Expression of DRG during murine embryonic development.

We had previously characterised a cDNA which encodes a novel GTP-binding protein DRG. The expression of drg gene is down-regulated during the embryonic development of murine central nervous system. Further analysis of drg mRNA and protein in adult mouse tissues and various cell lines of different origins indicated that it is expressed widely, albeit at low and variable levels. In situ hybridisation analysis of mRNA expression in sections of mouse embryos indicated that drg is expressed strongly in various embryonic tissues. The expression of drg mRNA is greatly reduced in newborn animals. At cellular level, DRG protein can be detected in the cytoplasm. These observations suggest that DRG may play multiple roles in development and normal cell metabolism.     

Expression of murine Ia antigens during embryonic development.

An immunochemical analysis of the kinetics of appearance of Ia antigens during embryonic development was performed. Ia antigens first appear on the surface of embryonic cells 11 days postconception and their expression between days 11 and 16 of gestation is confined to the fetal liver. Ia antigen synthesis by fetal liver cells is detectable at day 14. Ia seems to precede Ig as a surface marker of embryonic liver cells, since Ig cannot be detected until day 16 of gestation. H-2 antigens may be immunoprecipitated from day 10 whole embryo cells. F9 primitive teratocarcinoma cells are Ia negative and H-2 negative.     

Expression of specific clones during B cell development.

The expression of DNP- and TNP-specific B cells in spleens of neonatal BALB/c mice was analyzed by the in vitro splenic focus technique. B cells of these specificities were found to be present in slightly higher frequency in neonatal than in adult spleens. The parameters of stimulation of neonatal B cells were similar to those of adult B cells but the antibody-forming cell progeny of neonatal B cells produce predominantly gammaM rather than gammaG antibody and produce less antibody than the progeny of adult B cells. Isoelectric focusing analyses of monoclonal antibodies derived from neonatal B cells stimulated in vitro with DNP or TNP revealed that over 90 per cent of the antibodies could be identified as belonging to one of six predominant clonotypes, three specific for DNP and three for TNP. While individual neonates rarely expressed all of the predominant clonotypes, B cells of each of the six clonotypes were found in several donors. When B cells of a given predominant clonotype were present in an individual many such B cells could be found and in many cases the entire DNP- or TNP-specific B cell population of an individual could be accounted for by B cells of a single clonotype. These findings are discussed in terms of the diversity of clonotype specificities available in neonates, the kinetics of development of cells within a clonotype, and factors that may play a role in controlling the expression of B cell clones.     

Expression of link protein during mouse follicular development.

To gain insight into the role of link protein in ovarian follicle development, we used immunohistochemistry to determine the patterns of link protein expression in mouse ovary in response to gonadotropin stimulation. Polyclonal antibodies were raised against link protein purified from bovine cartilage. Stimulation of immature mice with gonadotropins increased link protein expression in the granulosa layer of large preovulatory follicles. The number and intensity of immunostained cells increased over 2 hr after hCG injection. Cumulus cells stained link protein mainly in the extracellular matrix, whereas mural granulosa cells showed marked deposits of link protein in the cytoplasm. Link protein expression persisted in luteinized granulosa cells after ovulation and in corpora lutea. Link protein staining was also present in the theca cells and oocytes, which was a consistent finding regardless of gonadotropin treatment. The staining intensity was negated by treatment with hyaluronidase, suggesting that the link protein is bound to hyaluronic acid. On Western blotting, a reacting protein species of about 42 kD was seen in the gonadotropin-treated ovarian extract. The precise cellular distribution of link protein in mouse ovary was determined for the first time by an immunohistochemical method in this study. (J Histochem Cytochem 47:1433-1442, 1999)     

Specific expression of lactase in the jejunum and colon during postnatal development and hormone treatments in the rat.

The effects of hyperinsulinaemia imposed on normal rats on the subsequent insulin-responsiveness in vivo of 2-deoxy-D-glucose uptake of white adipose tissue and of various muscle types were investigated. This was done by treating normal rats with insulin via osmotic minipumps, and by comparing them with saline-infused controls. Hyperinsulinaemia produced by prior insulin treatment resulted in a well-tolerated hypoglycaemia. At the end of the treatment, the glucose utilization index of individual tissues was determined by euglycaemic/hyperinsulinaemic clamps associated with the labelled 2-deoxy-D-glucose method. Prior insulin treatment resulted in increased insulin-responsiveness of the glucose utilization index of white adipose tissue, and in increased total lipogenesis in white adipose tissue and fat-pad weight. In contrast, prior insulin treatment resulted in a decreased glucose utilization index of several muscles. These opposite effects of hyperinsulinaemia on glucose utilization in white adipose tissue and muscles persisted when the hypoglycaemia-induced catecholamine output was prevented (adrenomedullectomy, propranolol treatment), as well as when hypoglycaemia was normalized by concomitant insulin treatment and glucose infusion. Insulin suppressed hepatic glucose production during the clamps in insulin-treated rats as in the respective controls, whereas total hepatic lipid synthesis and liver fat content were greater in rats treated with insulin than in controls. It is concluded that hyperinsulinaemia itself could be one of the driving forces responsible for producing increased glucose utilization by white adipose tissue, increased total lipid synthesis with fat accumulation in adipose tissue and the liver, together with an insulin-resistant state at the muscular level.     

Expression of the Dan gene during chicken embryonic development.

The Dan gene was first identified as the putative rat tumor suppressor gene and encodes a protein structurally related to Cerberus and Gremlin in vertebrates. Xenopus DAN, as with Cerberus and Gremlin, was demonstrated to block bone morphogenetic protein (BMP) signaling by binding BMPs, and to be capable of inducing additional anterior structures by ectopic overexpression in Xenopus embryos. DAN, thus, is suggested to play pivotal roles in early patterning and subsequent organ development, as in the case of other BMP antagonists. In this report, we isolated the chicken counterpart of Dan. Chicken Dan is mainly expressed in the cephalic and somitic mesoderm and several placodes during organ development.     

Expression of the EMILIN-1 gene during mouse development.

Expression of EMILIN-1, the first member of a newly discovered family of extracellular matrix genes, has been investigated during mouse development. EMILIN-1 mRNA is detectable in morula and blastocyst by RT-PCR. First expression of the gene is found by in situ hybridization in ectoplacental cone in embryos of 6.5 days and in extraembryonic visceral endoderm at 7.5 days. The allantois is also labeled. Staining of ectoplacental cone-derived secondary trophoblast giant cells and spongiotrophoblast is strong up to 11.5 days and then declines. In the embryo, high levels of mRNA are initially expressed in blood vessels, perineural mesenchyme and somites at 8.5 days. Later on, intense labeling is identified in the mesenchymal component of organs anlage (i.e. lung and liver) and different mesenchymal condensations (i.e. limb bud and branchial arches). At late gestation staining is widely distributed in interstitial connective tissue and smooth muscle cell-rich tissues. The data suggest that EMILIN-1 may have a function in placenta formation and initial organogenesis and a later role in interstitial connective tissue.     

Expression of Wnt signalling pathway genes during tooth development.

We have carried out comparative in situ hybridisation analysis of six Wnt genes Wnts-3, -4, -5a, -6, -7b, and 10b together with Wnt receptor MFz6 and receptor agonist/antagonists MFrzb1 and Mfrp2 during murine odontogenesis from the earliest formation of the epithelial thickening to the early bell stage. Expression of Wnt-4, Wnt-6, and one Wnt receptor MFz6 was observed in the facial, oral and dental epithelium. Wnt10b was localised specifically to the presumptive dental epithelium. Wnts-3 and -7b were expressed in oral epithelium but showed no expression in the presumptive dental epithelium. Wnt-3 also showed no expression in the epithelial cells of the molar bud stage tooth germs, but showed restricted expression in the enamel knots which are signalling centres believed to be involved in regulating tooth shape. Wnts -6, -10b and MFz6 were also detected in the primary and secondary enamel knots. Wnt-5a and agonist/antagonists MFrzb1 and Mfrp2 were expressed in a graded proximo-distal (P-D) manner in mesenchymal cells during the early stages of tooth development with no overlying expression in the oral or dental epithelium. Wnt-5a and MFrzb1 show strong expression in the dental papilla mesenchyme.     

Expression of gap junction genes during postnatal neural development.

The timing of appearance of mRNAs encoding gap junction proteins was examined during development of the rat and mouse brain. Complementary DNAs (cDNAs) specific for the mRNA for the liver-type gap junction protein, connexin32, and the heart-type gap junction protein, connexin43, were used to probe Northern blots of total RNA isolated from the forebrain and hindbrain of mice and rats at various times before and after birth. Prior to postnatal day 10, connexin32 mRNA is detectable only at low levels. By postnatal days 10 to 16, a sharp increase occurs in the level of this mRNA. This increase is detectable first in the hindbrain, and subsequently in the forebrain. In contrast, connexin43 mRNA is readily detectable at birth, and the level of this mRNA also increases during subsequent development. The developmental appearance of the gap junction proteins, connexin32 and connexin43, was similar to that of their respective mRNAs. These results indicate that the genes encoding connexin32 and connexin43 are differentially expressed during neural development.     

Expression of the disconnected gene during development of Drosophila melanogaster.

Proper development of the larval visual nerve, Bolwig's nerve, of Drosophila melanogaster requires the wild type function of the disconnected (disco) gene. In disco mutants, the nerve does not make stable connections with its targets in the larval brain. We have begun to explore the role of disco in the formation of the nervous system by examining the distribution of disco mRNA and protein in embryos and third instar larvae using in situ hybridization and antibody staining respectively. No differences between the distribution patterns of the two products are detected; disco is expressed in many tissues including both neural and non-neural cells. Many of the cells which express disco undergo extensive movement during development as they participate in major morphogenetic movements. Antibody staining shows that the protein is found in the cell nucleus. Products of the disco gene are detected in cells near the terminus of the growing Bolwig's nerve. In embryos homozygous for either of two mutant alleles of disco, the disco protein is absent near the nerve terminus, although protein distribution elsewhere is indistinguishable from wild type.     

Expression of the T-box gene Eomesodermin during early mouse development.

We report the expression pattern of a murine homolog of the Xenopus laevis T-box gene Eomesodermin. mEomes expression is first detected in the extra-embryonic ectoderm prior to gastrulation, and persists there until head-fold stages. In the embryo proper, mEomes is expressed throughout the early primitive streak, nascent mesoderm and in the anterior visceral endoderm. Although mEomes expression disappears from the embryo at late-streak stages, a second domain of mEomes expression is observed in the telencephalon beginning around E10.5.