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Summary: Background: Peeling skin syndrome type 1 (PSS1) is a rare and severe autosomal recessive form of congenital ichthyosis. Patients are affected by pronounced erythroderma accompanied by pruritus and superficial generalized peeling of the skin. The disease is caused by nonsense mutations or complete deletion of the CDSN gene encoding for corneodesmosin (CDSN). PSS1 severely impairs quality of life and therapeutic approaches are totally unsatisfactory. Objectives: The objective of this study was to develop the first steps towards a specific protein replacement therapy for CDSN deficiency. Using this approach, we aimed to restore the lack of CDSN and improve cell–cell cohesion in the transition area of the stratum granulosum (SG) to the stratum corneum. Methods: Human CDSN was recombinantly expressed in Escherichia coli. A liposome‐based carrier system, prepared with a cationic lipopeptide to mediate the transport to the outer membrane of keratinocytes, was developed. This formulation was chosen for CDSN delivery into the skin. The liposomal carrier system was characterized with respect to size, stability and toxicity. Furthermore, the interaction with primary keratinocytes and human epidermal equivalents was investigated. Results: The liposomes showed an accumulation at the membranes of keratinocytes. CDSN‐deficient epidermal equivalents that were treated with liposomal encapsulated CDSN demonstrated presence of CDSN in the SG. Finally, the penetration assay and histological examinations revealed an improved epidermal integrity for CDSN‐deficient epidermal equivalents, if they were treated with liposomal encapsulated CDSN. Conclusions: This study presents the first preclinical in vitro experiments for a future specific protein replacement therapy for patients affected by PSS1. What is already known about this topic?Peeling skin syndrome type 1 (PSS1) refers to Mendelian disorders of cornification and is characterized by pruritus, pronounced erythroderma with skin abnormalities and lifelong patchy peeling of the skin.The disease is caused by autosomal recessive nonsense mutations in the gene encoding corneodesmosin (CDSN). Histopathologically, the absence of CDSN is characterized by subcorneal splitting and enhanced detachment of corneocytes. What does this study add?We present a liposomal formulation, which can be used for topical delivery of recombinant CDSN.Our in vitro study shows that the impaired barrier of CDSN‐deficient human epidermal equivalents can be improved when treated with liposomal human CDSN. What is the translational message?This study presents, for the first time, preclinical in vitro experiments for a specific protein replacement therapy in patients with PSS1. Linked Comment: Schmuth. Br J Dermatol 2021; 184:998–999. [ABSTRACT FROM AUTHOR]

Corrosion and dissolution reactions of Na2O.xSiO2 glasses (x=2·0, 2·5, and 3·3) were investigated by static and dynamic corrosion tests at pH values between 7 and 14 and temperatures of 30 and 50°C. The investigated glass compositions are close to those used for water glass production. Several types of leachants with varied pH and SiO2 concentrations were applied to induce corrosion effects. One characteristic feature of the corrosion process is the development of a reaction layer on top of the corroding glasses, a process depending on leachant pH. At intermediate pH values reaction layers were observed, but not at high pH values (>12). To generate altered glass surfaces with and without reaction layers sodium silicate glasses were corroded in deionized H2O, 0·1 M NaOH, and diluted liquid sodium water glasses. The resulting samples or layer materials were investigated by scanning electron microscopy, Raman microscopy, infrared spectroscopy, or 29Si-MAS-NMR spectroscopy. Especially Raman microscopy was applied to characterize depth profiles of structural changes of silicate (Qn) groups and of concentration profiles of molecular H2O within reaction layers. The results are discussed with respect to influences of pH and leachant composition in order to evaluate possible corrosion mechanisms. [ABSTRACT FROM AUTHOR]

The corrosion and dissolution of Na2O.xSiO2 glasses (x=2·0, 2·5, and 3·3) was investigated by static and dynamic corrosion tests at pH values between 7 and 14 with a focus on saturation effects. The corrosion temperatures were 30 and 50°C. The investigated glass compositions are close to those used for water glass production. Two types of leachants with varied concentrations have been applied to study saturation effects. The first type was deionized H2O enriched with up to 2·5 mM SiO2 by dispersing and equilibrating pyrogenic silica. The second type of leachants were liquid sodium water glasses with increasing concentrations and a SiO2:Na2O ratio similar to the dissolving glasses forming dilution series. Two different saturation effects which influence corrosion kinetics were recognised: first, the enrichment of deionised H2O with pyrogenic silica reduced the dissolution rate of reaction layers. An extrapolation of mass loss rates yielded silica saturation levels of 2·8 to 3·8 mM SiO2. Second, the dilution series of sodium water glasses demonstrated the shielding of glass surfaces when pH conditions hindered reaction layer formation. The corrosion rates of a sodium silicate glass with a molar ratio SiO2:Na2O=2·0 were reduced but the respective corrosion rates of a sodium silicate glass with a molar ratio SiO2:Na2O=3·3 were increased with increasing sodium water glass concentration of the initial leachants. The sodium silicate glass with a molar ratio SiO2:Na2O=2·5 showed first an increase and at higher sodium water glass concentrations a decrease of the corrosion rate with increasing concentration of the liquid sodium water glass and thus displayed an intermediate dependence on water glass concentration. [ABSTRACT FROM AUTHOR]

One basic process for the production of liquid sodium water glasses is the dissolution of sodium silicate glasses which can be regarded as an extreme case of glass corrosion. The dissolution processes occur at high pH values and achieve highly concentrated liquid sodium water glasses. To better understand the dissolution process, the methods of glass corrosion studies were adapted and applied. The corrosion and dissolution of Na2O.xSiO2 glasses (x=2·0, 2·5, and 3·3) was investigated by static and dynamic corrosion tests at pH values between 7 and 14. The corrosion temperatures were 30 and 50°C. The investigated glass compositions are close to those used for water glass production. The corrosion of sodium silicate glasses generally follows linear time laws at short corrosion times. The release rates of Na2O and SiO2 are correlated with the glass composition and to the thermodynamics of glass hydrolysis. Two distinct pH regimes exist. Between pH 7 and a certain pH value between 11 and 13 (depends on glass composition and corrosion temperature) the corrosion rate increases slightly with pH and the glasses develop extensive reaction layers. Generally, published corrosion models predict such a rate increase, but at a higher extent than observed here. Above a pH of 11 to 13 the corrosion rates decline with pH by a factor up to 100 and the glasses develop almost no reaction layer. A possible explanation of the rate decrease at higher pH is the shielding of the silicate surface by alkalis. The shielding effect increases with alkali concentration. The pH dependence of reaction layer formation is discussed in terms of solubility or diffusion through a boundary layer with pH and silica concentration gradients in the leachate near the glass surface. [ABSTRACT FROM AUTHOR]

The influence of experimental parameters on the dissolution of sodium silicate glasses with SiO2:Na2O ratios of 2·0 and 3·3 has been investigated in order to establish reliable corrosion conditions. Two principal experimental setups were tested; static (without exchange of leachant) and dynamic (with leachants continuously flowing through). Deionized water and 0·1 M NaOH were used as initial leachants. Surface pretreatment, temperature and time of corrosion, flow rate, surface area to solution volume ratio were varied and optimized. Balancing of glass samples and chemical analysis of leachates were used to describe the dissolution kinetics. Etching with HF was proven to be a reliable surface pretreatment. The chosen optimized corrosion conditions were temperatures of 30 and 50°C, flow rates of 2 l/d, and surface area to solution ratios of 10 m-1. [ABSTRACT FROM AUTHOR]

Many antimicrobial peptides bear arginine (R)- and tryptophan (W)-rich sequence motifs. Based on the sequence Ac-RRWWRF-NH2, sets of linear and cyclic peptides were generated by changes in the amino acid sequence,l-d-amino acid exchange and naphthylalanine substituted for tryptophan. Linear RW-peptides displayed moderate activity towards Gram-positiveBacillus subtilis(15 < MIC < 31 μm) and were inactive against Gram-negativeEscherichia coliat peptide concentrations<100 μm. Cyclization induced high antimicrobial activity. The effect of cyclization was most pronounced for peptides with three adjacent aromatic residues. Incorporation ofd-amino acid residues had minor influence on the biological activity. The haemolytic activity of all RW-peptides at 100 μmconcentration was low (<7% lysis for linear R/W-rich peptides and<28% for the cyclic analogues). Introduction of naphthylalanine enhanced the biological activities of both the linear and cyclic peptides. All peptides induced permeabilization of large unilamellar vesicles (LUVs) composed of lipids of the membrane ofB. subtilisand erythrocytes, but surprisingly had no effect on LUVs composed of lipids of theE. coliinner membrane. The profiles of peptide activity againstB. subtilisand red blood cells correlated with the permeabilizing effects on the corresponding model membranes and were related to hydrophobicity parameters as derived from reversed phase high-performance liquid chromatography (HPLC). The results underlined the importance of amphipathicity as a driving force for cell lytic activity and suggest that conformational constraints and an appropriate position of aromatic residues allowing the formation of hydrophobic clusters are highly favourable for antimicrobial activity and selectivity. [ABSTRACT FROM AUTHOR]

Labeling biomolecules with fluorescent labels is an established tool for structural, biochemical, and biophysical studies; however, it remains underused for small peptides. In this work, an amino acid bearing a 3-hydroxychromone fluorophore, 2-amino-3-(2-(furan-2-yl)-3-hydroxy-4-oxo-4H-chromen-6-yl)propanoic acid (FHC), was incorporated in a known hexameric antimicrobial peptide, cyclo[RRRWFW] (cWFW), in place of aromatic residues. Circular dichroism spectropolarimetry and antibacterial activity measurements demonstrated that the FHC residue perturbs the peptide structure depending on labeling position but does not modify the activity of cWFW significantly. FHC thus can be considered an adequate label for studies of the parent peptide. Several analytical and imaging techniques were used to establish the activity of the obtained labeled cWFW analogues toward animal cells and to study the behavior of the peptides in a multicellular organism. The 3-hydroxychromone fluorophore can undergo excited-state intramolecular proton transfer (ESIPT), resulting in double-band emission from its two tautomeric forms. This feature allowed us to get insights into conformational equilibria of the labeled peptides, localize the cWFW analogues in human cells (HeLa and HEK293) and zebrafish embryos, and assess the polarity of the local environment around the label by confocal fluorescence microscopy. We found that the labeled peptides efficiently penetrated cancerous cells and localized mainly in lipid-containing and/or other nonpolar subcellular compartments. In the zebrafish embryo, the peptides remained in the bloodstream upon injection into the cardinal vein, presumably adhering to lipoproteins and/or microvesicles. They did not diffuse into any tissue to a significant extent during the first 3 h after administration. This study demonstrated the utility of fluorescent labeling by double-emission labels to evaluate biologically active peptides as potential drug candidates in vivo ., Competing Interests: The authors SK, AS, and IK, are employers of the company Enamine Co, Ltd. SA, OB, and IK are co-founders of the company Lumobiotics GmbH. The remaining authors declare that the research was conducted without any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Afonin, Koniev, Préau, Takamiya, Strizhak, Babii, Hrebonkin, Pivovarenko, Dathe, le Noble, Rastegar, Strähle, Ulrich and Komarov.)

The influence of several antimicrobial trivalent cyclic hexapeptides on the mixing behavior of bilayer lipid membranes containing phosphatidylglycerol (PG) and phosphatidylethanolamine (PE) with varying composition was studied using DSC and ITC. The peptides contained three arginines and three aromatic amino acids and had different sequences. All of them induce clustering of PG-rich clusters with bound peptides after binding. In a previous publication we could show that a correlation between clustering efficacy and the antimicrobial activity of the peptides exists (S. Finger et al., Biochim. Biophys. Acta 1848 (2015) 2998-3006). In the current study we investigated whether the non-ideality of the lipid mixture had any effect on the clustering efficacy and the critical peptide/lipid clustering ratio. We could show that for PG/PE membranes containing 1:1 M ratios and lipids with equal or unequal chain lengths, the amount of clustered PG depended only slightly on the absolute chain length and on the chain length difference between PG and PE. Much larger differences were observed when the PG/ PE mixing ratio was changed. In mixtures of DPPG/DPPE with high PG content, the amount of clustered PG per added peptide was much higher than in PE-rich mixtures. The ITC experiments showed that the critical peptide/lipid ratio for cluster formation is also strongly dependent on the PG/PE ratio in the mixture. In the PG/PE 3:1 mixture, the formation of clusters with bound peptide is much more likely than for mixtures with less PG. For 1:1 and 1:3 lipid mixtures, the critical peptide/lipid ratio for demixing is between 0.002 and 0.004. Therefore, even in these mixtures clustering occurs way below charge saturation of the PG in the mixture and the PG-rich clusters are not charge compensated either. The peptide concentration necessary for inducing clustering amounts to ~8 μM, a value well within the range of minimal inhibitory concentration values observed for the cyclic peptides studied here. Our results show that not only the structure of the cyclic peptide influences the clustering efficacy but also the mixing behavior of the lipids in the bilayers has an influence on the amount of clustering induced by binding of cyclic peptides., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier B.V. All rights reserved.)

The influence of side chain residue and phospholipid characteristics of the cytoplasmic membrane upon the fibrillation and bacterial aggregation of arginine (Arg) and tryptophan (Trp) rich antimicrobial peptides (AMPs) has not been well described to date. Here, we utilized the structural advantages of HHC-10 and 4Har HHC-10 (Har, l-homoarginine) that are highly active Trp-rich AMPs and investigated their fibril formation and activity behavior against bacteria. The peptides revealed time-dependent self-assembly of polyproline II (PPII) α-helices, but by comparison, 4Har HHC-10 tended to form higher ordered fibrils due to relatively strong cation-π stacking of Trp with Har residue. Both peptides rapidly killed S. aureus and E. coli at their MICs and caused aggregation of bacteria at higher concentrations. This bacterial aggregation was accompanied by the formation of morphologically distinct electron-dense nanostructures, likely including but not limited to peptides alone. Both HHC-10-derived peptides caused blebs and buds in the E. coli membrane that are rich in POPE phospholipid that promotes negative curvature. However, the main population of S. aureus cells retained their cocci structure upon treatment with HHC peptides even at concentration higher than the MICs. In contrast, the cell aggregation was not induced by HHC fibrils that were most likely stabilized through intra-/intermolecular cation-π stacking. It is proposed that masking of these interactions might have resulted in diminished membrane association/insertion of the HHC nanostructures. The peptides caused aggregation of POPC/POPG (1/3) and POPE/POPG (3/1) liposomes. Nonetheless, disaggregation of the former vesicles was observed at ratios of lipid to peptide of greater than 6 and 24 for HHC-10 and 4Har HHC-10, respectively. Collectively, our results revealed dose-dependent bacterial aggregation mediated by Trp-rich AMPs that was profoundly influenced by the degree of peptide's self-association and the composition and intrinsic curvature of the cytoplasmic membrane lipids.

Improving diagnostic imaging and therapy by targeted compound delivery to pathological areas and across biological barriers is of urgent need. A lipopeptide, P-CrA-A2, composed of a highly cationic peptide sequence (A2), an N-terminally attached palmitoyl chain (P) and cryptophane molecule (CrA) for preferred uptake into blood-brain barrier (BBB) capillary endothelial cells, was generated. CrA allows reversible binding of Xe for NMR detection with hyperpolarized nuclei. The lipopeptide forms size-optimized micelles with a diameter of about 11 nm at low micromolar concentration. Their high local CrA payload has a strong and switchable impact on the bulk magnetization through Hyper-CEST detection. Covalent fixation of CrA does not impede micelle formation and does not hamper its host functionality but simplifies Xe access to hosts for inducing saturation transfer. Xe Hyper-CEST magnetic resonance imaging (MRI) allows for distinguishing BBB endothelial cells from control aortic endothelial cells, and the small micelle volume with a sevenfold improved CrA-loading density compared to liposomal carriers allows preferred cell labelling with a minimally invasive volume (≈16 000-fold more efficient than 19 F cell labelling). Thus, these nanoscopic particles combine selectivity for human brain capillary endothelial cells with great sensitivity of Xe Hyper-CEST MRI and might be a potential MRI tool in brain diagnostics., (© 2020 The Authors. Published by WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)