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B Reichelt

Publications and source records attributed to B Reichelt.

7 recordsLinked to original sources
PubMed1995

Ratio of inner cell mass and trophoblastic cells in demi- and intact pig embryos.

Pig morulae, early blastocysts and blastocysts were microsurgically bisected to produce zona-free demi-embryos or remained nonbisected with or without zona pellucida, and the presence of inner cell mass cells was determined using a differential fluorochrome staining technique. After 24 h of in vitro culture, all demi-embryos were classified into three categories, based on morphological criteria: 1, excellent; 2, fair; and 3, degenerated. The average number of total cells and inner cell mass cells in intact embryos cultured without zona pellucida for 24 h was higher (P < 0.05) than that for those with zona pellucida in morulae and early blastocysts. The percentage of demi-embryos without inner cell mass cells in these different morphological categories was 18.7%, 22.2% and 29.8% for morulae, respectively; 3.8%, 16.7% and 30.8% for early blastocysts, respectively; and 3.7%, 32.0% and 36.4% for blastocysts, respectively. The percentage of demi-embryos without inner cell mass cells was lower (P < 0.01) in demi-embryos classified in category 1 compared with category 3 in early blastocysts and in category 1 compared with categories 2 and 3 in blastocysts. Significant differences in the total number of cells and the number of inner cell mass cells were apparent among the three morphological categories of demi-embryos derived from morulae, early blastocysts and blastocysts. The ratio of total cells to inner cell mass cells was similar among intact pig embryos and the different morphological categories of demi-embryos derived from morulae, early blastocysts and blastocysts, with the exception of that between demi-blastocysts of category 1 and the other groups.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1994

Negatively cis-acting elements in the distal part of the promoter of Epstein-Barr virus trans-activator gene BZLF1.

Epstein-Barr virus (EBV) replicates in a latent or a lytic way in the infected organism, depending on the type and level of differentiation of the host cell. The switch between latency and lytic replication was previously shown, for Burkitt's lymphoma cell lines, to depend on the viral BZLF1 gene product. Protein-DNA assays were used to identify the cis-acting elements that represent the link between regulating signal transduction pathways and the viral cascade of gene expression. Specific binding of proteins to several sites of the BZLF1 promoter during latency was shown. Induction of the lytic cycle by stimulation with 12-O-tetradecanoyl-phorbol 13-acetate abolished the binding of these proteins to the distal promoter (positions -227 to -551), suggesting a functional role for the down-regulation of promoter activity during latency. Computer analysis identified a multiply repeated sequence motif, HI, in this region and exonuclease III footprints confirmed that these sites act as specific protein recognition sites. Using a set of reporter plasmids we were able to demonstrate a negative regulatory effect of the HI motif in some B lymphoid cell lines, in contrast to epithelial HeLa cells. The HI silencer elements are different from other silencer elements described so far in respect of their sequence and protein-binding pattern during the activation of BZLF1.

Base Sequence↗
PubMed1994

Generation of identical twin piglets following bisection of embryos at the morula and blastocyst stage.

Porcine morulae and blastocysts were microsurgically bisected and the resulting zona pellucida-free demi-embryos were either cultured in vitro for 48 h or transferred after 24 h of culture into--24 h asynchronous recipients. All demi-embryos were evaluated according to morphological criteria and classified into three categories (excellent, fair or degenerated). The average diameter and the number of cells were determined. Of 1162 bisected embryos, 764 pairs (66%) were evaluated as transferable after 24 h of culture in vitro. The average diameter after 48 h of culture in vitro was different (P < 0.01) among demi-embryos of the three morphological categories as was the number of cells. The greatest diameter and the greatest number of cells were found in demi-embryos classified as morphologically excellent. A total of 22 of 27 recipients (81.5%) remained pregnant and 21 recipients delivered 126 piglets of which six were stillborn. The survival rate of demi-embryos in farrowing recipients was 21.2% (126 of 594). Litter size was significantly reduced in recipients after transfer of demi-embryos compared with that of mated controls (6.0 +/- 2.5 versus 10.8 +/- 2.1 piglets). Similarly, the postpartum losses of piglets were higher in the experimental than in the control gilts (26.7% versus 11.6%). Duration of gestation, average birth weight and daily weight gain were not affected. Among the 126 piglets, seven pairs of identical twins (2.3% of 311 transferred pairs) were identified using several genetic markers in blood (blood groups, polymorphic enzymes and plasma proteins) in a total of 25 gene loci. DNA fingerprinting revealed an identical banding pattern between the two partners of each of the seven pairs. Birth and weaning weight as well as daily weight gain varied considerably between monozygotic partners.

Animals↗
PubMed1993

The lytic transition of Epstein-Barr virus is imitated by recombinant B-cells.

Lytic transition of Epstein-Barr virus (EBV) is initiated by distinct immediate early regulators of the viral cycle, in synchronization to temporary, permissive conditions during host cell differentiation. We developed eukaryotic vectors suitable to imitate the processes involved in lytic transition in cell culture systems. Two stable B cell lines were established: R59Z activator cells were used to induce lytic EBV expression in a constitutive manner by the production of the BZLF 1 trans-activator (Zta). R7-57 reporter cells, on the other hand, signaled induced activity of the lytic origin of EBV replication (ori Lyt). Different modes, like chemical induction, lytic superinfection with EBV and single gene trans-activation converted the recombinant ori Lyt element in R7-57 reporter cells. BZLF 1, transiently expressed in R7-57 reporter cells, was the only EBV trans-activator found, sufficient in inducing the viral lytic cycle. Basing on these experiments, trans-cellular activation of EBV was tested by cocultivation of BZLF 1-expressing R59Z activator cells with the R7-57 reporter line. No lytic effect on the reporter cells could be measured, neither by cocultivation of activator cells nor by coincubation of BZLF 1-containing cell lysates. Latency breaking activity, however, was transferred from activator to reporter cells when active, exogenous virus was added. The cell system described in these experiments provides a tool for the detection of EBV reactivation and demonstrates the potential of the lytic regulatory gene BZLF 1.

Animals↗
PubMed1993

Manipulating early pig embryos.

On the basis of established surgical procedures for embryo recovery and transfer, the early pig embryo can be subjected to various manipulations aimed at a long-term preservation of genetic material, the generation of identical multiplets, the early determination of sex or the alteration of the genetic make-up. Most of these procedures are still at an experimental stage and despite recent considerable progress are far from practical application. Normal piglets have been obtained after cryopreservation of pig blastocysts hatched in vitro, whereas all attempts to freeze embryos with intact zona pellucida have been unsuccessful. Pig embryos at the morula and blastocyst stage can be bisected microsurgically and the resulting demi-embryos possess a high developmental potential in vitro, whereas their development in vivo is impaired. Pregnancy rates are similar (80%) but litter size is reduced compared with intact embryos and twinning rate is approximately 2%. Pig blastomeres isolated from embryos up to the 16-cell stage can be grown in culture and result in normal blastocysts. Normal piglets have been born upon transfer of blastocysts derived from isolated eight-cell blastomeres, clearly underlining the totipotency of this developmental stage. Upon nuclear transfer the developmental capacity of reconstituted pig embryos is low and < 10% develop to morulae or blastocysts in vitro. Pig oocytes can be stimulated parthenogenetically and up to 10% grow to blastocysts in the in vitro culture. Sex determination can be achieved either by separation of X and Y chromosome bearing spermatozoa by flow cytometry or by analysing the expression of the HY antigen in pig embryos from the eight-cell to morula stage. Microinjection of foreign DNA has been successfully used to alter growth and development of transgenic pigs, and to produce foreign proteins in the mammary gland or in the bloodstream, indicating that pigs can be used as donors for valuable human pharmaceutical proteins. Another promising area of gene transfer is the increase of disease resistance in transgenic lines of pigs. Approximately 30% of pig spermatozoa bind considerable amounts of foreign DNA preferably at the post-acrosomal region, suggesting that transgenic animals can be obtained more efficiently than with the usual microinjection procedure. To increase gene transfer efficiency, considerable research efforts have been made to establish embryonic stem (ES) cells, but so far there is no definite proof of totipotency of the generated pig ES-like cells through viable chimaeras. In general, biotechnological procedures are much less advanced in pigs than in cows.

Animals↗
PubMedDate not supplied

[Production of monozygotic multiple offspring in agricultural domestic animals].

The production of monozygotic twins/multiplets in livestock animal can be achieved either by microsurgical bisection of embryos at the morular- or blastocyst stage, isolation and proliferation of blastomeres from early cleavage stages or nuclear transfer. While the success rates of micro-surgical bisection are high in ruminants (pregnancy rates approximately 50%, twinning rates 20-40%) in polyovulatory species such as swine, the efficiency is low with an average of 20% embryonic survival and 2% monozygotic twins that can positively identified via DNA-fingerprinting. Isolated blastomeres from multicellular embryos still possess great developmental capacity in vitro to progress to the blastocyst stage. However, their development in vivo is markedly reduced. This article summarizes the results obtained by the authors during several years of investigation. The results show for the first time that identical twins can be obtained in pigs which have been demonstrated to be a useful tool in biomedical research.

Animals↗