Search Results (10)

Arsenic, a potent metalloid contaminant of drinking water, is known for its ability to act as an initiator and modulator of disease in a variety of human tissues. Upon ingestion, arsenic is bio-transformed in the liver into a variety of metabolites, including arsenite. Arsenite permeates the blood–brain barrier (BBB), inducing oxidative stress that can be detrimental to brain neurons. As the primary glial cell at the BBB interface, astrocytes play a pivotal role in detoxifying xenobiotics such as arsenite via the production of the tripeptide antioxidant γ-glutamylcysteine, or glutathione (GSH). In this study, we assessed the mRNA levels of key components of the GSH synthetic pathway in astrocytes exposed to arsenite compared to vehicle controls. These components included xCT [substrate-specific light chain of the substrate importing transporter, system x_c⁻ (Sx_c⁻)], glutamate-cysteine ligase [both catalytic (GCL_C) and modifying (GCL_M) subunits], and glutathione synthetase (GS). Additionally, we analyzed protein levels of some components by Western blotting and evaluated functional activity of Sx_c⁻ using a fluorescence-based cystine uptake assay. Finally, we utilized a luminescence-based glutathione assay to determine the intracellular and extracellular GSH content in arsenite-treated cells. Arsenite significantly increased xCT, GCL_C, GCL_M, and GS mRNA levels, an effect blocked by the transcriptional inhibitor actinomycin D (ActD). A corresponding increase in Sx_c⁻ activity was also observed in the arsenite treatment groups, along with significant increases in GCL_C and GCL_M protein expression. However, no increase in GS protein expression was detected. Finally, arsenite treatment significantly increased extracellular GSH levels, an effect which was also prevented by the inclusion of ActD. Overall, our study provides evidence that arsenite transcriptionally regulates several cellular processes necessary for GSH synthesis in primary cortical astrocyte cultures, thereby contributing to a better understanding of how this environmental toxicant influences antioxidant defenses in the brain. However, these results should be interpreted with caution regarding their applicability to vivo systems. Full article

The present study aimed to create a more sustainable and controlled delivery system based on natural biopolymer bacterial nanocellulose (BNC) and bacterial natural product actinomycin (Act), with the applicative potential in the biomedical field. In order to provide improved interaction between BNC and the active compound, and thus to modulate the release kinetics, the TEMPO oxidation of BNC support was carried out. A mix of actinomycins from bacterial fermentation (ActX) were used as natural antimicrobial agents with an established bioactivity profile and clinical use. BNC and TEMPO-oxidized BNC films with incorporated active compounds were obtained and analyzed by FTIR, SEM, XPS, and XRD. The ActX release profiles were determined in phosphate-buffer solution, PBS, at 37 °C over time. FTIR analysis confirmed the improved incorporation and efficiency of ActX adsorption on oxidized BNC due to the availability of more active sites provided by oxidation. SEM analysis indicated the incorporation of ActX into the less-dense morphology of the TEMPO-oxidized BNC in comparison to pure BNC. The release kinetics of ActX were significantly affected by the BNC structure, and the activated BNC sample indicated the sustained release of active compounds over time, corresponding to the Fickian diffusion mechanism. Antimicrobial tests using Staphylococcus aureus NCTC 6571 confirmed the potency of this BNC-based system for biomedical applications, taking advantage of the capacity of modified BNC to control and modulate the release of bioactive compounds. Full article

Actinobacteria produce a broad spectrum of bioactive substances that are used in the pharmaceutical, agricultural, and biotechnology industries. This study investigates the production of bioactive substances in Streptomyces, isolated from soil under five tropical plants, focusing on their potential as natural antibacterial dyes for silk fabrics. Out of 194 isolates, 44 produced pigments on broken rice as a solid substrate culture. Eight antibacterial pigmented isolates from under Magnolia baillonii (TBRC 15924, TBRC 15927, TBRC 15931), Magnolia rajaniana (TBRC 15925, TBRC 15926, TBRC 15928, TBRC 15930), and Cinnamomum parthenoxylon (TBRC 15929) were studied in more detail. TBRC 15927 was the only isolate where all the crude extracts inhibited the growth of the test organisms, Staphylococcus epidermidis TISTR 518 and S. aureus DMST 4745. The bioactive compounds present in TBRC 15927 were identified through LC-MS/MS analysis as belonging to the actinomycin group, actinomycin D (or X₁), X₂, and X_0β. Also, the ethyl acetate crude extract exhibited non-toxicity at an IC₅₀ value of 0.029 ± 0.008 µg/mL on the mouse fibroblast L-929 assay. From the 16S rRNA gene sequence analysis, TBRC 15927 had 100% identity with Streptomyces gramineus JR-43^T. Raw silk dyed with the positive antimicrobial TBRC 15927 extract (8.35 mg/mL) had significant (>99.99%) antibacterial properties. Streptomyces gramineus TBRC 15927 is the first actinomycin-producing strain reported to grow on broken rice and shows promise for antibacterial silk dyeing. Full article

Actinomycin is a family of chromogenic lactone peptides that differ in their peptide portions of the molecule. An antimicrobial peptide, actinomycin X2 (Ac.X2), was produced through the fermentation of a Streptomyces cyaneofuscatus strain. Immobilization of Ac.X2 onto a prepared silk fibroin (SF) film was done through a carbodiimide reaction. The physical properties of immobilized Ac.X2 (antimicrobial films, AMFs) were analyzed by ATR-FTIR, SEM, AFM, and WCA. The findings from an in vitro study showed that AMFs had a more broad-spectrum antibacterial activity against both S. aureus and E. coli compared with free Ac.X2, which showed no apparent strong effect against E. coli. These AMFs showed a suitable degradation rate, good hemocompatibility, and reduced cytotoxicity in the biocompatibility assay. The results of in vivo bacterially infected wound healing experiments indicated that wound inflammation was prevented by AMFs, which promoted wound repair and improved the wound microenvironment. This study revealed that Ac.X2 transformation is a potential candidate for skin wound healing. Full article

The opportunistic fungus Aspergillus fumigatus causes a set of diseases ranging from allergy to lethal invasive mycosis. Within the human airways, A. fumigatus is embedded in a biofilm that forms not only a barrier against the host immune defense system, but also creates a physical barrier protecting the fungi from chemicals such as antifungal drugs. Novel therapeutic strategies aim at combining drugs that inhibit biofilm synthesis or disrupt existing biofilm with classical antimicrobials. One of the major constituents of A. fumigatus biofilm is the polysaccharide galactosaminogalactan (GAG) composed of α1,4-linked N-acetylgalactosamine, galactosamine, and galactose residues. GAG is synthesized on the cytosolic face of the plasma membrane and is extruded in the extracellular space, where it is partially deacetylated. The deacetylase Agd3 that mediates this last step is essential for the biofilm formation and full virulence of the fungus. In this work, a previously described enzyme-linked lectin assay, based on the adhesion of deacetylated GAG to negatively charged plates and quantification with biotinylated soybean agglutinin was adapted to screen microbial natural compounds, as well as compounds identified in in silico screening of drug libraries. Actinomycin X2, actinomycin D, rifaximin, and imatinib were shown to inhibit Agd3 activity in vitro. At a concentration of 100 µM, actinomycin D and imatinib showed a clear reduction in the biofilm biomass without affecting the fungal growth. Finally, imatinib reduced the virulence of A. fumigatus in a Galleria mellonella infection model in an Agd3-dependent manner. Full article

Actinomycetes are natural architects of numerous secondary metabolites including antibiotics. With increased multidrug-resistant (MDR) pathogens, antibiotics that can combat such pathogens are urgently required to improve the health care system globally. The characterization of actinomycetes available in Nepal is still very much untouched which is the reason why this paper showcases the characterization of actinomycetes from Nepal based on their morphology, 16S rRNA gene sequencing, and metabolic profiling. Additionally, antimicrobial assays and liquid chromatography-high resolution mass spectrometry (LC-HRMS) of ethyl acetate extracts were performed. In this study, we employed a computational-based dereplication strategy for annotating molecules which is also time-efficient. Molecular annotation was performed through the GNPS server, the SIRIUS platform, and the available databases to predict the secondary metabolites. The sequencing of the 16S rRNA gene revealed that the isolates BN6 and BN14 are closely related to Streptomyces species. BN14 showed broad-spectrum antibacterial activity with the zone of inhibition up to 30 mm against Staphylococcus aureus (MIC: 0.3051 µg/mL and MBC: 9.7656 µg/mL) and Shigella sonnei (MIC: 0.3051 µg/mL and MBC: 4.882 µg/mL). Likewise, BN14 also displayed significant inhibition to Acinetobacter baumannii, Klebsiella pneumoniae, and Salmonella typhi. GNPS approach suggested that the extracts of BN6 and BN14 consisted of diketopiperazines ((cyclo(D-Trp-L-Pro), cyclo(L-Leu-L-4-hydroxy-Pro), cyclo(L-Phe-D-Pro), cyclo(L-Trp-L-Pro), cyclo(L-Val-L-Pro)), and polypeptide antibiotics (actinomycin D and X2). Additional chemical scaffolds such as bacterial alkaloids (bohemamine, venezueline B, and G), anthramycin-type antibiotics (abbeymycin), lipase inhibitor (ebelactone B), cytocidal (oxopropaline D), antifungal and antitumor antibiotics (reductiomycin, streptimidone, deoxynybomycin), alaremycin, fumaramidmycin, anisomycin, and others were also annotated, which were further confirmed by using the SIRIUS platform, and literature survey. Thus, the bioprospecting of natural products from Streptomyces species from Nepal could be a potential source for the discovery of clinically significant and new antimicrobial agents in the future. Full article

Transcriptional regulation is a major mechanism by which organisms integrate gene x environment interactions. It can be achieved by coordinated interplay between cis-regulatory elements (CREs) and transcription factors (TFs). Euryhaline tilapia (Oreochromis mossambicus) tolerate a wide range of salinity and thus are an appropriate model to examine transcriptional regulatory mechanisms during salinity stress in fish. Quantitative proteomics in combination with the transcription inhibitor actinomycin D revealed 19 proteins that are transcriptionally upregulated by hyperosmolality in tilapia brain (OmB) cells. We searched the extended proximal promoter up to intron1 of each corresponding gene for common motifs using motif discovery tools. The top-ranked motif identified (STREME1) represents a binding site for the Forkhead box TF L1 (FoxL1). STREME1 function during hyperosmolality was experimentally validated by choosing two of the 19 genes, chloride intracellular channel 2 (clic2) and uridine phosphorylase 1 (upp1), that are enriched in STREME1 in their extended promoters. Transcriptional induction of these genes during hyperosmolality requires STREME1, as evidenced by motif mutagenesis. We conclude that STREME1 represents a new functional CRE that contributes to gene x environment interactions during salinity stress in tilapia. Moreover, our results indicate that FoxL1 family TFs are contribute to hyperosmotic induction of genes in euryhaline fish. Full article

Actinomycins as clinical medicine have been extensively studied, while few investigations were conducted to discover the feasibility of actinomycins as antimicrobial natural dye contributing to the medical value of the functional fabrics. This study was focused on the application of actinomycin X2 (Ac.X2), a peptide pigment cultured from marine-derived Streptomyces cyaneofuscatus, in the dyeing and finishing of silk fabric. The dyeing potential of Ac.X2 with silk vs. cotton fabrics was assessed. As a result, the silk fabric exhibited greater uptake and color fastness with Ac.X2. Through Fourier transform infrared spectroscopy (FT-IR), scanning electron microscopy (SEM), and X-ray diffraction (XRD) analyses, some changes of chemical property for the dyed fabric and Ac.X2 were studied. The silk fabric dyed with Ac.X2 exhibited good UV protection ability. The antibacterial properties of dyed and finished silk were also evaluated, which exhibited over 90% antibacterial activity even after 20 washing cycles. In addition, the brine shrimp assay was conducted to evaluate the general toxicity of the tested fabric, and the results indicated that the dyed silk fabrics had a good biological safety property. Full article

Leishmaniasis, a Neglected Tropical Parasitic Disease (NTPD), is induced by several Leishmania species and is disseminated through sandfly (Lutzomyia longipalpis) bites. The parasite has developed resistance to currently prescribed antileishmanial drugs, and it has become pertinent to the search for new antileishmanial agents. The current study aimed to investigate the in vitro and in silico antileishmanial activity of two newly sourced actinomycins, X₂ and D, produced by the novel Streptomyces smyrnaeus strain UKAQ_23. The antileishmanial activity conducted on promastigotes and amastigotes of Leishmania major showed actinomycin X₂ having half-maximal effective concentrations (EC₅₀), at 2.10 ± 0.10 μg/mL and 0.10 ± 0.0 μg/mL, and selectivity index (SI) values of 0.048 and 1, respectively, while the actinomycin D exhibited EC₅₀ at 1.90 ± 0.10 μg/mL and 0.10 ± 0.0 μg/mL, and SI values of 0.052 and 1. The molecular docking studies demonstrated squalene synthase as the most favorable antileishmanial target protein for both the actinomycins X₂ and D, while the xanthine phosphoribosyltransferase was the least favorable target protein. The molecular dynamics simulations confirmed that both the actinomycins remained stable in the binding pocket during the simulations. Furthermore, the MMPBSA (Molecular Mechanics Poisson-Boltzmann Surface Area) binding energy calculations established that the actinomycin X₂ is a better binder than the actinomycin D. In conclusion, both actinomycins X₂ and D from Streptomyces smyrnaeus strain UKAQ_23 are promising antileishmanial drug candidates and have strong potential to be used for treating the currently drug-resistant leishmaniasis. Full article

An extract prepared from the culture of a marine-derived actinomycete Streptomyces sp. ZZ338 was found to have significant antimicrobial and antiproliferative activities. A chemical investigation of this active extract resulted in the isolation of three known bioactive actinomycins (1–3) and two new metabolites (4 and 5). The structures of the isolated compounds were identified as actinomycins D (1), V (2), X_0β (3), 2-acetylamino-3-hydroxyl-4-methyl-benzoic acid methyl ester (4), and N-1S-(4-methylaminophenylmethyl)-2-oxo-propyl acetamide (5) based on their nuclear magnetic resonance (NMR) and high resolution electrospray ionization mass spectroscopy (HRESIMS) data as well as their optical rotation. This class of new compound 5 had never before been found from a natural resource. Three known actinomycins showed activities in inhibiting the proliferation of glioma cells and the growth of methicillin-resistant Staphylococcus aureus, Escherichia coli, and Candida albicans and are responsible for the activity of the crude extract. Actinomycin D (1) was also found to downregulate several glioma metabolic enzymes of glycolysis, glutaminolysis, and lipogenesis, suggesting that targeting multiple tumor metabolic regulators might be a new anti-glioma mechanism of actinomycin D. This is the first report of such a possible mechanism for the class of actinomycins. Full article