epigenetic marking correlates with developmental potential in cloned bovine preimplantation embryos. Current biology : CB 2003; 13:1116-1121

Schernthaner W, Wenigerkind H, Stojkovic M, Palma GA, Modl J, Wolf E, Brem G. Pregnancy rate after ultrasound-guided follicle aspiration in nonlactating cows from different breeds. Zentralblatt fur Veterinarmedizin Reihe A 1999; 46:33-37

Shi W, Dirim F, Wolf E, Zakhartchenko V, Haaf T. Methylation reprogramming and chromosomal aneuploidy in in vivo fertilized and cloned rabbit preimplantation embryos. Biology of reproduction 2004; 71:340-347

Shi W, Hoeflich A, Flaswinkel H, Stojkovic M, Wolf E, Zakhartchenko V. Induction of a senescent-like phenotype does not confer the ability of bovine immortal cells to support the development of nuclear transfer embryos. Biology of reproduction 2003a; 69:301-309

Shi W, Zakhartchenko V, Wolf E. Epigenetic reprogramming in mammalian nuclear transfer. Differentiation; research in biological diversity 2003b; 71:91-113

Steinborn R, Schinogl P, Zakhartchenko V, Achmann R, Schernthaner W, Stojkovic M, Wolf E, Muller M, Brem G. Mitochondrial DNA heteroplasmy in cloned cattle produced by fetal and adult cell cloning. Nature genetics 2000; 25:255-257

Steinborn R, Zakhartchenko V, Jelyazkov J, Klein D, Wolf E, Muller M, Brem G. Composition of parental mitochondrial DNA in cloned bovine embryos. FEBS letters 1998a; 426:352-356

Steinborn R, Zakhartchenko V, Wolf E, Muller M, Brem G. Non-balanced mix of mitochondrial DNA in cloned cattle produced by cytoplast-blastomere fusion. FEBS letters 1998b; 426:357-361

Stojkovic M, Buttner M, Zakhartchenko V, Brem G, Wolf E. A reliable procedure for differential staining of in vitro produced bovine blastocysts: comparison of tissue culture medium 199 and Menezo's B2 medium. Animal reproduction science 1998; 50:1-9

Stojkovic M, Buttner M, Zakhartchenko V, Riedl J, Reichenbach HD, Wenigerkind H, Brem G, Wolf E. Secretion of interferon-tau by bovine embryos in long-term culture: comparison of in vivo derived, in vitro produced, nuclear transfer and demi-embryos. Animal reproduction science 1999a; 55:151-162

Stojkovic M, Kolle S, Peinl S, Stojkovic P, Zakhartchenko V, Thompson JG, Wenigerkind H, Reichenbach HD, Sinowatz F, Wolf E. Effects of high concentrations of hyaluronan in culture medium on development and survival rates of fresh and frozen-thawed bovine embryos produced in vitro. Reproduction 2002; 124:141-153

Stojkovic M, Krebs O, Kolle S, Prelle K, Assmann V, Zakhartchenko V, Sinowatz F, Wolf E. Developmental regulation of hyaluronan-binding protein (RHAMM/IHABP) expression in early bovine embryos. Biology of reproduction 2003; 68:60-66

Stojkovic M, Machado SA, Stojkovic P, Zakhartchenko V, Hutzler P, Goncalves PB, Wolf E. Mitochondrial distribution and adenosine triphosphate content of bovine oocytes before and after in vitro maturation: correlation with morphological criteria and developmental capacity after in vitro fertilization and culture. Biology of reproduction 2001; 64:904-909

Stojkovic M, Westesen K, Zakhartchenko V, Stojkovic P, Boxhammer K, Wolf E. Coenzyme Q(10) in submicron-sized dispersion improves development, hatching, cell proliferation, and adenosine triphosphate content of in vitro-produced bovine embryos. Biology of reproduction 1999b; 61:541-547

Stojkovic M, Wolf E, Buttner M, Berg U, Charpigny G, Schmitt A, Brem G. Secretion of biologically active interferon tau by in vitro-derived bovine trophoblastic tissue. Biology of reproduction 1995; 53:1500-1507

Stojkovic M, Wolf E, Van Langendonckt A, Vansteenbrugge A, Charpigny G, Reinaud P, Gandolfi F, Brevini TA, Mermillod P, Terqui M, Brem G, Massip A. Correlations between chemical parameters, mitogenic activity and embryotrophic activity of bovine oviduct-conditioned medium. Theriogenology 1997a; 48:659-673

Stojkovic M, Zakhartchenko V, Brem G, Wolf E. Support for the development of bovine embryos in vitro by secretions of bovine trophoblastic vesicles derived in vitro. Journal of reproduction and fertility 1997b; 111:191-196

Suttner R, Zakhartchenko V, Stojkovic P, Muller S, Alberio R, Medjugorac I, Brem G, Wolf E, Stojkovic M. Intracytoplasmic sperm injection in bovine: effects of oocyte activation, sperm pretreatment and injection technique. Theriogenology 2000; 54:935-948

Wolf E. Cloning of animals--perspectives for biomedical research. Experimental and toxicologic pathology : official journal of the Gesellschaft fur Toxikologische Pathologie 2002; 54:161-162

Wolf E, Arnold GJ, Bauersachs S, Beier HM, Blum H, Einspanier R, Frohlich T, Herrler A, Hiendleder S, Kolle S, Prelle K, Reichenbach HD, Stojkovic M, Wenigerkind H, Sinowatz F. Embryo-maternal communication in bovine - strategies for deciphering a complex cross-talk. Reproduction in domestic animals = Zuchthygiene 2003; 38:276-289

Wolf E, Schernthaner W, Zakhartchenko V, Prelle K, Stojkovic M, Brem G. Transgenic technology in farm animals--progress and perspectives. Experimental physiology 2000; 85:615-625

Wolf E, Zakhartchenko V, Brem G. Nuclear transfer in mammals: recent developments and future perspectives. Journal of biotechnology 1998; 65:99-110

Zakhartchenko V, Alberio R, Stojkovic M, Prelle K, Schernthaner W, Stojkovic P, Wenigerkind H, Wanke R, Duchler M, Steinborn R, Mueller M, Brem G, Wolf E. Adult cloning in cattle: potential of nuclei from a permanent cell line and from primary cultures. Molecular reproduction and development 1999a; 54:264-272

Zakhartchenko V, Durcova-Hills G, Schernthaner W, Stojkovic M, Reichenbach HD, Mueller S, Steinborn R, Mueller M, Wenigerkind H, Prelle K, Wolf E, Brem G. Potential of fetal germ cells for nuclear transfer in cattle. Molecular reproduction and development 1999b; 52:421-426

Zakhartchenko V, Durcova-Hills G, Stojkovic M, Schernthaner W, Prelle K, Steinborn R, Muller M, Brem G, Wolf E. Effects of serum starvation and re-cloning on the efficiency of nuclear transfer using bovine fetal fibroblasts. Journal of reproduction and fertility 1999c; 115:325-331

Zakhartchenko V, Mueller S, Alberio R, Schernthaner W, Stojkovic M, Wenigerkind H, Wanke R, Lassnig C, Mueller M, Wolf E, Brem G. Nuclear transfer in cattle with non-transfected and transfected fetal or cloned transgenic fetal and postnatal fibroblasts. Molecular reproduction and development 2001; 60:362-369

Zakhartchenko V, Reichenbach HD, Riedl J, Palma GA, Wolf E, Brem G. Nuclear transfer in cattle using in vivo-derived vs. in vitro-produced donor embryos: effect of developmental stage. Molecular reproduction and development 1996; 44:493-498

Zakhartchenko V, Stojkovic M, Brem G, Wolf E. Karyoplast-cytoplast volume ratio in bovine nuclear transfer embryos: effect on developmental potential. Molecular reproduction and development 1997; 48:332-338