Synthesis of C3-symmetric star shaped amphiphiles for drug delivery applications.

C 3-symmetric star-shaped aromatic compounds are known to possess unique characteristics which facilitate their industrial and biomedical applications. Herein, we report the design, synthesis, self-assembly and drug/dye delivery capabilities of C3-symmetric, hexa-substituted benzene-based amphiphiles. The synthesis of the hexa-substituted C3-symmetric core involves C-acetylation of phloroglucinol to yield the corresponding tri-acetyl derivative. This was further subjected to O-propargylation, followed by the carbonyl reduction of acetyl groups to yield the central core. Various hydrophilic (mPEG) and lipophilic units were then incorporated into this core via click and esterification reactions, respectively, to produce a new type of star shaped amphiphiles. So the obtained amphiphilic architectures have a tendency to aggregate in an aqueous medium forming nanosized assemblies with an inner hydrophobic core, allowing the substituents to control the tension-active properties. The critical aggregation concentration of the amphiphiles was evaluated by fluorescence measurement using the dye Nile red as a fluorescent probe. The hydrodynamic diameter of self-assembled aggregates in aqueous solution was studied by dynamic light scattering, while the actual size and morphology were determined by cryo-transmission electron microscopy (cryo-TEM) analysis. The physicochemical properties of the amphiphiles suggested their suitability for exploring their drug delivery applications. In this endeavor, the amphiphiles were utilized for the encapsulation of model hydrophobic entities and studying their subsequent release from their hydrophobic core in a controlled manner. The transport potential of the synthesised amphiphiles was explored for transdermal drug delivery. Furthermore, cytotoxicity studies were conducted using MCF7 and HeLa cells, which indicated that the nanocarriers had no toxic effect on the cells.

Comparing the effects of three different multilayer dressings for pressure ulcer prevention on sacral skin after prolonged loading: An exploratory crossover trial.

Evidence suggests that preventive dressings applied on sacral skin help to prevent pressure ulcers. However, possible performance differences of different dressing types are unclear. An exploratory randomized crossover trial with intra-individual comparisons was conducted to compare the effects of three different multi-layer foam dressings (Mepilex Border Sacrum, ALLEVYN Life Sacrum and Optifoam Gentle Sacrum) compared to no dressing on the sacral skin. Healthy female volunteers (n = 12, mean age 72 years) wore three different dressings on their sacral skin for 3.5 hours while lying supine on a standard hospital mattress. At regular intervals, subjects performed standardized movements to enhance shear loads. Skin surface temperature, stratum corneum hydration, erythema, skin roughness and the interleukin 1 alpha (IL-1α) concentration per total protein were measured at baseline and after the lying periods. After 3.5 hours, the median skin temperature increased in all four groups between 3.0°C and 3.8°C with only minor differences between the no dressing and the dressing groups. Median stratum corneum hydration increased during the lying period in all groups with highest increases in the Optifoam Gentle Sacrum (7.3 arbitrary units) and no dressing group (7.0 arbitrary units). There was a median decrease of the mean roughness (Rz) in the Optifoam Gentle Sacrum group of -6.3 µm but no relevant changes in the other groups. After loading, the erythema index was highest in the ALLEVYN Life Sacrum and no dressing groups. Highest releases of IL-1α were observed in the ALLEVYN Life Sacrum and Optifoam Gentle Sacrum groups, in the Mepilex Border Sacrum group changes were minor. Study results indicate, that the application of preventive dressings on sacral skin during loading do not cause additional occlusion compared to loading without dressings when lying supine. Different dressings cause different cutaneous responses during loading.

A niche in the spotlight: Could external factors critically disturb hair follicle homeostasis and contribute to inflammatory hair follicle diseases?

The anatomy of the hair follicle and the dynamics of its barrier provide a special space for interactions between macromolecules and the underlying tissue. Translocation across the hair follicle epithelium and immune recognition have been confirmed for proteins, nucleic acids, engineered particles, virus particles and others. Tissue responses can be modulated by pro-inflammatory stimuli as demonstrated in penetration and transcutaneous immunization studies. Even under physiological conditions, hair follicle openings are filled with exogenous material ranging from macromolecules, engineered particles to natural particles including diverse communities of microbes. The exposed position of the infundibulum suggests that local inflammatory insults could disturb the finely tuned balance and may trigger downstream responses that initiate or facilitate local outbreaks of inflammatory hair diseases typically occurring in close spatial association with the infundibulum as observed in cicatricial alopecia. The question as to how microbial colonization or deposition of contaminants on the surface of the hair follicle epithelium interact with the barrier status under the influence of individual predisposition, may help us understand local flare-ups of inflammatory hair diseases. Specifically, learning more about skin barrier alterations in the different types of inflammatory hair diseases and cross-talk with exogenous compounds could give new insights in this less explored aspect of hair follicle homeostasis. Such knowledge may not only be used to develop supportive measures to maintain a healthy scalp. It may have wider implications for our understanding on how external factors influence inflammation and immunological responses in the skin.

Data-based modeling of drug penetration relates human skin barrier function to the interplay of diffusivity and free-energy profiles.

Based on experimental concentration depth profiles of the antiinflammatory drug dexamethasone in human skin, we model the time-dependent drug penetration by the 1D general diffusion equation that accounts for spatial variations in the diffusivity and free energy. For this, we numerically invert the diffusion equation and thereby obtain the diffusivity and the free-energy profiles of the drug as a function of skin depth without further model assumptions. As the only input, drug concentration profiles derived from X-ray microscopy at three consecutive times are used. For dexamethasone, skin barrier function is shown to rely on the combination of a substantially reduced drug diffusivity in the stratum corneum (the outermost epidermal layer), dominant at short times, and a pronounced free-energy barrier at the transition from the epidermis to the dermis underneath, which determines the drug distribution in the long-time limit. Our modeling approach, which is generally applicable to all kinds of barriers and diffusors, allows us to disentangle diffusivity from free-energetic effects. Thereby we can predict short-time drug penetration, where experimental measurements are not feasible, as well as long-time permeation, where ex vivo samples deteriorate, and thus span the entire timescales of biological barrier functioning.

Modeling of Drug Diffusion Based on Concentration Profiles in Healthy and Damaged Human Skin.

Based on experimental drug concentration profiles in healthy as well as tape-stripped ex vivo human skin, we model the penetration of the antiinflammatory drug dexamethasone into the skin layers by the one-dimensional generalized diffusion equation. We estimate the position-dependent free-energy and diffusivity profiles by solving the conjugated minimization problem, in which the only inputs are concentration profiles of dexamethasone in skin at three consecutive penetration times. The resulting free-energy profiles for damaged and healthy skin show only minor differences. In contrast, the drug diffusivity in the first 10 µm of the upper skin layer of damaged skin is 200-fold increased compared to healthy skin, which reflects the corrupted barrier function of tape-stripped skin. For the case of healthy skin, we examine the robustness of our method by analyzing the behavior of the extracted skin parameters when the number of input and output parameters are reduced. We also discuss techniques for the regularization of our parameter extraction method.

Qualifying X-ray and Stimulated Raman Spectromicroscopy for Mapping Cutaneous Drug Penetration.

Research on topical drug delivery relies on reconstructed human skin (RHS) in addition to ex vivo human and animal skin, each with specific physiological features. Here, we compared the penetration of dexamethasone from an ethanolic hydroxyethyl cellulose gel into ex vivo human skin, murine skin, and RHS. For comprehensive insights into skin morphology and penetration enhancing mechanisms, scanning transmission X-ray microscopy (STXM), liquid chromatography tandem-mass spectrometry (LC-MS/MS), and stimulated Raman spectromicroscopy (SRS) were combined. STXM offers high spatial resolution with label-free drug detection and is therefore sensitive to tissue damage. Despite differences in sample preparation and data analysis, the amounts of dexamethasone in RHS, detected and quantified by STXM and LC-MS/MS, were very similar and increased during the first 100 min of exposure. SRS revealed interactions between the gel and the stratum corneum or, more specifically, its protein and lipid structures. Similar to both types of ex vivo skin, higher protein-to-lipid ratios within the stratum corneum of RHS indicated reduced lipid amounts after 30 min of ethanol exposure. Extended ethanol exposure led to a continued reduction of lipids in the ex vivo matrixes, while protein integrity appeared to be compromised in RHS, which led to declining protein signals. In conclusion, LC-MS/MS proved the predictive capability of STXM for label-free drug detection. Combining STXM with SRS precisely dissected the penetration enhancing effects of ethanol. Further studies on topical drug delivery should consider the potential of these complementary techniques.

Shape-Dependent Dissolution and Cellular Uptake of Silver Nanoparticles.

The cellular uptake and dissolution of trigonal silver nanoprisms (edge length 42 ± 15 nm, thickness 8 ± 1 nm) and mostly spherical silver nanoparticles (diameter 70 ± 25 nm) in human mesenchymal stem cells (hMSC's) and human keratinocytes (HaCaT cells) were investigated. Both particles are stabilized by polyvinylpyrrolidone (PVP), with the prisms additionally stabilized by citrate. The nanoprisms dissolved slightly in pure water but strongly in isotonic saline or at pH 4, corresponding to the lowest limit for the pH during cellular uptake. The tips of the prisms became rounded within minutes due to their high surface energy. Afterward, the dissolution process slowed down due to the presence of both PVP stabilizing Ag{100} sites and citrate blocking Ag{111} sites. On the contrary, nanospheres, solely stabilized by PVP, dissolved within 24 h. These results correlate with the finding that particles in both cell types have lost >90% of their volume within 24 h. hMSC's took up significantly more Ag from nanoprisms than from nanospheres, whereas HaCaT cells showed no preference for one particle shape. This can be rationalized by the large cellular interaction area of the plateletlike nanoprisms and the bending stiffness of the cell membranes. hMSC's have a highly flexible cell membrane, resulting in an increased uptake of plateletlike particles. HaCaT cells have a membrane with a 3 orders of magnitude higher Young's modulus than for hMSC. Hence, the energy gain due to the larger interaction area of the nanoprisms is compensated for by the higher energy needed for cell membrane deformation compared to that for spheres, leading to no shape preference.

Comparison of different methods to study effects of silver nanoparticles on the pro- and antioxidant status of human keratinocytes and fibroblasts.

In times of antibiotic-resistant bacteria new strategies to avoid the septic-inducing threat of dangerous microorganisms are needed. Silver ions (Ag+) in the forms of silver nitrate or silver sulfadiazine have been used as antimicrobial agents for years. A step further was the development of micro and silver particles (AgNP). In contrast to other Ag+ ion sources, AgNP allow a sustained release of Ag+ ions, due to their high surface to volume ratio. However, AgNP are also toxic to eukaryotic cells and the mechanisms of cytotoxicity have not yet been fully elucidated. In this study, the impact of different AgNP preparations on a human keratinocyte cell line was investigated. The intracellular radical formation was confirmed by the 2',7'-dichlorodihydrofluorescein di-acetate (H2DCF-DA) assay on two cell types (HaCaT cells and normal human dermal fibroblasts) as well as by electron paramagnetic resonance (EPR) spectroscopy, which showed comparable results. EPR spectroscopy was performed for the first time for 24h in experiments using keratinocytes. Drastic changes in the mitochondrial activity were induced in cells incubated with AgNP containing high concentrations of Ag+ ions. It was also possible to show that the quantitative uptake of AgNP was dependent on the AgNP concentration. In addition, the effects of AgNP on the GSH/GSSG system were elucidated. The results showed a batch- and concentration-dependent decrease of the total glutathione concentration which correlated well with the decrease of cell viability. Furthermore, the results suggest a direct reaction of GSH molecules with Ag+ ions. In conclusion, this study proves the efficacy of the H2DCF-DA assay and the EPR spectroscopy. The investigations show that AgNP formulations containing high amounts of released Ag+ ions induce radicals in human keratinocytes and deplete them of their natural anti-oxidative molecules. On the contrary, nanoparticles prepared and stored under argon did not induce significant adverse effects, suggesting that slowing down the release of Ag+ may help to reduce AgNP-related side effects without affecting the antibacterial impact.

Hair follicle targeting, penetration enhancement and Langerhans cell activation make cyanoacrylate skin surface stripping a promising delivery technique for transcutaneous immunization with large molecules and particle-based vaccines.

Transcutaneous immunization (TCI) requires targeting of a maximum number of skin antigen-presenting cells as non-invasive as possible on small skin areas. In two clinical trials, we introduced cyanoacrylate skin surface stripping (CSSS) as a safe method for TCI. Here, using ex vivo human skin, we demonstrate that one CSSS procedure removed only 30% of stratum corneum, but significantly increased the penetration of 200 nm polystyrene particles deep into vellus and intermediate hair follicles from where they could not been retrieved by conventional tape stripping. Two subsequent CSSS had no striking additional effect. CSSS increased particle penetration in superficial stratum corneum and induced Langerhans cell activation. Formulation in amphiphilic ointment or massage did not substantially influences the interfollicular penetration profiles. Hair follicle (HF) targeting by CSSS could become a highly effective tool for TCI when combined with carrier-based delivery and is gaining new attention as our understanding on the HF immune system increases.

Application of single molecule fluorescence microscopy to characterize the penetration of a large amphiphilic molecule in the stratum corneum of human skin.

We report here on the application of laser-based single molecule total internal reflection fluorescence microscopy (TIRFM) to study the penetration of molecules through the skin. Penetration of topically applied drug molecules is often observed to be limited by the size of the respective drug. However, the molecular mechanisms which govern the penetration of molecules through the outermost layer of the skin are still largely unknown. As a model compound we have chosen a larger amphiphilic molecule (fluorescent dye ATTO-Oxa12) with a molecular weight >700 Da that was applied to excised human skin. ATTO-Oxa12 penetrated through the stratum corneum (SC) into the viable epidermis as revealed by TIRFM of cryosections. Single particle tracking of ATTO-Oxa12 within SC sheets obtained by tape stripping allowed us to gain information on the localization as well as the lateral diffusion dynamics of these molecules. ATTO-Oxa12 appeared to be highly confined in the SC lipid region between (intercellular space) or close to the envelope of the corneocytes. Three main distinct confinement sizes of 52 ± 6, 118 ± 4, and 205 ± 5 nm were determined. We conclude that for this amphiphilic model compound several pathways through the skin exist.

Effects of a leave-on product on the strength of the dermoepidermal junction: An exploratory, intraindividual, randomized controlled trial in older adults with dry skin.

Background and Aims: Skin aging is associated with dry skin and a decrease of the strength of the dermoepidermal adhesion, which increases the risk for lacerations (skin tears). Application of leave-on products improves dry skin and seems to reduce skin tear incidence. The aim of this study was to measure the effects of a humectant containing leave-on product on the strength of the dermoepidermal junction in older adult participants with dry skin. Methods: A randomized controlled trial using a split body design was conducted. One forearm was randomly selected and treated with a lipophilic leave-on product containing 5% urea for 8 weeks. The other forearm was the control. The parameters stratum corneum hydration (SCH), transepidermal water loss, pH, roughness, epidermal thickness and skin stiffness were measured at the baseline, Weeks 4 and 8. At Week 8, suction blisters were created and time to blistering was measured. Blister roofs and interstitial fluid were analyzed for Interleukin-1α, 6 and 8. Results: Twelve participants were included. After 8 weeks treatment, SCH was higher (median difference 11.6 AU), and the overall dry skin score (median difference -1) and median roughness (Rz difference -12.2 µm) were lower compared to the control arms. The median group difference for Interleukin-1α was -452 fg/µg total protein (TP) in the blister roofs and -2.2 fg/µg TP in the blister fluids. The median time to blister formation was 7.7 min higher compared to the control arms. Conclusion: The regular application of humectant containing leave-on products improves dry skin and seems to lower inflammation and contribute to the strengthening of the dermoepidermal adhesion. This partly explains how the use of topical leave-on products helps to prevent skin tears.

Modulating the immune system towards a functional chronic wound healing: A biomaterials and Nanomedicine perspective.

Chronic non-healing wounds persist as a substantial burden for healthcare systems, influenced by factors such as aging, diabetes, and obesity. In contrast to the traditionally pro-regenerative emphasis of therapies, the recognition of the immune system integral role in wound healing has significantly grown, instigating an approach shift towards immunological processes. Thus, this review explores the wound healing process, highlighting the engagement of the immune system, and delving into the behaviors of innate and adaptive immune cells in chronic wound scenarios. Moreover, the article investigates biomaterial-based strategies for the modulation of the immune system, elucidating how the adjustment of their physicochemical properties or their synergistic combination with other agents such as drugs, proteins or mesenchymal stromal cells can effectively modulate the behaviors of different immune cells. Finally this review explores various strategies based on synthetic and biological nanostructures, including extracellular vesicles, to finely tune the immune system as natural immunomodulators or therapeutic nanocarriers with promising biophysical properties.

Topical Delivery of Tofacitinib in Dermatology: The Promise of a Novel Therapeutic Class Using Biodegradable Dendritic Polyglycerol Sulfates.

Inflammatory skin diseases, such as psoriasis, atopic dermatitis, and alopecia areata, occur when the regulatory tolerance of the innate immune system is disrupted, resulting in the activation of the Janus kinase-signal transducer and activator of transcription (JAK-STAT) inflammatory signaling pathway by interleukin 6 (IL-6) and other key inflammatory cytokines. JAK inhibitors, such as tofacitinib, bind to these enzymes which are coupled to receptors on cell surfaces and block the transcription of inflammatory cytokine-induced genes. The first topical applications are being marketed, yet insufficient effects regarding indications, such as alopecia areata, suggest that improved delivery technologies could help increase the efficacy. In this study, we used sulfated dendritic polyglycerol with caprolactone segments integrated in its backbone (dPGS-PCL), with a molecular weight of 54 kDa, as a degradable carrier to load and solubilize the hydrophobic drug tofacitinib (TFB). TFB loaded in dPGS-PCL (dPGS-PCL@TFB), at a 11 w/w% loading capacity in aqueous solution, showed in an ex-vivo human skin model better penetration than free TFB in a 30:70 (v/v) ethanol/water mixture. We also investigated the anti-inflammatory efficacy of dPGS-PCL@TFB (0.5 w/w%), dPGS-PCL, and free TFB in the water/ethanol mixture by measuring their effects on IL-6 and IL-8 release, and STAT3 and STAT5 activation in ex vivo skin models of simulated inflamed human skin. Our results suggest that dPGS-PCL@TFB reduces the activation of STAT3 and STAT5 by increasing the penetration of the tofacitinib. However, no statistically significant differences with respect to the inhibition of IL-6 and IL-8 were observed in this short incubation time.

Efficient skin interactions of graphene derivatives: challenge, opportunity or both?

Interactions between graphene, with its wide deployment in consumer products, and skin, the body's largest organ and first barrier, are highly relevant with respect to toxicology and dermal delivery. In this work, interaction of polyglycerol-functionalized graphene sheets, with 200 nm average lateral size and different surface charges, and human skin was studied and their potential as topical delivery systems were investigated. While neutral graphene sheets showed no significant skin interaction, their positively and negatively charged counterparts interacted with the skin, remaining in the stratum corneum. This efficient skin interaction bears a warning but also suggests a new topical drug delivery strategy based on the sheets' high loading capacity and photothermal property. Therefore, the immunosuppressive drug tacrolimus was loaded onto positively and negatively charged graphene sheets, and its release measured with and without laser irradiation using liquid chromatography tandem-mass spectrometry. Laser irradiation accelerated the release of tacrolimus, due to the photothermal property of graphene sheets. In addition, graphene sheets with positive and negative surface charges were loaded with Nile red, and their ability to deliver this cargo through the skin was investigated. Graphene sheets with positive surface charge were more efficient than the negatively charged ones in enhancing Nile red penetration into the skin.

Ciprofloxacin-Loaded Polyvinylpyrrolidone Foils for the Topical Treatment of Wound Infections with Methicillin-Resistant Staphylococcus aureus (MRSA).

Bacterial infections are a constant challenge in the management of acute and chronic wounds. Chronic wounds, such as diabetic foot ulcers, have increased significantly in the last few years due to the rise of an aging population. A better understanding of the infectious pathophysiological mechanisms is urgently needed along with new options for the treatment of wound infections and wound-healing disorders. New advances in the preparation of biocompatible dressing materials that can be loaded with antimicrobial drugs may improve the topical treatment of infected wounds. In this study, we investigated the antimicrobial activity of polyvinylpyrrolidone (PVP) foils loaded with ciprofloxacin (Cipro-foils) in the presence of acetic acid as a co-solvent. We used ex vivo human wounds that were infected with two bacterial strains: methicillin-resistant Staphylococcus aureus (MRSA) or Pseudomonas aeruginosa (PAO1). The effectiveness of the treatment was demonstrated by the quantification of the living bacteria extracted from the wound and the detection of released immunological mediators in skin extracts and in the skin culture media. We found that Cipro-foils effectively treated the infection with both PAO1 and MRSA. Other than PAO1, MRSA had no lytic activity toward skin proteins. MRSA infections increased cytokines' expression and release. Interestingly, treatment with Cipro-foils could partially counteract these effects.

Surface functionalization of silica nanoparticles supports colloidal stability in physiological media and facilitates internalization in cells.

The influence of the surface functionalization of silica particles on their colloidal stability in physiological media is studied and correlated with their uptake in cells. The surface of 55 ± 2 nm diameter silica particles is functionalized by amino acids or amino- or poly(ethylene glycol) (PEG)-terminated alkoxysilanes to adjust the zeta potential from highly negative to positive values in ethanol. A transfer of the particles into water, physiological buffers, and cell culture media reduces the absolute value of the zeta potential and changes the colloidal stability. Particles stabilized by L-arginine, L-lysine, and amino silanes with short alkyl chains are only moderately stable in water and partially in PBS or TRIS buffer, but aggregate in cell culture media. Nonfunctionalized, N-(6-aminohexyl)-3-aminopropyltrimethoxy silane (AHAPS), and PEG-functionalized particles are stable in all media under study. The high colloidal stability of positively charged AHAPS-functionalized particles scales with the ionic strength of the media, indicating a mainly electrostatical stabilization. PEG-functionalized particles show, independently from the ionic strength, no or only minor aggregation due to additional steric stabilization. AHAPS stabilized particles are readily taken up by HeLa cells, likely as the positive zeta potential enhances the association with the negatively charged cell membrane. Positively charged particles stabilized by short alkyl chain aminosilanes adsorb on the cell membrane, but are weakly taken up, since aggregation inhibits their transport. Nonfunctionalized particles are barely taken up and PEG-stabilized particles are not taken up at all into HeLa cells, despite their high colloidal stability. The results indicate that a high colloidal stability of nanoparticles combined with an initial charge-driven adsorption on the cell membrane is essential for efficient cellular uptake.

Effects of Adipose-Derived Stem Cells and Their Conditioned Medium in a Human Ex Vivo Wound Model.

Adult stem cells have been extensively investigated for tissue repair therapies. Adipose-derived stem cells (ASCs) were shown to improve wound healing by promoting re-epithelialization and vascularization as well as modulating the inflammatory immune response. In this study, we used ex vivo human skin cultured in a six-well plate with trans-well inserts as a model for superficial wounds. Standardized wounds were created and treated with allogeneic ASCs, ASCs conditioned medium (ASC-CM), or cell culture medium (DMEM) supplemented with fetal calf serum (FCS). Skin viability (XTT test), histology (hematoxylin and eosin, H and E), ß-catenin expression as well as inflammatory mediators and growth factors were monitored over 12 days of skin culture. We observed only a moderate time-dependent decrease in skin metabolic activity while skin morphology was preserved, and re-epithelialization occurred at the wound edges. An increase in ß-catenin expression was observed in the newly formed epithelia, especially in the samples treated with ASC-CM. In general, increased growth factors and inflammatory mediators, e.g., hepatocytes growth factor (HGF), platelet-derived growth factor subunit AA (PDGF-AA), IL-1α, IL-7, TNF-α, and IL-10, were observed over the incubation time. Interestingly, different expression profiles were observed for the different treatments. Samples treated with ASC-CM significantly increased the levels of inflammatory cytokines and PDGF-AA with respect to control, whereas the treatment with ASCs in DMEM with 10% FCS resulted in significantly increased levels of fibroblast growth factor-basic (FGF-basic) and moderate increases of immunomodulatory cytokines. These results confirm that the wound microenvironment can influence the type of mediators secreted by ASCs and the mode as to how they improve the wound healing process. Comparative investigations with pre-activated ASCs will elucidate further aspects of the wound healing mechanism and improve the protocols of ACS application.

Fabrication of hydrolase responsive diglycerol based Gemini amphiphiles for dermal drug delivery applications.

Since biocatalysts manoeuvre most of the physiological activities in living organisms and exhibit extreme selectivity and specificity, their use to trigger physicochemical change in polymeric architectures has been successfully used for targeted drug delivery. Our major interest is to develop lipase responsive nanoscale delivery systems from bio-compatible and biodegradable building blocks. Herein, we report the synthesis of four novel non-ionic Gemini amphiphiles using a chemo-enzymatic approach. A symmetrical diglycerol has been used as a core that is functionalised with alkyl chains for the creation of a hydrophobic cavity, and for aqueous solubility (polyethylene glycol) monomethyl ether (mPEG) is incorporated. Such systems can exhibit a varied self-assembly behaviour leading to the observance of different morphological structures. The aggregation behaviour of the synthesised nanocarrier was studied by dynamic light scattering (DLS) and critical aggregation concentration (CAC) measurements. The nanotransport potential of amphiphiles was investigated for hydrophobic guest molecules, i.e. Nile red, nimodipine and curcumin. Cytotoxicity of the amphiphiles was studied using HeLa and MCF7 cell lines at different concentrations, i.e. 0.05, 0.1, and 0.5 mg mL-1. All nanocarriers were found to be non-cytotoxic up to a concentration of 0.1 mg mL-1. Confocal laser scanning microscopy (cLSM) study suggested the uptake of encapsulated dye in the cytosol of the cancer cells within 4 h, thus implying that amphiphilic systems can efficiently transport hydrophobic drug molecules into cells. The biomedical application of the synthesised Gemini amphiphiles was also investigated for dermal drug delivery. In addition, the enzyme-mediated release study was performed that demonstrated 90% of the dye is released within three days. All these results supported the capability of nanocarriers in drug delivery systems.

Improved Skin Permeability after Topical Treatment with Serine Protease: Probing the Penetration of Rapamycin by Scanning Transmission X-ray Microscopy.

Drug penetration in human skin ex vivo following a modification of skin barrier permeability is systematically investigated by scanning transmission X-ray microscopy. Element-selective excitation is used in the O 1s regime for probing quantitatively the penetration of topically applied rapamycin in different formulations with a spatial resolution reaching <75 nm. The data were analyzed by a comparison of two methods: (i) two-photon energies employing the Beer-Lambert law and (ii) a singular value decomposition approach making use of the full spectral information in each pixel of the X-ray micrographs. The latter approach yields local drug concentrations more reliably and sensitively probed than the former. The present results from both approaches indicate that rapamycin is not observed within the stratum corneum of nontreated skin ex vivo, providing evidence for the observation that this high-molecular-weight drug inefficiently penetrates intact skin. However, rapamycin is observed to penetrate more efficiently the stratum corneum when modifications of the skin barrier are induced by the topical pretreatment with the serine protease trypsin for variable time periods ranging from 2 to 16 h. After the longest exposure time to serine protease, the drug is even found in the viable epidermis. High-resolution micrographs indicate that the lipophilic drug preferably associates with corneocytes, while signals found in the intercellular lipid compartment were less pronounced. This result is discussed in comparison to previous work obtained from low-molecular-weight lipophilic drugs as well as polymer nanocarriers, which were found to penetrate the intact stratum corneum exclusively via the lipid layers between the corneocytes. Also, the role of the tight junction barrier in the stratum granulosum is briefly discussed with respect to modifications of the skin barrier induced by enhanced serine protease activity, a phenomenon of clinical relevance in a range of inflammatory skin disorders.

Screening of Surfactants for Improved Delivery of Antimicrobials and Poly-Lactic-co-Glycolic Acid Particles in Wound Tissue.

Topical wound management is often a challenge due to the poor penetration of antimicrobials in wound tissue and across the biofilm matrix where bacteria are embedded. Surfactants have been used for decades to improve the stability of formulations, increase drug solubility, and enhance penetration. In this study, we screened different detergents with respect to their cytotoxicity and their ability to improve the penetration of poly-lactic-co-glycolic acid (PLGA) particles in wound tissue. Among the tested surfactants, Kolliphor SLS and Tween 80 increased the penetration of PLGA particles and had a limited cytotoxicity. Then, these surfactants were used to formulate PLGA particles loaded with the poorly water-soluble antibiotic ciprofloxacin. The antimicrobial efficacy of the formulations was tested in a wound infection model based on human ex vivo skin. We found that even though PLGA particles had the same antimicrobial efficiency than the particle-free drug formulation, thanks to their solubilizing and anti-biofilm properties, the surfactants remarkably improved the antimicrobial activity of ciprofloxacin with respect to the drug formulation in water. We conclude that the use of Tween 80 in antimicrobial formulations might be a safe and efficient option to improve the topical antimicrobial management of chronic wound infections.