Cytoprotective effects of α-linolenic acid, eicosapentaenoic acid, docosahexaenoic acid, oleic acid and α-tocopherol on 7-ketocholesterol - Induced oxiapoptophagy: Major roles of PI3-K / PDK-1 / Akt signaling pathway and glutathione peroxidase activity in cell rescue.

On murine N2a cells, 7-ketocholesterol induced an oxiapotophagic mode of cell death characterized by oxidative stress (reactive oxygen species overproduction on whole cells and at the mitochondrial level; lipid peroxidation), apoptosis induction (caspase-9, -3 and -7 cleavage, PARP degradation) and autophagy (increased ratio LC3-II / LC3-I). Oxidative stress was strongly attenuated by diphenyleneiodonium chloride which inhibits NAD(P)H oxidase. Mitochondrial and peroxisomal morphological and functional changes were also observed. Down regulation of PDK1 / Akt signaling pathways as well as of GSK3 / Mcl-1 and Nrf2 pathways were simultaneously observed in 7-ketocholesterol-induced oxiapoptophagy. These events were prevented by α-linolenic acid, eicosapentaenoic acid, docosahexaenoic acid, oleic acid and α-tocopherol. The inhibition of the cytoprotection by LY-294002, a PI3-K inhibitor, demonstrated an essential role of PI3-K in cell rescue. The rupture of oxidative stress in 7-ketocholesterol-induced oxiapoptophagy was also associated with important modifications of glutathione peroxidase, superoxide dismutase and catalase activities as well as of glutathione peroxidase-1, superoxide dismutase-1 and catalase level and expression. These events were also counteracted by α-linolenic acid, eicosapentaenoic acid, docosahexaenoic acid, oleic acid and α-tocopherol. The inhibition of the cytoprotection by mercaptosuccinic acid, a glutathione peroxidase inhibitor, showed an essential role of this enzyme in cell rescue. Altogether, our data support that the reactivation of PI3-K and glutathione peroxidase activities by α-linolenic acid, eicosapentaenoic acid, docosahexaenoic acid, oleic acid and α-tocopherol are essential to prevent 7KC-induced oxiapoptophagy.

Cyclodextrins permeabilize DPPC liposome membranes: a focus on cholesterol content, cyclodextrin type, and concentration.

Cyclodextrins (CDs) are known for their ability to extract lipid components from synthetic and biological membranes and therefore to induce an increase of membrane permeability. However, the effect of cholesterol (CHOL) content in the membrane on the CD permeabilizing effect was not considered yet. Given that an increase in CHOL content reduces the membrane permeability, the aim of this work was to reveal how CHOL would modulate the CDs effect on the membrane. Hence, liposomes made of dipalmitoyl phosphatidylcholine (DPPC) and various CHOL contents (DPPC/CHOL 100:10, 100:25, 100:50, and 100:100) encapsulating the hydrophilic fluorophore, sulforhodamine B (SRB), were prepared and exposed to the native CDs (α-CD, ß-CD, γ-CD) and four ß-CD derivatives: the randomly methylated-ß-CD (RAMEB), the low methylated-ß-CD (CRYSMEB), the hydroxypropyl-ß-CD (HP-ß-CD) and the sulfobutyl ether-ß-CD (SBE-ß-CD) at different CD/DPPC molar ratios (1:1, 10:1, and 100:1). The membrane permeability was monitored following the release of SRB with time. The results demonstrated that the CDs effect on the membrane depends on the CD type, CD concentration, and membrane CHOL content. The investigated CDs exhibited an instantaneous permeabilizing effect promoting vesicle leakage of SRB from the various membranes; this effect increased with CDs concentration. Among the studied CDs, α-CD, ß-CD, and RAMEB were the most permeabilizing CDs on the different membranes. Similar modifications of SRB release from the various liposomal formulations were obtained with HP-ß-CD, CRYSMEB, and SBE-ß-CD. γ-CD was the less potent CD in affecting the membrane permeability. The CDs effect also depended on the CHOL content: at the CD/DPPC molar ratio (100:1), RAMEB and ß-CD considerably permeabilized the membrane of high CHOL content (50%, 100%) while the remaining CDs showed a decreasing permeabilizing effect upon CHOL content membrane increase.

Activity of Na+/K+- and Ca2+-ATPases in human erythrocyte membranes: Protocol improvement, relation to cholesterol content, and effects of polyphenols.

It is known that the activities of Na+/K+- and Ca2+-ATPases in the plasma membrane with an excess of cholesterol are compromised. Our main goal was to find out whether quercetin, resveratrol, or caffeic acid, in the nano- and low micromolar concentration ranges, can improve the ATPase activity in human erythrocyte membranes with excess cholesterol. These molecules belong to different chemical classes of polyphenols and are widely present in plant foods. Also, due to some variations in the protocol for determining the ATPase activity, we first analyzed several key parameters of the protocol to improve the accuracy of the results. The activities of Na+/K+- and Ca2+-ATPases were reduced in membranes with moderate and high cholesterol levels compared to membranes from normocholesterolemic subjects (p < 0.01). All three polyphenols affected the ATPase activity in a similar biphasic manner. Namely, the ATPase activity gradually increased with increasing polyphenol concentration up to 80-200 nM, and then gradually decreased with further increase in polyphenol concentration. Moreover, the stimulating effect of the polyphenols was highest in membranes with high cholesterol content, making ATPase activity values close/equal to those in normal cholesterol membranes. In other words, quercetin, resveratrol, and caffeic acid at nanomolar concentrations were able to improve/restore the functioning of Na+/K+- and Ca2+-ATPases in erythrocyte membranes with high cholesterol levels. This suggests a common membrane-mediated mechanism of action for these polyphenols, related to the content of membrane cholesterol.

Encapsulation of Rosmarinus officinalis essential oil and of its main components in cyclodextrin: application to the control of the date moth Ectomyelois ceratoniae (Pyralidae).

BACKGROUND: Synthetic insecticides are the most useful tools for preventing losses caused by insect pest's infestation during storage. However, the use of pesticides should be limited because of the development of insect resistance and their adverse effects on human health and environment. In the last decades, natural insecticidal products, principally essential oils (EOs) and their active components, exhibited potential alternatives for pest control. Nevertheless, due to their volatile nature, encapsulation could be considered as the most appropriate solution. Therefore, this work aims to investigate the fumigant ability of inclusion complexes of Rosmarinus officinalis EO and its major constituents (1,8-cineole, α-pinene and camphor) with 2-hydroxypropyl-beta-cyclodextrin (HP-ß-CD) against Ectomyelois ceratoniae (Pyralidae) larvae. RESULTS: The encapsulation within HP-ß-CD reduced greatly the release rate of the encapsulated molecules. Therefore, free compounds were more toxic than those encapsulated. Moreover, results revealed that encapsulated volatiles exhibited interesting insecticidal toxicity towards E. ceratoniae larvae. In fact, after 30 days mortality rates were 53.85, 94.23, 3.85 and 42.31% for α-pinene, 1,8-cineole, camphor and EO, respectively, encapsulated within HP-ß-CD. In addition, results showed also that 1,8-cineole free and encapsulated was more effective toward E. ceratoniae larvae than the other tested volatiles. Additionally, the HP-ß-CD/volatiles complexes exhibited best persistence compared to the volatiles components. The half-life of the encapsulated α-pinene, 1,8-cineole, camphor and EO (7.83, 8.75, 6.87 and 11.20 days) was significantly longer than that of the free ones (3.46, 5.02, 3.38 and 5.58 days). CONCLUSION: These results sustain the utility of R. officinalis EO and its main components encapsulated in CDs as treatment to stored-date commodities. © 2023 Society of Chemical Industry.

Bicelles as a carrier for bioactive compounds in beverages: application to nerolidol, an active sesquiterpene alcohol.

ABSTRACT: Nerolidol is a natural sesquiterpene alcohol with promising but limited application in food and pharmaceutical fields due to several factors including low photostability and low aqueous solubility. Recently, several carriers loading nerolidol were prepared and tested in fresh orange juice. Lipid vesicles loading nerolidol did not exhibit satisfactory organoleptic properties in this beverage. Hence, DMPC/DHPC bicelles were prepared as a new phospholipid-based carrier for nerolidol at different molar ratios. The bicelle suspensions were characterized in terms of homogeneity, particles size, and morphology. The optimal formulation (phospholipid:nerolidol molar ratio 100:1) was selected based on transparent appearance, homogeneity, and particle size (~ 45 nm). Besides, it showed a high encapsulation efficiency of nerolidol and a high incorporation rate of phospholipids. Transmission electron microscopy analysis demonstrated the formation of bicelles. The bicelles membrane fluidity was assessed by 1,6-diphenyl-1,3,5-hexatriene fluorescence anisotropy and differential scanning calorimetry analysis. The membrane fluidity of bicelles appeared to increase in the presence of nerolidol in a concentration dependent manner. To our knowledge this is the first study dealing with the encapsulation of an essential oil component in bicelles.

Effect of cysteamine hydrochloride-loaded liposomes on skin depigmenting and penetration.

Skin hyperpigmentation is caused by an excessive production of melanin. Cysteamine, an aminothiol compound physiologically synthetized in human body cells, is known as depigmenting agent. The aim of this study was to evaluate the depigmenting activity and skin penetration of liposome formulations encapsulating cysteamine hydrochloride. First, cysteamine hydrochloride-loaded liposomes were prepared and characterized for their size, polydispersity index, zeta potential and the encapsulation efficiency of the active molecule. The stability of cysteamine hydrochloride in the prepared liposome formulations in suspension and freeze-dried forms was then assessed. The in vitro cytotoxicity of cysteamine and cysteamine-loaded liposome suspensions (either original or freeze-dried) was evaluated in B16 murine melanoma cells. The measurement of melanin and tyrosinase activities was assessed after cells treatment with free and encapsulated cysteamine. The antioxidant activity of the free and encapsulated cysteamine was evaluated by the measurement of ROS formation in treated cells. The ex vivo human skin penetration study was also performed using Franz diffusion cell. The stability of cysteamine hydrochloride was improved after encapsulation in liposomal suspension. In addition, for the liposome re-suspended after freeze-drying, a significant increase of vesicle stability was observed. The free and the encapsulated cysteamine in suspension (either original or freeze-dried) did not show any cytotoxic effect, inhibited the melanin synthesis as well as the tyrosinase activity. An antioxidant activity was observed for the free and the encapsulated cysteamine hydrochloride. The encapsulation enhanced the skin penetration of cysteamine hydrochloride. The penetration of this molecule was better for the re-suspended freeze-dried form than the original liposomal suspension where the drug was found retained in the epidermis layer of the skin.

Pentacyclic triterpenes modulate liposome membrane fluidity and permeability depending on membrane cholesterol content.

Since the membrane-related processes represent an integral part of the biological activities of drugs, their effect on the membrane dynamics is actually considered. In this study, we investigated the effect of pentacyclic triterpenes (TTPs), oleanolic acid (OA) and erythrodiol (ER), on the fluidity and permeability of liposomes membranes differing by their cholesterol content. All liposomes were prepared by reverse phase evaporation technique (REV). Spin-labeled liposomes exposed or not to TTPs were used for fluidity studies by using 5- and 16-doxyl stearic acids (DSA). TTPs-loaded liposomes (phospholipid:cholesterol of 1:1), and preformed vesicles exposed to TTPs were used for permeability studies by monitoring the release of sulforhodamine B (SRB) at 37 °C. The apparent release constants of SRB were determined by Higuchi model based on a biphasic curve shape (0-10 h; 10-48 h). TTPs-loaded liposomes were characterized for their size and homogeneity. Results showed that ER increased the membrane fluidity at the upper region of the membrane while the both TTPs produced a condensing effect at the deeper region of the membrane. The membrane composition was a critical parameter modulating the effect of TTPs on the membrane permeability. Also, this study consolidated the fact that a fluidizing membrane agent is not necessarily a permeabilizing-membrane compound.

Encapsulation of α-Pinene in Delivery Systems Based on Liposomes and Cyclodextrins.

The essential oil component α-pinene has multiple biological activities. However, its application is limited owing to its volatility, low aqueous solubility, and chemical instability. For the aim of improving its physicochemical properties, α-pinene was encapsulated in conventional liposomes (CLs) and drug-in-cyclodextrin-in-liposomes (DCLs). Hydroxypropyl-ß-cyclodextrin/α-pinene (HP-ß-CD/α-pinene) inclusion complexes were prepared in aqueous solution, and the optimal solubilization of α-pinene occurred at HP-ß-CD:α-pinene molar ratio of 7.5:1. The ethanol-injection method was applied to produce different formulations using saturated (Phospholipon 90H) or unsaturated (Lipoid S100) phospholipids in combination with cholesterol. The size, the phospholipid and cholesterol incorporation rates, the encapsulation efficiency (EE), and the loading rate (LR) of α-pinene were determined, and the storage stability of liposomes was assessed. The results showed that α-pinene was efficiently entrapped in CLs and DCLs with high EE values. Moreover, Lipoid S100 CLs displayed the highest LR (22.9 ± 2.2%) of α-pinene compared to the other formulations. Both carrier systems HP-ß-CD/α-pinene inclusion complex and Lipoid S100 CLs presented a gradual release of α-pinene. Furthermore, the DPPH radical scavenging activity of α-pinene was maintained upon encapsulation in Lipoid S100 CLs. Finally, it was found that all formulations were stable after three months of storage at 4 °C.

Apiaceae Essential Oils: Boosters of Terbinafine Activity against Dermatophytes and Potent Anti-Inflammatory Effectors.

Dermatophyte infections represent an important public health concern, affecting up to 25% of the world's population. Trichophyton rubrum and T. mentagrophytes are the predominant dermatophytes in cutaneous infections, with a prevalence accounting for 70% of dermatophytoses. Although terbinafine represents the preferred treatment, its clinical use is hampered by side effects, drug-drug interactions, and the emergence of resistant clinical isolates. Combination therapy, associating terbinafine and essential oils (EOs), represents a promising strategy in the treatment of dermatophytosis. In this study, we screened the potential of selected Apiaceae EOs (ajowan, coriander, caraway, and anise) to improve the antifungal activity of terbinafine against T. rubrum ATCC 28188 and T. mentagrophytes ATCC 9533. The chemical profile of EOs was analyzed by gas chromatography. The minimal inhibitory concentration (MIC) and minimal fungicidal concentration (MFC) of EOs/main compounds were determined according to EUCAST-AFST guidelines, with minor modifications. The checkerboard microtiter method was used to identify putative synergistic combinations of EOs/main constituents with terbinafine. The influence of EOs on the viability and pro-inflammatory cytokine production (IL-1ß, IL-8 and TNF-α) was determined using an ex vivo human neutrophils model. The binary associations of tested EOs with terbinafine were found to be synergistic against T. rubrum, with FICI values of 0.26-0.31. At the tested concentrations (6.25-25 mg/L), EOs did not exert cytotoxic effects towards human neutrophils. Anise EO was the most potent inhibitor of IL-1ß release (46.49% inhibition at 25 mg/L), while coriander EO displayed the highest inhibition towards IL-8 and TNF-α production (54.15% and 54.91%, respectively). In conclusion, the synergistic combinations of terbinafine and investigated Apiaceae EOs could be a starting point in the development of novel topical therapies against T. rubrum-related dermatophytosis.

Honokiol and Magnolol: Insights into Their Antidermatophytic Effects.

Dermatophyte infections represent a significant public health concern, with an alarming negative impact caused by unsuccessful therapeutic regimens. Natural products have been highlighted as a promising alternative, due to their long-standing traditional use and increasing scientific recognition. In this study, honokiol and magnolol, the main bioactives from Magnolia spp. bark, were investigated for their antidermatophytic activity. The antifungal screening was performed using dermatophyte standard strains and clinical isolates. The minimal inhibitory concentration (MIC) and the minimal fungicidal concentration (MFC) were determined in accordance with EUCAST-AFST guidelines, with minor modifications. The effects on ergosterol biosynthesis were assessed in Trichophyton rubrum cells by HPLC-DAD. Putative interactions with terbinafine against T. rubrum were evaluated by the checkerboard method. Their impact on cells' viability and pro-inflammatory cytokines (IL-1ß, IL-8 and TNF-α) was shown using an ex vivo human neutrophils model. Honokiol and magnolol were highly active against tested dermatophytes, with MIC and MFC values of 8 and 16 mg/L, respectively. The mechanism of action involved the inhibition of ergosterol biosynthesis, with accumulation of squalene in T. rubrum cells. Synergy was assessed for binary mixtures of magnolol with terbinafine (FICI = 0.50), while honokiol-terbinafine combinations displayed only additive effects (FICI = 0.56). In addition, magnolol displayed inhibitory effects towards IL-1ß, IL-8 and TNF-α released from lipopolysaccharide (LPS)-stimulated human neutrophils, while honokiol only decreased IL-1ß secretion, compared to the untreated control. Overall, honokiol and magnolol acted as fungicidal agents against dermatophytes, with impairment of ergosterol biosynthesis.

Liposome Permeability to Essential Oil Components: A Focus on Cholesterol Content.

Encapsulation in liposomes has been an efficient strategy to improve the stability of sensitive bioactive compounds such as essential oils (EOs). However, the stability of liposomal formulations remains a key parameter controlling the delivery of encapsulated ingredients. Cholesterol (Chol) modulates the membrane properties conferring stability to the lipid bilayer. Thus, the Chol content in the liposome formulations encapsulating EO components should be carefully chosen. In this work, various liposome formulations differing by Chol content (DPPC:Chol 100:10; 100:25; 100:50; 100:75; 100:100) were exposed to a series of 22 EO components at DPPC/EO 100/25. The formulations were characterized for their final composition and their permeability to the hydrophilic fluorophore, sulforhodamine B (SRB), was monitored. Results showed that the Chol content experimentally determined for the various formulations (above 10% Chol) was below the theoretical weighed Chol. Among the tested components, 13 molecules displayed a significant permeabilizing effect on 10% Chol membranes. Most of these possess a hydroxyl group. The EO induced permeability was dependent on the Chol content which affects the membrane phase: their effect was reduced upon increasing Chol content keeping five EOs components effective at 40% Chol. The EO's effect was also linked to the hydrophobicity of the molecule. Hence, the DPPC:Chol ratio of the formulation is chosen considering the structure of the compound, its hydrophobicity and its effect on the permeability at different Chol content: a formulation comprising 40% Chol is suggested for highly hydrophobic molecules whereas a formulation with higher Chol content could be selected for less hydrophobic compounds.

Effect of quercetin on lipid membrane rigidity: assessment by atomic force microscopy and molecular dynamics simulations.

Quercetin (3,3',4',5,7-pentahydroxyl-flavone) is a natural flavonoid with many valuable biological effects, but its solubility in water is low, posing major limitations in applications. Quercetin encapsulation in liposomes increases its bioavailability; the drug effect on liposome elastic properties is required for formulation development. Here, we quantify the effect of quercetin molecules on the rigidity of lipoid E80 liposomes using atomic force microscopy (AFM) and molecular dynamics (MD) simulations. AFM images show no effect of quercetin molecules on liposomes morphology and structure. However, AFM force curves suggest that quercetin softens lipid membranes; the Young modulus measured for liposomes encapsulating quercetin is smaller than that determined for blank liposomes. We then used MD simulations to interpret the effect of quercetin on membrane rigidity in terms of molecular interactions. The decrease in membrane rigidity was confirmed by the simulations, which also revealed that quercetin affects structural and dynamic properties: membrane thickness is decreased, acyl chains disorder is increased, and diffusion coefficients of lipid molecules are also increased. Such changes appear to be related to the preferential localization of quercetin within the membrane, near the interface between the hydrophobic core and polar head groups of the lipids.

Elastic moduli of lipid membranes: Reproducibility of AFM measures.

Membrane elastic properties play a major role in membrane remodeling events, such as vesicle fusion and fission. They are also crucial in drug delivery by liposomes. Different experimental techniques are available to measure elastic properties. Among them, atomic force microscopy (AFM) presents the unique advantage of being directly applicable to nano-sized liposomes. Unfortunately, different AFM measures reported in the literature show little agreement among each other and are difficult to compare with measures of bending modulus obtained by other experimental techniques or by molecular simulations. In this work we determine the bending rigidity of Egg PC liposomes in terms of Young modulus via AFM measurements, using two different tip shapes and different cantilever force constants. We interpret the measures using the Hertz and Shell models, and observe a clear dependency of the Young modulus values on the tip properties and on the interpretative theory. The effect of the AFM tip shape is less important than the effect of the cantilever force constant, and the mathematical model has a major effect on the interpretation of the data. The Shell theory provides the closest agreement between AFM data and other experimental data for the membrane bending modulus. Finally, we compare the results to calculations of bending modulus from molecular dynamics simulations of membrane buckles. Simulations provide values of bending modulus consistent with literature data, but the agreement with AFM experiments is reasonable only for some specific experimental conditions.

Prevention by Dietary Polyphenols (Resveratrol, Quercetin, Apigenin) Against 7-Ketocholesterol-Induced Oxiapoptophagy in Neuronal N2a Cells: Potential Interest for the Treatment of Neurodegenerative and Age-Related Diseases.

The Mediterranean diet is associated with health benefits due to bioactive compounds such as polyphenols. The biological activities of three polyphenols (quercetin (QCT), resveratrol (RSV), apigenin (API)) were evaluated in mouse neuronal N2a cells in the presence of 7-ketocholesterol (7KC), a major cholesterol oxidation product increased in patients with age-related diseases, including neurodegenerative disorders. In N2a cells, 7KC (50 µM; 48 h) induces cytotoxic effects characterized by an induction of cell death. When associated with RSV, QCT and API (3.125; 6.25 µM), 7KC-induced toxicity was reduced. The ability of QCT, RSV and API to prevent 7KC-induced oxidative stress was characterized by a decrease in reactive oxygen species (ROS) production in whole cells and at the mitochondrial level; by an attenuation of the increase in the level and activity of catalase; by attenuating the decrease in the expression, level and activity of glutathione peroxidase 1 (GPx1); by normalizing the expression, level and activity of superoxide dismutases 1 and 2 (SOD1, SOD2); and by reducing the decrease in the expression of nuclear erythroid 2-like factor 2 (Nrf2) which regulates antioxidant genes. QCT, RSV and API also prevented mitochondrial dysfunction in 7KC-treated cells by counteracting the loss of mitochondrial membrane potential (ΨΔm) and attenuating the decreased gene expression and/or protein level of AMP-activated protein kinase α (AMPKα), sirtuin 1 (SIRT1) and peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α) implicated in mitochondrial biogenesis. At the peroxisomal level, QCT, RSV and API prevented the impact of 7KC by counteracting the decrease in ATP binding cassette subfamily D member (ABCD)3 (a peroxisomal mass marker) at the protein and mRNA levels, as well as the decreased expresssion of genes associated with peroxisomal biogenesis (Pex13, Pex14) and peroxisomal ß-oxidation (Abcd1, Acox1, Mfp2, Thiolase A). The 7KC-induced decrease in ABCD1 and multifunctional enzyme type 2 (MFP2), two proteins involved in peroxisomal ß-oxidation, was also attenuated by RSV, QCT and API. 7KC-induced cell death, which has characteristics of apoptosis (cells with fragmented and/or condensed nuclei; cleaved caspase-3; Poly(ADP-ribose) polymerase (PARP) fragmentation) and autophagy (cells with monodansyl cadaverine positive vacuoles; activation of microtubule associated protein 1 light chain 3-I (LC3-I) to LC3-II, was also strongly attenuated by RSV, QCT and API. Thus, in N2a cells, 7KC induces a mode of cell death by oxiapoptophagy, including criteria of OXIdative stress, APOPTOsis and autoPHAGY, associated with mitochondrial and peroxisomal dysfunction, which is counteracted by RSV, QCT, and API reinforcing the interest for these polyphenols in prevention of diseases associated with increased 7KC levels.

Development of cysteamine loaded liposomes in liquid and dried forms for improvement of cysteamine stability.

Despite the high aqueous solubility of cysteamine, its unpleasant organoleptic properties, hygroscopicity, instability in solutions, and poor pharmacokinetic profile are the main drawbacks that limit its use for medical and cosmetic purposes. In this study, cysteamine-loaded liposomes were prepared using the ethanol injection method. Liposomes were characterized for their size, homogeneity, surface charge, and morphology. The incorporation ratios of cholesterol and phospholipids, the encapsulation efficiency and the loading ratio of cysteamine in liposomes were determined. Moreover, the stability of free and encapsulated cysteamine was assessed at different temperatures (4, 25, and 37 °C) in the presence and absence of light. Cysteamine-loaded liposomes were freeze-dried and reconstituted liposomes were characterized. Finally, the storage stability of the freeze-dried cysteamine-loaded liposomes was studied. Liposomes were nanometric, oligolamellar, and spherical. The encapsulation efficiency and the loading ratio of cysteamine varied between 12 and 40% in the different formulations. The encapsulation improved the stability of cysteamine in the various storage conditions. The dried form of cysteamine-loaded liposomes conserved the size of the vesicles and retained 33% of cysteamine present in the liposomal suspension before lyophilization. The freeze-dried liposomes formulations were stable after four months of storage at 4 °C.

Recent advances in tackling microbial multidrug resistance with essential oils: combinatorial and nano-based strategies.

The emergence of multidrug resistant pathogens is a great challenge to the medical field and a major global health threat requiring novel therapeutic strategies. Among plant products, essential oils have significant antimicrobial properties that make them promising agents in the fight against drug resistant human pathogens. The aim of the present review was to highlight the most important essential oil-based antimicrobial strategies as revealed by recent studies. Synergistic interactions between essential oils or their bioactive compounds in combination with known antibiotics are presented. Also, nanoformulation approaches to boost the antimicrobial activity of essential oils are reviewed in terms of bioefficiency, stability and design of the nanostructured delivery systems. The focus was mainly put on the antimicrobial activity against multi-drug resistant pathogens, also called "ESKAPE" organisms (Enterococcus spp., Staphylococcus aureus, Klebsiella spp., Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter spp.). Thus, essential oils in combinatorial and nano-based strategies may cope with infections caused by drug-resistant bacteria and may offer possibilities for reducing antibiotic use. Research on the in vivo efficacy and safety of such strategies is required for further clinical antimicrobial chemotherapy. In this regard, the understanding of the interactions between essential oil-based strategies and biological interface is essential.

Prevention of 7-Ketocholesterol-Induced Overproduction of Reactive Oxygen Species, Mitochondrial Dysfunction and Cell Death with Major Nutrients (Polyphenols, ω3 and ω9 Unsaturated Fatty Acids) of the Mediterranean Diet on N2a Neuronal Cells.

The brain, which is a cholesterol-rich organ, can be subject to oxidative stress in a variety of pathophysiological conditions, age-related diseases and some rare pathologies. This can lead to the formation of 7-ketocholesterol (7KC), a toxic derivative of cholesterol mainly produced by auto-oxidation. So, preventing the neuronal toxicity of 7KC is an important issue to avoid brain damage. As there are numerous data in favor of the prevention of neurodegeneration by the Mediterranean diet, this study aimed to evaluate the potential of a series of polyphenols (resveratrol, RSV; quercetin, QCT; and apigenin, API) as well as ω3 and ω9 unsaturated fatty acids (α-linolenic acid, ALA; eicosapentaenoic acid, EPA; docosahexaenoic acid, DHA, and oleic acid, OA) widely present in this diet, to prevent 7KC (50 µM)-induced dysfunction of N2a neuronal cells. When polyphenols and fatty acids were used at non-toxic concentrations (polyphenols: ≤6.25 µM; fatty acids: ≤25 µM) as defined by the fluorescein diacetate assay, they greatly reduce 7KC-induced toxicity. The cytoprotective effects observed with polyphenols and fatty acids were comparable to those of α-tocopherol (400 µM) used as a reference. These polyphenols and fatty acids attenuate the overproduction of reactive oxygen species and the 7KC-induced drop in mitochondrial transmembrane potential (ΔΨm) measured by flow cytometry after dihydroethidium and DiOC6(3) staining, respectively. Moreover, the studied polyphenols and fatty acids reduced plasma membrane permeability considered as a criterion for cell death measured by flow cytometry after propidium iodide staining. Our data show that polyphenols (RSV, QCT and API) as well as ω3 and ω9 unsaturated fatty acids (ALA, EPA, DHA and OA) are potent cytoprotective agents against 7KC-induced neurotoxicity in N2a cells. Their cytoprotective effects could partly explain the benefits of the Mediterranean diet on human health, particularly in the prevention of neurodegenerative diseases.

New generation of supramolecular mixtures: Characterization and solubilization studies.

In this work, a series of novel low melting mixtures (LMM) based on cyclodextrins (CD) and levulinic acid and inspired by the deep eutectic solvents (DES), were prepared. These supramolecular mixtures are the first reported CD-based mixtures that are liquid at room temperature. Density, viscosity and rheological measurements as well as differential scanning calorimetry and thermogravimetric analysis were performed to characterize these new LMM. Nuclear magnetic resonance (NMR) spectroscopy was used to monitor their stability. Furthermore, their ability to solubilize trans-anethole (AN) and related essentials oils were evaluated by static headspace-gas chromatography (SH-GC), in comparison with water. AN was up to 1300 times more soluble in the CD-based LMM than in water. Finally, multiple headspace extraction (MHE) was used to monitor the release of AN from these LMM. After 10 extractions, 20 to 40% of AN was released from the studied LMM, while 70% was released from water. The new CD-based LMM have potential applications for solubilization and delivery of poorly soluble drugs.

Liposomal membrane permeability assessment by fluorescence techniques: Main permeabilizing agents, applications and challenges.

Liposomes are lipid vesicles made of one or multiple lipid bilayers surrounding an internal aqueous core. They are broadly employed as models to study membrane structure and properties. Among these properties, liposome membrane permeability is crucial and widely assessed by fluorescence techniques. The first part of this review is devoted to describe the various techniques used for membrane permeability assessment. Attention is paid to fluorescence techniques based on vesicle leakage of self-quenching probes, dye/quencher pair or cation/ligand pair. Secondly, the membrane-active agents inducing membrane permeabilization is presented and details on their mechanisms of action are given. Emphasis is also laid on the intrinsic and extrinsic factors that can modulate the membrane permeability. Hence, a suitable liposomal membrane should be formulated according to the aim of the study and its application.

Cyclodextrins: from solute to solvent.

A supramolecular solvent based on cyclodextrin (CD) is presented here for the first time. Indeed, a low melting mixture was obtained by mixing levulinic acid and a CD derivative, which retained its inclusion ability in the resulting solvent. This new system gives rise to a new family of solvents that could be called SUPRADES (supramolecular deep eutectic solvents).