Insight Into the Anti-staphylococcal Activity of JBC 1847 at Sub-Inhibitory Concentration

Multidrug-resistant pathogens constitute a serious global issue and, therefore, novel antimicrobials with new modes of action are urgently needed. Here, we investigated the effect of a phenothiazine derivative (JBC 1847) with high antimicrobial activity on Staphylococcus aureus, using a wide range of in vitro assays, flow cytometry, and RNA transcriptomics. The flow cytometry results showed that JBC 1847 rapidly caused depolarization of the cell membrane, while the macromolecule synthesis inhibition assay showed that the synthesis rates of DNA, RNA, cell wall, and proteins, respectively, were strongly decreased. Transcriptome analysis of S. aureus exposed to sub-inhibitory concentrations of JBC 1847 identified a total of 78 downregulated genes, whereas not a single gene was found to be significantly upregulated. Most importantly, there was downregulation of genes involved in adenosintrifosfat (ATP)-dependent pathways, including histidine biosynthesis, which is likely to correlate with the observed lower level of intracellular ATP in JBC 1847–treated cells. Furthermore, we showed that JBC 1847 is bactericidal against both exponentially growing cells and cells in a stationary growth phase. In conclusion, our results showed that the antimicrobial properties of JBC 1847 were primarily caused by depolarization of the cell membrane resulting in dissipation of the proton motive force (PMF), whereby many essential bacterial processes are affected. JBC 1847 resulted in lowered intracellular levels of ATP followed by decreased macromolecule synthesis rate and downregulation of genes essential for the amino acid metabolism in S. aureus. Bacterial compensatory mechanisms for this proposed multi-target activity of JBC 1847 seem to be limited based on the observed very low frequency of resistance toward the compound.

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In Vitro Cellular Uptake Studies of Self-Assembled Fluorinated Nanoparticles Labelled with Antibodies

Nanomaterials ◽

10.3390/nano11081906 ◽

2021 ◽

Vol 11 (8) ◽

pp. 1906

Author(s):

Mona Atabakhshi-Kashi ◽

Mónica Carril ◽

Hossein Mahdavi ◽

Wolfgang J. Parak ◽

Carolina Carrillo-Carrion ◽

...

Keyword(s):

Flow Cytometry ◽

Cellular Uptake ◽

Hydrophobic Interactions ◽

Intrinsic Properties ◽

Wide Range ◽

Self Assembled ◽

Uptake Studies ◽

Targeting Ability ◽

Fluorinated Nanoparticles

Nanoparticles (NPs) functionalized with antibodies (Abs) on their surface are used in a wide range of bioapplications. Whereas the attachment of antibodies to single NPs to trigger the internalization in cells via receptor-mediated endocytosis has been widely studied, the conjugation of antibodies to larger NP assemblies has been much less explored. Taking into account that NP assemblies may be advantageous for some specific applications, the possibility of incorporating targeting ligands is quite important. Herein, we performed the effective conjugation of antibodies onto a fluorescent NP assembly, which consisted of fluorinated Quantum Dots (QD) self-assembled through fluorine–fluorine hydrophobic interactions. Cellular uptake studies by confocal microscopy and flow cytometry revealed that the NP assembly underwent the same uptake procedure as individual NPs; that is, the antibodies retained their targeting ability once attached to the nanoassembly, and the NP assembly preserved its intrinsic properties (i.e., fluorescence in the case of QD nanoassembly).

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Membrane Channel Interactions Underlying Rat Subthalamic Projection Neuron Rhythmic and Bursting Activity

Journal of Neurophysiology ◽

10.1152/jn.00525.2005 ◽

2006 ◽

Vol 95 (4) ◽

pp. 2352-2365 ◽

Cited By ~ 31

Author(s):

Andrew Gillies ◽

David Willshaw

Keyword(s):

Calcium Channel ◽

High Voltage ◽

Low Voltage ◽

Low Frequency ◽

Projection Neuron ◽

Morphological Data ◽

Primary Role ◽

Small Current ◽

Wide Range

A computational model of the rat subthalamic nucleus projection neuron is constructed using electrophysiological and morphological data and a restricted set of channel specifications. The model cell exhibits a wide range of electrophysiological behaviors characteristic of rat subthalamic neurons. It reveals that a key set of three channels play a primary role in distinguishing behaviors: a high-voltage-activated calcium channel (Cav1.2.-1.3), a low-voltage-activated calcium channel (Cav3.-), and a small current calcium-activated potassium channel (KCa2.1–2.3). Short and long posthyperpolarization rebound responses, low-frequency rhythmic bursting (<1 Hz), higher-frequency rhythmic bursting (4–7 Hz), and slow action and depolarizing potentials are behaviors all mediated by the interaction of these channels. This interaction can generate a robust calcium-dependent extended depolarization in the dendrites (a depolarizing plateau). The diversity observed in the rat subthalamic physiology (such as short or long rebounds, or the presence of low-frequency rhythmic busting) can arise from alterations in both the density and distributions of these channel types and, consequently, their ability to generate this depolarizing plateau. A number of important predictions arise from the model. For example, blocking or disrupting the low-voltage-activated Cav3.- calcium current should mute the emergence of rebound responses and rhythmic bursting. Conversely, increasing this channel current via large hyperpolarizing potentials in combination with partial blockade of the high-voltage-activated calcium channels should lead to the more experimentally elusive in vitro high-frequency bursting.

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In-Vitro Evaluation of Biological Activity of New Series of 1, 3, 4-Oxadiazole Derivatives Against Bap Gene Expression in Acinetobacter bumanni Biofilm

Gene Cell and Tissue ◽

10.5812/gct.111894 ◽

2021 ◽

Vol In Press (In Press) ◽

Author(s):

Sedigheh Akbarnezhad Ghareh Lar ◽

Nakisa Zarrabi Ahrabi ◽

Yasin SarveAhrabi

Keyword(s):

Biofilm Formation ◽

Functional Group ◽

Biological Activities ◽

Antimicrobial Properties ◽

Housekeeping Gene ◽

Bacterial Biofilm ◽

Wide Range ◽

Oxadiazole Derivatives ◽

The Impact

Background: Acinetobacter bumanni is one of the most common opportunistic pathogens in health centers that is resistant to many antibiotics due to biofilm production. 1, 3, 4-oxadiazoles have a wide range of biological activities. Objectives: The aim of this research was to examine the impact of new 1, 3, 4-oxadiazole derivatives on the expression of biofilm-associated surface protein (Bap), playing an important role in promoting the biofilm formation ability of A. baumannii strains. Methods: Derivatives of 1, 3, 4-oxadiazole were synthesized through a one-step synthesis. A. baumannii strains were identified and isolated in the laboratory. The antimicrobial properties of the synthesized materials against the isolated strains were investigated. DNA, RNA, and cDNA were extracted, and the relative expression of BAP gene in A. baumannii isolates was evaluated by real-time polymerase chain reaction. Results: The compound with methoxyphenyl functional group with MIC = 62.50 mg/mL had the best inhibitory performance among all derivatives. Also, the combination of 4i reduced the expression of the Bap gene by about 24 times, but it had no effect on the expression of the 16srRNA housekeeping gene. Conclusions: 1, 3, 4-oxadiazole derivatives, especially the methoxyphenyl functional group, act as an inhibitor of bacterial biofilm formation and have the potential to be used in the pharmaceutical and biological industries.

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Engineering and Characterisation of Bacterial Phosphopantetheinyl Transferases and their Peptide Substrates

10.26686/wgtn.17060873 ◽

2021 ◽

Author(s):

◽

Jack Alexander Sissons

Keyword(s):

Single Gene ◽

Specific Protein ◽

Rapid Identification ◽

Phosphopantetheinyl Transferase ◽

Post Translational Modification ◽

Diverse Range ◽

Peptide Motifs ◽

Wide Range

<p>Throughout all domains of life, phosphopantetheinyl transferase (PPTase) enzymes catalyse a post-translational modification that is important in both primary and secondary metabolism; the transfer of a phosphopantetheine (PPant) group derived from Coenzyme A to specific protein domains within large, multi-modular biosynthetic enzymes, thereby activating each module for biosynthesis. The short peptide motif of the protein to which this group is attached is known as a ‘tag’, and can be fused to other proteins, making them also substrates for post-translational modification by a PPTase. Additionally, it has been demonstrated that PPTases can utilise a diverse range of CoA analogues, such as biotin-linked or click-chemistry capable CoA derivatives, as substrates for tag attachment. Together, these characteristics make post-translational modification by PPTases an attractive system for many different biotechnological applications. Perhaps the most significant application is in vivo and in vitro site-specific labelling of proteins, for which current technologies are hindered by cumbersome fusion protein requirements, toxicity of the process, or limited reporter groups that can be attached. Confoundingly, most PPTases exhibit a high degree of substrate promiscuity which limits the number of PPTase-tag pairs that can be used simultaneously, and therefore the number of protein targets that can be simultaneously labelled. To address this, directed evolution at a single gene level was used in an attempt to generate multiple PPTase variants that have non-overlapping tag specificity which have applications in orthogonal labelling. Furthermore, assays for the rapid identification, characterisation and evolution of short, novel peptide motifs that are recognised by PPTases has further diversified the labelling toolkit. These developments have enhanced the utility of the PPTase system and potentially have a wide range of applications in a number of fields.</p>

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The prediction of the spectrum of biological activity and antimicrobial properties of diaminoazoles

Biomeditsinskaya Khimiya ◽

10.18097/pbmc20196502099 ◽

2019 ◽

Vol 65 (2) ◽

pp. 99-102 ◽

Cited By ~ 1

Author(s):

Yu.V. Butina ◽

T.V. Kudayarova ◽

E.A. Danilova ◽

M.K. Islyaikin

Keyword(s):

Antibacterial Activity ◽

Biological Activity ◽

Antimicrobial Properties ◽

Biological Properties ◽

Diffusion Method ◽

Bacterial Strains ◽

Wide Range ◽

Alternative Drugs

The work is devoted to predicting and studying biological properties of N-substituted analogs of 3,5-diamino-1,2,4-thiadiazole, which, in their turn, include in the composition of many drugs that exhibit a wide range of pharmacological actions. For searching of new alternative drugs with an antibacterial activity, but lacking resistance of microorganism strains to them, a computer screening of 2N-alkyl-substituted 5-amino-3-imino-1,2,4-thiadiazolines previously synthesized by us was carried out. The prediction of the spectrum of biological activity, as well as the determination of the probable toxicity of these compounds, was performed using the freely available computer programs PASS, Anti-Bac-Pred, and GUSAR. The study of the antibacterial activity in vitro against gram-positive (Staphylococcus aureus, Staphylococcus saprophyticus, Staphylococcus epidermidis) and gram-negative (Escherichia coli, Pseudomonas aeruginosae) bacterial strains was performed by the disco-diffusion method. Experimental data roughly correspond to the predictions.

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Vγ9Vδ2 T cells strengthen cisplatin inhibition activity against breast cancer MDA-MB-231 cells by disrupting mitochondrial function and cell ultrastructure

Cancer Cell International ◽

10.1186/s12935-021-01815-0 ◽

2021 ◽

Vol 21 (1) ◽

Author(s):

Xin Huang ◽

Cunchuan Wang ◽

Ningxia Wang

Keyword(s):

Breast Cancer ◽

Flow Cytometry ◽

T Cells ◽

Cell Membrane ◽

Cancer Cells ◽

Cancer Patients ◽

Mitochondrial Function ◽

Inhibition Activity ◽

Vγ9vδ2 T Cells

Abstract Background Breast cancer ranks second of new cases and fifth of death in 2018 worldwide. Cis-platinum (CDDP) has been used as a chemotherapy to treat breast cancer for years. However, CDDP can adversely disrupt immune function of host. Thus, development of new protocol that can minimize side effect and meanwhile elevate clinical efficacy of CDDP will eventually benefit cancer patients. Since Vγ9Vδ2 T cells can up-regulate immune function of cancer patients, therefore, our hypothesis is that introduction of Vγ9Vδ2 T cells could potentiate CDDP efficacy against breast cancer. Methods We used breast cancer cell line MDA-MB-231 as model cell to test our hypothesis. The cancer cell viability in vitro in the context of different dose of CDDP was analyzed by flow cytometry. The cytoskeleton alteration was visualized by confocal microscopy, and the ultrastructure of cell membrane was observed by atomic force microscopy. The mitochondrial function of MDA-MB-231 cells was detected as well by flow cytometry. Results Comparing to either Vγ9Vδ2 T cells or CDDP alone, Vγ9Vδ2 T cells plus CDDP could more strikingly induce MDA-MB-231 cell membrane ultrastructure disruption and cytoskeleton disorder, and more significantly enhance the inhibition of CDDP on proliferation of MDA-MB-231 cells. At the same time, Vγ9Vδ2 T cells strengthened CDDP-induced mitochondrial dysfunction of cancer cells. Conclusion This work revealed that Vγ9Vδ2 T cells could synergistically enhance the inhibition activity of CDDP against breast cancer cells. Meanwhile, this in vitro proof-of-concept study implied the clinical prospect of the combining application of Vγ9Vδ2 T cells and CDDP in breast cancer therapy.

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Secretory sphingomyelinase in health and disease

Biological Chemistry ◽

10.1515/hsz-2015-0109 ◽

2015 ◽

Vol 396 (6-7) ◽

pp. 707-736 ◽

Cited By ~ 56

Author(s):

Johannes Kornhuber ◽

Cosima Rhein ◽

Christian P. Müller ◽

Christiane Mühle

Keyword(s):

Current Knowledge ◽

Clinical Chemistry ◽

Single Gene ◽

Enzyme Activation ◽

Acid Sphingomyelinase ◽

Sphingolipid Metabolism ◽

Glycosylation Pattern ◽

Wide Range

Abstract Acid sphingomyelinase (ASM), a key enzyme in sphingolipid metabolism, hydrolyzes sphingomyelin to ceramide and phosphorylcholine. In mammals, the expression of a single gene, SMPD1, results in two forms of the enzyme that differ in several characteristics. Lysosomal ASM (L-ASM) is located within the lysosome, requires no additional Zn2+ ions for activation and is glycosylated mainly with high-mannose oligosaccharides. By contrast, the secretory ASM (S-ASM) is located extracellularly, requires Zn2+ ions for activation, has a complex glycosylation pattern and has a longer in vivo half-life. In this review, we summarize current knowledge regarding the physiology and pathophysiology of S-ASM, including its sources and distribution, molecular and cellular mechanisms of generation and regulation and relevant in vitro and in vivo studies. Polymorphisms or mutations of SMPD1 lead to decreased S-ASM activity, as detected in patients with Niemann-Pick disease B. Thus, lower serum/plasma activities of S-ASM are trait markers. No genetic causes of increased S-ASM activity have been identified. Instead, elevated activity is the result of enhanced release (e.g., induced by lipopolysaccharide and cytokine stimulation) or increased enzyme activation (e.g., induced by oxidative stress). Increased S-ASM activity in serum or plasma is a state marker of a wide range of diseases. In particular, high S-ASM activity occurs in inflammation of the endothelium and liver. Several studies have demonstrated a correlation between S-ASM activity and mortality induced by severe inflammatory diseases. Serial measurements of S-ASM reveal prolonged activation and, therefore, the measurement of this enzyme may also provide information on past inflammatory processes. Thus, S-ASM may be both a promising clinical chemistry marker and a therapeutic target.

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Inhibitory activity of stilbenes against filamentous fungi

Italian Journal of Food Safety ◽

10.4081/ijfs.2021.8461 ◽

2021 ◽

Vol 10 (1) ◽

Author(s):

Luce Mattio ◽

Giorgia Catinella ◽

Marcello Iriti ◽

Lisa Vallone

Keyword(s):

Antifungal Activity ◽

Filamentous Fungi ◽

Antimicrobial Properties ◽

Microbial Infection ◽

Wide Range ◽

Mycotoxigenic Fungi ◽

Wild Strains ◽

Diffusion Assay ◽

Inflammatory Action

Stilbenoids (resveratrol and its derivatives) are secondary metabolites produced by plants as defence mechanism to microbial infection. These compounds are known for their anti-inflammatory action and health benefits in preventing a wide range of disorders (e.g. cancer and cardiovascular diseases). However, their antimicrobial properties are less investigated. A series of 8 stilbenoid compounds were synthesized and their antifungal activity against 19 wild strains of filamentous fungi and yeasts (isolated from the environment and food) was tested in vitro. Using an agar diffusion assay, compounds were tested at the concentration of 100 μg/ml on filamentous fungi and yeasts at 104 CFU/ml. The results showed that tested derivatives possess moderate antifungal activity: in particular, monomeric stilbenoids 3’-hydroxy-pterostilbene and piceatannol, and dimeric stilbenoids (±)-trans-δ-viniferin and pallidol were active against mycotoxigenic fungi.

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Antimicrobial properties of phytohormone (gibberellins) against phytopathogens and clinical pathogens

Access Microbiology ◽

10.1099/acmi.0.000278 ◽

2021 ◽

Vol 3 (10) ◽

Author(s):

Paoirse Toner ◽

David Nelson ◽

Juluri R. Rao ◽

Madeleine Ennis ◽

John E. Moore ◽

...

Keyword(s):

Pathogenic Bacteria ◽

Antimicrobial Properties ◽

Inhibitory Effect ◽

Phytopathogenic Fungi ◽

Microbial Pathogens ◽

Inhibitory Effects ◽

Plant Host ◽

Therapeutic Value ◽

Wide Range

The in vitro antimicrobial potential of physiologically active diterpenoid plant-derived gibberellins (gibberellic acids; GAs) was tested on microbial pathogens of significance to plant and human health. The racemic enantiomer GA3 produced varying inhibitory effects against a wide range of plant host disease causal agents (phytopathogens) comprising fungi, oomycetes and bacteria. The results showed that GA3 effected either strong growth arrest of phytopathogenic fungi or holistic biocidal effects on oomycete and phytopathogenic fungi at higher concentration (>10–50 mM) and increased hyphal extension growth when the concentration of GA3 was lowered (<10−0.1 mM). When human clinical pathogenic bacteria cohorts were challenged with gibberellin enantiomers, viz GA1, GA4, GA5, GA7, GA9 and GA13 (50 mM), employing Kirby–Bauer disc bioassay methods for assessment of their efficacies, no inhibitory effect was seen with gibberellin enantiomers, viz GA1, GA3, GA5 and GA13, while GA4 inhibited all human clinical bacterial organisms examined, with GA7 and GA9 showing limited activity. The antibiotic effects of enantiomeric diterpenoid phytohormones evinced in our preliminary study raise prospects for further studies to fully examine their potential therapeutic value for human healthcare and their compliance with cytotoxicity and other ethical considerations in the future.

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Effect of Cytokinins on Micropropagation of Moringa Oleifera Lam. and Genomic Stability Assessment Using Flow-Cytometry

10.21203/rs.3.rs-1238384/v1 ◽

2022 ◽

Author(s):

Rohit Bharati ◽

Moses Okao ◽

Katerina Hamouzová ◽

Eloy Fernandez-Cusimamani

Keyword(s):

Flow Cytometry ◽

Moringa Oleifera ◽

Genetic Fidelity ◽

Limited Range ◽

Surface Sterilization ◽

Wide Range ◽

The Family ◽

Livestock Feeding ◽

Planting Materials

Abstract Moringa oleifera Lam. is a multipurpose medicinal plant of the family Moringaceae which has been widely utilized as a pharmaceutical remedy to treat a wide range of diseases. In addition, the tree has several applications in human nutrition as well as livestock feeding. M. oleifera is easily multiplied through epigeal germination (recalcitrant) but seed propagated plants are heterogeneous and take longer to reach fruit-bearing age. As an alternative, branch cuttings have been used but their establishment is erratic and often leads to reduced growth of the mother plant. Thus, to produce superior planting materials, in-vitro propagation has become paramount. As a result, several studies using a limited range of cytokinin have been undertaken to multiply M. oleifera through tissue culture. Otherwise, a study was conducted to examine the effect of five different cytokinins on in-vitro regeneration of this tree species. Results showed that nodal explants cultured on Murashige and Skoog (MS) medium supplemented with 1.0 mg/L 6-Benzylaminopurine (BAP) and subsequently rooted on basic MS media was the most optimal treatment. Furthermore, acclimatization of plantlets in sterile soil substrate and perlite (1:3;v/v) under transparent polythene sheet for 7 days resulted in survival rate of 100%. Assessment of genetic fidelity using flow cytometry revealed that surface sterilization alongside cytokinin treatments produced plantlets that were genetically stable regardless of the growth regulator used. Thus, the in-vitro protocol developed in this study can be utilized for in-vitro studies and mass propagation of this imperative plant species.

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