Prospectively isolated CD133/CD24-positive ependymal cells from the adult spinal cord and lateral ventricle wall differ in their long-term in vitro self-renewal and in vivo gene expression.

In contrast to ependymal cells located above the subventricular zone (SVZ) of the adult lateral ventricle wall (LVW), adult spinal cord (SC) ependymal cells possess certain neural stem cell characteristics. The molecular basis of this difference is unknown. In this study, antibodies against multiple cell surface markers were applied to isolate pure populations of SC and LVW ependymal cells, which allowed a direct comparison of their in vitro behavior and in vivo gene expression profile. Isolated CD133(+)/CD24(+)/CD45(-)/CD34(-) ependymal cells from the SC displayed in vitro self-renewal and differentiation capacity, whereas those from the LVW did not. SC ependymal cells showed a higher expression of several genes involved in cell division, cell cycle regulation, and chromosome stability, which is consistent with a long-term self-renewal capacity, and shared certain transcripts with neural stem cells of the embryonic forebrain. They also expressed several retinoic acid (RA)-regulated genes and responded to RA exposure. LVW ependymal cells showed higher transcript levels of many genes regulated by transforming growth factor-ß family members. Among them were Dlx2, Id2, Hey1, which together with Foxg1 could explain their potential to turn into neuroblasts under certain environmental conditions.

Tissue-specific regulatory network extractor (TS-REX): a database and software resource for the tissue and cell type-specific investigation of transcription factor-gene networks.

The prediction of transcription factor binding sites in genomic sequences is in principle very useful to identify upstream regulatory factors. However, when applying this concept to genomes of multicellular organisms such as mammals, one has to deal with a large number of false positive predictions since many transcription factor genes are only expressed in specific tissues or cell types. We developed TS-REX, a database/software system that supports the analysis of tissue and cell type-specific transcription factor-gene networks based on expressed sequence tag abundance of transcription factor-encoding genes in UniGene EST libraries. The use of expression levels of transcription factor-encoding genes according to hierarchical anatomical classifications covering different tissues and cell types makes it possible to filter out irrelevant binding site predictions and to identify candidates of potential functional importance for further experimental testing. TS-REX covers ESTs from H. sapiens and M. musculus, and allows the characterization of both presence and specificity of transcription factors in user-specified tissues or cell types. The software allows users to interactively visualize transcription factor-gene networks, as well as to export data for further processing. TS-REX was applied to predict regulators of Polycomb group genes in six human tumor tissues and in human embryonic stem cells.

CD133 is not present on neurogenic astrocytes in the adult subventricular zone, but on embryonic neural stem cells, ependymal cells, and glioblastoma cells.

Human brain tumor stem cells have been enriched using antibodies against the surface protein CD133. An antibody recognizing CD133 also served to isolate normal neural stem cells from fetal human brain, suggesting a possible lineage relationship between normal neural and brain tumor stem cells. Whether CD133-positive brain tumor stem cells can be derived from CD133-positive neural stem or progenitor cells still requires direct experimental evidence, and an important step toward such investigations is the identification and characterization of normal CD133-presenting cells in neurogenic regions of the embryonic and adult brain. Here, we present evidence that CD133 is a marker for embryonic neural stem cells, an intermediate radial glial/ependymal cell type in the early postnatal stage, and for ependymal cells in the adult brain, but not for neurogenic astrocytes in the adult subventricular zone. Our findings suggest two principal possibilities for the origin of brain tumor stem cells: a derivation from CD133-expressing cells, which are normally not present in the adult brain (embryonic neural stem cells and an early postnatal intermediate radial glial/ependymal cell type), or from CD133-positive ependymal cells in the adult brain, which are, however, generally regarded as postmitotic. Alternatively, brain tumor stem cells could be derived from proliferative but CD133-negative neurogenic astrocytes in the adult brain. In the latter case, brain tumor development would involve the production of CD133.

Tracheal remodeling: comparison of different composite cultures consisting of human respiratory epithelial cells and human chondrocytes.

The reconstruction of extensive tracheal defects is still an unsolved challenge for thoracic surgery. Tissue engineering is a promising possibility to solve this problem through the generation of an autologous tracheal replacement from patients' own tissue. Therefore, this study investigated the potential of three different coculture systems, combining human respiratory epithelial cells and human chondrocytes. The coculture systems were analyzed by histological staining with alcian blue, immunohistochemical staining with the antibodies, 34betaE12 and CD44v6, and scanning electron microscopy. The first composite culture consisted of human respiratory epithelial cells seeded on human high-density chondrocyte pellets. For the second system, we used native articular cartilage chips as base for the respiratory epithelial cells. The third system consisted of a collagen membrane, seeded with respiratory epithelial cells and human chondrocytes onto different sides of the membrane, which achieved the most promising results. In combination with an air-liquid interface system and fibroblast-conditioned medium, an extended epithelial multilayer with differentiated epithelial cells could be generated. Our results suggest that at least three factors are necessary for the development towards a tracheal replacement: (1) a basal lamina equivalent, consisting of collagen fibers for cell-cell interaction and cell polarization, (2) extracellular factors of mesenchymal fibroblasts, and (3) the presence of an air-liquid interface system for proliferation and differentiation of the epithelial cells.

Definition of genetic events directing the development of distinct types of brain tumors from postnatal neural stem/progenitor cells.

Although brain tumors are classified and treated based upon their histology, the molecular factors involved in the development of various tumor types remain unknown. In this study, we show that the type and order of genetic events directs the development of gliomas, central nervous system primitive neuroectodermal tumors, and atypical teratoid/rhabdoid-like tumors from postnatal mouse neural stem/progenitor cells (NSC/NPC). We found that the overexpression of specific genes led to the development of these three different brain tumors from NSC/NPCs, and manipulation of the order of genetic events was able to convert one established tumor type into another. In addition, loss of the nuclear chromatin-remodeling factor SMARCB1 in rhabdoid tumors led to increased phosphorylation of eIF2α, a central cytoplasmic unfolded protein response (UPR) component, suggesting a role for the UPR in these tumors. Consistent with this, application of the proteasome inhibitor bortezomib led to an increase in apoptosis of human cells with reduced SMARCB1 levels. Taken together, our findings indicate that the order of genetic events determines the phenotypes of brain tumors derived from a common precursor cell pool, and suggest that the UPR may represent a therapeutic target in atypical teratoid/rhabdoid tumors.