Permeability Of Phospholipid Membranes Research Articles

Stability of Immobilized Chemosensor‐Filled Vesicles on Anti‐Fouling Polymer Brush Surfaces

AbstractThe characterization of phospholipid membrane permeability for small molecules is crucial for many applications in drug discovery and biomedical research in general. Here, chemosensor‐laden vesicles offer an attractive platform for permeability assays. In this work, the stability of immobilized chemosensor‐filled vesicles is explored on anti‐fouling polymer brush surfaces for potential use in monitoring small molecule membrane permeability. The study focuses on the development of a method for immobilizing sensor‐loaded vesicles into arbitrary patterns on surfaces and characterizing their stability under changing temperatures and compositions. As substrate to enable intact, long‐term stable vesicle immobilization reactive polymer brushes with anti‐fouling properties are used. Utilizing microchannel cantilever spotting, biotin moieties are introduced on the polymer brush surface, enabling stable tethering of vesicles through streptavidin‐biotin interactions. The immobilized vesicles are monitored through fluorescence microscopy for their response to analytes under changing environmental parameters and vesicle composition. A higher stability of immobilized vesicles compared to free‐floating ones is observed, with permeability only at elevated temperatures. By tuning vesicle compositions, permeability at lower temperatures can be raised again. Overall, the study provides insights into a novel approach for vesicle immobilization, showcasing the potential of surface‐bound vesicles for applications in microfluidic systems and as biosensors in various assays.

Permeability of phospholipid membranes and human red blood cell membranes to hydrogen peroxide

Permeability of phospholipid membranes and human red blood cell membranes to hydrogen peroxide

Analogs of a Natural Peptaibol Exert Anticancer Activity in Both Cisplatin- and Doxorubicin-Resistant Cells and in Multicellular Tumor Spheroids.

Peptaibols, by disturbing the permeability of phospholipid membranes, can overcome anticancer drug resistance, but their natural hydrophobicity hampers their administration. By a green peptide synthesis protocol, we produced two water-soluble analogs of the peptaibol trichogin GA IV, termed K6-Lol and K6-NH2. To reduce production costs, we successfully explored the possibility of changing the naturally occurring 1,2-aminoalcohol leucinol to a C-terminal amide. Peptaibol activity was evaluated in ovarian cancer (OvCa) and Hodgkin lymphoma (HL) cell lines. Peptaibols exerted comparable cytotoxic effects in cancer cell lines that were sensitive—and had acquired resistance—to cisplatin and doxorubicin, as well as in the extrinsic-drug-resistant OvCa 3-dimensional spheroids. Peptaibols, rapidly taken up by tumor cells, deeply penetrated and killed OvCa-spheroids. They led to cell membrane permeabilization and phosphatidylserine exposure and were taken up faster by cancer cells than normal cells. They were resistant to proteolysis and maintained a stable helical structure in the presence of cancer cells. In conclusion, these promising results strongly point out the need for further preclinical evaluation of our peptaibols as new anticancer agents.

Unveiling a Hidden Event in Fluorescence Correlative Microscopy by AFM Nanomechanical Analysis.

Fluorescent imaging combined with atomic force microscopy (AFM), namely AFM-fluorescence correlative microscopy, is a popular technology in life science. However, the influence of involved fluorophores on obtained mechanical information is normally underestimated, and such subtle changes are still challenging to detect. Herein, we combined AFM with laser light excitation to perform a mechanical quantitative analysis of a model membrane system labeled with a commonly used fluorophore. Mechanical quantification was additionally validated by finite element simulations. Upon staining, we noticed fluorophores forming a diffuse weakly organized overlayer on phospholipid supported membrane, easily detected by AFM mechanics. The laser was found to cause a degradation of mechanical stability of the membrane synergically with presence of fluorophore. In particular, a 30 min laser irradiation, with intensity similar to that in typical confocal scanning microscopy experiment, was found to result in a ∼40% decrease in the breakthrough force of the stained phospholipid bilayer along with a ∼30% reduction in its apparent elastic modulus. The findings highlight the significance of analytical power provided by AFM, which will allow us to “see” the “unseen” in correlative microscopy, as well as the necessity to consider photothermal effects when using fluorescent dyes to investigate, for example, the deformability and permeability of phospholipid membranes.

Analysis of α-synuclein species enriched from cerebral cortex of humans with sporadic dementia with Lewy bodies.

Since researchers identified α-synuclein as the principal component of Lewy bodies and Lewy neurites, studies have suggested that it plays a causative role in the pathogenesis of dementia with Lewy bodies and other ‘synucleinopathies’. While α-synuclein dyshomeostasis likely contributes to the neurodegeneration associated with the synucleinopathies, few direct biochemical analyses of α-synuclein from diseased human brain tissue currently exist. In this study, we analysed sequential protein extracts from a substantial number of patients with neuropathological diagnoses of dementia with Lewy bodies and corresponding controls, detecting a shift of cytosolic and membrane-bound physiological α-synuclein to highly aggregated forms. We then fractionated aqueous extracts (cytosol) from cerebral cortex using non-denaturing methods to search for soluble, disease-associated high molecular weight species potentially associated with toxicity. We applied these fractions and corresponding insoluble fractions containing Lewy-type aggregates to several reporter assays to determine their bioactivity and cytotoxicity. Ultimately, high molecular weight cytosolic fractions enhances phospholipid membrane permeability, while insoluble, Lewy-associated fractions induced morphological changes in the neurites of human stem cell-derived neurons. While the concentrations of soluble, high molecular weight α-synuclein were only slightly elevated in brains of dementia with Lewy bodies patients compared to healthy, age-matched controls, these observations suggest that a small subset of soluble α-synuclein aggregates in the brain may drive early pathogenic effects, while Lewy body-associated α-synuclein can drive neurotoxicity.

Understanding of Relationship between Phospholipid Membrane Permeability and Self-Diffusion Coefficients of Some Drugs and Biologically Active Compounds in Model Solvents.

In this work we measured self-diffusion coefficients of 5 drugs (aspirin, caffeine, ethionamide, salicylic acid, and paracetamol) and 11 biologically active compounds of similar structure in deuterated water and 1-octanol by NMR. It has been found that an increase in the van der Waals volume of the molecules of the studied substances result in reduction of their diffusion mobility in both solvents. The analysis of the experimental data showed the influence of chemical nature and structural isomerization of the molecules on the diffusion mobility. Apparent permeability coefficients of the studied compounds were determined using an artificial phospholipid membrane made of egg lecithin as a model of in vivo absorption. Distribution coefficients in 1-octanol/buffer pH 7.4 system were measured. For the first time the model of the passive diffusion through the phospholipid membrane was validated based on the experimental data. To this end, the passive diffusion was considered as an additive process of molecule passage through the aqueous boundary layer before the membrane and 1-octanol barrier simulating the lipid layer of the membrane.

Colistin-entrapped liposomes driven by the electrostatic interaction: Mechanism of drug loading and in vivo characterization

The potential in vivo application of liposome for polycationic colistin has been hindered by the poor entrapment efficiency (EE) due to their phospholipid membrane permeability. The objective of this study is to investigate the loading mechanism and validity of applying electrostatic attraction for the colistin entrapment and delivery in liposomes. Anionic lipids with various structures were used for colistin entrapment, and the properties of resulting liposomes (i.e. zeta-potential, EE and release rate) were highly dependent on the structure of anionic lipids. Based on consideration of intermolecular interactions, the retention of electrostatically entrapped colistin is essentially determined by the balance of interfacial hydrophobic attraction and electrostatic repulsion. The liposomal colistin showed the reduced bacterial killing rate, but did not compromise the in vitro antibacterial activity. Specially, the PEGylated liposomal colistin of sodium cholesteryl sulfate (Chol-SO4−) showed the best drug retention, resulting in the significantly increased maximum-tolerated dose, prolonged blood circulation and decreased colistin distribution in kidney after intravenous administration in mice. These results highlight the potential utility of electrostatically entrapped liposome for polycationic colistin delivery.

Electrostatically entrapped colistin liposomes for the treatment of Pseudomonas aeruginosa infection

The potential use of liposomes for the pulmonary delivery of colistin has been hindered by their phospholipid membrane permeability resulting in a very low entrapment of colistin in the liposomes. To increase the entrapment capacity of colistin in liposomes, the anionic lipid sodium cholesteryl sulfate (Chol-SO4−) was used to enhance the electrostatic attraction between colistin and the lipid membrane. The resulting colistin-entrapped liposomes of Chol-SO4− (CCL) showed significantly greater entrapment efficiency in comparison with liposomes without Chol-SO4−. A time-kill kinetics study showed that colistin could redistribute from the liposomes into a new bacterial cell membrane to exert bactericidal activity. After intratracheal instillation, the CCL exhibited prolonged colistin retention in the lung with less colistin being transferred to the bloodstream and kidney, and the improved biodistribution further resulted in the enhanced therapeutic efficacy in a murine pulmonary Pseudomonas aeruginosa infection model compared with the colistin solution. These results highlight the suitability of applying an electrostatic attraction to entrap colistin in liposomes for pulmonary delivery by increasing colistin retention in the lung, while reducing the systemic exposure.

Preparation and characterisation of the colistin-entrapped liposome driven by electrostatic interaction for intravenous administration

Potential use of liposome for polycationic colistin is hindered by their phospholipid membrane permeability. In this study, liposomes were modified with sodium cholesteryl sulphate (Chol-SO4−) for improving the colistin loading by enhancing the colistin-bilayer electrostatic attraction. We have evaluated two liposomes: colistin-entrapped liposome of Chol-SO4− (CCL) and coated Chol-SO4−/colistin complex liposome (CCCL). In comparison with CCL which formed large aggregates at Chol-SO4−/colistin charge ratio below 2:1, CCCL showed a smaller size less dependent on the charge ratio, probably arising from more colistin entrapped on the inner leaflet of bilayer. Both liposomes exhibited significantly increased entrapment efficiency as compared with the liposome without Chol-SO4−. But colistin released upon dilution, implying free transfer of colistin through bilayers. Pharmacokinetics results showed the approximately four-fold increase in the plasma AUC0–8 h for CCCL and CCL as compared with colistin solution, showing potential benefit for infectious target localisation by prolonging the systemic circulation of colistin.

Investigating the Double-Peak Phenomenon in Oral Pharmacokinetics of Eurycoma longifolia quassinoids in Lipid-Based Solid Dispersion

α(21)-Epoxyeurycomanone (EP) and eurycomanone (EN), the polar quassinoids of Eurycoma longifolia extract (TAF2) formulated as a lipid-based solid dispersion (TAF2-SD) to increase sperm count for oligospermic infertility, displayed improvement of oral pharmacokinetics and bioavailability in rats over the non-formulated TAF2 extract. The plasma concentration-time curves of the quassinoids in both preparations displayed double-peak phenomenon, similarly observed in BCS Class 3 drugs that are highly hydrophilic and of low phospholipid membrane permeability, which therefore prompted investigation of the contributing factors. The results following the oral co-administration of TAF2-SD with the gastrointestinal transit markers indicated that less than 50% w/w of EP and EN were absorbed in the rat small intestine prior to 2.5 h, the gastric emptying time of 50% w/w theophylline marker absorption. More than 70% w/w of the quassinoids were absorbed in the colon after 8 h, the detection of 50% w/w sulfapyridine, the hydrolyzed metabolite of sulfasalazine marker. The non-formulated TAF2 extract at 0.028 ppm and its major quassinoid, EN at 0.005 ppm reduced acetylcholine-and histamine-induced contractions of the guinea pig ileum, suggesting that the muscle relaxant effect contributed to the delay in gastric-emptying and absorption in the intestine. In conclusion, the formulated quassinoids improved the oral bioavailability without changing the double-peak pharmacokinetic profile, and reduced gut motility causing delayed gastric emptying and intestinal absorption.

Confocal Raman Microscopy Probing of Temperature-Controlled Release from Individual, Optically-Trapped Phospholipid Vesicles

Control of permeability of phospholipid vesicle (liposome) membranes is critical to their applications in analytical sensing, in fundamental studies of chemistry in small volumes, and in encapsulation and release of payloads for site-directed drug delivery. Applications of liposome formulations in drug delivery often take advantage of the enhanced permeability of phospholipid membranes at their gel-to-fluid phase transition, where the release of encapsulated molecules can be initiated by an increase in temperature. Despite numerous successful liposome formulations for encapsulation and release methods to study the kinetics, this process has been limited to investigations of bulk vesicle dispersions, which provide little or no information about the vesicle membrane structure and its relationship to the kinetics of trans-membrane transport. In this work, confocal Raman microscopy is adapted to study temperature-dependent release of a model compound, 3-nitrobenzene sulfonate (3-NBS), from individual optically trapped phospholipid vesicles, while simultaneously monitoring structural changes in the vesicle membrane reported by vibrational modes of phospholipid acyl chains and the local environment of the encapsulated compound. The confocal geometry allows efficient excitation and collection of Raman scattering from a single vesicle, while optical trapping allows more than hour-long observations of the same vesicle. With window factor analysis to resolve component spectra, temperature-controlled release of 3-NBS through vesicle membranes composed of pure 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) was measured and compared to transport through a lysolipid-containing membrane specifically formulated for efficient drug delivery.

Effect of Hydrophobic Surfactant Proteins SP-B and SP-C on the Permeability of Phospholipid Membranes

Pulmonary surfactant is a complex mixture of lipids and proteins whose main function is to reduce surface tension at the alveolar air-liquid interface in order to avoid alveolar collapse at the end of expiration and facilitate the work of breathing. It is composed by around 90% lipids and 8-10% specific proteins, including the hydrophobic SP-B and SP-C. In this study, we have analyzed the effect of hydrophobic surfactant proteins on the permeability of phospholipid membranes by two different approaches: fluorescence microscopy of giant vesicles (GV) and electroconduction in planar lipid membranes. The effect of surfactant proteins on the permeability of GV membranes was assessed under the microscope using the fluorescent water-soluble probes FM®1-43 and calcein. Membrane-sensitive FM®1-43 only labels the external leaflet of membranes, and calcein emits green fluorescence in aqueous media. Neither can permeate through pure lipid membranes. In the presence of physiological amounts of SP-B and SP-C, giant oligolamellar POPC vesicles incorporated FM®1-43 in every single membrane when added to the external medium and were also permeable to calcein. These results suggest the existence of direct connections between aqueous compartments of GV in the presence of these proteins. On the other hand, planar lipid membranes (PLM) have been widely used to study ionic permeation through phospholipid bilayers mediated by membrane proteins. Permeability of bilayers incorporating small amounts of hydrophobic surfactant proteins has been analyzed in PLMs prepared by the dual monolayer technique. Conductance and selectivity experiments as well as noise analysis of the signals were performed. All our measurements indicate the formation of channel-like structures with defined properties in terms of ionic conductance and selectivity. Possible implications of membrane-permeabilizing structures in the biological context of the pulmonary surfactant system will be discussed.

Structural Elucidation of Light Activated Vesicles

In the present study, synchrotron small-angle X-ray scattering (SAXS) combined with a UV light source (in situ SAXS-UV irradiation) was used to determine the structure response of gold nanoparticle (NP)-loaded vesicles. The investigated system consisted of multilamellar vesicles of N-methylated dioleoylphosphatidylethanolamine (DOPE-Me) containing hydrophilic gold NPs with a size of 40 Å. Our results indicate drastic optothermically induced structural changes in these gold NP-loaded aqueous dispersions, which are accompanied by strong alterations in the phospholipid membrane permeability. A structural mechanism from well-ordered multilamellar vesicles in the fluid lamellar (Lα) phase to an inverted type hexagonal liquid crystalline (H2) phase via the formation of an intermediate phase of uncorrelated fluid bilayers is proposed. In general, these investigations are also crucial for the understanding of the potential of gold NP-loaded model phospholipid systems as efficient drug nanocarriers aiming to improve drug targeting and releasing on demand.

The Temperature Dependence And Quantized Nature Of The Lipid Membrane Permeability

We investigate the permeability of phospholipid membranes for fluorescence dyes and ions. We find that the permeability is strongly enhanced close to the chain melting transition of the membranes. Close to transitions, fluctuations in area and compressibility are high, leading to an increased likelihood of spontaneous lipid pore formation. Fluorescence Correlation Spectroscopy (FCS) reveals the permeability for rhodamine dyes across 100 nm vesicles. Using FCS, we find that the permeability of vesicle membranes for fluorescence dyes is within error proportional to the excess heat capacity. To estimate defect size we measure the conductance of solvent-free planar lipid bilayer. Microscopically, we show that permeation events appear as quantized current events. Furthermore, we demonstrate that general anaesthetics lead to a change in membrane permeability that can be predicted from their effect on heat capacity profiles. Depending on temperature, the permeability can be enhanced or reduced. We demonstrate that anaesthetics decrease channel conductance and ultimately lead to ‘blocking’ of the lipid pores in experiments performed at or above the chain melting transition. Our data suggests that the macroscopic increase in permeability close to transitions and microscopic lipid channel formation are the same physical process.

Comparative Study of Effects of Various Sterols and Triterpenoids on Permeability of Model Lipid Membranes

Using permeability to labeled glucose as a criterion of stability for liposomal membranes, a comparative study on stabilizing properties of different sterols and triterpenes in phospholipid bilayer has been carried out as well as on structural peculiarities of sterols responsible for membranolytic properties of cucumarioside G1 from the cucumaria Eupentacta fraudatris. Stabilizing action of the studied sterols and triterpenoides incorporated in the bilayer decreases in the following order: cholesterol sulfate > cholesterol > Δ 5-sterols > β-sitosterol > ergosterols > Δ 7-sterols > epicholesterol > pregnane > androstane > coprosterol > 14α-methylcholest-9(11)-en-3β-ol > 4α, 14α-dimethylcholest-9(11)-en-3β-ol > holothurinogenin A1 > glucoside of cholesterol > β-xylosidase of Δ 7-sterols > betulin > protopanaxatriol > phosphatidylcholine liposomes without sterol > protopanaxadiol > oleanolic acid. Sterol-dependent membranolytic cucumarioside G1 practically loses its ability to increase permeability of phospholipid membranes containing sterols obtained from this holothuria as well as coprosterol, epicholestrol, sulfated and glycosylated forms of sterols. The obtained results confirm the “sterol” hypothesis of the mechanism of membranotropic action of holothuria glycosides and of resistance to them of holothuria cell membranes.

Effect of the exposure to gentamicin and diltiazem on the permeability of model membranes

Preliminary observations showed that the calcium-antagonist diltiazem enhances the ‘in vitro’ bactericidal action of the aminoglycoside gentamicin, especially against Gram-positive bacteria. To verify if a non-specific interaction of these two drugs with biomembranes may play a role in their synergic effect on bacterial cells, we have studied the effect of exposure to gentamicin, in the absence or presence of diltiazem, on the release of carboxyfluorescein (CF) trapped in phosphatidylcholine (PC) unilamellar vesicles (LUVs) used as model membranes. A significant leakage of trapped CF from PC LUVs was registered when liposomes were treated with gentamicin and diltiazem together, employed at doses (50 and 100 μg/ml, respectively) unable to affect CF release if applied alone; the combined effect of gentamicin and diltiazem was synergic and not cumulative. The present findings demonstrate that the simultaneous exposure to gentamicin and diltiazem may induce significant alterations in the permeability of phospholipid membranes and, so, very likely, in functional properties of bacterial membranes, targets of their action.

RNAs that bind and change the permeability of phospholipid membranes.

The RNA world hypothesis presumes that RNA will be competent for varied essential cellular functions. One such indispensable cell function is regulation of membrane permeability. Though this was not a known RNA activity, selection-amplification yielded RNAs that bound phosphatidylcholine:cholesterol liposomes. At least eight distinct, approximately 95-mer sequences bind well to the outside of the lipid bilayer, though randomized sequences had no such activity. No distinct sequence motif for lipid binding was found. However, truncation of one such RNA shows that a smaller, 44-nucleotide irregular RNA hairpin is an active membrane binding domain. Bound RNA increases the permeability of liposomes to (22)Na(+). In addition, using voltage clamp technique, four individual RNAs increase the ion permeability of the plasma membrane of cultured human cells. The existence of multiple sequences that bind membranes and provoke permeability changes suggests that these may be elementary RNA functions that could be selected in vivo.

Post mortem changes in Ca2+ transporting proteins of sarcoplasmic reticulum in dependence on malignant hyperthermia status in pigs.

Meat quality of pigs is dependent on biochemical and biophysical processes in the time course post mortem (p.m.) and is associated with the intracellular Ca2+ homeostasis. However, there is little known about changes in the Ca2+ transporting proteins controlling the Ca2+ uptake of sarcoplasmic reticulum (SR) in the time course p.m. In this study changes in the Ca2+ transporting proteins were investigated in homogenates of longissimus muscles of 4 malignant hyperthermia susceptible (MHS) and 6 malignant hyperthermia resistant (MHR) Pietrain pigs. Muscle samples were obtained at different time intervals: biopsy 2 h prior slaughtering and from the carcass immediately after exsanguination (0 h), 45 min, 4 h, and 22 h p.m. The SR Ca2+ uptake rate was measured immediately after homogenization with closed calcium release channel (CRC), with opened CRC and without manipulation of CRC. Additionally the SR Ca2+ ATPase activity was determined. The results show: (i) The ability of SR to sequester Ca2+ declined to about 60% in the first 45 min p.m. in MHS samples irrespective of CRC state, whereas in MHR samples this decline was about 5%; (ii) Ca2+ uptake and Ca2+ ATPase activity were not different between the biopsy and 0 h samples, i.e. the stress of slaughter was of no immediate influence; (iii) The Ca2+ ATPase activity of the SR declined at about the same rate as the Ca2+ uptake in both MHS and MHR pig samples in the course of time p.m.; (iv) In samples, taken immediately after exsanguination, the Ca2+ ATPase activity of MHS pigs was higher than that of MHR pigs. However, in samples taken 4 h p.m. Ca2+ ATPase activity of MHS pigs has declined to about 30% of the value at 0 h; (v) The CRC can be closed and opened in all samples up to 22 h p.m. and seems to be fully functional at all sampling times; (vi) The CRC of MHS pigs is almost fully open, whereas the CRC of MHR pigs is only partially open at all sampling times; (vii) The permeability of the SR membrane to Ca2+ (determined as the ratio of SR Ca2+ ATPase with and without ionophore A23187) is the same in both MHS and MHR and did not change with ongoing time; (viii) No uncoupling of uptake from ATP hydrolysis occurred up to 4 h p.m., but the coupling differed between MHS and MHR for all time intervals with lower values for MHS pigs. The results suggest that the decreasing Ca2+ uptake rate of homogenates, sampled at different times p.m., is essentially caused by changes in the Ca2+ pump and not by changes in the CRC or an increased phospholipid membrane permeability to Ca2+.

Phospholipid membrane permeability of peptide nucleic acid

Phospholipid vesicles (liposomes) as membrane models have been used to study the penetration properties of peptide nucleic acid (PNA), a new DNA analog in which the nucleobases are attached to a pseudo-peptide backbone. The liposomes were characterised by carboxyfluorescein efflux, light-scattering and cryogenic transmission electron microscopy. The liposome structure was found not to be affected by the incorporation of PNA or an oligonucleotide. Two 10-mer fluorescein-labelled PNAs were found to have low efflux rates (half-times of 5.5 and 11 days), comparable to a 10-mer oligonucleotide (half-time of 7 days). We conclude that passive diffusion of unmodified PNA is not an effective way of transport into biological cells.

Effects of hexagonal phase induction by dolichol on phospholipid membrane permeability and morphology

The effect of H 11-phase induction by dolichol (DOL) (C 80–105, 0–2 wt%) within the bilayers of multilamellar liposomes (MLV, DOPE:DOPC 2:1 or 3:1 w/w) on their permeability, lipid mixing and morphology was determined. Low-angle X-ray diffraction patterns were consistent with mixtures of bilaycr and H 11 phases, the latter increasing with increasing DOL or DOPE content and temperature. Efflux rate constants for 6-carboxyfluorescein (6-CF) from 2:1 DOPE:DOPC vesicles depended on temperature and DOPE/DOL content, increasing as much as 200-fold over DOL-free controls at 2% w/w DOL. Fluorescence resonance energy transfer assays detected lipid mixing with unlabeled target MLV. It was appreciable only when target MLV contained DOPE and increased with DOL content. Confocal scanning fluorescence microscopy was applied to study the morphological structure of fully-hydrated samples and field scanning electron microscopy the ultrastructure of cryo-stabilized samples. 3:1 DOPE/DOPC MLV, stable at pH 9.5, underwent rapid morphological changes at pH 7.4. Within minutes filaments formed and large areas of membrane surface became studded with 10–15 nm bumps and 5 nm holes, resembling in size and shape unilamellarly covered intcrlamellar micellar intermediates and interlamellar attachments (ILA) previously associated with H 11-phase transitions. The filaments, seen in MLV with and without DOL, may represent extensions of IMI into coaxial assemblages of rod micellar intermediates (RMI). These phenomena may have implications for liposomal delivery of therapeutic peptides/proteins if they can be made to trigger the convective release of liposomal contents via controlled formation of ILA between adjacent lamellae of MLV.