Secretion of interferon-tau by bovine embryos in long-term culture: comparison of in vivo derived, in vitro produced, nuclear transfer and demi-embryos

1999 ◽  
Vol 55 (3-4) ◽  
pp. 151-162 ◽  
Author(s):  
M Stojkovic ◽  
M Büttner ◽  
V Zakhartchenko ◽  
J Riedl ◽  
H.-D Reichenbach ◽  
...  
Keyword(s):  
Nuclear Transfer ◽  
Bovine Embryos ◽  
Interferon Tau ◽  
Long Term Culture

From the germinal cells to the newborn animal: The transmission of genes and life through the generations

2003 ◽  
Vol 51 (3) ◽  
pp. 371-384
Author(s):  
P. V. Drion ◽  
O. Szenci ◽  
F. Ectors ◽  
D. Wirth ◽  
Zs. Perényi ◽  
...  
Keyword(s):  
Nuclear Transfer ◽  
Fetal Mortality ◽  
Newborn Animal ◽  
Multiple Ovulation ◽  
Early Appearance ◽  
Early Embryos ◽  
Germinal Cells

The technology of reproduction progressed considerably during the last decade, leading to a certain availability of in vitro methods for fertilisation, oocyte maturation and embryo culture. The most spectacular manipulations are cloning and transgenesis. This review focuses on the early appearance of germinal cell precursors and the long-standing fate of gametes in mammals. The evident complexity and long-term programming of events in gametes and early embryos explain part of the difficulties encountered during the development of in vitro and in vivo methods such as multiple ovulation and embryo transfer (MOET), oestrus synchronisation, ovulation induction, superovulation, in vitro maturation and fertilisation, cryopreservation, transgenesis, nuclear transfer and cloning) and the occurrence of unexpected alterations of development, e.g. embryonic or fetal mortality, large-weight newborn syndrome and other dysregulations in imprinting or DNA transmission.

Distinct Requirements for Optimal Growth and In Vitro Expansion of Human CD34+CD38− Bone Marrow Long-Term Culture-Initiating Cells (LTC-IC), Extended LTC-IC, and Murine In Vivo Long-Term Reconstituting Stem Cells

Blood ◽  
1999 ◽  
Vol 94 (12) ◽  
pp. 4093-4102 ◽  
Author(s):  
Veslemøy Ramsfjell ◽  
David Bryder ◽  
Helga Björgvinsdóttir ◽  
Sten Kornfält ◽  
Lars Nilsson ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Stem Cell ◽  
Cell Activity ◽  
Optimal Growth ◽  
Cell Manipulation ◽  
Long Term Culture ◽  
Human Cd34

Abstract Recently, primitive human bone marrow (BM) progenitors supporting hematopoiesis in extended (>60 days) long-term BM cultures were identified. Such extended long-term culture-initiating cells (ELTC-IC) are of the CD34+CD38− phenotype, are quiescent, and are difficult to recruit into proliferation, implicating ELTC-IC as the most primitive human progenitor cells detectable in vitro. However, it remains to be established whether ELTC-IC can proliferate and potentially expand in response to early acting cytokines. Here, CD34+CD38− BM ELTC-IC (12-week) were efficiently recruited into proliferation and expanded in vitro in response to early acting cytokines, but conditions for expansion of ELTC-IC activity were distinct from those of traditional (5-week) LTC-IC and murine long-term repopulating cells. Whereas c-kit ligand (KL), interleukin-3 (IL-3), and IL-6 promoted proliferation and maintenance or expansion of murine long-term reconstituting activity and human LTC-IC, they dramatically depleted ELTC-IC activity. In contrast, KL, flt3 ligand (FL), and megakaryocyte growth and development factor (MGDF) (and KL + FL + IL-3) expanded murine long-term reconstituting activity as well as human LTC-IC and ELTC-IC. Expansion of LTC-IC was most optimal after 7 days of culture, whereas optimal expansion of ELTC-IC activity required 12 days, most likely reflecting the delayed recruitment of quiescent CD34+CD38− progenitors. The need for high concentrations of KL, FL, and MGDF (250 ng/mL each) and serum-free conditions was more critical for expansion of ELTC-IC than of LTC-IC. The distinct requirements for expansion of ELTC-IC activity when compared with traditional LTC-IC suggest that the ELTC-IC could prove more reliable as a predictor for true human stem cell activity after in vitro stem cell manipulation.

Distinct Requirements for Optimal Growth and In Vitro Expansion of Human CD34+CD38− Bone Marrow Long-Term Culture-Initiating Cells (LTC-IC), Extended LTC-IC, and Murine In Vivo Long-Term Reconstituting Stem Cells

Blood ◽  
1999 ◽  
Vol 94 (12) ◽  
pp. 4093-4102 ◽  
Author(s):  
Veslemøy Ramsfjell ◽  
David Bryder ◽  
Helga Björgvinsdóttir ◽  
Sten Kornfält ◽  
Lars Nilsson ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Stem Cell ◽  
Cell Activity ◽  
Optimal Growth ◽  
Cell Manipulation ◽  
Long Term Culture ◽  
Human Cd34

Recently, primitive human bone marrow (BM) progenitors supporting hematopoiesis in extended (>60 days) long-term BM cultures were identified. Such extended long-term culture-initiating cells (ELTC-IC) are of the CD34+CD38− phenotype, are quiescent, and are difficult to recruit into proliferation, implicating ELTC-IC as the most primitive human progenitor cells detectable in vitro. However, it remains to be established whether ELTC-IC can proliferate and potentially expand in response to early acting cytokines. Here, CD34+CD38− BM ELTC-IC (12-week) were efficiently recruited into proliferation and expanded in vitro in response to early acting cytokines, but conditions for expansion of ELTC-IC activity were distinct from those of traditional (5-week) LTC-IC and murine long-term repopulating cells. Whereas c-kit ligand (KL), interleukin-3 (IL-3), and IL-6 promoted proliferation and maintenance or expansion of murine long-term reconstituting activity and human LTC-IC, they dramatically depleted ELTC-IC activity. In contrast, KL, flt3 ligand (FL), and megakaryocyte growth and development factor (MGDF) (and KL + FL + IL-3) expanded murine long-term reconstituting activity as well as human LTC-IC and ELTC-IC. Expansion of LTC-IC was most optimal after 7 days of culture, whereas optimal expansion of ELTC-IC activity required 12 days, most likely reflecting the delayed recruitment of quiescent CD34+CD38− progenitors. The need for high concentrations of KL, FL, and MGDF (250 ng/mL each) and serum-free conditions was more critical for expansion of ELTC-IC than of LTC-IC. The distinct requirements for expansion of ELTC-IC activity when compared with traditional LTC-IC suggest that the ELTC-IC could prove more reliable as a predictor for true human stem cell activity after in vitro stem cell manipulation.

In vitro culture of stem-like cells derived from somatic cell nuclear transfer bovine embryos of the Korean beef cattle species, HanWoo

2016 ◽  
Vol 28 (11) ◽  
pp. 1762 ◽  
Author(s):  
Daehwan Kim ◽  
Sangkyu Park ◽  
Yeon-Gil Jung ◽  
Sangho Roh
Keyword(s):  
Beef Cattle ◽  
Somatic Cell ◽  
Somatic Cell Nuclear Transfer ◽  
Nuclear Transfer ◽  
Embryoid Bodies ◽  
Marker Genes ◽  
Bovine Embryos ◽  
In Vivo Differentiation

We established and maintained somatic cell nuclear transfer embryo-derived stem-like cells (SCNT-eSLCs) from the traditional Korean beef cattle species, HanWoo (Bos taurus coreanae). Each SCNT blastocyst was placed individually on a feeder layer with culture medium containing three inhibitors of differentiation (3i). Primary colonies formed after 2–3 days of culture and the intact colonies were passaged every 5–6 days. The cells in each colony showed embryonic stem cell-like morphologies with a distinct boundary and were positive to alkaline phosphatase staining. Immunofluorescence and reverse transcription–polymerase chain reaction analyses also confirmed that these colonies expressed pluripotent markers. The colonies were maintained over 50 passages for more than 270 days. The cells showed normal karyotypes consisting of 60 chromosomes at Passage 50. Embryoid bodies were formed by suspension culture to analyse in vitro differentiation capability. Marker genes representing the differentiation into three germ layers were expressed. Typical embryonal carcinoma was generated after injecting cells under the testis capsule of nude mice, suggesting that the cultured cells may also have the potential of in vivo differentiation. In conclusion, we generated eSLCs from SCNT bovine embryos, using a 3i system that sustained stemness, normal karyotype and pluripotency, which was confirmed by in vitro and in vivo differentiation.

scholarly journals 63INTERFERON-TAU EXPRESSION FROM PRIMARY TROPHECTODERM OUTGROWTHS OF BOVINE BLASTOCYSTS: COMPARISON BETWEEN IVP, NT, AND PARTHENOGENIC EMBRYOS

2004 ◽  
Vol 16 (2) ◽  
pp. 153
Author(s):  
O.M. Ocon ◽  
N.C. Talbot ◽  
A.M. Powell ◽  
A.D. Ealy
Keyword(s):  
Nuclear Transfer ◽  
Culture Medium ◽  
Mixed Linear Model ◽  
Bovine Embryos ◽  
Interferon Tau ◽  
Vitro Fertilization ◽  
Fetal Bovine ◽  
Conditioned Cell Culture Medium ◽  
Bovine Blastocyst

Interferon-tau (IFN-tau) is expressed soon after bovine blastocyst formation and might be useful as a marker of appropriate biological function in embryos produced by nuclear transfer. To assess this possibility we have compared IFN-tau levels in the conditioned medium of primary trophectoderm cultures derived from IVP, nuclear transfer (NT), or parthenogenic bovine embryos. Embryos were produced from in vitro-matured cumulus-oocyte complexes processed from local slaughterhouse ovaries or obtained from Bomed, Inc. (Madison, WI, USA). In vitro fertilization, NT, and parthenogensis were as previously described (Talbot et al., 2000 Tissue and Cell, 32, 9–27) except that embryo culture was in G1/G2 medium in 5% oxygen (Lane et al., 2003 Theriogenology, 60, 407–419). Each 8–11-day embryo was cultured individually in a 4-well plate well (Nunc) with STO feeder cells using DMEM medium containing 10% fetal bovine serum as previously described (Talbot et al., 2000 Biol. Reprod. 62, 235–247). Any contaminating epiblast or endoderm was physically dissected and discarded so as to produce pure trophectoderm outgrowths. The success/failure ratio for colony formation was similar for IVP and NT embryos (IVP=155/29; NT=104/25), but was significantly different (P<0.05) for parthenogenic embryos (54/43). Trophectoderm colonies reached diameters of 1 to 1.5cm in 3–4wk, and, at this time, 72-h-conditioned cell culture medium was harvested, frozen, and measured for IFN-tau anti-viral activity as previously described (Talbot et al., 2000 Biol. Reprod. 62, 235–247). From 313 observations, IFN-tau production was analyzed as a two-factor mixed linear model. Differences in IFN-tau production by type of embryo were statistically significant (F=42.61; P<0.0001; df=2). Mean comparisons were done with Sidak adjusted P-values so that the experiment-wise error was 0.05. IFN-tau production means for IVP-, NT-, and parthenogenic-derived trophectoderm were 4311IUmL−1 (n=155), 626IUmL−1 (n=104), and 1595IUmL−1 (n=54), respectively. The results show that mean IFN-tau production from trophectoderm cultures derived from NT embryos is significantly reduced in comparison to IVP- and parthenogenote-derived cultures. Parthenogenote-derived cultures also produced significantly less IFN-tau than IVP embryos on average. IFN-tau production from trophectoderm outgrowths may be a useful measure of NT reprogramming success.

scholarly journals 243TEMPORAL DIVERGENCE IN THE PATTERN OF MRNA EXPRESSION IN BOVINE EMBRYOS CULTURED FROM THE ZYGOTE TO BLASTOCYST STAGE IN VITRO OR IN VIVO

2004 ◽  
Vol 16 (2) ◽  
pp. 242
Author(s):  
P. Lonergan ◽  
D. Rizos ◽  
A. Gutierrez-Adan ◽  
P.M. Moreira ◽  
B. Pintado ◽  
...  
Keyword(s):  
Growth Factor ◽  
Relative Abundance ◽  
Transcript Abundance ◽  
Blastocyst Stage ◽  
Insulin Like Growth Factor ◽  
Bovine Embryos ◽  
Interferon Tau ◽  
Cell Stage

The objective of this study was to examine the time during the post-fertilization culture period that gene expression patterns of in vitro cultured bovine embryos diverge from those of their in vivo cultured counterparts. Presumptive bovine zygotes were produced by IVM/IVF of immature oocytes collected from the ovaries of slaughtered animals. At approximately 20h post-insemination (hpi), presumptive zygotes were randomly divided into two culture groups, either in vitro in synthetic oviduct fluid or in vivo, and transferred into the ewe oviduct. Embryos were recovered from both systems at approximately 30hpi (2-cell), two (4-cell), three (8-cell), four (16-cell), five (early morula), six (compact morula) or seven (blastocyst) days pi and snap-frozen for the analysis of transcript abundance using real-time PCR. The transcripts studied were interferon-tau, apoptosis regulator box-a (Bax), connexin 43, sarcosine oxidase, glucose transporter 5, mitochondrial Mn-superoxide dismutase, insulin-like growth factor II, and insulin-like growth factor-I receptor, most of which are known from our previous work to be differentially transcribed in blastocysts derived from culture in vitro or in vivo. Analysis was done on pools of 10 embryos. Data were analyzed using one-way repeated measures ANOVA. The relative abundance of the transcripts studied varied throughout the preimplantation period and was strongly influenced by the culture environment. For example, transcripts for interferon-tau were detected from the 8-cell stage onwards in in vitro-cultured embryos but not until the early morula stage in those cultured in vivo. Levels of this transcript increased significantly at the compact morula and blastocyst stages in both groups but were significantly higher (P<0.05) in in vitro-cultured embryos at both stages. mRNA for Bax was not detected before the 8-cell stage in in vitro cultured embryos and not until the 16-cell stage in in vivo cultured embryos. The abundance of this transcript increased significantly thereafter up to the blastocyst stage in both groups. The level of expression was significantly higher (P<0.05) at all stages of development in in vitro-cultured embryos than those cultured in vivo. The relative abundance of Cx43 transcripts decreased in both in vitro- and in vivo-cultured embryos at the 8- to 16-cell stage. Levels remained low thereafter in the in vitro-cultured embryos but significantly increased in those cultured in vivo. Transcript abundance was significantly higher in in vivo cultured embryos from Day 4 onwards with a ten-fold difference presence at the blastocyst stage. Differences also existed for the other transcripts studied. These data demonstrate that changes in transcript abundance in blastocyst stage embryos are in many cases a consequence of perturbed transcription earlier in development. Depending on the transcript, these differences may be evident in as short as 10h of culture.

Developmental ability of bovine embryos after nuclear transfer based on the nuclear source: In vivo versus in vitro

1994 ◽  
Vol 42 (4) ◽  
pp. 695-702 ◽  
Author(s):  
Y. Heyman ◽  
P. Chesné ◽  
D. Lebourhis ◽  
N. Peynot ◽  
J.P. Renard
Keyword(s):  
Nuclear Transfer ◽  
Bovine Embryos

scholarly journals Extended long-term culture reveals a highly quiescent and primitive human hematopoietic progenitor population

Blood ◽  
1996 ◽  
Vol 88 (9) ◽  
pp. 3306-3313 ◽  
Author(s):  
QL Hao ◽  
FT Thiemann ◽  
D Petersen ◽  
EM Smogorzewska ◽  
GM Crooks
Keyword(s):  
Cord Blood ◽  
Hematopoietic Progenitors ◽  
Generative Capacity ◽  
Long Term Culture ◽  
Clonal Proliferation ◽  
Unit Cells ◽  
Functional Hierarchy

Long-term culture-initiating cells (LTC-IC) are hematopoietic progenitors able to generate colony-forming unit-cells (CFU) after 5 to 8 weeks (35 to 60 days) of culture on bone marrow (BM) stroma and represent the most primitive progenitors currently detectable in vitro. We have recently reported that long-term cultures initiated with CD34+CD38- cells from BM or cord blood are able to continue generating CFU for at least 100 days, ie, beyond the standard LTC-IC period. In this report, single-cell cultures from cord blood and retroviral marking of cord blood and BM were used to study whether the subpopulation of CD34+CD38- cells able to generate CFU beyond 60 days (“extended long-term culture-initiating cells” or ELTC-IC) are functionally distinct from LTC-IC in terms of timing of initial clonal proliferation and generative capacity. All cord blood LTC-IC formed clones of greater than 50 cells by day 30. In contrast, cord blood ELTC-IC proliferated later in culture, 50% forming clones after day 30. Although efficient retroviral marking of LTC-IC was seen (25% to 45%), marking of ELTC-IC was inefficient (< 1%), consistent with a more quiescent progenitor population. There was a positive correlation between time of clonal proliferation and generative capacity. ELTC-IC generated threefold to fourfold more progeny than did LTC-IC (P < .002). These studies show that there is a functional hierarchy of progenitors in long-term culture which correlates with their level of quiescence. By extending the LTC-IC assay, a more primitive progenitor may be studied that may be functionally closer to the human long-term repopulation stem cell in vivo.

scholarly journals Chromatin Modifying Agents in theIn VitroProduction of Bovine Embryos

2011 ◽  
Vol 2011 ◽  
pp. 1-9 ◽  
Author(s):  
Fabio Morato Monteiro ◽  
Clara Slade Oliveira ◽  
Letícia Zoccolaro Oliveira ◽  
Naiara Zoccal Saraiva ◽  
Maria Eugênia Zerlotti Mercadante ◽  
...  
Keyword(s):  
Gene Expression ◽  
Nuclear Transfer ◽  
Epigenetic Reprogramming ◽  
Nuclear Reprogramming ◽  
Bovine Embryos ◽  
Aberrant Gene Expression ◽  
Low Efficiency ◽  
Aberrant Gene

The low efficiency observed in cloning by nuclear transfer is related to an aberrant gene expression following errors in epigenetic reprogramming. Recent studies have focused on further understanding of the modifications that take place in the chromatin of embryos during the preimplantation period, through the use of chromatin modifying agents. The goal of these studies is to identify the factors involved in nuclear reprogramming and to adjustin vitromanipulations in order to better mimicin vivoconditions. Therefore, proper knowledge of epigenetic reprogramming is necessary to prevent possible epigenetic errors and to improve efficiency and the use ofin vitrofertilization and cloning technologies in cattle and other species.

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