Biomechanical and biochemical investigation of erythrocytes in late stage human leptospirosis

Leptospirosis is a zoonotic disease caused by bacteria of the genus Leptospira, which can cause lipid changes in the erythrocyte membrane. Optical tweezers were used to characterize rheological changes in erythrocytes from patients with leptospirosis in the late stage. Biochemical methods were also used for quantification of plasma lipid, erythrocyte membrane lipid, and evaluation of liver function. Our data showed that the mean elastic constant of erythrocytes from patients with leptospirosis was around 67% higher than the control (healthy individuals), indicating that patient's erythrocytes were less elastic. In individuals with leptospirosis, several alterations in relation to control were observed in the plasma lipids, however, in the erythrocyte membrane, only phosphatidylcholine showed a significant difference compared to control, increasing around 41%. With respect to the evaluation of liver function of individuals with leptospirosis, there was a significant increase in levels of alanine transaminase (154%) and aspartate transaminase (150%), whereas albumin was 43.8% lower than control (P<0.01). The lecithin-cholesterol acyltransferase fractional activity was 3.6 times lower in individuals with leptospirosis than in the healthy individuals (P<0.01). The decrease of the erythrocyte elasticity may be related to the changes of erythrocyte membrane phospholipids composition caused by disturbances that occur during human leptospirosis, with phosphatidylcholine being a strong candidate in the erythrocyte rheological changes.

Preparation and pharmacokinetic investigation of propranolol-loaded elastic liposomes composed of DP-PC and SPC / 中国药学杂志

OBJECTIVE: To enhance the percutaneous absorption of propranolol by modifying the lipid composition of elastic liposomes. METHODS: Elastic liposomes composed of both 1, 2-dipalmitoyl-m-glycero-3-phosphacholme(DPPC) and soy phosphatidylcholine (SPC) were prepared and propranolol was encapsulated into liposomes by ammonium sulfate gradient loading method. Then the pharmaceutical and pharmacokinetic properties of propranolol-loaded elastic liposomes (PEL) with different lipid compositions were compared. RESULTS: The highest deformability was obtained with PEL composed of binary mixtures of DPPC and SPC (6:4, molar ratio). The encapsulation efficiency(EE) values of PEL composed of binary mixtures of DPPC and SPC(6:4, molar ratio) and SPC a-lone were 78.97 ± 2.94% and 68.67 ± 0.58%, respectively. And the drug release values were (23.23 ± 0.90)% and (39.84 ± 0.51)%, respectively. Following transdermal administration, the bioavailability of PEL composed of binary mixtures of DPPC and SPC (6:4, molar ratio) was significantly increased(14.73-fold) compared with that of PEL composed of SPC alone. CONCLUSION: The membrane stability and percutaneous absorption of PEL can be significantly improved by the application of binary mixtures of DPPC and SPC.

Structural properties of lipid reconstructs and lipid composition of normotensive and hypertensive rat vascular smooth muscle cell membranes

Multiple cell membrane alterations have been reported to be the cause of various forms of hypertension. The present study focuses on the lipid portion of the membranes, characterizing the microviscosity of membranes reconstituted with lipids extracted from the aorta and mesenteric arteries of spontaneously hypertensive (SHR) and normotensive control rat strains (WKY and NWR). Membrane-incorporated phospholipid spin labels were used to monitor the bilayer structure at different depths. The packing of lipids extracted from both aorta and mesenteric arteries of normotensive and hypertensive rats was similar. Lipid extract analysis showed similar phospholipid composition for all membranes. However, cholesterol content was lower in SHR arteries than in normotensive animal arteries. These findings contrast with the fact that the SHR aorta is hyporeactive while the SHR mesenteric artery is hyperreactive to vasopressor agents when compared to the vessels of normotensive animal strains. Hence, factors other than microviscosity of bulk lipids contribute to the vascular smooth muscle reactivity and hypertension of SHR. The excess cholesterol in the arteries of normotensive animal strains apparently is not dissolved in bulk lipids and is not directly related to vascular reactivity since it is present in both the aorta and mesenteric arteries. The lower cholesterol concentrations in SHR arteries may in fact result from metabolic differences due to the hypertensive state or to genes that co-segregate with those that determine hypertension during the process of strain selection.

Mechanisms of Selective Antimicrobial Activity of Gaegurin 4

Gaegurin 4 (GGN4), an antimicrobial peptide isolated from a Korean frog, is five times more potent against Gram-positive than Gram-negative bacteria, but has little hemolytic activity. To understand the mechanism of such cell selectivity, we examined GGN4-induced K+ efflux from target cells, and membrane conductances in planar lipid bilayers. The K+ efflux from Gram-positive M. luteus (2.5microgram/ml) was faster and larger than that from Gram-negative E. coli (75microgram/ml), while that from RBC was negligible even at higher concentration (100microgram/ml). GGN4 induced larger conductances in the planar bilayers which were formed with lipids extracted from Gram-positive B. subtilis than in those from E. coli (p<0.01), however, the effects of GGN4 were not selective in the bilayers formed with lipids from E. coli and red blood cells. Addition of an acidic phospholipid, phosphatidylserine to planar bilayers increased the GGN4-induced membrane conductance (p<0.05), but addition of phosphatidylcholine or cholesterol reduced it (p<0.05). Transmission electron microscopy revealed that GGN4 induced pore-like damages in M. luteus and dis-layering damages on the outer wall of E. coli. Taken together, the present results indicate that the selectivity of GGN4 toward Gram-positive over Gram-negative bacteria is due to negative surface charges, and interaction of GGN4 with outer walls. The selectivity toward bacteria over RBC is due to the presence of phosphatidylcholine and cholesterol, and the trans-bilayer lipid asymmetry in RBC. The results suggest that design of selective antimicrobial peptides should be based on the composition and topology of membrane lipids in the target cells.

Membrane fluidity and lipid composition of fluconazole resistant and susceptible strains of Candida albicans isolated from diabetic patients / Fluidez e composição lipídica da membrana de cepas de Candida albicans resistentes e sensíveis ao fluconazol, isoladas de pacientes diabéticos

Ten clinical isolates of Candida albicans, five strains belonging to each of fluconazole resistant and susceptible groups isolated from diabetic patients, were studied for the membrane fluidity and lipid composition. Compared to fluconazole susceptible strains, fluconazole resistant ones exhibited enhanced membrane fluidity as measured by fluorescence polarization technique. The increased membrane fluidity was reflected in the decreased p-values exhibited by the resistant strains. On the other hand, susceptible isolates contained higher amount of ergosterol, almost twice as compared to resistant isolates which might have contributed to their lower membrane fluidity. However, no significant alteration was observed in the phospholipid and fatty acid composition of these isolates. Labeling experiments with fluorescamine dye revealed that the percentage of the exposed aminophospholipid, phosphatidylethanolamine was highest in the resistant strains as compared to the susceptible strains, indicating a possible overexpression of CDR1 and CDR2 genes in resistant strains. The results presented here suggest that the changes in the ergosterol content and overexpression of ABC transporter genes CDR1 and CDR2 could contributeto fluconazole resistance in C. albicans isolated from diabetic patients.

Dez isolados clínicos, sendo cinco resistentes e cinco sensíveis ao fluconazol, obtidos de pacientes diabéticos, foram estudados quanto à fluidez e composição química da membrana. Quando comparados aos isolados sensíveis ao fluconazol, os isolados resistentes apresentaram fluidez de membrana aumentada, conforme mensurado pela técnica de polarização fluorescente. A fluidez de membrana aumentada refletiu-se pelos valores mais baixos de p. Por outro lado, os isolados sensíveis continham quantidades mais elevadas de ergosterol, quase o dobro dos isolados resistentes, o que pode ter contribuído para a fluidez de membrana mais baixa. Entretanto, não se observou alteração significativa na composição fosfolipídica e de ácidos graxos nesses isolados. Experimentos de marcação com corante fluorescamina indicaram que a porcentagem de aminofosfolípides e fosfatidiletanolamina expostos foi mais elevada nos isolados resistentes do que nos sensíveis, indicando uma possível superexpressão dos genes CDR1 e CDR2 nos isolados resistentes. Os resultados aqui apresentados sugerem que alterações no teor de ergosterol e superexpressão dos genes ABC transportadores CDR1 e CDR2 podem contribuir na resistência ao fluconazol em isolados de C. albicans de pacientes diabéticos.

Lipids Analysis of Epidermis and Stratum corneum Using Circumcised Prepuce / 대한피부과학회지

BACKGROUND: The changes in lipid composition during epidermal differentiation has been reported in human and animal models. Because of the difficulties in getting adeguate specimens from human subjects, the authors used easily obtainable circumcised prepuce for lipid analysis. OBJECTIVE: To investigate the changes in lipid composition duriig cornification of the epidermis, the lipid compositions of whole epidermis and stratum corneum were analyzed by thin layer chromatography(TLC). METHODS: From circumcied prepuce whole epidermis and stratum orneum were separated by 10mM EDTA(ethylene diamine tatraacetate) in PBS(phosphate-buffered saline) or heat(60C), and 0.5% trypsin in PBS respectively. Lipids were extracted with methanolctloroform-HO mixture(4:2:1.6, v/ v, Bligh-Dyer solvent), TLC was performed and lipid composition was quantitated by photodensitometer. RESULTS: In the composition of stratum corneum lipids, sphingoliids were the highest(33.3+2.9%) followed by cholesterol, free fatty acids and cholesterol esters in cleceasing order, there were small percentages of triglycerides, cholesterol sulfate and squalene. CONCLUSION: In this study the lipid composition of epidermis was similar to that of stratum corneum rather than those of previous reports on epidermal lipids, which may indicate the regional characteristics of epidermal/stratum orneum lipids in hyperkeratotic prepuie.

Effect of environmental factors on the limpid composition, membrane structure and permeability of Microsporum gypseum.

Supplementation with unsaturated fatty acids, substitution of glucose by glycerol as carbon source and lowered growth temperature (20°C) increased the total phospholipid content of Microsporum gypseum spheroplasts. Levels of sterols increased with glycerol substitution and decreased in other growth conditions. Substantial changes were seen in the ratios of unsaturated to saturated fatty acids and phosphatidylcholine to phosphatidylethanolafne under all the experimental conditions. Changed lipid composition resulted in altered uptake of amino acids (L-lysine, L-aspartic acid and L-glycine) and increased number of binding sites for a fluorescent probe, 1-anilinonaphthalene-8-sulfonate.

Lipid Composition of Ear Wax in Hircismus

To investigate the difference of dry ear wax and wet ear wax, the lipid composition of wet ear wax was analyzed and compared with that of dry ear wax. In dry ear wax, squalene, steryl esters, wax esters, triglycerides free fatty acids and cholesterol were found. Squalene, triglycerides, free fatty acids and cholesterol formed the main demonstrable fractions in wet ear wax. In addition, three unidentified spots were always present in wet ear wax. Our results indicate that wet ear wax is due to the difference of quantity and composition of ear wax lipids.