Effects of the extracellular matrix on fetal choroid plexus epithelial cells: changes in morphology and multicellular organization do not affect gene expression.
We have developed a primary culture system for fetal mouse choroid plexus epithelial cells which maintains their differentiated phenotype. When grown on a reconstituted basement membrane substrate (Matrigel) epithelial cells formed aggregates which became embedded in the matrix and developed into characteristic and highly reproducible multicellular vesicular structures. These vesicles consisted of a squamous layer of epithelial cells with extensive attachment to the matrix substrate, surrounding a fluid-filled lumen. Electron microscopy showed that cells comprising these vesicles had a high degree of membrane specialization and polarized morphology which in many respects mimicked the in vivo morphology. Biochemical analyses demonstrated that under these culture conditions the tissue-specific pattern of gene expression of fetal choroid plexus epithelium was maintained. After 6 days in culture these cells contained approximately the same amount of transthyretin mRNA as the 12.5-day choroid plexus in vivo, and the level of total RNA per cell, which is proportional to the protein synthetic capability of the cells, was also maintained. The pattern of protein secretion was also very similar to that generated by fetal mouse choroid plexus cells in vivo. In contrast choroid plexus epithelial cells attached poorly to collagen I gels. Heterogeneous aggregates were formed in which cell-cell interactions were more extensive than cell-substrate interactions, and in no cases was a central lumen observed. Cells on the surface of large aggregates showed some evidence of membrane polarization, while the majority of cells in the cultures exhibited little evidence of polarized morphology. Despite the striking difference in morphology and multicellular organization these cells still expressed high levels of transthyretin mRNA and maintained the same pattern of protein synthesis as cells cultured on Matrigel. These results indicate that the basement membrane is important for the organization of choroid plexus epithelial cells into a functional epithelium in vitro and thus presumably the maintenance of the integrity of the blood-brain barrier in vivo. In contrast to several other epithelial systems which have been studied, the type of extracellular matrix does not appear to directly influence tissue-specific gene expression by choroid plexus epithelial cells. Thus the level of gene expression is not dependent on the cytoarchitecture and multicellular organization of this cell type.
Thomas
T
T
Centre for Early Human Development, Monash Medical Centre, Victoria, Australia.
Stadler
E
E
Dziadek
M
M
D000818:Animals; D001483:Base Sequence; D001672:Biocompatible Materials; D015152:Blotting, Northern; D002448:Cell Adhesion; D002452:Cell Count; D002478:Cells, Cultured; D002831:Choroid Plexus / Q000166:cytology* / Q000502:physiology*; D003094:Collagen; D015342:DNA Probes; D004338:Drug Combinations; D004847:Epithelial Cells; D004848:Epithelium / Q000502:physiology / Q000648:ultrastructure; D005109:Extracellular Matrix / Q000502:physiology*; D005333:Fetus; D015870:Gene Expression; D007700:Kinetics; D007797:Laminin; D008715:Methionine / Q000378:metabolism; D051379:Mice; D008854:Microscopy, Electron; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000096:biosynthesis; D015345:Oligonucleotide Probes; D011228:Prealbumin / Q000096:biosynthesis / Q000235:genetics; D011509:Proteoglycans; D012313:RNA / Q000235:genetics / Q000302:isolation & purification; D012333:RNA, Messenger / Q000378:metabolism; D013997:Time Factors
D001672:Biocompatible Materials; D015342:DNA Probes; D004338:Drug Combinations; D007797:Laminin; D009419:Nerve Tissue Proteins; D015345:Oligonucleotide Probes; D011228:Prealbumin; D011509:Proteoglycans; D012333:RNA, Messenger; C059485:matrigel; D012313:RNA; D003094:Collagen; D008715:Methionine