Heterogeneous and anomalous diffusion inside lipid tubules

Self-assembled lipid tubules with crystalline bilayer walls are promising candidates for controlled drug delivery vehicles on the basis of their ability to release preloaded biological molecules in a sustained manner. While a previous study has shown that the release rate of protein molecules from lipid tubules depends on the associated molecular mass, suggesting that the pertinent diffusion follows the well-known Stokes-Einstein relationship, only a few attempts have been made toward investigating the details of molecular diffusion in the tubule interior. Herein, we have characterized the diffusion rates of several molecules encapsulated in lipid tubules formed by 1,2-bis(10,12-tricosadiynoyl)-sn-glycero-3-phosphocholine (DC8,9PC) using the techniques of fluorescence recovery after photobleaching (FRAP) and fluorescence correlation spectroscopy (FCS). Our results show that the mobility of these molecules depends not only on their positions in the DC8,9PC tubules but also on their respective concentrations. While the former indicates that the interior of the DC8,9PC tubules is heterogeneous in terms of diffusion, the latter further highlights the possibility of engineering specific conditions for achieving sustained release of a "drug molecule" over a targeted period of time. In addition, our FCS results indicate that the molecular diffusions inside the crystalline bilayer walls of the DC8,9PC tubules strongly deviate from the normal, stochastic processes, with features characterizing not only anomalous subdiffusions but also motions that are superdiffusive in nature.     

Design of liposomes containing photopolymerizable phospholipids for triggered release of contents

We describe a novel class of light-triggerable liposomes prepared from a photo-polymerizable phospholipid DC_8,9PC (1,2-bis (tricosa-10,12-diynoyl)-sn-glycero-3-phosphocholine) and DPPC (1,2-Dipalmitoyl-sn-Glycero-3-Phosphocholine). Exposure to UV (254 nm) radiation for 0–45 min at 25 °C resulted in photo-polymerization of DC_8,9PC in these liposomes and the release of an encapsulated fluorescent dye (calcein). Kinetics and extents of calcein release correlated with mol% of DC_8,9PC in the liposomes. Photopolymerization and calcein release occurred only from DPPC/DC_8,9PC but not from Egg PC/DC_8,9PC liposomes. Our data indicate that phase separation and packing of polymerizable lipids in the liposome bilayer are major determinants of photo-activation and triggered contents release.     

Membrane orientation of MSI-78 measured by sum frequency generation vibrational spectroscopy

Antimicrobial peptides (AMPs) selectively disrupt bacterial cell membranes to kill bacteria whereas they either do not or weakly interact with mammalian cells. The orientations of AMPs in lipid bilayers mimicking bacterial and mammalian cell membranes are related to their antimicrobial activity and selectivity. To understand the role of AMP-lipid interactions in the functional properties of AMPs better, we determined the membrane orientation of an AMP (MSI-78 or pexiganan) in various model membranes using sum frequency generation (SFG) vibrational spectroscopy. A solid-supported single 1,2-dipalmitoyl-an-glycero-3-[phospho-rac-(1-glycerol)] (DPPG) bilayer or 1-palmitoyl-2-oleoyl-sn-glycero-3-[phospho-rac-(1-glycerol)] (POPG) bilayer was used as a model bacterial cell membrane. A supported 1,2-dipalmitoyl-an-glycero-3-phosphocholine (DPPC) bilayer or a 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) bilayer was used as a model mammalian cell membrane. Our SFG results indicate that the helical MSI-78 molecules are associated with the bilayer surface with ～70° deviation from the bilayer normal in the negatively charged gel-phase DPPG bilayer at 400 nM peptide concentration. However, when the concentration was increased to 600 nM, MSI-78 molecules changed their orientation to make a 25° tilt from the lipid bilayer normal whereas multiple orientations were observed for an even higher peptide concentration in agreement with toroidal-type pore formation as reported in a previous solid-state NMR study. In contrary, no interaction between MSI-78 and a zwitterionic DPPC bilayer was observed even at a much higher peptide concentration (～12,000 nM). These results demonstrate that SFG can provide insights into the antibacterial activity and selectivity of MSI-78. Interestingly, the peptide exhibits a concentration-dependent membrane orientation in the lamellar-phase POPG bilayer and was also found to induce toroidal-type pore formation. The deduced lipid flip-flop from SFG signals observed from lipids also supports MSI-78-induced toroidal-type pore formation.     

Atomic force microscopy studies of ganglioside GM1alpha in dioleoylphosphatidylcholine/dipalmitoylphosphatidylcholine mixed monolayers and hybrid bilayers

The membrane states of the alpha-series ganglioside GM1alpha in 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC)/1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) mixed monolayers and hybrid bilayers were investigated using atomic force microscopy (AFM). The AFM image for the GM1alpha/DOPC/DPPC ternary monolayers showed the formation of GM1alpha-raft in the DOPC matrix. As increase of the surface pressure, GM1alpha are condensed in DPPC-rich domains; long and slender GM1alpha-rafts are separated from the DPPC-rich domains into the DOPC matrix. The GM1alpha/DOPC/DPPC ternary monolayers were deposited on mica coated with the first layer (1,2-dipalmitoyl-sn-glycero-3-phosphoethanolamine: DPPE) using the Langmuir-Schaeffer technique. The AFM image for the hybrid bilayers showed that same molecules were heterogeneously concentrated according to increase of the surface pressure to form GM1alpha-raft, DPPC-rich domain and DOPC matrix, being in agreement with the observation on the monolayer experiment. The found phenomenon implies that a binding of lectin to GM1alpha causes the increase of the surface pressure, the localization of GM1alpha and the succeeding formation of the raft as a first step of a specific signal transduction.     

Photopolymerization of mixed monolayers and black lipid membranes containing gramicidin A and diacetylenic phospholipids

We formed monolayers and black lipid membranes (BLMs) of photopolymerizable lipids mixed with the channel-forming protein gramicidin A to evaluate their miscibility and the potential for improved stability of the BLM scaffold through polymerization. Analyses of surface pressure vs area isotherms indicated that gramicidin A dispersed with three different synthetic, polymerizable, diacetylene-containing phospholipids, 1,2-di-10,12-tricosadiynoyl-sn-glycero-3-phosphocholine (DTPC), 1,2-di-10,12-tricosadiynoyl-sn-glycero-3-phosphoethanolamine (DTPE), and 1-palmitoyl-2,10,12-tricosadiynoyl-sn-glycero-3-phosphoethanolamine (PTPE) to form mixed monolayers at the air-water interface on a Langmuir-Blodgett (LB) trough. Conductance measurements across a diacetylenic lipid-containing BLM confirmed dispersion of the gramicidin channel with the lipid layer and demonstrated gramicidin ion-channel activity before and after UV exposure. Polymerization kinetics of the diacetylenic films were monitored by film pressure changes at constant LB trough area and by UV-vis absorption spectroscopy of polymerized monolayers deposited onto quartz. An initial increase in film pressure of both the pure diacetylene lipid monolayers and mixed films upon exposure to UV light indicated a change in the film structure. Over the time scale of the pressure increase, an absorbance peak indicative of polymerization evolved, suggesting that the structural change in the lipid monolayer was due to polymerization. Film pressure and absorbance kinetics also revealed degradation of the polymerized chains at long exposure times, indicating an optimum time of UV irradiation for maximized polymerization in the lipid layer. Accordingly, exposure of polymerizable lipid-containing black lipid membranes to short increments of UV light led to an increase in the bilayer lifetime.     

DOPC/DPPC单层膜中分子间的相互作用及形态特征

利用Langmuir-Blodgett(LB)技术制备了不同表面压力下的1,2-二油酸-甘油-3-磷脂酰胆碱(DOPC)/1,2-二棕榈酸甘油-3-磷脂酰胆碱(DPPC)(摩尔比为1:1)和DOPC/DPPC/Chol(摩尔比为2:2:1)单层膜, 对单层膜内分子间的相互作用进行了热力学分析, 并用荧光显微镜和原子力显微镜对其形态进行了观测.热力学分析表明, DOPC与DPPC分子在单层膜结构中相互作用为排斥力, 诱导单层膜出现相变; DOPC, DPPC与胆固醇(Chol)间的相互作用均为吸引力, 当表面压力(π)大于18 mN/m时, DPPC与胆固醇的作用力大于DOPC.荧光显微镜观测表明, DOPC/DPPC单层膜出现明显相分离现象, 富含DPPC微区成“花形”结构, 且随着表面压力的升高微区逐渐增大, “花瓣”增多; 当胆固醇加入到DOPC/DPPC体系时, 单层膜相态由液相与凝胶相共存转变为液态无序相与液态有序相共存结构, 富含DPPC的微区形状从“花形”转变成“圆形”.原子力显微镜对单层膜的表征验证了荧光显微镜的观测结果, 表明胆固醇加入到DOPC/DPPC体系中对单层膜排列具有明显的影响, 压力和溶液状态等是影响脂膜结构的重要因素.     

Micropatterned composite membranes of polymerized and fluid lipid bilayers

Micropatterned composite membranes of polymerized and fluid lipid bilayers were constructed on solid substrates. Lithographic photopolymerization of a diacetylene-containing phospholipid, 1,2-bis(10,12-tricosadiynoyl)-sn-glycero-3-phosphocholine (DiynePC), and subsequent removal of nonreacted monomers by a detergent solution (0.1 M sodium dodecyl sulfate (SDS)) yielded a patterned polymeric bilayer matrix on the substrate. Fluid lipid bilayers of phosphatidylcholine from egg yolk (egg-PC) were incorporated into the lipid-free wells surrounded by the polymeric bilayers through the process of fusion and reorganization of suspended small unilamellar vesicles. Spatial distribution of the fluid bilayers in the patterned bilayer depended on the degree of photopolymerization that in turn could be modulated by varying the applied UV irradiation dose. The polymeric bilayer domains blocked lateral diffusion of the fluid lipid bilayers and confined them in the defined areas (corrals), if the polymerization was conducted with a sufficiently large UV dose. On the other hand, lipid molecules of the fluid bilayers penetrated into the polymeric bilayer domains, if the UV dose was relatively small. A direct correlation was observed between the applied UV dose and the lateral diffusion coefficient of fluorescent marker molecules in the fluid bilayers embedded within the polymeric bilayer domains. Artificial control of lateral diffusion by polymeric bilayers may lead to the creation of complex and versatile biomimetic model membrane arrays.     

Phase transition of a single lipid bilayer measured by sum-frequency vibrational spectroscopy

In this communication, we demonstrate the first use of sum-frequency generation (SFG) vibrational spectroscopy to measure directly the phase transition temperature (Tm) of a single planar supported lipid bilayer (PSLB). Three saturated phospholipids, 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC), 1,2-diheptadecanoyl-sn-glycero-3-phosphocholine (DHPC), and 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC), were studied. Lipid bilayer films were prepared by the the Langmuir-Blodgett method at a surface pressure of 30 nN/m. The symmetric nature of the bilayer was used to determine the Tm of bilayers by measuring the intensity of the symmetric methyl stretch at 2875 cm-1 from the lipid fatty acid chains as a function of temperature. A maximum in the CH3 symmetric stretch transition was observed at the Tm of the lipid film due to the reduction of symmetry in the bilayer. The SFG measured Tm for DPPC, DHPC, and DSPC were 41.0 +/- 0.4, 52.4 +/- 0.7, and 57.9 +/- 0.5 degrees C, respectively. These values correlate well with the literature values of 41.3 +/- 1.8, 49 +/- 3, and 54.5 +/- 1.5 degrees C for DPPC, DHPC, and DSPC, respectively obtained by differential scanning calorimetry (DSC) of lipid vesicles in solution. The high degree of correlation between the SFG spectroscopic measurements and the DSC results suggests the Tm of these lipids is not significantly altered upon immobilization on a surface.     

Phosphonate lipid tubules II

We describe a new chiral tubule-forming lipid in which the C-O-P phosphoryl linkage of the archetypal tubule-forming molecule, 1,2-bis(10,12-tricosadiynoyl)-sn-glycero-3-phosphocholine, "DC(8,9)PC", is replaced by a C-P linkage. Tubule formation with this phosphonate analogue proceeds under the same mild conditions as with DC(8,9)PC and produces similar yields, but synchrotron small-angle X-ray scattering, atomic force microscopy, and optical microscopy show the new tubules to have diameters 1.94 times as great, to be significantly shorter, and to be thinner-walled. A significant portion of the enantiomerically pure chiral phosphonate precipitate is in the form of stable open helices, and these helices are divided almost evenly between left- and right-handed members.     

Interfacial behavior of chroman-6 and chroman-6 palmitoyl ester and their interaction with phospholipids

The surface activity of chroman-6 (CR-6) and chroman-6 palmitoyl ester (PCR-6) and the interactions with lipid membranes, using 1,2-dipamitoyl-sn-glycero-3-phosphocholine (DPPC) and 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) monolayers, were determined. 8-Anilino-1-naphthalenesulfonic acid titration indicates that none of these molecules was able to form aggregates in aqueous media. The presence of a palmitoyl chain in PCR-6 increases strongly the surface activity of the parent compound (CR-6), rendering a molecule to form stable monomolecular layers. The interaction of both compounds with DPPC and DOPC, measured at constant area or in a compression isotherm model, follows the same trend. Miscibility studies performed with DPPC/CR-6 or PCR-6 indicate that the energies involved are small. The presence of CR-6 and PCR-6 has a soft influence on the compressibility of the Langmuir mixed films. Differential scanning calorimetry studies indicate that CR-6 and PCR-6 modify the temperature and cooperativity of the transition from gel to liquid crystal process in DPPC vesicles.     

Interaction of antimicrobial arginine-based cationic surfactants with liposomes and lipid monolayers

The present work examines the relationship between the antimicrobial activity of novel arginine-based cationic surfactants and the physicochemical process involved in the perturbation of the cell membrane. To this end, the interaction of these surfactants with two biomembrane models, namely, 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) multilamellar lipid vesicles (MLVs) and monolayers of DPPC, 1,2-dipalmitoyl-sn-glycero-3-[phospho-rac-(1-glycerol)] sodium salt (DPPG), and Escherichia coli total lipid extract, was investigated. For the sake of comparison, this study included two commercial antimicrobial agents, hexadecyltrimethylammonium bromide and chlorhexidine dihydrochloride. Changes in the thermotropic phase transition parameters of DPPC MLVs in the presence of the compounds were studied by differential scanning calorimetry analysis. The results show that variations in both the transition temperature (Tm) and the transition width at half-height of the heat absorption peak (deltaT1/2) were consistent with the antimicrobial activity of the compounds. Penetration kinetics and compression isotherm studies performed with DPPC, DPPG, and E. coli total lipid extract monolayers indicated that both steric hindrance effects and electrostatic forces explained the antimicrobial agent-lipid interaction. Overall, in DPPC monolayers single-chain surfactants had the highest penetration capacity, whereas gemini surfactants were the most active in DPPG systems. The compression isotherms showed an expansion of the monolayers compared with that of pure lipids, indicating an insertion of the compounds into the lipid molecules. Owing to their cationic character, they are incorporated better into the negatively charged DPPG than into zwitterionic DPPC lipid monolayers.     

Filipin orientation revealed by linear dichroism. Implication for a model of action

The organization of the polyene antibiotic filipin in membranes containing cholesterol is a controversial matter of debate. Two contradictory models exist, one suggesting a parallel and the other perpendicular organization of filipin with respect to the plane of the membrane. UV-vis linear dichroism, ATR-FTIR, and fluorescence anisotropy decay techniques were combined to study the orientation of filipin in model systems of membranes composed of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) or 1,2-palmitoyl-sn-glycero-3-phosphocholine (DPPC) with and without cholesterol. Filipin's orientation is determined by the presence/absence of cholesterol when it is inserted in gel crystalline phase model membranes. When cholesterol (33%) is present in DPPC bilayers, filipin stands perpendicular to the membrane surface as expected in "pore-forming" models. At variance, absence of cholesterol leaves filipin in an essentially random organization in the lipidic matrix. In liquid crystalline phase bilayers (POPC) filipin's orientation is perpendicular to the membrane surface even in absence of cholesterol. Thus filipin's activity/organization depends not only on cholesterol presence but also in the lipid phase domain it is inserted in. These findings were combined with spectroscopy and microscopy data in the literature, solving controversial matters of debate.     

A facile approach for assembling lipid bilayer membranes on template-stripped gold

Lipid vesicles are designed with functional chemical groups to promote vesicle fusion on template-stripped gold (TS Au) surfaces that does not spontaneously occur on unfunctionalized Au surfaces. Three types of vesicles were exposed to TS Au surfaces: (1) vesicles composed of only 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) lipids; (2) vesicles composed of lipid mixtures of 2.5 mol % of 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-poly(ethylene glycol)-2000-N-[3-(2-pyridyldithio)propionate] (DSPE-PEG-PDP) and 97.5 mol % of POPC; and (3) vesicles composed of 2.5 mol % of 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[methoxy(poly(ethylene glycol))-2000] (DSPE-PEG) and 97.5 mol % POPC. Atomic force microscopy (AFM) topography and force spectroscopy measurements acquired in a fluid environment confirmed tethered lipid bilayer membrane (tLBM) formation only for vesicles composed of 2.5 mol % DSPE-PEG-PDP/97.5 mol % POPC, thus indicating that the sulfur-containing PDP group is necessary to achieve tLBM formation on TS Au via Au-thiolate bonds. Analysis of force-distance curves for 2.5 mol % DSPE-PEG-PDP/97.5 mol % POPC tLBMs on TS Au yielded a breakthrough distance of 4.8 ± 0.4 nm, which is about 1.7 nm thicker than that of POPC lipid bilayer membrane formed on mica. Thus, the PEG group serves as a spacer layer between the tLBM and the TS Au surface. Fluorescence microscopy results indicate that these tLBMs also have greater mechanical stability than solid-supported lipid bilayer membranes made from the same vesicles on mica. The described process for assembling stable tLBMs on Au surfaces is compatible with microdispensing used in array fabrication.     

Effects of cholesterol component on molecular interactions between paclitaxel and phospholipid within the lipid monolayer at the air-water interface

Cholesterol is a main component of the cell membrane and could have significant effects on drug-cell membrane interactions and thus the therapeutic efficacy of the drug. It also plays an important role in liposomal formulation of drugs for controlled and targeted delivery. In this research, Langmuir film technique, atomic force microscopy (AFM) and Fourier transform infrared spectroscopy (FTIR) are employed for a systematic investigation on the effects of cholesterol component on the molecular interactions between a prototype antineoplastic drug (paclitaxel) and 1,2-dipalmitoyl-sn-glycerol-3-phosphocholine (DPPC) within the cell membrane by using the lipid monolayer at the air-water interface as a model of the lipid bilayer membrane and the biological cell membrane. Analysis of the measured surface pressure (pi) versus molecular area (a) isotherms of the mixed DPPC/paclitaxel/cholesterol monolayers at various molar ratios shows that DPPC, paclitaxel and cholesterol can form a non-ideal miscible system at the air-water interface. Cholesterol enhances the intermolecular forces between paclitaxel and DPPC, produces an area-condensing effect and thus makes the mixed monolayer more stable. Investigation of paclitaxel penetration into the mixed DPPC/cholesterol monolayer shows that the existence of cholesterol in the DPPC monolayer can considerably restrict the drug penetration into the monolayer, which may have clinical significance for diseases of high cholesterol. FTIR and AFM investigation on the mixed monolayer deposited on solid surface confirmed the obtained results.     

Effects of lipid chain unsaturation and headgroup type on molecular interactions between paclitaxel and phospholipid within model biomembrane

Molecular interactions between paclitaxel, an anticancer drug, and phospholipids of various chain unsaturations and headgroup types were investigated in the present study by Langmuir film balance and differential scanning calorimetry. Both the lipid monolayer at the air-water interface and the lipid bilayer vesicles (liposomes) were employed as model cell membranes. It was found that, regardless of the difference in molecular structure of the lipid chains and headgroup, the drug can form nonideal, miscible systems with the lipids at the air-water interface over a wide range of paclitaxel mole fractions. The interaction between paclitaxel and phospholipid within the monolayer was dependent on the molecular area of the lipids at the interface and can be explained by intermolecular forces or geometric accommodation. Paclitaxel is more likely to form thermodynamically stable systems with 1,2-dipalmitoyl-sn-glycerol-3-phosphocholine (DPPC) and 1,2-dielaidoyl-sn-glycero-3-phosphocholine (DEPC) than with 1,2-dipalmitoyl-sn-glycero-3-phosphoethanolamine (DPPE) and 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC). Investigation of the drug penetration into the lipid monolayer showed that DPPC and DEPC have higher incorporation abilities for the drug than DPPE and DSPC. A similar trend was also evidenced by DSC investigation with liposomes. While little change of DSC profiles was observed for the DPPE/paclitaxel and DSPC/paclitaxel liposomes, paclitaxel caused noticeable changes in the thermographs of DPPC and DEPC liposomes. Paclitaxel was found to cause broadening of the main phase transition without significant change in the peak melting temperature of the DPPC bilayers, which demonstrates that paclitaxel was localized in the outer hydrophobic cooperative zone of the bilayer, i.e., in the region of the C1-C8 carbon atoms of the acyl chain or binding at the polar headgroup site of the lipids. However, it may penetrate into the deeper hydrophobic zone of the DEPC bilayers. These findings provide useful information for liposomal formulation of anticancer drugs as well as for understanding drug-cell membrane interactions.     

Interaction of 3β-amino-5-cholestene with phospholipids in binary and ternary bilayer membranes

3β-Amino-5-cholestene (aminocholesterol) is a synthetic sterol whose properties in bilayer membranes have been examined. In fluid palmitoyl sphingomyelin (PSM) bilayers, aminocholesterol and cholesterol were equally effective in increasing acyl chain order, based on changes in diphenylhexatriene (DPH) anisotropy. In fluid 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) bilayers, aminocholesterol ordered acyl chains, but slightly less efficiently than cholesterol. Aminocholesterol eliminated the PSM and DPPC gel-to-liquid crystalline phase transition enthalpy linearly with concentration, and the enthalpy approached zero at 30 mol % sterol. Whereas cholesterol was able to increase the thermostability of ordered PSM domains in a fluid bilayer, aminocholesterol under equal conditions failed to do this, suggesting that its interaction with PSM was not as favorable as cholesterols. In ternary mixed bilayers, containing 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC), PSM or DPPC, and cholesterol at proportions to contain a liquid-ordered phase (60:40 by mol of POPC and PSM or DPPC, and 30 mol % cholesterol), the average lifetime of trans-parinaric acid (tPA) was close to 20 ns. When cholesterol was replaced with aminocholesterol in such mixed bilayers, the average lifetime of tPA was only marginally shorter (about 18 ns). This observation, together with acyl chain ordering data, clearly shows that aminocholesterol was able to form a liquid-ordered phase with saturated PSM or DPPC. We conclude that aminocholesterol should be a good sterol replacement in model membrane systems for which a partial positive charge is deemed beneficial.     

Nanoscale ripples in self-assembled lipid tubules

Self-assembled cylindrical tubules of 1,2-bis(tricosa-10,12-diynoyl)-sn-glycero-3-phosphocholine (DC(8,9)PC) have been studied by atomic force microscopy in both the height and amplitude modes. Nanoscale ripple structures in the cylindrical lipid tubules are clearly resolved in amplitude mode images. The periodicity of the ripples is found to be 200 +/- 30 nm for tubules with diameters in the range from 200 to 650 nm. The angle of the ripples with respect to the equator of the tubules shows a bimodal distribution with centers at approximately 28 degrees and approximately 5 degrees.     

Effects of albumin and erythrocyte membranes on spread monolayers of lung surfactant lipids

Dipalmitoyl phosphatidylcholine (DPPC), one of the main constituents of lung surfactant is mainly responsible for reduction of surface tension to near 0 mN/m during expiration, resisting alveolar collapse. Other unsaturated phospholipids like palmitoyloleoyl phosphatidylglycerol (PG), palmitoyloleoyl phosphatidylcholine (POPC) and neutral lipids help in adsorption of lung surfactant to the air-aqueous interface. Lung surfactant lipids may interact with plasma proteins and hematological agents flooding the alveoli in diseased states. In this study, we evaluated the effects of albumin and erythrocyte membranes on spread films of DPPC alone and mixtures of DPPC with each of PG, POPC, palmitoyloleoyl phosphatidylethanolamine (PE), cholesterol (CHOL) and palmitic acid (PA) in 9:1 molar ratios. Surface tension-area isotherms were recorded using a Langmuir-Blodgett (LB) trough at 37 degrees C with 0.9% saline as the sub-phase. In the presence of erythrocyte membranes, DPPC and DPPC+PA monolayers reached minimum surface tensions of 7.3+/-0.9 and 9.6+/-1.4 mN/m, respectively. Other lipid combinations reached significantly higher minimum surface tensions >18 mN/m in presence of membranes (Newman Keul's test, p<0.05). The relative susceptibility to membrane inhibition was [(DPPC+PG, 7:3)=(DPPC+PG, 9:1)=(DPPC+POPC)=(DPPC+PE)=(DPPC+CHOL)]>[(DPPC+PA)=(DPPC)]. The differential response was more pronounced in case of albumin with DPPC and DPPC+PA monolayers reaching minimum surface tensions less than 2.4 mN/m in presence of albumin, whereas DPPC+PG and DPPC+POPC reached minimum surface tensions of around 20 mN/m in presence of albumin. Descending order of susceptibility of the spread monolayers of lipid mixtures to albumin destabilization was as follows: [(DPPC+PG, 7:3)=(DPPC+PG, 9:1)=(DPPC+POPC)]>[(DPPC+PE)=(DPPC+CHOL)]>[(DPPC+PA)=(DPPC)] The increase in minimum surface tension in presence of albumin and erythrocyte membranes was accompanied by sudden increases in compressibility at surface tensions of 15-30 mN/m. This suggests a monolayer destabilization and could be indicative of phase transitions in the mixed lipid films due to the presence of the hydrophobic constituents of erythrocyte membranes.     

Ethanol-induced reorganization of the liquid-ordered phase: enhancement of cholesterol-phospholipid association

This paper records what is believed to be the first evidence for the reorganization of the liquid-ordered phase by ethanol. Specifically, ethanol has been found to significantly enhance sterol-phospholipid association in liquid-ordered bilayers derived from 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) plus cholesterol and also 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC) plus cholesterol. The evidence for such reorganization comes from a series of nearest-neighbor recognition (NNR) experiments that have been carried out, where low concentrations of equilibrating lipid dimers (i.e., "reporter molecules") have been used to detect changes in the phase composition of host membranes made from varying mixtures of DPPC/cholesterol, and also DSPC/cholesterol, in the presence and in the absence of ethanol. These findings have important biological implications, which are briefly discussed.     

HIV fusion inhibitor peptide T-1249 is able to insert or adsorb to lipidic bilayers. Putative correlation with improved efficiency

T-1249 is a HIV fusion inhibitor peptide under clinical trials. Its interaction with biological membrane models (large unilamellar vesicles) was studied using fluorescence spectroscopy. A gp41 peptide that includes one of the hydrophobic terminals of T-1249 was also studied. Both peptides partition extensively to liquid-crystalline POPC (1-palmitoyl-2-oleyl-sn-glycero-3-phosphocholine) (DeltaG = -7.0 kcal/mol and -8.7 kcal/mol, for T-1249 and terminal peptide, respectively) and are located at the interface of the membrane. T-1249 is essentially in a random coil conformation in this lipidic medium, although a small alpha-helix contribution is present. When other lipid compositions are used (DPPC, POPG + POPC, and POPC + cholesterol) (DPPC (1,2-dipalmitoyl-sn-glycero-3-phosphocholine) and POPG (1-palmitoyl-2-oleyl-sn-glycero-3-[phospho-rac-(1-glycerol)), partition decreases, the most severe effect being the presence of cholesterol. Partition experiments and fluorescence resonance energy transfer analysis show that T-1249 adsorbs to cholesterol-rich membranes. The improved clinical efficiency of T-1249 relative to enfuvirtide (T20) may be related to its bigger partition coefficient and ability to adsorb to rigid lipidic areas on the cell surface, where most receptors are inserted. Moreover, adsorption to the sterol-rich viral membrane helps to increase the local concentration of the inhibitor peptide at the fusion site.