Molecular mechanisms at the basis of the protective effect exerted by EPPS on neurodegeneration induced by prefibrillar amyloid oligomers.

It has been shown recently, without an explanation of the possible molecular mechanisms involved, that 4-(2-hydroxyethyl)-1-piperazinepropanesulphonic (EPPS) acid effectively protects from the neurotoxicity induced by oligomers and plaques formed by the protein amyloid-β protein. Here we report the same protective effect, obtained in vitro (HT22-diff cell line) and ex vivo (hippocampal slices) models, against amyloid neurotoxicity induced by oligomers of salmon Calcitonin (sCT), which has been shown to be a good model for the study of neurodegenerative diseases. Based on biophysical studies focusing on the protein aggregation kinetic and the interaction of the aggregates with model membranes, we propose a possible molecular mechanism underlying the protective effects.

Resveratrol and Resveratrol-Loaded Galactosylated Liposomes: Anti-Adherence and Cell Wall Damage Effects on and MRSA.

Antibiotic resistance due to bacterial biofilm formation is a major global health concern that makes the search for new therapeutic approaches an urgent need. In this context,, -resveratrol (RSV), a polyphenolic natural substance, seems to be a good candidate for preventing and eradicating biofilm-associated infections but its mechanism of action is poorly understood. In addition, RSV suffers from low bioavailability and chemical instability in the biological media that make its encapsulation in delivery systems necessary.

Quantification of particle number concentration in liposomal suspensions by Laser Transmission Spectroscopy (LTS).

Laser Transmission Spectroscopy (LTS) is an experimental technique able to determine the particle number concentration and the size of colloidal suspensions by a single measurement of the transmittance of a laser beam through the suspension of particles as a function of the wavelength. In this protocol, we show that LTS represents a unique and powerful tool to investigate suspensions of liposomes, where the precise quantification of the number concentration is particularly relevant for the complete definition of the colloidal properties of the suspension. We study a model formulation of Soy-PC:Chol liposomes and we validate LTS results by comparison with High-Performance Liquid Chromatography determination of lipid mass.

Electrochemical Bottom-Up Synthesis of Chiral Carbon Dots from L-Proline and Their Application as Nano-Organocatalysts in a Stereoselective Aldol Reaction.

Chirality is undoubtedly a fundamental property of nature since the different interactions of optically active molecules in a chiral environment are essential for numerous applications. Thus, in the field of asymmetric synthesis, the search for efficient, sustainable, cost-effective and recyclable chiral catalysts is still the main challenge in organic chemistry. The field of carbon dots (CDs) has experienced tremendous development in the last 15 years, including their applications as achiral catalysts.

Transport of cationic liposomes in a human blood brain barrier model: Role of the stereochemistry of the gemini amphiphile on liposome biological features.

Hypothesis: The positive charge on liposome surface is known to promote the crossing of the Blood brain barrier (BBB). However, when diastereomeric cationic gemini amphiphiles are among lipid membrane components, also the stereochemistry may affect the permeability of the vesicle across the BBB.

Experiments: Liposomes featuring cationic diasteromeric gemini amphiphiles were formulated, characterized, and their interaction with cell culture models of BBB investigated.

Resveratrol loaded in cationic glucosylated liposomes to treat Staphylococcus epidermidis infections.

Glucosylated liposomes composed of the natural saturated phospholipid 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC), cholesterol (Chol) and a cationic amphiphile featuring a glucosyl moiety (GL4), have been developed for delivering the antimicrobial trans-Resveratrol (RSV) to S. epidermidis, characterized by carbohydrate-specific adhesins able to recognize glucose. The cationic derivative of cholesterol, DC-Chol, was also included in liposome formulations, alone or in combination with GL4, in order to explore the role of both cationic charge and sugar moiety in the interaction of liposomes with bacterial cells.

How stereochemistry of lipid components can affect lipid organization and the route of liposome internalization into cells.

Though liposome-based drugs are in clinical use, the mechanism of cell internalization of liposomes is yet an object of controversy. The present experimental investigation, carried out on human glioblastoma cells, indicated different internalization routes for two diastereomeric liposomes. Molecular dynamics simulations of the lipid bilayers of the two formulations indicated that the different stereochemistry of a lipid component controls some parameters such as area per lipid molecule and fluidity of lipid membranes, surface potential and water organization at the lipid/water interface, all of which affect the interaction with biomolecules and cell components.

Time-programmable pH: decarboxylation of nitroacetic acid allows the time-controlled rising of pH to a definite value.

In this report it is shown that nitroacetic acid (ONCHCOH) can be conveniently used to control the pH of a water solution over time. Time-programmable sequences of the kind pH-pH-pH can be achieved, where both the extent of the initial pH jump (pH-pH) and the time required for the subsequent pH rising (pH-pH) can be predictably controlled by a judicious choice of the absolute and relative concentrations of the reagents (acid and NaOH). Successive pH-pH-pH sequences can be obtained by subsequent additions of acid .

Synthesis and Characterization of Mitochondria-Targeted Triphenylphosphonium Bolaamphiphiles.

In this chapter we describe: (1) the procedure for the synthesis of four single chain bolaamphiphiles, displaying chains of 12, 16, 20 and 30 methylene units and triphenylphosphonium moieties as headgroups (TPP1-TPP4); (2) the methods used to characterize TPP1-TPP4 spontaneous aggregation in aqueous solution. We illustrate the determination of Krafft point and cac by conductivity measurements and the procedures used to investigate dimensions, morphology, and stability by dynamic and dielectrophoretic laser light scattering, dialysis, transmission electron microscopy, and Raman spectroscopy measurements.

The Interaction between Amyloid Prefibrillar Oligomers of Salmon Calcitonin and a Lipid-Raft Model: Molecular Mechanisms Leading to Membrane Damage, Ca-Influx and Neurotoxicity.

To investigate the interaction between amyloid assemblies and "lipid-rafts", we performed functional and structural experiments on salmon calcitonin (sCT) solutions rich in prefibrillar oligomers, proto- and mature-fibers interacting with liposomes made of monosialoganglioside-GM1 (4%), DPPC (48%) and cholesterol (48%). To focus on the role played by electrostatic forces and considering that sCT is positive and GM1 is negative at physiologic pH, we compared results with those relative to GM1-free liposomes while, to assess membrane fluidity effects, with those relative to cholesterol-free liposomes. We investigated functional effects by evaluating Ca-influx in liposomes and viability of HT22-DIFF neurons.

Aggregation behaviour of triphenylphosphonium bolaamphiphiles.

Hypothesis: Bolaamphiphiles are characterized by wide polymorphism of their aggregates, due to the connection of the headgroups that renders their investigation very intriguing in several technological applications. Some bolaamphiphiles displaying the triphenylphosphonium motif (TPP-bolaamphiphiles) were previously explored for their ability in crossing the mitochondrial membranes but their colloidal features, which are crucial for the potential development of an effective drug delivery system, were never investigated.

Experiments: Single chain TPP-bolaamphiphiles, featuring chains of 12, 16, 20 and 30 methylene units, were synthesized and their aggregation features (Krafft point, cac, dimensions, morphology, stability) were investigated by conductivity, dialysis, transmission electron microscopy, Raman spectroscopy, dynamic and dielectrophoretic laser light scattering measurements.

Monosialoganglioside-GM1 triggers binding of the amyloid-protein salmon calcitonin to a Langmuir membrane model mimicking the occurrence of lipid-rafts.

GM1 ganglioside is known to be involved in the amyloid-associated diseases and it is a crucial factor for the assembly of amyloid proteins on lipid-rafts, which are lipid structures located on the synaptic plasma membranes. Due to its slow aggregation rate, we employed salmon calcitonin (sCT) as a suitable probe representative of amyloid proteins, to study the interaction between this class of proteins and a membrane model. Here, we prepared a neuronal membrane model by depositing onto mica two Langmuir-Blodgett films in liquid-condensed phase: the outer monolayer was characterized by high content of GM1 (50%) and minority parts of cholesterol and POPC (25-25%), while the inner one by plain POPC.

Remote loading of aloe emodin in gemini-based cationic liposomes.

Anthraquinone compound aloe-emodin (AE) has shown antineoplastic, antibacterial, antiviral, and anti-inflammatory properties and scavenging activity on free radicals. Because of these therapeutic features, AE has been attracting increasing interest and could be applied in the curing of many diseases. However, until now the physicochemical features of this compound have not been fully investigated; furthermore, its wide application might be hindered by its scarce solubility in aqueous media (∼19 μM).

Synthesis and physicochemical characterization of pyrrolidinium based surfactants.

Three new pyrrolidinium based surfactants were synthesized and characterized as pure aggregate components and in mixtures with 1,2-dimyristoyl-sn-glycero-3-phosphocholine to understand how the molecular structure of the cationic amphiphile and its mole percentage might affect the physicochemical properties of the resulting aggregates. The thermotropic behavior of the mixed liposomes was investigated by differential scanning calorimetry on multilamellar vesicles, whereas their size and surface potential were investigated on large unilamellar vesicles by dynamic laser light scattering and fluorescence experiments, respectively. The presence of either methoxy or hydroxy groups in the positions 3 and 4 of the pyrrolidinium ring as well as the presence of a second alkyl chain on pyrrolidinium nitrogen, controls the aggregates features.

Fusion of gemini based cationic liposomes with cell membrane models: implications for their biological activity.

The interaction of neutral and anionic phospholipid liposomes, used as cell models, with cationic liposomes formulated with 1,2-dimyristoyl-sn-glicero-3-phosphocholine and stereomeric cationic gemini surfactants was investigated by differential scanning calorimetry, fluorescence experiments and dynamic laser light scattering. This study was aimed at rationalizing the different biological features shown by liposomes based on different gemini stereoisomers observed in previous investigations. In fact, to correlate the observed biological activity of liposomes with the molecular structure of their components is critical for a rational and systematic approach to the design of new carriers for drug delivery.

Influence of lipid composition on the thermotropic behavior and size distribution of mixed cationic liposomes.

Cationic liposomes are studied mainly as nonviral nucleic acid delivery systems and to a lesser extent as carriers/adjuvants of vaccines and as low-molecular-weight drug carriers. It is well established that the performance and the biological activity of liposomes in general are strongly related to their physicochemical properties. We investigated the thermotropic behavior and the size distribution of mixed cationic liposomes formulated with different percentages of 1,2 dimyristoyl-sn-glycero-3-phosphatidylcholine and one of four cationic amphiphiles characterized by a pyrrolidinium headgroup with the aim of achieving a better understanding of how the molecular structure of the cationic amphiphile and its mole percentage affect the physicochemical properties of the liposomes.

Extended cardiolipin anchorage to cytochrome c: a model for protein-mitochondrial membrane binding.

Two models have been proposed to explain the interaction of cytochrome c with cardiolipin (CL) vesicles. In one case, an acyl chain of the phospholipid accommodates into a hydrophobic channel of the protein located close the Asn52 residue, whereas the alternative model considers the insertion of the acyl chain in the region of the Met80-containing loop. In an attempt to clarify which proposal offers a more appropriate explanation of cytochrome c-CL binding, we have undertaken a spectroscopic and kinetic study of the wild type and the Asn52Ile mutant of iso-1-cytochrome c from yeast to investigate the interaction of cytochrome c with CL vesicles, considered here a model for the CL-containing mitochondrial membrane.

Efficiency of liposomes in the delivery of a photosensitizer controlled by the stereochemistry of a gemini surfactant component.

Liposomes formulated with dimyristoyl-sn-glycero-phosphatidylcholine, DMPC, and either one of the cationic gemini surfactants (S,S)-2,3-dimethoxy-1,4-bis(N-hexadecyl-N,N-dimethylammonio)butane bromide, 1a, and (S,R)-2,3-dimethoxy-1,4-bis(N-hexadecyl-N,N-dimethylammonio)butane bromide, 1b, were investigated as vehicles of the photosensitizer m-tetrahydroxyphenylchlorin, m-THPC, to cell models of malignant glioma. The delivery efficiency of DMPC/1a and DMPC/1b liposome formulations were evaluated on the murine glioblastoma cell line C6 and on the human glioblastoma cell line LN229 by flow cytometry and laser scanning confocal microscopy. The stereochemistry of the spacer of the gemini was found to strongly influence the delivery efficiency of m-THPC to cells, the mode of interaction with the cell membrane, and the intracellular distribution of m-THPC.

New cationic liposomes as vehicles of m-tetrahydroxyphenylchlorin in photodynamic therapy of infectious diseases.

Antimicrobial photodynamic therapy is emerging as a promising therapeutic modality for bacterial infections. For optimizing the antibacterial activity of the photosensitizer m-tetrahydroxyphenylchlorin, it has been encapsulated in mixed cationic liposomes composed of different ratios of dimyristoyl- sn-glycero-phosphatidylcholine and any of four cationic surfactants derived from l-prolinol. The delivery efficiency of the different liposomes formulations has been evaluated on a methicillin-resistant Staphylococcus aureus bacterial strain (MRSA), and one of the tested formulations shows a biological activity comparable to that of the free chlorin.

Concentration as the switch for chiral recognition in biomembrane models.

Chiral recognition was observed in a biomembrane model. Micellar aggregates formed by enantiopure N-alkyl-N,N-dimethyl-N-(1-phenyl)ethylammonium bromide were in fact able to convert the racemic mixture of bilirubin-IXalpha into an enantiomerically enriched mixture. The stereochemical preference and the extent of enantiomeric enrichment depend on the length of the hydrophobic portion of the surfactant and on the concentration conditions, and changes in the stereochemical bias are reversible.

PEGylated lipoplexes: preparation protocols affecting DNA condensation and cell transfection efficiency.

The inclusion of poly(ethylene glycol) monolaurate in liposomes formulated with dimyristoyl-sn-glycero-3-phosphocholine and certain cationic gemini surfactants improves their capability of condensing DNA into a psi phase and transfecting it into cells. Both the condensation, observed by circular dichroism, and the transfection efficiency are strongly effected by the protocol of inclusion of the polymer in the formulations. The highest transfection efficiency is observed in correspondence of the highest extent of DNA condensation.

Influence of the spacer of cationic gemini amphiphiles on the hydration of lipoplexes.

The impact of the length of gemini surfactant spacer on complexation and condensation of calf thymus DNA by cationic mixed phospholipid/gemini liposomes was investigated by monitoring the conformational changes of DNA by circular dichroism and the lipid hydration level by the emission characteristics of the fluorescent probe laurdan included in the lipid bilayer. The length of the spacer was shown to influence, on one hand, the hydration level and the organization of the corresponding liposomes and, on the other, the variation of lipid hydration level and the DNA conformation upon complexation. In fact, in correspondence with the longest spacer we observed more hydrated liposomes, probably organized in domains, a higher extent of dehydration promoted by the addition of DNA, and a minor extent of DNA conformational change.

m-THPC-mediated photodynamic therapy of malignant gliomas: assessment of a new transfection strategy.

Malignant gliomas represent the most common primary brain tumor: more than 50% of them are glioblastoma multiforme (GBM). Photodynamic therapy may offer a very good chance of targeted destruction of infiltrating GBM cells, thus increasing the survival time and recurrence-free interval of GBM patients. Among photosensitizing agents, meta-tetrahydroxyphenylchlorin (m-THPC) is promising for the treatment of brain tumors.

Role of the spacer stereochemistry on the aggregation properties of cationic gemini surfactants.

The preparation and characterization of three stereoisomeric cationic gemini surfactants, 2,3-dimethoxy-1,4-bis(N-hexadecyl-N,N-dimethylammonio)butane dibromide, are described. The aggregation properties have been studied by fluorescence, electrical conductivity, and quasi-elastic light scattering. A conformational study of the surfactant headgroups has been performed by molecular mechanics calculations to investigate the effect of the stereogenic centers on the surfactant molecular shape and therefore on the different organizations of the monomers in the aggregates.