Genes expressed after retinoic acid-mediated differentiation of embryoid bodies are likely to be expressed during embryo development

In order to test if retinoic acid-mediated differentiation of embryoid bodies can be used as an in vitro preselection method for ES cell lines generated by gene trap, we correlated gene expression after in vitro differentiation and in 11.5-day embryos. Fifty-two genes captured by gene trap and expressed in undifferentiated embryonic stem cells were analyzed. Most genes expressed after differentiation in vitro were also expressed during embryo development. In order to correlate the expression patterns in vitro and in vivo, the in vitro expression in the center and in the periphery of the embryoid body outgrowths was observed. This allowed us to distinguish, according to in vitro expression, not expressed genes from those expressed widely in 11.5-day embryos. Consequently, with this parameter we increased the probability to obtain the restricted expression patterns in vivo. This study demonstrates the potential of the differentiation procedure in combination with the gene trap to select in vitro for genes expressed during embryo development.     

Pollen development in the anthers of several cereal strains and hybrids during cultivation in vitro

In order to induce androgenesis in vitro anthers of some cereals were cultivated. The highest number of proembryos was obtained in the hybrid Triticale in F3 generation on Blayder's medium supplemented with 3 and 12% sucrose. Proembryos represented multi-nuclear and multicellular formations which stagnated at the globular stage of development. Origin of roots from calluses was not accompanied by formation of buds. There was no formation of embryos from pollen grains in case of lines Triticum aestivum and Secale cereale.     

Dipeptidyl peptidase I and granzyme A are coordinately expressed during CD8+ T cell development and differentiation

Dipeptidyl peptidase I (DPPI) is a granule protease that plays a requisite role in processing the proenzyme form of the CTL granule serine proteases (granzymes). This study assesses DPPI mRNA and enzyme expression during T lymphocyte ontogeny and CTL differentiation. The most immature CD3- CD4- CD8- thymocytes were found to express >40-fold higher levels of DPPI mRNA, although levels of DPPI enzymatic activity in CD3- CD4- CD8- thymocytes were only modestly higher than those seen for CD4+ CD8+ or CD4+ CD8- thymocytes. More mature CD8+ CD4- thymocytes and CD8+ splenocytes expressed significantly higher levels of DPPI mRNA and enzymatic activity than CD4+ CD8+ or CD4+ CD8- thymocytes. Granzyme A mRNA expression was observed in DPPI expressing CD3- CD4- CD8- and CD8+ CD4- thymocytes and was also observed in CD8+ CD4- splenocytes; however, expression was not observed in CD4+ CD8+ or CD4+ CD8- thymocytes. Both DPPI mRNA and granzyme A mRNA expression in CD8+ T cells decreased to very low or undetectable levels during the first 48 h after allostimulation in MLCs. However, peak levels of both DPPI and granzyme A expression were observed later in the course of CD8+ T cell responses to alloantigen, with DPPI mRNA expression peaking on either day 3 or day 4 and granzyme A expression peaking at the end of a 5-day MLR. These data indicate that DPPI is expressed at all stages of T cell ontogeny and differentiation in which granzyme A mRNA is detected; consequently, DPPI appears to be available for the processing and activation of granzyme A during both CD8+ T cell development and differentiation.     

Connexins and E-cadherin are differentially expressed during trophoblast invasion and placenta differentiation in the rat

We have characterized the spatial and temporal expression pattern of six different connexin genes and E-cadherin during trophectoderm development in the rat. During the initial phase of trophoblast invasion at 6 days postcoitum (dpc), the trophoblast expressed E-cadherin but no connexin expression could be observed. With progressing invasion of the polar trophoblast into the maternal decidua, from 7 dpc onwards E-cadherin expression in the ectoplacental cone cells was lost and was now restricted to the extraembryonic ectoderm. In the ectoplacental cone and extraembryonic ectoderm instead connexin31 mRNA and protein could be found. This pattern was maintained up to day 10 postcoitum. The start of labyrinthine trophoblast differentiation from day 11 postcoitum onwards was characterized by persisting expression of E-cadherin in the extraembryonic ectoderm and its derivative, the chorionic plate. In addition to E-cadherin, from 10 dpc onwards, connexin26 started to be expressed in the chorionic plate, and both molecules remained coexpressed in the labyrinthine trophoblast of the mature placenta. During this differentiation process connexin31 remained expressed mainly in the proliferating spongiotrophoblast. From day 14 postcoitum onwards, the expression of connexin31 in the spongiotrophoblastic cells decreased, and in parallel they started to express connexin43. The trophoblastic giant cells, first characterized by connexin31, lost all of the investigated connexins during midgestation on day 12 postcoitum but started to express connexin43 from day 18 postcoitum onwards. Our studies suggest that loss of E-cadherin and induction of connexin31 expression is correlated with the proliferative and invasive stages of the ectoplacental cone, whereas appearance of connexin26, E-cadherin and connexin43 reflects the switch to the differentiated phenotypes of the mature placenta.