Clonal origin of the mammalian forebrain from widespread oriented mixing of early regionalized neuroepithelium precursors

The forebrain is formed by remodeling and growth of the anterior neural plate. This morphogenesis occurs in response to inductive signals during gastrulation and neurulation but is poorly understood at the cellular level. Here, we have used the LaacZ method of single cells labeling to visualize, at E12.5, clones originated at early stages of mouse forebrain development. The largest clones show that single progenitors can give rise to neuroepithelial cells dispersed across the forebrain. A significant fraction of the clones, and even relatively small ones, populated both the diencephalon and the telencephalon, indicating that the clonal separation between diencephalic and telencephalic progenitors is transient and/or partial. However, two groups of large clones, populating either the diencephalon or the telencephalon, dispersed within their respective domains, suggesting an early regionalization between some diencephalic and telencephalic progenitors. Widespread oriented mixing within these territories and then clonal expansion into smaller domains probably follow this initial regionalization. These data are consistent with a model of progressive specification of forebrain domains. We propose that the ordered expansion of early regionalized progenitor pools for the diencephalon and telencephalon could establish a potential link between early inductive signals and forebrain morphogenesis.     

Regionalization of cell fates and cell movement in the endoderm of the mouse gastrula and the impact of loss of Lhx1(Lim1) function

Investigation of the developmental fates of cells in the endodermal layer of the early bud stage mouse embryo revealed a regionalized pattern of distribution of the progenitor cells of the yolk sac endoderm and the embryonic gut. By tracing the site of origin of cells that are allocated to specific regions of the embryonic gut, it was found that by late gastrulation, the respective endodermal progenitors are already spatially organized in anticipation of the prospective mediolateral and anterior-posterior destinations. The fate-mapping data further showed that the endoderm in the embryonic compartment of the early bud stage gastrula still contains cells that will colonize the anterior and lateral parts of the extraembryonic yolk sac. In the Lhx1(Lim1)-null mutant embryo, the progenitors of the embryonic gut are confined to the posterior part of the endoderm. In particular, the prospective anterior endoderm was sequestered to a much smaller distal domain, suggesting that there may be fewer progenitor cells for the anterior gut that is poorly formed in the mutant embryo. The deficiency of gut endoderm is not caused by any restriction in endodermal potency of the mutant epiblast cells but more likely the inadequate allocation of the definitive endoderm. The inefficient movement of the anterior endoderm, and the abnormal differentiation highlighted by the lack of Sox17 and Foxa2 expression, may underpin the malformation of the head of Lhx1 mutant embryos.     

Genetic interaction of Gsc and Dkk1 in head morphogenesis of the mouse

Mouse embryos lacking Gsc and Dkk1 function display severe deficiencies in craniofacial structures which are not found in either Dkk1 homozygous null or Gsc homozygous null mutant embryos. Loss of Gsc has a dosage-related effect on the severity of head truncation phenotype in Dkk1 heterozygous embryos. The synergistic effect of these mutations in enhancing head truncation provides direct evidence of a genetic interaction between Gsc and Dkk1, which display overlapping expression in the prechordal mesoderm. In the absence of Gsc activity, the expression of Dkk1, WNT genes and a transgenic reporter for WNT signalling are altered. Our results show that Gsc and Dkk1 functions are non-redundant in the anterior mesendoderm for normal anterior development and Gsc may influence Wnt signalling as a negative regulator.     

Regulation of anchor cell invasion and uterine cell fates by the egl-43 Evi-1 proto-oncogene in Caenorhabditis elegans

Cell invasion is a tightly controlled process occurring during development and tumor progression. The nematode Caenorhabditis elegans serves as a genetic model to study cell invasion during normal development. In the third larval stage, the anchor cell in the somatic gonad first induces and then invades the adjacent epidermal vulval precursor cells. The homolog of the Evi-1 oncogene, egl-43, is necessary for basement membrane destruction and anchor cell invasion. egl-43 is part of a regulatory network mediating cell invasion downstream of the fos-1 proto-oncogene. In addition, EGL-43 is required to specify the cell fates of ventral uterus cells downstream of or in parallel with LIN-12 NOTCH. Comparison with mammalian Evi-1 suggests a conserved pathway controlling cell invasion and cell fate specification.     

Immunocytochemical localization of cell type-specific markers in reaggregating cell cultures of mouse cerebellum

Summary Reaggregate cultures were obtained from single-cell suspensions of fetal and early postnatal cerebellum, and fetal telencephalon and mesencephalon from C57BL/6J and NMRI mice and maintained in suspension under constant rotation as described previously (Seeds 1971). The percentage of dead cells in the aggregates as measured by the uptake of the fluorescent dye propidium iodide was always less than 5% of all cells. During the initial phase of reaggregation up to 20 h in vitro (hiv) several immunocytochemically defined cell types had a random distribution within the aggregate. Astrocytes were identified by indirect immunofluorescence by the use of the markers glial fibrillary acidic protein (GFAP), C1 and M1 antigens; neurons by NS-4 antigen and tetanus-toxin receptors; fibroblasts or fibroblast-like cells by fibronectin and laminin; and oligodendrocytes by myelin basic protein (MBP). Choleratoxin receptors and M 2 antigen served to distinguish the more mature from the less mature neurons. In reaggregates of early postnatal cerebellar cells neurons had started to redistribute after 40 hiv, forming an outer region containing more immature neurons and a core with more mature neurons. After 5 days in vitro (div) immature neurons were no longer detectable. From 3–8 div M1-and GFAP-positive astrocytic processes in the outer region showed a tendency for radial orientation. At later stages the processes appeared more randomly distributed and formed a dense glial network. Few oligodendrocytes and fibronectin-positive cells were present in the reaggregates. When reaggregates were prepared from 15 day-old embryonic cerebella, formation of radially oriented astrocytic processes and redistribution of neurons proceeded more slowly, but in a similar pattern as described for early postnatal cerebellum. GFAP was detectable at earlier ages than in situ. In reaggregates of 15 to 17 day old embryonic telencephalic anlage or midbrain, radially oriented astrocytic processes were not detectable. Similar to cerebellar reaggregates, accumulation of neurons in the inner region was observed.     

Marker genes identify three somatic cell types in the fetal mouse ovary

The two main functions of the ovary are the production of oocytes, which allows the continuation of the species, and secretion of female sex hormones, which control many aspects of female development and physiology. Normal development of the ovaries during embryogenesis is critical for their function and the health of the individual in later life. Although the adult ovary has been investigated in great detail, we are only starting to understand the cellular and molecular biology of early ovarian development. Here we show that the adult stem cell marker Lgr5 is expressed in the cortical region of the fetal ovary and this expression is mutually exclusive to FOXL2. Strikingly, a third somatic cell population can be identified, marked by the expression of NR2F2, which is expressed in LGR5- and FOXL2 double-negative ovarian somatic cells. Together, these three marker genes label distinct ovarian somatic cell types. Using lineage tracing in mice, we show that Lgr5-positive cells give rise to adult cortical granulosa cells, which form the follicles of the definitive reserve. Moreover, LGR5 is required for correct timing of germ cell differentiation as evidenced by a delay of entry into meiosis in Lgr5 loss-of-function mutants, demonstrating a key role for LGR5 in the differentiation of pre-granulosa cells, which ensure the differentiation of oogonia, the formation of the definitive follicle reserve, and long-term female fertility.     

Fluorescent latex microparticles: A non-invasive short-term cell lineage marker suitable for use in the mouse early embryo

Summary We have examined the potential of fluorescent latex microparticles for use as a short term cell lineage marker in the mouse preimplantation embryo. Isolated blastomeres and intact embryos rapidly adsorb and subsequently endocytose the particles (0.2 m diameter) from a monodisperse suspension in normal medium, so that cytoplasmic endocytic organelles, but not the cytosol itself, becomes labelled. Latex fluorescence, either within intact embryos, disaggregated cells or thick resin sections, is stable during UV irradiation. The development of labelled embryos, both in terms of sequential morphological changes and their time of expression, was comparable to controls and resulted in blastocysts with normal cell numbers and capacity for tissue differentiation. Latex fluorescence is preserved within all the progeny of labelled blastomeres over several cell cycles (e.g. from 8-cell stage to 64-cell stage) and is not transmitted to unlabelled cells either by exocytosis or via midbodies. The particles are particularly suitable for labelling exclusively the entire population of outside cells in the intact embryo from the 16-cell stage onwards.     

Association of matrix metalloproteinase-9 and Purkinje cell degeneration in mouse cerebellum caused by Angiostrongylus cantonensis

Angiostrongylosis is a neurological disorder caused by invasion of the central nervous system by developing larvae of Angiostrongylus cantonensis. Purkinje cells in infected mouse cerebellums are small and irregular with degenerative atrophy or partial loss. Ultrastructural changes in degenerative cells included enlarged vacuolar structures and swollen mitochondria within the cytoplasm. The matrix metalloproteinase-9 mRNA which is low in normal cerebellums was expressed in A. cantonensis-infected mice cerebellum prior to Purkinje cell degeneration. Matrix metalloproteinase-9 protein level and enzyme activity increased when the Purkinje cells appeared degenerated. Using immunohistochemistry, matrix metalloproteinase-9 was localised within degenerative Purkinje cells. In addition, when the specific matrix metalloproteinase inhibitor, GM6001, was added, matrix metalloproteinase-9 enzyme activity was reduced by 41.6%. The numbers of degenerative Purkinje cells increased significantly upon establishment of infection but subsided upon inhibition. These results suggested that the expression of matrix metalloproteinase-9 may be associated with degeneration of Purkinje cells in mouse cerebellum infected by A. cantonensis.     

Comparative analysis of embryonic cell lineage between Caenorhabditis briggsae and Caenorhabditis elegans

Comparative genomic analysis of important signaling pathways in Caenorhabditis briggsae and Caenorhabditis elegans reveals both conserved features and also differences. To build a framework to address the significance of these features we determined the C. briggsae embryonic cell lineage, using the tools StarryNite and AceTree. We traced both cell divisions and cell positions for all cells through all but the last round of cell division and for selected cells through the final round. We found the lineage to be remarkably similar to that of C. elegans. Not only did the founder cells give rise to similar numbers of progeny, the relative cell division timing and positions were largely maintained. These lineage similarities appear to give rise to similar cell fates as judged both by the positions of lineally equivalent cells and by the patterns of cell deaths in both species. However, some reproducible differences were seen, e.g., the P4 cell cycle length is more than 40% longer in C. briggsae than that in C. elegans (p < 0.01). The extensive conservation of embryonic development between such divergent species suggests that substantial evolutionary distance between these two species has not altered these early developmental cellular events, although the developmental defects of transpecies hybrids suggest that the details of the underlying molecular pathways have diverged sufficiently so as to not be interchangeable.