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Biomedical subjects

B E Brown

Publications and source records attributed to B E Brown.

At least 19 recordsLinked to original sources
PubMed2007

The impact of glycation on apolipoprotein A-I structure and its ability to activate lecithin:cholesterol acyltransferase.

AIMS/HYPOTHESIS: Hyperglycaemia, one of the main features of diabetes, results in non-enzymatic glycation of plasma proteins, including apolipoprotein A-I (apoA-I), the most abundant apolipoprotein in HDL. The aim of this study was to determine how glycation affects the structure of apoA-I and its ability to activate lecithin:cholesterol acyltransferase (LCAT), a key enzyme in reverse cholesterol transport. MATERIALS AND METHODS: Discoidal reconstituted HDL (rHDL) containing phosphatidylcholine and apoA-I ([A-I]rHDL) were prepared by the cholate dialysis method and glycated by incubation with methylglyoxal. Glycation of apoA-I was quantified as the reduction in detectable arginine, lysine and tryptophan residues. Methylglyoxal-AGE adduct formation in apoA-I was assessed by immunoblotting. (A-I)rHDL size and surface charge were determined by non-denaturing gradient gel electrophoresis and agarose gel electrophoresis, respectively. The kinetics of the LCAT reaction was investigated by incubating varying concentrations of discoidal (A-I)rHDL with a constant amount of purified enzyme. The conformation of apoA-I was assessed by surface plasmon resonance. RESULTS: Methylglyoxal-mediated modifications of the arginine, lysine and tryptophan residues in lipid-free and lipid-associated apoA-I were time- and concentration-dependent. These modifications altered the conformation of apoA-I in regions critical for LCAT activation and lipid binding. They also decreased (A-I)rHDL size and surface charge. The rate of LCAT-mediated cholesterol esterification in (A-I)rHDL varied according to the level of apoA-I glycation and progressively decreased as the extent of apoA-I glycation increased. CONCLUSIONS/INTERPRETATION: It is concluded that glycation of apoA-I may adversely affect reverse cholesterol transport in subjects with diabetes.

Apolipoprotein A-I↗
PubMed2006

Hydrazine compounds inhibit glycation of low-density lipoproteins and prevent the in vitro formation of model foam cells from glycolaldehyde-modified low-density lipoproteins.

AIMS/HYPOTHESIS: Previous studies have shown that glycation of LDL by methylglyoxal and glycolaldehyde, in the absence of significant oxidation, results in lipid accumulation in macrophage cells. Such 'foam cells' are a hallmark of atherosclerosis. In this study we examined whether LDL glycation by methylglyoxal or glycolaldehyde, and subsequent lipid loading of cells, can be inhibited by agents that scavenge reactive carbonyls. Such compounds may have therapeutic potential in diabetes-associated atherosclerosis. MATERIALS AND METHODS: LDL was glycated with methylglyoxal or glycolaldehyde in the absence or presence of metformin, aminoguanidine, Girard's reagents P and T, or hydralazine. LDL modification was characterised by changes in mobility (agarose gel electrophoresis), cross-linking (SDS-PAGE) and loss of amino acid residues (HPLC). Accumulation of cholesterol and cholesteryl esters in murine macrophages was assessed by HPLC. RESULTS: Inhibition of LDL glycation was detected with equimolar or greater concentrations of the scavengers over the reactive carbonyl. This inhibition was structure-dependent and accompanied by a modulation of cholesterol and cholesteryl ester accumulation. With aminoguanidine, Girard's reagent P and hydralazine, cellular sterol levels returned to control levels despite incomplete inhibition of LDL modification. CONCLUSIONS/INTERPRETATION: Inhibition of LDL glycation by interception of the reactive aldehydes that induce LDL modification prevents lipid loading and model foam cell formation in murine macrophage cells. Carbonyl-scavenging reagents, such as hydrazines, may therefore help inhibit LDL glycation in vivo and prevent diabetes-induced atherosclerosis.

Acetaldehyde↗
PubMed2005

Glycation of low-density lipoproteins by methylglyoxal and glycolaldehyde gives rise to the in vitro formation of lipid-laden cells.

AIMS/HYPOTHESIS: Previous studies have implicated the glycoxidative modification of low-density lipoprotein (LDL) by glucose and aldehydes (apparently comprising both glycation and oxidation), as a causative factor in the elevated levels of atherosclerosis observed in diabetic patients. Such LDL modification can result in unregulated cellular accumulation of lipids. In previous studies we have characterized the formation of glycated, but nonoxidized, LDL by glucose and aldehydes; in this study we examine whether glycation of LDL, in the absence of oxidation, gives rise to lipid accumulation in arterial wall cell types. METHODS: Glycated LDLs were incubated with macrophage, smooth muscle, or endothelial cells. Lipid loading was assessed by HPLC analysis of cholesterol and individual esters. Oxidation was assessed by cholesterol ester loss and 7-ketocholesterol formation. Cell viability was assessed by lactate dehydrogenase release and cell protein levels. RESULTS: Glycation of LDL by glycolaldehyde and methylglyoxal, but not glucose (in either the presence or absence of copper ions), resulted in cholesterol and cholesterol ester accumulation in macrophage cells, but not smooth muscle or endothelial cells. The extent of lipid accumulation depends on the degree of glycation, with increasing aldehyde concentration or incubation time, giving rise to greater extents of particle modification and lipid accumulation. Modification of lysine residues appears to be a key determinant of cellular uptake. CONCLUSIONS/INTERPRETATION: These results are consistent with LDL glycation, in the absence of oxidation, being sufficient for rapid lipid accumulation by macrophage cells. Aldehyde-mediated "carbonyl-stress" may therefore facilitate the formation of lipid-laden (foam) cells in the artery wall.

Acetaldehyde↗
PubMed2001

Hypochlorite- and hypobromite-mediated radical formation and its role in cell lysis.

Activated leukocytes generate the potent oxidants HOCl and HOBr via the formation of H(2)O(2) and the release of peroxidase enzymes (myeloperoxidase, eosinophil peroxidase). HOCl and HOBr are potent microbiocidal agents, but excessive or misplaced production can cause tissue damage and cell lysis. In this study it is shown that HOBr induces red blood cell lysis at approximately 10-fold lower concentrations than HOCl, whereas with monocyte (THP1) and macrophage (J774) cells HOCl and HOBr induce lysis at similar concentrations. The role of radical formation during lysis has been investigated by EPR spin trapping, and it is shown that reaction of both oxidants with each cell type generates cell-derived radicals. Red blood cells exposed to nonlytic doses of HOCl generate novel nitrogen-centered radicals whose formation is GSH dependent. In contrast, HOBr gives rise to nitrogen-centered, membrane-derived protein radicals. With lytic doses of either oxidant, protein (probably hemoglobin)-derived, nitrogen-centered radicals are observed. Unlike the red blood cells, treatment of monocytes and macrophages with HOCl gives significant radical formation only under conditions where cell lysis occurs concurrently. These radicals are nitrogen-centered, cell-protein-derived species and have parameters identical to those detected with red blood cells and HOBr. Exposure of these cells to HOBr did not give detectable radicals. Overall these experiments demonstrate that HOCl and HOBr react with different selectivity with cellular targets, and that this can result in radical formation. This radical generation can precede, and may play a role in, cell lysis.

Bromates↗
PubMed2001

Desmoglein isoform distribution affects stratum corneum structure and function.

Desmogleins are desmosomal cadherins that mediate cell-cell adhesion. In stratified squamous epithelia there are two major isoforms of desmoglein, 1 and 3, with different distributions in epidermis and mucous membrane. Since either desmoglein isoform alone can mediate adhesion, the reason for their differential distribution is not known. To address this issue, we engineered transgenic mice with desmoglein 3 under the control of the involucrin promoter. These mice expressed desmoglein 3 with the same distribution in epidermis as found in normal oral mucous membranes, while expression of other major differentiation molecules was unchanged. Although the nucleated epidermis appeared normal, the epidermal stratum corneum was abnormal with gross scaling, and a lamellar histology resembling that of normal mucous membrane. The mice died shortly after birth with severe dehydration, suggesting excessive transepidermal water loss, which was confirmed by in vitro and in vivo measurement. Ultrastructure of the stratum corneum showed premature loss of cohesion of corneocytes. This dysadhesion of corneocytes and its contribution to increased transepidermal water loss was confirmed by tape stripping. These data demonstrate that differential expression of desmoglein isoforms affects the major function of epidermis, the permeability barrier, by altering the structure of the stratum corneum.

Animals↗
PubMed1998

The secretory granular cell: the outermost granular cell as a specialized secretory cell.

The contents of epidermal lamellar bodies (LB) are delivered selectively to the intercellular spaces at the stratum granulosum (SG)-stratum corneum (SC) interface. We assessed the subcellular basis for LB secretion first by confocal microscopy, following labeling with Nile red or NBD-ceramide, which reveals a tubulo-reticular membrane system within the apical cytosol of the outermost SG cell layer under basal conditions, changing to a more peripheral staining pattern when secretion is stimulated. Ultrastructural study demonstrates that this network is composed of a widely disbursed trans-Golgi-like network (TGN), associated with arrays of contiguous LB, and deep invaginations of the SG-SC interface. Under basal conditions, limited fusion of apically directed LB leads to deep, interconnected invaginations of the apical plasma membrane, resulting in the formation of an extensive, honeycomb extension of the SG-SC interface. Still deeper invaginations and more extensive organelle fusion develop after the epidermis is acutely permeabilized by either acetone treatment, sonophoresis, or iontophoresis. Finally, nascent LB appear to bud off cisternae of the TGN, a process that appears to accelerate after barrier disruption. The deep invaginations of the SG-SC interface; the wide distribution of the TGN within the apical cytosol; the association of nascent LB with the TGN; and the rapid fusion of LB with these invaginations, deep within the cytosol, account for (i) the polarized secretion of LB from the apex of the outermost SG cell, and (ii) the rapid LB-secretory response to barrier perturbations. Finally, our results point to the outermost SG cell as a uniquely specialized secretory cell. We propose the term "secretory granulocyte" to encompass the specialized features of these cells.

4-Chloro-7-nitrobenzofurazan↗
PubMed1998

Glucosylceramides stimulate mitogenesis in aged murine epidermis.

Glucosylceramides (GlcCer) and ceramides (Cer) appear to have opposite effects on epidermal growth and differentiation. Whereas Cer inhibit mitosis and induce terminal differentiation and apoptosis in cultured keratinocytes, GlcCer is mitogenic in young murine epidermis. Using a recently described murine model of chronologic senescence we explored whether GlcCer is mitogenic in aged epidermis. Epidermal GlcCer content increases following topical applications of either conduritol-B epoxide (CBE), an inhibitor of GlcCer hydrolysis, or exogenous GlcCer in a penetration-enhancing vehicle. During chronologic aging in the hairless mouse, baseline epidermal DNA synthesis rates remain normal until 18 mo, but decline significantly at 24 mo. Topical CBE stimulates a 1.5- to 1.9-fold increase in epidermal DNA synthesis in all age groups (i.e., 1-2, 18, and 24 mo). Although the CBE induced increase in [3H]thymidine incorporation in 24 mo old animals is significant (p < 0.01), it is not sufficient to reach the absolute levels reached in similarly treated, younger mouse epidermis. Moreover, topical GlcCer induced mitogenesis is both dose dependent and hexose specific in young (1-2 mo old) animals, and remains effective in aged (< or = 24 mo old) animals. Furthermore, the CBE induced increase in DNA synthesis in aged epidermis is sufficient to produce epidermal hyperplasia. Finally, although an increased GlcCer:Cer ratio can alter stratum corneum barrier function and membrane structure, neither stratum corneum function nor extracellular membrane structure change under these experimental conditions, and therefore the mitogenic effects of increased epidermal GlcCer cannot be attributed to effects on the stratum corneum. These results show that: (i) elevations in endogenous GlcCer are mitogenic for aged as well as young murine epidermis; (ii) topical GlcCer is also mitogenic when delivered in an enhancing vehicle; and (iii) despite the putative importance of epidermal DNA synthesis for barrier homeostasis, these mitogenic alterations do not alter stratum corneum function.

Administration, Topical↗
PubMed1997

Epidermal injury stimulates prenylation in the epidermis of hairless mice.

Isoprenylation is the covalent attachment of isoprenyl groups, intermediates of the cholesterol biosynthesis pathway, to carboxyl terminal cysteine residues of proteins. Numerous proteins are isoprenylated including small GTP binding proteins, trimeric G proteins, and nuclear lamins, and these prenylated proteins regulate a variety of cell functions, including cell growth, cytokinesis, and differentiation. Here, we quantitated protein prenylation and determined which proteins are prenylated in the epidermis of hairless mice by radiolabeling with 3H-mevalonolactone following acute or chronic epidermal injury. In normal epidermis, four major radiolabeled bands, with molecular weights of 17-26, 48, 54, and 68 kDa, were observed. The levels of each of these bands increased by 24-63% 16 h following acute epidermal injury induced by topical acetone treatment or tape stripping, returning to normal by 24 h. On 2D gel electrophoresis, there were no major differences between the patterns of labeling following barrier disruption. Subacute epidermal injury induced by either acetone or tape stripping twice a day for 7 days and chronic injury induced by feeding an essential fatty acid-deficient (EFAD) diet, also resulted in a significant increase in protein prenylation. As with an acute injury, SDS-PAGE and 2D gel electrophoresis did not reveal marked differences in the pattern of protein prenylation. These results demonstrate that the prenylation of proteins in the epidermis is stimulated by injury, suggesting that one or more of these prenylated species may be important in epidermal proliferation or differentiation.

Alkyl and Aryl Transferases↗
PubMed1996

Are there task specific performance effects for differently configured numeric keypads?

Previous studies of numeric keypad user preference and performance have indicated that the telephone layout (TEL) was superior to the layout seen on computer keyboards and adding machines (ADD). A recent study (Straub and Granaas, 1993) suggested that the TEL preference was subject to task specific effects. To investigate the possibility of task specific performance in using keypads, 24 subjects were tested on four different keypad layouts (TEL, zero at top; TEL, zero at bottom; ADD, zero at top; ADD, zero at bottom) using three different tasks (four digit strings, seven digit strings, and seven digit strings depicted like standard North American telephone numbers). Results indicated that differences in rate of performance across the four keypad layouts were the result of zero placement, with the zero in the bottom position yielding the fastest keypad use. No significant differences were found for error rate across the different keypads. No task specific performance effects were found. These findings suggest that either the ADD or TEL layouts could be adopted universally for numeric keypads, with the stipulation that the zero key be placed below the other keys.

Journal Article↗
PubMed1996

Decreased epidermal lipid synthesis accounts for altered barrier function in aged mice.

The epidermis of aged mice displays decreased stratum corneum (SC) lipid content and decreased extracellular bilayers, which result in impaired barrier recovery following the solvent treatment or tape stripping. We assessed the role of altered lipid synthesis as the cause of the abnormal barrier and lipid content in aged epidermis, both under basal conditions and in response to acute barrier perturbations. In aged epidermis ( > or = 18 months), synthesis of one of the three key lipid classes (cholesterol) is decreased under basal conditions, and sterologenesis fails to attain the levels reached in young epidermis following comparable acute perturbations. In contrast, fatty acid and sphingolipid synthesis in aged epidermis increase sufficiently to approach the levels attained in stimulated young epidermis. The abnormalities in sterologenesis in aged epidermis are paralleled by a decrease in activity of its rate-limiting enzyme, 3-hydroxy-3-methylglutaryl-coenzyme A reductase, under basal conditions, and enzyme activity also fails to increase as much as in young epidermis after barrier disruption. That defective lipid generation contributes to the barrier defect is shown directly by the ability of either a cholesterol-containing mixture of SC lipids or cholesterol alone to enhance barrier recovery. Finally, lipid-induced acceleration of barrier recovery in aged epidermis correlates with repletion of the extracellular spaces with normal lamellar structures. Thus, a deficiency in lipid synthesis, particularly in cholesterologenesis, accounts for the barrier abnormality in aged epidermis.

Aging↗
PubMed1995

Exogenous nonphysiologic vs physiologic lipids. Divergent mechanisms for correction of permeability barrier dysfunction.

BACKGROUND AND DESIGN: Although barrier function requires cholesterol, free fatty acids, and ceramides, applications of one or two of these lipids to damaged skin impedes barrier recovery, while equimolar mixtures allow normal recovery. Both incomplete and complete mixtures appear to be internalized within the epidermal nucleated layers, followed by the secretion of abnormal vs normal lamellar body contents, respectively. We compared the ability of complete physiologic lipid mixtures vs a nonmetabolized hydrophobic lipid, petrolatum, to repair the barrier and the requirement for intracellular processing of these lipids within the epidermis. RESULTS: Neat petrolatum, which remains restricted to the stratum corneum, produces more rapid improvement in barrier function than the solvent-dispersed physiologic lipids, and its effects are not altered by coapplication of either monensin or brefeldin A (both from Sigma Chemical Co, St Louis, Mo), known inhibitors of exocytosis and organellogenesis, respectively. In contrast, the physiologic lipids enter the nucleated layers in substantial amounts and require longer to produce barrier recovery. Whereas monensin blocks their ability to facilitate barrier recovery, the physiologic lipids overcome brefeldin A-induced delays in barrier recovery, bypassing the subcellular site of brefeldin A blockade, normalizing both lamellar body contents and intercellular bilayers. CONCLUSIONS: While petrolatum remains restricted to the stratum corneum, physiologic lipid mixtures influence barrier recovery after transport to subjacent, nucleated layers, followed by internalization, apparent transport to the distal Golgi apparatus, and incorporation into nascent lamellar bodies.

Animals↗
PubMed1995

The aged epidermal permeability barrier. Structural, functional, and lipid biochemical abnormalities in humans and a senescent murine model.

Aged epidermis displays altered drug permeability, increased susceptibility to irritant contact dermatitis, and often severe xerosis, suggesting compromise of the aged epidermal barrier. To delineate the functional, structural, and lipid biochemical basis of epidermal aging, we compared barrier function in young (20-30 yr) vs aged (> 80 yr) human subjects, and in a murine model. Baseline transepidermal water loss in both aged humans and senescent mice was subnormal. However, the aged barrier was perturbed more readily with either acetone or tape stripping (18 +/- 2 strippings vs 31 +/- 5 strippings in aged vs young human subjects, respectively). Moreover, after either acetone treatment or tape stripping, the barrier recovered more slowly in aged than in young human subjects (50 and 80% recovery at 24 and 72 h, respectively, in young subjects vs 15% recovery at 24 h in aged subjects), followed by a further delay over the next 6 d. Similar differences in barrier recovery were seen in senescent vs young mice. Although the total lipid content was decreased in the stratum corneum of aged mice (approximately 30%), the distribution of ceramides (including ceramide 1), cholesterol, and free fatty acids was unchanged. Moreover, a normal complement of esterified, very long-chain fatty acids was present. Finally, stratum corneum lamellar bilayers displayed normal substructure and dimensions, but were focally decreased in number, with decreased secretion of lamellar body contents. Thus, assessment of barrier function in aged epidermis under basal conditions is misleading, since both barrier integrity and barrier repair are markedly abnormal. These functional changes can be attributed to a global deficiency in all key stratum corneum lipids, resulting in decreased lamellar bilayers in the stratum corneum interstices. This constellation of findings may explain the increased susceptibility of intrinsically aged skin to exogenous and environmental insults.

Acetone↗
PubMed1992

Exogenous origin of n-alkanes in pathologic scale.

BACKGROUND: Although n-alkanes accumulate in some disorders of cornification, recent studies using radioactive carbon 14 content by accelerator mass spectrometry point to an exogenous origin for alkanes in normal stratum corneum, and their derivation in congenital ichthyosiform erythroderma remains controversial. DESIGN AND RESULTS: Using 14C content to measure sample age, the n-alkane fractions from two patients with congenital ichthyosiform erythroderma contained no detectable contemporary materials. By electron microscopy, alkane-enriched emollients (petrolatum [Vaseline]) permeated to all levels of stratum corneum of hairless mice, expanding the intercellular domains and distorting membrane bilayers. Similar ultrastructural changes were also observed in the stratum corneum of patients with congenital ichthyosiform erythroderma. When alkanes were excluded, no differences in lipid content were evident between two forms of autosomal recessive ichthyosis. CONCLUSIONS: These data demonstrate that scale n-alkanes in disorders of cornification derive from environmental sources and indicate the pervasiveness of petroleum-based emollients in skin. Therefore, epidermal lipid analyses must be interpreted with caution. However, these studies do not rule out an important therapeutic and/or pathogenic role for exogenous n-alkanes in skin.

Adult↗
PubMed1991

The lovastatin-treated rodent: a new model of barrier disruption and epidermal hyperplasia.

Recent studies have linked epidermal cholesterol synthesis with maintenance of the permeability barrier. To assess directly the importance of cholesterol synthesis, we applied lovastatin, a potent inhibitor of cholesterol synthesis, to hairless mouse skin. Transepidermal water loss (TEWL) began to increase after four to six daily applications. Co-application of cholesterol blocked the expected increase in TEWL, demonstrating the importance of cholesterol for development of the lesion. The histology of lovastatin-treated skin revealed epidermal hyperplasia, accompanied by accelerated DNA synthesis. Whereas cholesterol synthesis initially was reduced in lovastatin-treated epidermis, with further treatment cholesterol synthesis normalized, while fatty acid synthesis accelerated greatly. Although the total free sterol content of lovastatin-treated epidermis remained normal, the fatty acid content increased coincident with barrier disruption. Finally, morphologic abnormalities of both lamellar body structure and their deposited, intercellular contents occurred coincident with the emerging biochemical abnormalities. Thus, the abnormal barrier function in this model can be ascribed to an initial inhibition of epidermal sterol synthesis followed by an alteration in cholesterol and fatty acid synthesis, leading to an imbalance in stratum corneum lipid composition and abnormal membrane bilayer structure.

Animals↗
PubMed1990

Effect of prior heat shock on heat resistance of Listeria monocytogenes in meat.

The effect of prior heat shock on the thermal resistance of Listeria monocytogenes in meat was investigated. A sausage mix inoculated with approximately 10(7) L. monocytogenes per g was initially subjected to a heat shock temperature of 48 degrees C before being heated at a final test temperature of 62 or 64 degrees C. Although cells heat shocked at 48 degrees C for 30 or 60 min did not show a significant increase in thermotolerance as compared with control cells (non-heat shocked), bacteria heat shocked for 120 min did, showing an average 2.4-fold increase in the D64 degrees C value. Heat-shocked cells shifted to 4 degrees C appeared to maintain their thermotolerance for at least 24 h after heat shock.

Colony Count, Microbial↗
PubMed1990

Cholesterol synthesis is required for cutaneous barrier function in mice.

Previous studies have shown that topical acetone treatment results in the removal of stratum corneum lipids and disruption of the permeability barrier. This disruption stimulates epidermal lipid synthesis which is associated with the rapid restoration of stratum corneum lipids and barrier function. The aim of this study was to determine the role of cutaneous cholesterol synthesis in the barrier recovery. Here we show that topical lovastatin, a competitive inhibitor of HMG CoA reductase, inhibits cholesterol synthesis. After acetone disruption of the barrier, the normal rapid return of cholesterol to the stratum corneum and recovery of barrier function is impaired in animals treated topically with lovastatin. When lovastatin animals are simultaneously treated topically with either mevalonate, the immediate product of HMG CoA reductase, or cholesterol, the final end product of the pathway, the recovery of the barrier is normalized. Lovastatin resulted in the delayed secretion and abnormal appearance of lamellar bodies. These results provide the first evidence demonstrating that cholesterol synthesis is required for the maintenance of barrier structure and function and suggests a crucial role for cholesterol synthesis in allowing for terrestrial existence.

Animals↗