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K C Madison

Publications and source records attributed to K C Madison.

32 records · Page 2Linked to original sources
PubMed1988

Isolation of corneocyte envelopes from porcine epidermis.

Sheets of porcine stratum corneum were dispersed into individual corneocytes after 4 h in a solution consisting of 8 mM N,N-dimethyldodecylamine oxide and 2 mM sodium dodecylsulfate in phosphate-buffered isotonic saline, at 45 degrees C. With continued detergent treatment and moderate sonication, most of the cells lost their keratin contents and were then separated from the remaining intact cells by centrifugation in cesium chloride solution of density 1.280. Electron microscopy showed that the cell envelopes retained both the crosslinked protein envelope and its attached lipid envelope. The dry weight of envelopes was approximately 7% of the estimated dry weight of the original stratum corneum, while the corneocytes surviving intact also amounted to 7% of the starting weight. Mild alkaline hydrolysis of the corneocyte envelopes allowed the extraction of hydroxyceramides amounting to 10% of the dry weight of the envelopes. The procedure therefore provides isolated corneocyte envelopes suitable for studying both the protein and lipid components of this compound sheath.

Animals↗
PubMed1988

Fatty acids of acylceramides from comedones and from the skin surface of acne patients and control subjects.

Comedonal lipids and skin surface lipids were collected from six acne patients and surface lipids were collected from sex- and age-matched controls without acne. Six series of ceramides were found in each sample, the relative amounts of which were determined by thin-layer chromatography/photodensitometry. Acylceramides (ceramide 1) were isolated by preparative thin-layer chromatography and their ester-linked fatty acids were analyzed by gas-liquid chromatography. The comedonal acylceramides contained higher proportions of 16:0, 16:1 delta 6, and 18:1 delta 6 + delta 8 and much less linoleate (18:2 delta 9,12) than the acylceramides from the skin surface. In the surface lipids from legs, acylceramides from the acne patients contained less linoleate than the acylceramides from control subjects. Free fatty acids from the comedones were also isolated and analyzed, and had a composition very similar to the esterified fatty acids of comedonal acylceramides. The results confirm that fatty acids derived from sebum become incorporated into comedonal acylceramides, displacing linoleate, and show that this process even affects the acylceramides of surface epidermis, more so in acne patients than in normal subjects.

Acne Vulgaris↗
PubMed1988

Lamellar granule extrusion and stratum corneum intercellular lamellae in murine keratinocyte cultures.

Lamellar granules are specialized epidermal organelles containing stacks of membranous disks that are extruded into the intercellular spaces in the upper portion of the granular layer. The extruded disks are believed to undergo biochemical and biophysical changes to form the stratum corneum intercellular lipid sheets that constitute the epidermal permeability barrier. Little is known about this important component of epidermal differentiation, in part due to lack of a suitable in vitro model. We have demonstrated microscopically the presence of characteristic lipid membrane structures in a primary keratinocyte culture system which shows morphologic differentiation comparable to that seen in vivo. A basal cell-enriched fraction of isolated neonatal mouse keratinocytes was plated into Vitrogen-coated 30 mm Millicell (Millipore, Bedford, Massachusetts) wells, fed daily with Medium 199 containing 10% fetal bovine serum, 10 micrograms/ml each of insulin and hydrocortisone, and kept at 32 degrees C in a 5% CO2/95% air atmosphere in a humidified incubator. Three days after plating, cultures were placed on living, epidermis-free mouse dermis at the air/liquid interface. At 2 wk, histologic examination showed multiple well-organized cell layers, including a distinct granular layer and a well-developed stratum corneum. Transmission electron microscopy demonstrated numerous lamellar granules and extrusion of their contents into the intercellular space. After fixation with ruthenium tetroxide, stacked intercellular lamellae in the stratum corneum were seen. Both the presence of dermis and growth at the air/liquid interface were necessary to achieve complete differentiation. This system conclusively demonstrates the formation of complex epidermal lipid structures in vitro and should allow the mechanisms and regulation of their synthesis to be elucidated.

Animals↗
PubMed1987

Evidence that the corneocyte has a chemically bound lipid envelope.

The stratum corneum of mammalian epidermis contains a mixture of ceramides, free fatty acids, cholesterol, and cholesteryl sulfate, amounting to 14% of the dry weight of the tissue, that can be removed by exhaustive extraction with chloroform/methanol. Subsequent mild alkaline hydrolysis liberates additional lipid, consisting almost exclusively of C30-C34 omega-hydroxyacids in amide linkage with sphingosine, equal to 2% of the tissue mass. In the present study, transmission electron microscopy was used to demonstrate that the initial extraction removes the intercellular lamellae that constitute the epidermal water barrier but leaves the lucent band that has been termed the corneocyte plasma membrane. The subsequent alkaline hydrolysis and lipid extraction remove the lucent band, which must therefore contain the omega-hydroxyacylsphingosines. From the results of in situ derivatization of these lipids and the construction of molecular models, it is inferred that the bound lipids exist in ester linkage with protein on the surface of the corneocyte envelope. The tightly packed hydroxyacylsphingosine molecules thus form a lipid envelope for each corneocyte.

Animals↗
PubMed1987

Presence of intact intercellular lipid lamellae in the upper layers of the stratum corneum.

The epidermal permeability barrier necessary for terrestrial life resides in the intercellular spaces of the stratum corneum and is composed of lipids. Membrane coating granules (MCGs), small intracellular organelles found in the uppermost layers of the living epidermis, contain stacks of membranous disks which are extruded into the intercellular space and undergo both biochemical and physical changes to form the lipid sheets which constitute this barrier. Using ruthenium tetroxide as a secondary fixative, we are able to demonstrate stacks of lamellae filling the intercellular spaces in the uppermost layers of the stratum corneum. The structure of these lipid lamellae is consistent with the proposed derivation of MCG lipid disks and also suggests that the lipid bilayer adjacent to the corneocyte cell envelope may be assembled from lipids not derived from MCGs.

Animals↗
PubMed1987

Composition and morphology of epidermal cyst lipids.

The contents of epidermal cysts were used as a source of desquamated human keratinocytes uncontaminated by sebaceous, subcutaneous, or bacterial lipids. Lipids extracted with chloroform:methanol mixtures included six series of ceramides (41% of the total extractable lipid), cholesterol (27%), cholesteryl esters (10%), fatty acids (9%), cholesteryl sulfate (1.9%), a novel class of ceramide esters (3.8%), and a sterol diester (0.9%). Electron microscopy revealed that the lipids in the cyst contents existed as multiple intercellular lamellae, as in stratum corneum. One lamella, adjacent to the horny cell protein envelope, was resistant to lipid extraction and is thought to represent covalently bound lipid on the outer surface of the keratinocyte. The results indicate that the degradation of intercellular lipid lamellae is not required for desquamation.

Ceramides↗
PubMed1987

Essential fatty acids and epidermal integrity.

The intercellular spaces of the stratum corneum contain multilamellar lipid sheets derived from the extruded contents of lamellar granules. In the absence of linoleic acid, lamellar granules appear empty, and only fragmentary extracellular sheets are found. This defective differentiation is attributable to substitution of oleate for linoleate in O-acylsphingolipids. Normally, linoleate is ester-linked to 30- to 34-carbon omega-hydroxyacids, which, in turn, are amide-linked to sphingosine. Acylglucosylceramides, bearing a beta-D-glucosyl moiety on the sphingosine, may provide the driving force for lamellar granule assembly. The omega-hydroxyacyl chains are long enough to span a lipid bilayer, while the linoleate inserts into an adjacent bilayer. This interaction could promote assembly of lamellar granules. It has also been proposed that acylceramides may stabilize the extracellular sheets by a similar mechanism. In addition, the horny cell has been found to possess a covalently bound lipid envelope consisting principally of omega-hydroxyacylsphingosines derived from O-acylsphingolipids.

Animals↗
PubMed1986

Lipid composition of cultured murine keratinocytes.

The current study was undertaken to determine whether a previously reported murine culture system is an acceptable model for the study of epidermal lipid metabolism. The lipid composition of primary neonatal mouse keratinocyte cultures was determined and compared with that of freshly isolated keratinocytes and whole epidermis. 14C-Labeled arachidonic acid (AA) and linoleic acid (LA) were added to cultures and the incorporation into specific lipids was assessed. The lipid composition of the cultures indicated that they were partially differentiated, which parallels the well-known incomplete keratinization seen in many keratinocyte culture systems. Of particular importance, the LA-rich uniquely epidermal lipids which may be of importance in water barrier function, acylglucosylceramide (AGC) and acylceramide (AC), were made by the cultures. Fatty acid analysis of total lipid, phospholipid, and AGC extracts revealed a significant decrease in LA content compared with the parent epidermis; this may have resulted from the low level of LA in fetal bovine serum, which was the serum source for these cultures. Labeled AA and LA were incorporated into the lipids of cultured keratinocytes in distinct patterns that were consistent with the fatty acid content of the lipids. Both AGC and AC showed preferential uptake of LA compared with AA. There was minimal labeling of non-linoleate-containing lipids and a low degree of conversion of labeled LA to AA. Considering the grossly different environment of the in vitro system compared with the in vivo state, the overall lipid composition was remarkably well maintained. Keratinocyte cultures should be of great value in the study of epidermal lipid metabolism.

Animals↗
PubMed1985

A survey of polar and non-polar lipids of mouse organs.

Total lipid was extracted from mouse (Mus musculus) heart, kidney, lung, liver, intestine, brain, stomach, dermis and epidermis and analyzed by quantitative thin-layer chromatography. All of the tissues contained phospholipids, triglycerides, sterols and free fatty acids. All tissues except brain contained small amounts of steryl esters, and all except stomach contained some glycosylceramides. Wax diesters were found in both the dermis and epidermis. Only epidermis contained a high proportion of ceramides. Acylglucosylceramides were uniquely present in epidermis.

Animals↗
PubMed1984

Regulation of the expression of epidermal keratinocyte proliferation and differentiation by vitamin A analogs.

A number of vitamin A analogs (retinoids) were used to manipulate the growth of epidermal keratinocytes in culture. The retinoids used were the TMMP analog of ethyl retinoate (Ro 10-9359), 13-cis retinoic acid, all trans retinoic acid and retinol (trans). These were added to primarily neonatal mouse epidermal keratinocyte cultures that proliferate, stratify, and differentiate over 2-3 weeks. [3H]Tdr labeling technics were used to quantitate proliferation. A histologic stain, and a four buffer protein extraction protocol, used in conjunction with polyacrylamide gel electrophoresis and fluorographic technics, were used to assess the differentiation of the cultures. Our results showed that all of the vitamin A analogs we tested inhibited keratinocyte proliferation. Quantitation of specific differentiation proteins showed that Ro 10-9359 and 13-cis retinoic acid partially inhibited the differentiation of the cultures. The Ro 10-9359 retinoid was unusual in that it increased the synthesis of keratohyalin granule-related proteins. These studies showed that inhibition of basal cell proliferation did not result in the obligatory expression of cell differentiation and that at least one of the events that is a part of epidermal keratinocyte differentiation can be separately controlled.

Animals↗
PubMed1984

Isotretinoin therapy is associated with early skeletal radiographic changes.

Eight patients with disorders of keratinization (six with ichthyosis, one with Darier's disease, and one with palmar-plantar keratoderma) were treated with isotretinoin for 9 months (1 patient) to 1 year (7 patients). The patients ranged from 5 to 26 years of age. The average isotretinoin dose was 2 mg/kg/day (range, 1.0-2.9 mg/kg/day). Radiographic skeletal surveys were performed prior to therapy, and after 6 months and 1 year of therapy. After 1 year of isotretinoin treatment, six of the eight patients had small but unequivocal skeletal hyperostoses. Five of the patients had multiple hyperostoses. While only two patients were judged to have hyperostoses after 6 months of isotretinoin therapy during prospective evaluation, retrospective comparison with the radiographs obtained after 1 year revealed skeletal hyperostoses after 6 months of treatment in an additional three patients. Between 6 months and 1 year of therapy, some of the hyperostoses remained unchanged while others had progressed. In three patients, hyperostoses were seen at 12 months that were not detectable at 6 months. Based on this prospective study of skeletal changes during isotretinoin therapy, we recommend that patients taking high doses of isotretinoin for long periods be monitored radiographically.

Adolescent↗
PubMed1984

Early skeletal hyperostoses secondary to 13-cis-retinoic acid.

Prolonged therapy with retinoid drugs (chemically similar to vitamin A) often results in skeletal hyperostoses, similar to those seen in idiopathic skeletal hyperostosis. Eight patients, aged 5-26 years, with dermatologic disorders were treated with 13-cis-retinoic acid. Skeletal surveys were obtained before and during treatment. In 1 year, six of the eight patients had developed such skeletal hyperostoses in both axial and appendicular regions. The cervical spine was the most common site of involvement. None of the children demonstrated accelerated skeletal maturation. Two of the patients had mild musculoskeletal discomfort during this period. The findings indicate that high-dose 13-cis-retinoic acid therapy may cause skeletal hyperostoses, requiring radiographic monitoring during prolonged periods of treatment. An implication of these observations, relating to the etiology of idiopathic skeletal hyperostosis, is discussed.

Adolescent↗
PubMed1975

Inhibition of lipolysis and cyclic AMP accumulation in white fat cells by tetracycline.

The effects of tetracycline on the metabolism of isolated rat white fat cells were examined. Tetracycline at a concentration of 0.05 mg/ml inhibited lipolysis due to 0.075 or 0.15 muM norepinephrine, but not that due to adenosine deaminase, theophylline, dibutyryl cyclic AMP or 1.5 muM norepinephrine. Higher concentrations of tetracycline (1 mg/ml) inhibited lipolysis due to all added agents except dibutyryl cyclic AMP. The accumulation of cyclic AMP after 5 minutes incubation with 0.15 muM norepinephrine plus adenosine deaminase was inhibited by 0.05 mg/ml of tetracycline. The large rise in cyclic AMP accumulation at 5 minutes due to 1.5 muM norepinephrine in the presence of 100 muM theophylline was only slightly inhibited by 0.05 or 0.1 mg/ml of tetracycline. Tetracycline at 1 mg/ml did markedly inhibit cyclic AMP accumulation due to all added agents. The stimulation of adenylate cyclase activity of fat cell ghosts by norepinephrine or fluoride was inhibited by 0.05 mg/ml or greater concentration of tetracycline. Insulin-stimulated glucose oxidation by fat cells was inhibited by 1 mg/ml of tetracycline. These results suggest that the anti-lipolytic action of tetracycline on rat fat cells is secondary to inhibition of cyclic AMP accumulation.

Adenylyl Cyclase Inhibitors↗