SCL, GATA-2 and Lmo2 expression in neurogenesis.

SCL, Lmo2 and GATA factors form common transcription complexes during hematopoietic differentiation. The overlapping expression of SCL with GATA-2 and GATA-3 in the developing brain indicated that these factors might collaborate also in the course of neural tissue differentiation. The expression pattern of Lmo2 in the developing CNS, however, is not well understood. Here, we show that neural cells in the early embryonic chick mid- and hindbrain express SCL and GATA-2, while Lmo2 is expressed only in vascular elements. The lack of Lmo2 transcripts in neural cells demonstrated that SCL and GATA-2 cannot form common complexes with Lmo2 in the developing brain. In the course of neural tissue genesis, GATA-2 mRNA appeared prior to the SCL transcript. While GATA-2 expression decreased with maturation, SCL expression persisted at a high level also in post-neurogenic periods. The temporal pattern of SCL and GATA-2/3 expression was investigated also in vitro, in the course of induced neurogenesis by NE-4C neural stem cells. While GATA-2 expression increased from the very beginning of differentiation, SCL expression appeared only in more differentiated cells expressing proneural genes. GATA-3 expression, on the other hand, was detected only in advanced stages of the neuronal maturation, which were characterised by the activation of the Math2 neuronal gene. Similarly to the hematopoietic differentiation, GATA-2 expression precedes the activation of both SCL and GATA-3, and may play roles in the activation of the SCL gene in neuronal development. In contrast to hematopoietic differentiation, however, our results failed to demonstrate co-assembling of GATA factors or SCL with Lmo2. While overlapping expression of GATA-2/3 and SCL was detected, Lmo2 activation could not be demonstrated in neural cells in the investigated period of neuronal development.

In vitro pattern formation during neurogenesis in neuroectodermal progenitor cells immortalized by p53-deficiency.

In vitro neural differentiation was induced in a p53-deficient immortalized neuroectodermal progenitor cell line, NE-4C, by treatment with retinoic acid [K. Schlett and E. Madarász (1997) J. Neurosci. Res. 47, 405-416]. Rearrangement of nestin filaments was an early marker of neuron-formation. The increase in neurofilament protein content was accompanied by a decrease in the expression of nestin filaments in induced precursors. Cells with astroglial features appeared with a delay of 4-5 days compared to the appearence of neurons. Future neurons were sorted out from the substrate-attached population of apparently non-induced cells. The sorting out of future neurons resembled the separation of neural precursors in vivo. The continuous changes in the shape and also in the position of the cells resulted in the formation of characteristic morphological patterns. On the basis of morphological changes, five characteristic stages of in vitro neural differentiation were distinguished. The analysis of the morphological changes revealed that cell-to-cell interactions played an essential role in the cell fate decision made by induced precursors. Our observations indicate that the NE-4C cell line can serve as an in vitro model to investigate some early steps of neurogenesis.

Fate of cloned embryonic neuroectodermal cells implanted into the adult, newborn and embryonic forebrain.

NE-4C, one-cell derived neuroectodermal stem cells expressing a reporter gene--green fluorescent protein (GFP) or heat-resistant alkaline phosphatase (PLAP)--or prelabeled with bromodeoxyuridine (BrdU) were implanted into the forebrain of adult, new-born and fetal mice and into the mid- and forebrain vesicles of early chick embryos. The fate of implanted cells in the mouse and chick hosts was followed up to 6 and 2 weeks, respectively. Neural differentiation was monitored by detecting the expression of neuron-specific markers and GFAP. NE-4C cells integrated into the early embryonic brain tissue and developed into morphologically differentiated neurons. The same cells produced expanding tumor-like aggregates in the newborn forebrain and were expelled from the adult forebrain parenchyma. In the adult brain, long-term survival and integration of stem cells were revealed only in neurogenic zones. The data suggest that noncommitted, proliferating neuroectodermal progenitors can integrate into the brain tissue at time and site of tissue genesis.