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Biomedical subjects

E Stadler

Publications and source records attributed to E Stadler.

14 recordsLinked to original sources
PubMed1996

Extracellular matrix penetration by epithelial cells is influenced by quantitative changes in basement membrane components and growth factors.

We have previously shown that isolated mouse fetal choroid plexus epithelial (CPE) cells penetrate a basement membrane matrix (Matrigel) substrate in vitro to form single-layered epithelial vesicles embedded within the matrix. To determine which properties of the matrix are important for inducing or permitting cells to penetrate the substrate and organize into multicellular vesicles we have made quantitative changes to the basement membrane components and growth factors in cell cultures. Matrigel diluted to 33 or 10% with a collagen I gel was not permissive to cell invasion, and CPE cells formed a polarized epithelial monolayer on the substrate surface which had ultrastructural characteristics similar to those of CPE vesicles. Cells in these monolayers proliferated more rapidly than cells in epithelial vesicles. When deliberately embedded within a 33 or 10% Matrigel matrix, CPE cells were able to form vesicles, indicating that a dilute matrix is nonpermissive to cell invasion but promotes epithelial polarization and organization into vesicles. Cells embedded within a 100% collagen I matrix did not proliferate or form epithelial vesicles and the majority of cells did not remain viable. Addition of laminin to the collagen I gel promoted cell adhesion and cell survival, but did not promote the formation of extensive monolayers on the substrate nor the formation of epithelial vesicles within the matrix. Cell invasion into the 33% Matrigel matrix was induced by addition of laminin, nidogen, or a laminin-nidogen complex to the substrate or by addition of TGFbeta2 to the culture medium, but not TGFbeta1 or PDGF. These studies show that CPE cells are sensitive to quantitative changes in matrix composition, which influences their survival and proliferation and also their ability to penetrate the matrix and organize into multicellular epithelial vesicles.

Animals↗
PubMed1996

Electrochemical Properties of

[Ru(edta)(H2O)]- is strongly adsorbed on a zirconium(IV) oxide-coated silica gel surface. The immobilized complex showed an electrochemical response due to the Ru(II)/Ru(III) redox couple. By substituting the coordinated water molecule in the adsorbed complex, the midpoint potentials shifted in the order (in mV) water, -290; thiocyanate, -200; pyridine, -180; 4-cyanopyridine, -80; and pyrazine, -50 vs SCE.

Journal Article↗
PubMed1992

Three-dimensional structure of the Fab fragment of a neutralizing antibody to human rhinovirus serotype 2.

The crystal structure of the antigen-binding fragment of a monoclonal antibody (8F5) that neutralizes human rhinovirus serotype 2 has been determined by X-ray diffraction studies. Antibody 8F5, obtained by immunization with native HRV2 virions, cross-reacts with peptides of the viral capsid protein VP2, which contribute to the neutralizing immunogenic site B in this serotype. The structure was solved by the molecular replacement method and has been refined to an R-factor of 18.9% at 2.8 A resolution. The elbow angle, relating the variable and constant modules of the molecule is 127 degrees, representing the smallest elbow angle observed so far in an Fab fragment. Furthermore, the charged residues of the epitope can be well accommodated in the antigen-binding site. This is the first crystal structure reported for an antibody directed against an icosahedral virus.

Amino Acid Sequence↗
PubMed1992

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.

Animals↗
PubMed1989

Do cultured vascular smooth muscle cells resemble those of the artery wall? If not, why not?

Ultrastructural stereological techniques were used to quantitate the phenotype of vascular smooth muscle cells grown in primary culture on a plastic substrate (control), on and within matrices of collagen type I, and on collagen type III, collagen type IV, or basement membrane Matrigel. The volume fraction of myofilaments (Vv myo) of freshly isolated cells at day 0 was 54.2 +/- 2.0%. By day 5 in culture, the Vv myo of cells grown on plastic, on collagen type I, and collagen type III had decreased significantly to 14.2 +/- 2.1%, 16.5 +/- 2.4%, and 16.3 +/- 2.9%, respectively. In contrast, smooth muscle cells grown within matrices of collagen type I, on collagen type IV, or on basement membrane Matrigel had a Vv myo of 30.7 +/- 1.5%, 30.8 +/- 4.2%, and 32.1 +/- 3.1%, respectively. These findings demonstrate that the presence of extracellular matrix components prevents modulation of smooth muscle phenotype early in culture and suggests that the extracellular matrix synthesized and secreted by smooth muscle in the normal vessel wall contributes to the maintenance of their contractile functional state.

Animals↗
PubMed1977

Purification and characterization of animal porphobilinogen synthases. I. Bovine liver porphobilinogen synthase.

Porphobilinogen synthase was purified from ox liver by ammonium sulfate fractionation, heat denaturation and column chromatography (purification: 400-fold; specific activity 4.72 nkat). The molecular weight of the native enzyme obtained by thin-layer gel filtration is about 280 000. Using 8M urea in the presence of dithiothreitol as reducing agent, the molecule breaks down into 8 subunits of molecular weight 36 000 (dodecylsulfate gel electrophoresis); the preparation of aminoethylated subunit is described. According to the above-mentioned molecular weight and to the above-mentioned molecular weight and to the quantitative amino acid analysis after total hydrolysis, the following compositon of the enzymes subunit was calculated ASX23-25 Thr7 Ser23-24 Glx29-31 Pro22-23 Gly22-24 Ala36-37 Val23-26 Met7 Ile9 Leu34-35 Tyr10 Phe11-12 Lys11-12 Cys6-7 His6-8 Arg22 Trp1-2. The subunits, having two free sulfhydryl groups, therefore consists of a chain of about 306 amino acids. The Dansyl-Edman procedure did not enable identification of any free N-terminal amino acid. The acyl group blocking the N-terminus is an acetyl group. It was identified, after hydrazinolysis of the enzyme, by means of chromatographic comparison with 1-formyl-2-dansyl-hydrazine and 1-acetyl-2-dansylhydrazine, whose syntheses and UV spectra are described.

Amino Acids↗