Evaluation of a model photo-caged dehydropeptide as a stimuli-responsive supramolecular hydrogel

Short peptides capped on the N-terminus with aromatic groups are often able to form supramolecular hydrogels, via self-assembly, in aqueous media. The rheological properties of these readily tunable hydrogels resemble those of the extracellular matrix (ECM) and therefore have potential for various biological applications, such as tissue engineering, biosensors, 3D bioprinting, drug delivery systems and wound dressings. We herein report a new photo-responsive supramolecular hydrogel based on a “caged” dehydropeptide (CNB-Phe-ΔPhe-OH 2), containing a photo-cleavable carboxy-2-nitrobenzyl (CNB) group. We have characterized this hydrogel using a range of techniques. Irradiation with UV light cleaves the pendant aromatic capping group, to liberate the corresponding uncaged model dehydropeptide (H-Phe-ΔPhe-OH 3), a process which was investigated by 1H NMR and HPLC studies. Crucially, this cleavage of the capping group is accompanied by dissolution of the hydrogel (studied visually and by fluorescence spectroscopy), as the delicate balance of intramolecular interactions within the hydrogel structure is disrupted. Hydrogels which can be disassembled non-invasively with temporal and spatial control have great potential for specialized on-demand drug release systems, wound dressing materials and various topical treatments. Both 2 and 3 were found to be non-cytotoxic to the human keratinocyte cell line, HaCaT. The UV-responsive hydrogel system reported here is complementary to previously reported related UV-responsive systems, which are generally composed of peptides formed from canonical amino acids, which are susceptible to enzymatic proteolysis in vivo. This system is based on a dehydrodipeptide structure which is known to confer proteolytic resistance. We have investigated the ability of the photo-activated system to accelerate the release of the antibiotic, ciprofloxacin, as well as some other small model drug compounds. We have also conducted some initial studies towards skin-related applications. Moreover, this model system could potentially be adapted for on-demand “self-delivery”, through the uncaging of known biologically active dehydrodipeptides.This work was supported by the Portuguese Foundation for Science and Technology (FCT) in the framework of the Strategic Funding of CQUM (UID/QUI/00686/2019) and of IPC (UID/CTM/50025/2019). L.H. acknowledges funding from the FCT Investigator Programme through grant IF/00606/2014. FCT, FEDER, PORTUGAL2020 and COMPETE2020 are also acknowledged for funding under research project PTDC/QUI-QOR/29015/2017 (POCI-01-0145-FEDER-029015)

New eugenol derivatives with enhanced insecticidal activity

Eugenol, the generic name of 4-allyl-2-methoxyphenol, is the major component of clove essential oil, and has demonstrated relevant biological potential with well-known antimicrobial and antioxidant actions. New O-alkylated eugenol derivatives, bearing a propyl chain with terminals like hydrogen, hydroxyl, ester, chlorine, and carboxylic acid, were synthesized in the present work. These compounds were later subjected to epoxidation conditions to give the corresponding oxiranes. All derivatives were evaluated against their effect upon the viability of insect cell line Sf9 (Spodoptera frugiperda), demonstrating that structural changes elicit marked effects in terms of potency. In addition, the most promising molecules were evaluated for their impact in cell morphology, caspase-like activity, and potential toxicity towards human cells. Some molecules stood out in terms of toxicity towards insect cells, with morphological assessment of treated cells showing chromatin condensation and fragmentation, which are compatible with the occurrence of programmed cell death, later confirmed by evaluation of caspase-like activity. These findings point out the potential use of eugenol derivatives as semisynthetic insecticides from plant natural products.This research was funded by COMPETE 2020 program, co-financed by the FEDER and the European Union, PTDC/ASP-AGR/30154/2017 (POCI-01-0145-FEDER-030154). The authors also acknowledge the Foundation for Science and Technology (FCT; Portugal), and FEDER-COMPETE/QREN-EU for financial support to the research centers CQ/UM (UIDB/00686/2020), CF-UM-UP (UIDB/04650/2020) and REQUIMTE (UIDB/50006/2020). The NMR spectrometer Bruker Avance III 400 is part of the NationalNMRNetwork and was purchased within the framework of the National Program for Scientific Re-equipment, contract REDE/1517/RMN/2005, with funds from POCI 2010 (FEDER) and the FCT. The authors would also like to thank RIAIDT-USC for the use of their analytical facilities

Aryl-capped lysine-dehydroamino acid dipeptide supergelators as potential drug release systems

Employing amino acids and peptides as molecular building blocks provides unique opportunities for generating supramolecular hydrogels, owing to their inherent biological origin, bioactivity, biocompatibility, and biodegradability. However, they can suffer from proteolytic degradation. Short peptides (<8 amino acids) attached to an aromatic capping group are particularly attractive alternatives for minimalistic low molecular weight hydrogelators. Peptides with low critical gelation concentrations (CGCs) are especially desirable, as the low weight percentage required for gelation makes them more cost-effective and reduces toxicity. In this work, three dehydrodipeptides were studied for their self-assembly properties. The results showed that all three dehydrodipeptides can form self-standing hydrogels with very low critical gelation concentrations (0.050.20 wt%) using a pH trigger. Hydrogels of all three dehydrodipeptides were characterised by scanning tunnelling emission microscopy (STEM), rheology, fluorescence spectroscopy, and circular dichroism (CD) spectroscopy. Molecular modelling was performed to probe the structural patterns and interactions. The cytotoxicity of the new compounds was tested using human keratinocytes (HaCaT cell line). In general, the results suggest that all three compounds are non-cytotoxic, although one of the peptides shows a small impact on cell viability. In sustained release assays, the effect of the charge of the model drug compounds on the rate of cargo release from the hydrogel network was evaluated. The hydrogels provide a sustained release of methyl orange (anionic) and ciprofloxacin (neutral), while methylene blue (cationic) was retained by the network.This work was supported by the Portuguese Foundation for Science and Technology (FCT) in the framework of the Strategic Funding of CQUM (UID/QUI/00686/2019), IPC (UIDP/CTM/05256/2020 and UIDB/05256/2020) and REQUIMTE/LAQV (UIDB/50006/2020). L.H. acknowledges grant CEECINST/00156/2018. FCT, FEDER, PORTUGAL2020 and COMPETE2020 are also acknowl edged for funding under research project PTDC/QUI-QOR/29015/2017 (POCI-01-0145-FEDER 029015). TGC thanks FCT under the scope of the strategic funding of UIDB/04469/2020 unit, and LABBELS—Associate Laboratory in Biotechnology, Bioengineering and Microelectromechanical Systems, LA/P/0029/2020.info:eu-repo/semantics/publishedVersio

Toxicity and structure-activity relationship (SAR) of α, β-dehydroamino acids against human cancer cell lines

A library of N-protected dehydroamino acids, namely dehydroalanine, dehydroaminobutyric acid and dehydrophenylalanine derivatives, was screened in three human cancer cell lines [(lung (A549), gastric (AGS) and neuroblastoma (SH-SY5Y)] in order to characterize their toxicological profile and identify new molecules with potential anticancer activity. Results showed N-protected dehydrophenylalanine and dehydroaminobutyric acid derivatives have no or low toxicity for all tested cell lines. The N-protected dehydroalanines exhibit significant toxic effects and the AGS and SH-SY5Y cells were significantly more vulnerable than A549 cells. Four α,β- dehydroalanine derivatives, with IC50 < 62.5 μM, were selected to investigate the pathways by which these compounds promote cell death. All compounds, at their IC50 concentrations, were able to induce apoptosis in both AGS and SH-SY5Y cell lines. In both cell lines, loss of mitochondrial membrane potential (ΔΨm) was found and caspase activity was increased, namely endoplasmic reticulum-resident caspase-4 in AGS cells and caspase-3/7 in SH-SY5Y cells. When evaluated in a non-cancer cell line, the molecules displayed no to low toxicity, thus suggesting some degree of selectivity for cancer cells. The results indicate that α,β-dehydroalanine derivatives can be considered a future resource of compounds able to work as anticancer drugs.This work received financial support from National Funds (FCT/MEC) through Project UID/QUI/50006/2013, co-financed by European Union (FEDER under the Partnership Agreement PT2020); and from Norte Portugal Regional Operational Programme (NORTE 2020), under the PORTUGAL 2020 Partnership Agreement, through the European Regional Development Fund (ERDF) (project NORTE-01-0145-FEDER 000024).info:eu-repo/semantics/publishedVersio

Electron-Phonon Coupling in Highly-Screened Graphene

Photoemission studies of graphene have resulted in a long-standing controversy concerning the strength of the experimental electron-phonon interaction in comparison with theoretical calculations. Using high-resolution angle-resolved photoemission spectroscopy we study graphene grown on a copper substrate, where the metallic screening of the substrate substantially reduces the electron-electron interaction, simplifying the comparison of the electron-phonon interaction between theory and experiment. By taking the nonlinear bare bandstructure into account, we are able to show that the strength of the electron-phonon interaction does indeed agree with theoretical calculations. In addition, we observe a significant bandgap at the Dirac point of graphene.Comment: Submitted to Phys. Rev. Lett. on July 20, 201

Comparison of ratioing and RCNA methods in the detection of flooded areas using Sentinel 2 Imagery (case study: Tulun, Russia)

Climate change and natural disasters caused by hydrological, meteorological, and climatic phenomena have a significant impact on cities. Russia, a continental country with a vast territory of complex geographic–ecological environments and highly variable climatic conditions, is subject to substantial and frequent natural disasters. On 29 June 2019, an extreme precipitation event occurred in the city of Tulun in the Irkutsk oblast, Russian Federation, which caused flooding due to the increase in the water level of the Iya River that passes through the city, leaving many infrastructures destroyed and thousands of people affected. This study aims to determine the flooded areas in the city of Tulun based on two change detection methods: Radiometric Rotation Controlled by No-change Axis (RCNA) and Ratioing, using Sentinel 2 images obtained before the event (19 June 2019) and during the flood peak (29 June 2019). The results obtained by the two methodologies were compared through cross-classification, and a 98% similarity was found in the classification of the areas. The study was validated based on photointerpretation of Google Earth images. The methodology presented proved to be useful for the automatic precession of flooded areas in a straightforward, but rigorous, manner. This allows stakeholders to efficiently manage areas that are buffeted by flooding episodes.LA/P/0069/2020info:eu-repo/semantics/publishedVersio

Eugenol β-amino/β-alkoxy alcohols with selective anticancer activity

Eugenol, 4-allyl-2-methoxyphenol, is the main constituent of clove essential oil and has demonstrated relevant biological activity, namely anticancer activity. Aiming to increase this activity, we synthesized a series of eugenol β-amino alcohol and β-alkoxy alcohol derivatives, which were then tested against two human cancer cell lines, namely gastric adenocarcinoma cells (AGS) and lung adenocarcinoma cells (A549). An initial screening was performed to identify the most cytotoxic compounds. The results demonstrated that three β-amino alcohol derivatives had anticancer activity that justified subsequent studies, having been shown to trigger apoptosis. Importantly, the most potent molecules displayed no appreciable toxicity towards human noncancer cells. Structure-activity relationships show that changes in eugenol structure led to enhanced cytotoxic activity and can contribute to the future design of more potent and selective drugs.This research was funded by FCT under project PTDC/ASP-AGR/30154/2017 (PO-CI-01-0145- FEDER-030154) of COMPETE 2020, co-financed by FEDER and EU. FCT-Portugal and FEDERCOMPETE/ QREN-EU also gave financial support to the research centres CQ/UM (UIDB/00686/2020) and REQUIMTE (UIDB/50006/2020). The NMR spectrometer Bruker Avance III 400 (part of the National NMR Network) was financed by FCT and FEDER. Renato B. Pereira acknowledges PRIMA Foundation (H2020- PRIMA 2018—Section 2, Project MILKQUA) and FCT (PTDC/QUI-QFI/2870/2020) for the funding

Bolaamphiphilic Bis-Dehydropeptide hydrogels as potential drug release systems

The self-assembly of nanometric structures from molecular building blocks is an effective way to make new functional materials for biological and technological applications. In this work, four symmetrical bolaamphiphiles based on dehydrodipeptides (phenylalanyldehydrophenylalanine and tyrosyldehydrophenylalanine) linked through phenyl or naphthyl linkers (terephthalic acid and 2,6-naphthalenedicarboxylic acid) were prepared, and their self-assembly properties were studied. The results showed that all compounds, with the exception of the bolaamphiphile of tyrosyldehydrophenylalanine and 2,6-naphthalene dicarboxylic acid, gave self-standing hydrogels with critical gelation concentrations of 0.3 wt % and 0.4 wt %, using a pH trigger. The self-assembly of these hydrogelators was investigated using STEM microscopy, which revealed a network of entangled fibers. According to rheology, the dehydrodipeptide bolaamphiphilic hydrogelators are viscoelastic materials with an elastic modulus G' that falls in the range of native tissue (0.37 kPa brain-4.5 kPa cartilage). In viability and proliferation studies, it was found that these compounds were non-toxic toward the human keratinocyte cell line, HaCaT. In sustained release assays, we studied the effects of the charge present on model drug compounds on the rate of cargo release from the hydrogel networks. Methylene blue (MB), methyl orange (MO), and ciprofloxacin were chosen as cationic, anionic, and overall neutral cargo, respectively. These studies have shown that the hydrogels provide a sustained release of methyl orange and ciprofloxacin, while methylene blue is retained by the hydrogel network.Portugal2020, Compete2020, FEDER. PTDC/QUI-QOR/29015/2017. UIDB/04650/2020. UIDB/50006/2020. UID/QUI/00686/2019. UID/CTM/50025/2019. SFRH/BD/144017/201

The Effect of Transposable Element Insertions on Gene Expression Evolution in Rodents

Background:Many genomes contain a substantial number of transposable elements (TEs), a few of which are known to be involved in regulating gene expression. However, recent observations suggest that TEs may have played a very important role in the evolution of gene expression because many conserved non-genic sequences, some of which are know to be involved in gene regulation, resemble TEs. Results:Here we investigate whether new TE insertions affect gene expression profiles by testing whether gene expression divergence between mouse and rat is correlated to the numbers of new transposable elements inserted near genes. We show that expression divergence is significantly correlated to the number of new LTR and SINE elements, but not to the numbers of LINEs. We also show that expression divergence is not significantly correlated to the numbers of ancestral TEs in most cases, which suggests that the correlations between expression divergence and the numbers of new TEs are causal in nature. We quantify the effect and estimate that TE insertion has accounted for ~20% (95% confidence interval: 12% to 26%) of all expression profile divergence in rodents. Conclusions:We conclude that TE insertions may have had a major impact on the evolution of gene expression levels in rodents

Liposomal formulations loaded with a eugenol derivative for application as insecticides: encapsulation studies and In silico identification of protein targets

Supplementary Materials can be downloaded at: https://www.mdpi.com/article/10.3390/nano12203583/s1,A recently synthesized new eugenol derivative, ethyl 4-(2-methoxy-4-(oxiran-2-ylmethyl)phenoxy)butanoate, with a high insecticidal activity against Sf9 (Spodoptera frugiperda) insect cells, was encapsulated in the liposomal formulations of egg-phosphatidylcholine/cholesterol (Egg-PC:Ch) 70:30 and 100% dioleoylphosphatidylglycerol (DOPG), aiming at the future application as insecticides. Compound-loaded DOPG liposomes have sizes of 274 ± 12 nm, while Egg-PC:Ch liposomes exhibit smaller hydrodynamic diameters (69.5 ± 7 nm), high encapsulation efficiency (88.8% ± 2.7%), higher stability, and a more efficient compound release, thus, they were chosen for assays in Sf9 insect cells. The compound elicited a loss of cell viability up to 80% after 72 h of incubation. Relevantly, nanoencapsulation maintained the toxicity of the compound toward insect cells while lowering the toxicity toward human cells, thus showing the selectivity of the system. Structure-based inverted virtual screening was used to predict the most likely targets and molecular dynamics simulations and free energy calculations were used to demonstrate that this molecule can form a stable complex with insect odorant binding proteins and/or acetylcholinesterase. The results are promising for the future application of compound-loaded nanoliposome formulations as crop insecticides.This research was funded by project PTDC/ASP-AGR/30154/2017 (POCI-01-0145-FEDER 030154) of the COMPETE2020 program, co-financed by the FEDER and the European Union. The authors also acknowledge the Foundation for Science and Technology (FCT, Portugal) and FEDERCOMPETE QREN-EU for financial support to the research centers CQUM (UID/QUI/00686/2021), CF-UM-UP (UIDB/04650/2020) and REQUIMTE (UIDB/50006/2020). Renato B. Pereira acknowledges PRIMA Foundation (H2020-PRIMA 2018—Section 2, Project MILKQUA) and FCT (PTDC/QUI-QFI/2870/2020) for additional funding. The NMR spectrometer Bruker Avance III 400 is part of the National NMR Network and was purchased within the framework of the National Program for Scientific Re-equipment, contract REDE/1517/RMN/2005, with funds from POCI 2010 (FEDER) and FCT