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Comparative studies of the marginal band and plasma membrane of the epidermis
Abstract:
The membranous fraction isolated from stratum corneum by 8M urea-beta ME containing alkaline buffer (pH 9.0) is quite crude when observed by electron microscopy. However, this procedure may be useful for clinical samples, as one can isolate and compare both the soluble (interfilamentous) fraction and the keratin filament from the same sample in addition to the residues (membranous fraction). A further purified membranous fraction was isolated by a new method. Human stratum corneum was chopped and treated with 8M urea-50 mM Tris-HCl (pH 9.0), digested by the use of trypsin, and the product fractionated by a sucrose density gradient to obtain separate single cells without the cytoplasm. One sample was then treated with trypsin for 1 hour and another with urea buffer for 24 hours. Observations revealed a thickened inner membrane (marginal band) of approximately 150A. Each of the membranous samples contained a level of half-cystine markedly higher in amount (around 100/1,000) and involved mostly in the epsilon-(gamma-glutamyl) lysine cross-linkages (around 30%). In order to compare the membranous fraction of horny and living cells (marginal bands and plasma membranes), the fraction was then isolated from living cells. The relative amino acid composition of the membranous fraction of the plasma membrane resembled that of human erythrocytes, but was quite different from that of the marginal band. These comparative studies of biochemical and morphological features suggested the importance of S-S cross-linking enzymes and transglutaminase in the transformation mechanism of the marginal band.
Insights
Researchers developed a new method to isolate and analyze the membranous fraction of stratum corneum cells. This technique reveals insights into the marginal band
Area of Science:
- Biochemistry
- Cell Biology
- Dermatology
Background:
- The stratum corneum's membranous fraction is crucial for skin barrier function.
- Existing isolation methods yield crude fractions, limiting detailed analysis.
- Understanding the composition of cell membranes in horny and living cells is essential.
Purpose of the Study:
- To develop a refined method for isolating the membranous fraction from human stratum corneum.
- To characterize the biochemical and morphological features of the isolated membranous fraction.
- To compare the membranous fractions of horny cells (marginal bands) and living cells (plasma membranes).
Main Methods:
- Human stratum corneum was treated with urea and trypsin, followed by sucrose density gradient fractionation.
- Isolated single cells were further processed with trypsin or urea buffer.
- Membranous fractions from both horny and living cells were analyzed for amino acid composition and morphology.
Main Results:
- A purified membranous fraction was obtained, revealing a thickened inner membrane (marginal band) of approximately 150A.
- The membranous fraction showed a high content of half-cystine, primarily involved in epsilon-(gamma-glutamyl) lysine cross-linkages.
- The amino acid composition of the plasma membrane fraction differed significantly from that of the marginal band.
Conclusions:
- The new isolation method provides a purified membranous fraction for detailed study.
- The findings highlight the distinct biochemical properties of marginal bands compared to plasma membranes.
- S-S cross-linking enzymes and transglutaminase likely play key roles in the marginal band's transformation mechanism.