R. Preston Mason

Evidence for changes in the Alzheimer's disease brain cortical membrane structure mediated by cholesterol

Small angle X-ray diffraction analysis of Alzheimers disease (AD) lipid membranes extracted from cortical gray matter showed significant, reproducible structure changes relative to age-matched control samples. Specifically, there was an average 4 A reduction in the lipid bilayer width and significant changes in the membrane electron density profiles of AD cortical samples. There were no significant structure differences in the membrane bilayers isolated from an unaffected region (cerebellum) of the AD brain. Lipid and protein analysis of 6 AD and 6 age-matched controls showed that the phospholipid:protein mass ratio was unchanged but that the unesterified cholesterol:phospholipid (C:PL) mole ratio decreased by 30% in the AD temporal gyrus relative to age-matched controls. By contrast, the C:PL mole ratio in the cerebellum did not change significantly. X-ray diffraction analysis of a cholesterol enriched AD sample demonstrated a virtual restoration of the normal membrane bilayer width and electron density profile, suggesting that the cholesterol deficit played a major role in the AD lipid membrane structure perturbation. Alterations in the composition and structure of the membrane bilayer may play an important role in the pathophysiology of AD by altering the activity and catabolism of membrane-bound proteins, including the beta-amyloid precursor protein.

Atherosclerosis alters the composition, structure and function of arterial smooth muscle cell plasma membranes

The object of this study was to examine changes in plasma membranes of arterial smooth muscle (ASM) during atherogenesis obtained from cholesterol-fed (2%) rabbits. A microsomal fraction highly enriched with plasma membrane markers was prepared by subcellular organelle fractionation from ASM freshly isolated from the thoracic aorta. The membranes were analyzed for unesterified (free) cholesterol (FC) content, membrane bilayer structural parameters (X-ray diffraction), phospholipid (PL) composition, and Na+/K(+)-ATPase activity and kinetics. Following 8 weeks on diet, membrane FC content increased 67.1%. Small angle X-ray diffraction demonstrated an increase in membrane hydrocarbon core electron density and an increase in overall lipid bilayer width (56-62 A). This increase in bilayer width was highly correlated with the membrane FC content (r = 0.992). Both membrane FC content And bilayer width independently correlated with time on cholesterol diet. The phospholipid profile of the membrane revealed a 16.4% increase in phosphatidylcholine (PC), 19.3% decrease in phosphatidylethanolamine (PE) and 62.8% increase in sphingomyelin (SM) content with no change in total PL content. Na+/K(+)-ATPase activity was decreased 52.2% (P < 0.005), and [3H]ouabain binding kinetics demonstrated a 27.6% decrease in maximum binding sites (Bmax) (P < 0.01) while the dissociation constant (Kd) remained unaltered. Membranes obtained from control ASM cells enriched with FC in culture demonstrated changes similar to those in atherosclerotic ASM membranes including an increase in membrane FC content, an increase in bilayer width, and a decrease in Na+/K(+)-ATPase activity with decreased ouabain Bmax. These data demonstrate marked compositional, structural and functional changes in ASM cell membrane characteristics in dietary atherosclerosis. These changes were highly correlated with cholesterol accumulation in the plasma membrane bilayer and were observed before the appearance of visible lesions. We suggest that these membrane defects may be linked with early atherogenesis.

A membrane defect in the pathogenesis of the Smith-Lemli-Opitz syndrome

The Smith-Lemli-Opitz syndrome (SLOS) is an often lethal birth defect resulting from mutations in the gene responsible for the synthesis of the enzyme 3β-hydroxy-steroid-Δ7-reductase, which catalyzes the reduction of the double bond at carbon 7 on 7-dehydrocholesterol (7-DHC) to form unesterified cholesterol. We hypothesize that the deficiency in cholesterol biosynthesis and subsequent accumulation of 7-DHC in the cell membrane leads to defective composition, organization, dynamics, and function of the cell membrane. Using skin fibroblasts obtained from SLOS patients, we demonstrate that the SLOS membrane has increased 7-DHC and reduced cholesterol content and abnormal membrane fluidity. X-ray diffraction analyses of synthetic membranes prepared to mimic SLOS membranes revealed atypical membrane organization. In addition, calcium permeability is markedly augmented, whereas membrane-bound Na+/K+ATPase activity, folate uptake, inositol-1,4,5-trisphosphate signaling, and cell proliferation rates are markedly suppressed. These data indicate that the disturbance in membrane sterol content in SLOS, likely at the level of membrane caveolae, directly contributes to the widespread tissue abnormalities in this disease.

Membrane interaction of calcium channel antagonists modulated by cholesterol: Implications for drug activity

The interactions of lipophilic calcium channel antagonists with the membrane lipid bilayer are complex and highly dependent on membrane composition and structure. Variability in membrane lipid composition (e.g. cholesterol content, acyl chain saturation) can dramatically affect the membrane partitioning of calcium channel antagonists. The membrane binding properties of these drugs did not correlate with traditional measurements of drug lipophilicity, such as a simple hydrocarbon system (Table 2). These data indicate the need for directly measuring the binding of drug molecules to membranes in order to understand better certain pharmacological parameters, including bioavailability, under both normal and pathological conditions in which membrane composition is altered. The interaction of certain lipophilic amphipathic drugs with the membrane lipid bilayer may be an important component of their overall receptor binding mechanism. Specifically, the membrane bilayer may serve to concentrate and orient these drug molecules with respect to a hydrophobic receptor site at the protein receptor/membrane bilayer interface. Thus, the design of drugs which target membrane bound receptors should take into consideration the interaction of the drug molecule with the membrane lipid compartment. This understanding of drug/membrane interactions may lead to the development of drugs with more desirable pharmacokinetics, greater efficacy, and reduced side effects.

Membrane interactions of a phosphomonoester elevated early in Alzheimer's disease

Phosphoserine (L-PS) is among several phosphomonoesters found to be elevated in autopsied Alzheimers disease (AD) brain tissue. To investigate the molecular interactions of L-PS with membrane lipid bilayers, small angle X-ray diffraction and high resolution differential scanning calorimetry (DSC) approaches were used with liposomes composed of lecithin and cholesterol. A one-dimensional electron density profile of a control dimyristoyl phosphatidylcholine (DMPC)/cholesterol lipid bilayer with a unit cell dimension of 52 A at 37 degrees C was generated from the X-ray diffraction data. Following incubation with 2.0 mM L-PS, a broad decrease in electron density +/- 4-12 A from the lipid bilayer center was observed concomitant with an increase in the width of the phospholipid headgroup electron density and a 3 A reduction in lipid bilayer width. The interactions of L-PS with DMPC lipid bilayers were concentration-dependent, highly affected by cholesterol content and reproduced in egg phosphatidylcholine/cholesterol liposomes. DSC analysis showed that millimolar (1.0-5.0 mM) L-PS levels decrease the phase transition cooperative unit size of DMPC liposomes in a highly concentration-dependent manner which was significantly greater in preparations containing cholesterol. The endotherm width at half-maximum doubled at 5.0 mM and 1.25 mM L-PS, respectively, for DMPC and DMPC/cholesterol liposomes. These data provide direct evidence that elevated phosphomonoester levels modulate the biophysical properties of the membrane lipid bilayer which may, in turn, lead to altered structure/function relationships in membranes during AD.

Interactions biophysiques membranaires de l???amlodipine et propri??t??s antioxydantes

Objectif: Evaluer les benefices potentiels de leffet antioxydant de certains medicaments dans le traitement des maladies cardiovasculaires notamment les atteintes des arteres coronaires et linsuffisance cardiaque, par la diminution des lesions cellulaires irreversibles dues a latteinte oxydative. Methodes: Laction antioxydante de certains inhibiteurs calciques a ete etudiee et correlee aux effets moleculaires membranaires du compose. Celle-ci a ete mesuree a laide de tests de liaison utilisant des radioligands et par calorimetrie a balayage differentiel a haute resolution. Resultats: Les resultats de ces experiences montrent un lien direct entre les actions antioxydantes des inhibiteurs calciques et leur affinite pour la double couche lipidique de la membrane ainsi que leur capacite a moduler les proprietes thermodynamiques de la membrane (amlodipine > verapamil » diltiazem). Lamlodipine, inhibiteur calcique 1,4-dihydropyridine, a montre la plus forte affinite membranaire (K p 10 4 ) et a entraine les plus fortes modifications des proprietes thermodynamiques de la membrane, notamment une reduction de la temperature de transition de phase thermique (-11%), de lenthalpie (-14%) et du nombre de molecules passant simultanement la transition de phase (-59%), par rapport au controle par liposomes phosphatidylcholines. Conclusions: Ces donnees montrent que les inhibiteurs calciques lipophiles inhibent la peroxydation lipidique dans les membranes cellulaires du fait de leurs proprietes physico-chimiques de modulation de la double couche lipidique des membranes, independamment de linhibition des canaux calciques. Leffet antioxydant des inhibiteurs calciques fortement lipophiles tels que lamlodipine, pourrait contribuer a de nouveaux mecanismes cytoprotecteurs dans les maladies cardiovasculaires.

Atherogenic Activity caused by Excess Membrane Cholesterol in Arterial Smooth Muscle: Role of Calcium Channels

The concept of elevated serum cholesterol level as a principal risk factor in the etiology of atherosclerotic lesions has evolved from several large-scale epidemiological studies (1,2,3). These studies clearly link increased serum low-density lipoprotein (LDL) levels to mortality due to myocardial infarction. LDL is a cholesterol-rich, apo B100 containing particle which has been implicated in the forward transport of cholesterol to peripheral cells, including the smooth muscle cells of the arterial wall, by way of the classical LDL receptor (4). While the role of this particle, and its ligand (apo B100)-receptor complex, in the central hepatic metabolism of cholesterol has been well studied, little is known of the mechanism by which elevated LDL levels initiate and maintain lesion development and progression in the arterial wall.

Cholesterol Enrichment During Dietary Atherosclerosis Alters Smooth Muscle Plasma Membrane Width and Structure: Evidence for Reversal by the 1,4-Dihydropyridine Amlodipine

Free (unesterified) cholesterol is the single most abundant lipid species found in smooth muscle cell (SMC) plasma membrane and appears to have an important role in its membrane structure and physiology (1). In dietary atherosclerosis, the arterial SMC plasma membrane free cholesterol: phospholipid (C:PL) mole ration increased by 83% from 0.38 to 0.78 (1). Cholesterol enrichment in SMCs have been associated with the pathobiology of atherosclerosis, including marked changes in SMC transmembrane movements of cations and SMC proliferation (1-4).

Interactions of Lacidipine and other Calcium Channel Drugs with Biological Membranes: A Structural Model for Receptor/Drug Binding Utilizing the Membrane Bilayer

The analysis of calcium channel drugs that bind to membrane-associated receptors must take into account the local environment where the binding event occurs. The partitioning of these drugs in an isotropic two-phase bulk solvent system, such as octanol/buffer, apparently is not a good model for their partitioning into model or in native membranes where they exert their effects. Knowledge of these membrane-based partition coefficients then necessitates reanalysis of other physical, chemical, and functional parameters.

Probing Molecular Sites of Action for Alcohol's Acute and Chronic Effects on Synaptoneurosome Membranes: A Potential Tool for Studying Drug-Receptor Interactions

Ethanol is active in the central nervous system (CNS), producing a variety of effects when administered acutely. Long-tenn administration of ethanol produces tolerance and physical dependence. The study of alcohols effects on the CNS has been complicated by the absence of a specific binding site that would indicate the primary locus of action for this molecule on the cells of the CNS. Unlike other drugs of abuse such as opiates, psychomotor stimulants and hallucinogens, there is no receptor molecule, reuptake site or ion channel that possesses high affinxad ity binding for ethanol. Historically, Meyer (1906) and Overton (1901) described the solubility of ethanol and its ability to reach rapid equilibxad rium between the intra- and extracellular environment to explain some of ethanols actions on cellular processes and the biophysical state of cells and cellular organelles. The seemingly diverse effects of ethanol on the eNS were explained by a common underlying mechanism: through the distribution of ethanol within the matrix of biological membranes and subsequent alterations in the structure and function of these membranes. The literature on the effects of ethanol and membrane structure/funcxad tion demonstrates little consensus. Using electron spin resonance of probes, several laboratories have defined an order parameter that indicated