Lamellar Body-Enriched Fractions from Neonatal Mice: Preparative Techniques and Partial Characterization

Abstract

Several problems have frustrated the isolation of lamellar bodies (LB) from mammalian epidermis. We obtained pellets enriched in intact LB by utilizing the staphylococcal epidermolytic toxin to provide intact, outer epidermal sheets, by controlled homogenization in a cell disrupter, and by passage of homogenates through a graded series of nuclepore filters (Science 221:962, 1983). Such preparations contained more intact LB than did fractions prepared by a variety of differential or sucrose/metrizamide discontinuous centrifugation methods. Initial characterization of the enzymatic content of this fraction revealed it to be enriched in certain hydrolytic enzymes (acid phosphatase, carboxypeptidase, cathepsin B, acid lipase, sphingomyelinase, and phospholipase A), but strikingly depleted in all sulfatases, beta-glucuronidase, and the non-lysosomal protease, plasminogen activator. Thus, LB show some properties of lysosomes, although certain characteristic lysosomal enzymes are strikingly absent. Lamellar body fractions contained 2-3 times more lipid per unit weight than did homogenates, and were enriched in phospholipids, free sterols, and glycosphingolipids, but not in other neutral lipids or ceramides. In summary, whereas some of the enzymes in LB could participate in the metabolism of LB lipid precursors to hydrophobic barrier constituents, others may attack intercellular constituents, ultimately resulting in desquamation. The lipid profile of these organelles suggests that they deliver precursors of permeability barrier lipids to intercellular domains.

Inhibitors of 3-hydroxy-3-methylglutaryl coenzyme A reductase induce reductase accumulation and altered lamellar bodies in rat forestomach keratinocytes.

“…2130 a major site of steroidogenesis, 20 and 15-30% of the mass of epidermal lamellar bodies is composed of sterols or their derivatives. 31 - 32 As expected from these observations, disruption of the murine epidermal permeability barrier leads to increased keratinocyte sterol synthesis during barrier recovery. 33 - 35 In addition, certain epidermal alterations (neutral lipid storage disease and essential fatty acid deficiency) are characterized by keratinocyte hyperproliferation, increased levels of corneocyte desquamation, and breakdown of the epidermal permeability barrier.…”

High Throughput Quantitative Glycomics and Glycoform-focused Proteomics of Murine Dermis and Epidermis

“…LGs are a major source of stratum corneum lipid precursors and various hydrolytic enzymes such as lipases, proteases, acid phosphatases, and glycosidases (27,29,30) and other proteins including corneodesmosin (31). Among lysosomal glycoproteins identified in this study, glucosylceramidase and cathepsin D have been reported to be present in LGs (29,31).…”

“…Among lysosomal glycoproteins identified in this study, glucosylceramidase and cathepsin D have been reported to be present in LGs (29,31). Although further investigation is required, the high abundance of high mannose type oligosaccharides observed in epidermis may be attributable to the striking roles of lysosomal enzymes and/or the high abundance of lamellar granule in epidermis.…”

Ontogeny of the Epidermal Barrier to Water Loss in the Rat: Correlation of Function with Stratum Corneum Structure and Lipid Content

“…6) rather than into broad continuous lamellar bilayer unit structures. However, a failure to process lamellar body contents after secretion is likely to be too simplistic an interpretation for these observations because stratum corneum of 19-d pups was not enriched in glycosphingolipids or phospholipids as one would predict for unprocessed lamellar body-derived lipids (23,24). Coincident with establishment of a functional bamer, the quantity of lipid per unit area of stratum corneum increased -2-fold (Fig.…”

“…Lamellar bodies are enriched in phospholipids, free sterols, glucosylceramides, and a selected complement of hydrolytic enzymes, including lipases, glycosidases, and proteases (22)(23)(24)(25). Upon secretion of lamellar body contents at the stratum granulosum/stratum corneum interface, these enzymes apparently convert the polar lipids and glycosphingolipids to neutral lipids and ceramides.…”