New fluorescent dyes containing an assembled 1,4-dihydroazolo5,1-
1,2,4triazine (DAT) core and an isoxazole ring were synthesized through a reaction between diazopyrazole or diazoimidazoles and ...isoxazolyl-derived enamines in mild conditions. The photophysical characteristics (maxima absorption and emission, Stokes shifts, fluorescent quantum yields, and fluorescence lifetimes) of the new fluorophores were obtained. The prepared DATs demonstrated emission maxima ranging within 433-487 nm, quantum yields within 6.1-33.3%, and a large Stokes shift. The photophysical characteristics of representative DAT examples were studied in ten different solvents. Specific (hydrogen bonds) and non-specific (dipole-dipole) intermolecular and intramolecular interactions were analyzed using XRD data and spectral experiments. Solvatochromism was analyzed using Lippert-Mataga and Dimroth-Reichardt plots, revealing the relationship between the DAT structure and the nature of solute-solvent interactions. The significant advantages of DATs are the fluorescence of their powders (QY up to 98.7%). DAT-NMe
expressed bright aggregation-induced emission (AIE) behavior in DMSO and THF as the water content increased. The numerous possible variations of the structures of the heterocycles included in the DATs, as well as substituents, create excellent prospects for adjusting their photophysical and physicochemical properties.
Dysfunction of two structurally and functionally related proteins, FUS and TAR DNA-binding protein of 43 kDa (TDP-43), implicated in crucial steps of cellular RNA metabolism can cause amyotrophic ...lateral sclerosis (ALS) and certain other neurodegenerative diseases. The proteins are intrinsically aggregate-prone and form non-amyloid inclusions in the affected nervous tissues, but the role of these proteinaceous aggregates in disease onset and progression is still uncertain. To address this question, we designed a variant of FUS, FUS 1–359, which is predominantly cytoplasmic, highly aggregate-prone, and lacks a region responsible for RNA recognition and binding. Expression of FUS 1–359 in neurons of transgenic mice, at a level lower than that of endogenous FUS, triggers FUSopathy associated with severe damage of motor neurons and their axons, neuroinflammatory reaction, and eventual loss of selective motor neuron populations. These pathological changes cause abrupt development of a severe motor phenotype at the age of 2.5–4.5 months and death of affected animals within several days of onset. The pattern of pathology in transgenic FUS 1–359 mice recapitulates several key features of human ALS with the dynamics of the disease progression compressed in line with shorter mouse lifespan. Our data indicate that neuronal FUS aggregation is sufficient to cause ALS-like phenotype in transgenic mice.
Background: FUS inclusions are hallmarks of certain neurodegenerative diseases.
Results: Expression of a highly aggregate prone FUS variant in transgenic mice causes proteinopathy and severe motor phenotype.
Conclusion: Aggregation of FUS is sufficient to recapitulate motor pathology typical for amyotrophic lateral sclerosis.
Significance: Understanding the role of protein aggregation in the development of human neurodegenerative diseases is crucial for designing efficient therapeutic approaches.
A detailed study of the interaction of β-1,2,3-thiadiazolyl enamines with azoles containing diazo function allowed the difference in the reactivity of azole-5-diazonium salts and 5-diazoazoles to be ...established, which leads to the implementation of various mechanisms of reaction and the formation of aromatic or non-aromatic bicyclic systems. The monitoring of this transformation by NMR spectroscopy revealed the key unstable intermediate which was further isolated under low temperature and thus the evidence for the assigned structures was obtained. An efficient PASE approach to the preparation of new 1,4-dihydropyrazolo5,1-
c
1,2,4triazines and 1,4-dihydroimidazo5,1-
c
1,2,4triazines was developed which significantly expanded the scope of these compounds for further studies of their biological activity. The structure and photophysical properties of the synthesized bicyclic heterocycles containing a 1,2,3-thiadiazole ring and several functional groups have been studied. The features of their absorption and emission are revealed, and preliminary directions of their application as new fluorosensors were outlined.
An efficient PASE approach to 1,4-dihydroazolo5,1-
c
1,2,4triazines was developed and new details of the mechanism found. Spectral investigations revealed the dual fluorescence of these compounds both in a diluted solution and in a solid state.
The quality of soft tissue defect regeneration after dental surgeries largely determines their final success. Collagen membranes have been proposed for the healing of such defects, but in some cases, ...they do not guarantee a sufficient volume of the regenerated tissue and vascularization. For this purpose, lactoferrin, a protein with natural pro-regenerative, anti-inflammatory, and pro-angiogenic activity, can be added to collagen. In this article, we used a semipermeable barrier-assisted electrophoretic deposition (SBA-EPD) method for the production of collagen-lactoferrin membranes. The membrane structure was studied by SEM, and its mechanical properties were shown. The lactoferrin release kinetics were shown by ELISA within 75 h. When tested in vitro, we demonstrated that the collagen-lactoferrin membranes significantly increased the proliferation of keratinocytes (HaCaT) and fibroblasts (977hTERT) compared to blank collagen membranes. In vivo, on the vestibuloplasty and free gingival graft harvesting models, we showed that collagen-lactoferrin membranes decreased the wound inflammation and increased the healing rates and regeneration quality. In some parameters, collagen-lactoferrin membranes outperformed not only blank collagen membranes, but also the commercial membrane Mucograft
. Thus, we proved that collagen-lactoferrin membranes produced by the SBA-EPD method may be a valuable alternative to commercially used membranes for soft tissue regeneration in the oral cavity.
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PASE (pot, atom and step economic) synthesis of 1-azolyl-1H-1,2,4-triazole derivatives in up to 91% yield has been accomplished by addition of 5-diazoazoles to ethyl isocyanoacetate.
A comparative analysis of physicochemical properties and reactivity of 3-aryl-1
H
-pyrazole-5-diazonium tetrafluoroborates and 5-aryl-3-diazo-3
H
-pyrazoles in azo coupling reactions is presented. It ...is shown that diazonium salts have higher reactivity compared to the respective 3-diazopyrazoles, which is in agreement with their physicochemical properties. Heterocyclization of the synthesized azo compounds provided 2-arylpyrazolo5,1-
c
1,2,4benzotriazines, which were screened for antitumor activity against human uterine endothelium cancer cells (HeLa cell line) and human skin fibroblasts using the MTT assay and flow cytometry. It was found that all of the tested compounds exhibited moderate to high cytotoxic activity. The best results were obtained with 6,8-dimethoxy-2-phenylpyrazolo-5,1-
c
1,2,4benzotriazine.
5-Diazoimidazoles and 5-diazopyrazoles have been shown to react with acyl isothiocyanates yielding the imidazo- and pyrazolo5,1-d1,2,3,5thiatriazines stabilized by a nonbonded Sa=O interaction. In ...contrast to acyl isothiocyanates, alkyl-, aryl-, and arylsulfonyl isothiocyanates do not react with 5-diazoazoles. The nature and the strength of stabilizing intramolecular interaction between non-bonded S and O atoms have been studied by X-ray analysis for mono crystals and DFT calculations for selected azolo5,1-d1,2,3,5thiatriazines. The interaction was described in terms of Weinhold covalence ratio factors, NBO, and AIM schemes. The reaction discovered was used to develop an efficient approach toward the new 8-substituted 4-ethoxycarbonylimino-4-benzoyl- and 4-(3,4,5,6-tetrafluorobenzoyl)iminoimidazo(pyrazolo)5,1-d1,2,3, 5 thiatriazines.
Mutations to the RNA binding protein, fused in sarcoma (FUS) occur in ∼5% of familial ALS and FUS-positive cytoplasmic inclusions are commonly observed in these patients. Altered RNA metabolism is ...increasingly implicated in ALS, yet it is not understood how the specificity with which FUS interacts with RNA in the cytoplasm can affect its aggregation in vivo. To further understand this, we expressed, in mice, a form of FUS (FUS ΔRRMcyt) that lacked the RNA recognition motif (RRM), thought to impart specificity to FUS-RNA interactions, and carried an ALS-associated point mutation, R522G, retaining the protein in the cytoplasm. Here we report the phenotype and results of histological assessment of the brain of transgenic mice expressing this isoform of FUS. Results demonstrated that neuronal expression of FUS ΔRRMcyt caused early lethality often preceded by severe tremor. Large FUS-positive cytoplasmic inclusions were found in many brain neurons; however, neither neuronal loss nor neuroinflammatory response was observed. In conclusion, the extensive FUS proteinopathy and severe phenotype of these mice suggests that affecting the interactions of FUS with RNA in vivo may augment its aggregation in the neuronal cytoplasm and the severity of disease processes.
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