scholarly journals The processing of staphylococcal biofilm images from Fluorescence and Confocal Microscopy to assess in vitro efficacy of antiseptic molecules

2021 ◽  
Author(s):  
Adam Feliks Junka ◽  
Grzegorz Krasowski ◽  
Pawel Migdal ◽  
Marta Woroszylo ◽  
Karol Fijalkowski ◽  
...  
Keyword(s):  
Quantitative Methods ◽  
Live Cells ◽  
Parametric Image ◽  
Microscopic Images ◽  
Cellular Density ◽  
Parametric Data ◽  
Method Of Assessment ◽  
Syto 9 ◽  
The Impact

The in vitro efficacy of locally applied antiseptic molecules against staphylococcal biofilm is frequently assessed by a set of standard quantitative and semi-quantitative methods. The development of software for parametric image processing allowed to obtain parametric data also from microscopic images of biofilm dyed with a variety of dyes, especially with propidium iodine and SYTO-9, differentiating dead from live cells. In this work, using confocal/epifluorescent microscopy, we analyzed such major properties of staphylococcal biofilm in vitro as its thickness, cellular density and share of Live/Dead cells within its individual parts. We also scrutinized the impact of sample preparation and antiseptic introduction on the outcome obtained. As a result of our analyses we developed a revelatory method of assessment of the impact of antiseptic agents on staphylococcal biofilm in vitro, in which the microscopic images are processed with the use of ABE formula (Antiseptics Biofilm Eradication) which implements all the data and phenomena detected and revealed within the course of this study. We tested ABE with regard to polyhexanide, povidone-iodine and hypochlorous antiseptics and found a high correlation between this parameter and the results obtained by means of traditional techniques. Taking into account the fact that in vitro results of the efficacy of antiseptic agents against staphylococcal biofilm are frequently applied to back up their use in hospitals and ambulatory units, our work should be considered an important tool providing reliable, parametric data with this regard.

scholarly journals Application Solid Laser-Sintered or Machined Ti6Al4V Alloy in Manufacturing of Dental Implants and Dental Prosthetic Restorations According to Dentistry 4.0 Concept

Processes ◽  
2020 ◽  
Vol 8 (6) ◽  
pp. 664 ◽  
Author(s):  
Leszek A. Dobrzański ◽  
Lech B. Dobrzański ◽  
Anna Achtelik-Franczak ◽  
Joanna Dobrzańska
Keyword(s):  
Dental Implants ◽  
Computer Aided Design ◽  
Bending Strength ◽  
Ti6al4v Alloy ◽  
Cnc Machining ◽  
Live Cells ◽  
Computer Aided ◽  
Aided Design ◽  
The Impact

This paper presents a comparison of the impact of milling technology in the computer numerically controlled (CNC) machining centre and selective laser sintering (SLS) and on the structure and properties of solid Ti6Al4V alloy. It has been shown that even small changes in technological conditions in the SLS manufacturing variant significantly affect changes from two to nearly two and a half times in tensile and bending strengths. Both the tensile and bending strength obtained in the most favourable manufacturing variant by the SLS method is over 25% higher than in the case of cast materials subsequently processed by milling. Plug-and-play SLS conditions provide about 60% of the possibilities. Structural, tribological and electrochemical tests were carried out. In vitro biological tests using osteoblasts confirm the good tendency for the proliferation of live cells on the substrate manufactured under the most favourable SLS conditions. The use of SLS additive technology for the manufacturing of dental implants and abutments made of Ti6Al4V alloy in combination with the digitisation of dental diagnostics and computer-aided design and manufacture of computer-aided design/manufacturing (CAD/CAM) following the idea of Dentistry 4.0 is the best choice of technology for manufacturing of prosthetic and implant devices used in dentistry.

scholarly journals Direct Visualization of Amlodipine Intervention into Living Cells by Means of Fluorescence Microscopy

Molecules ◽  
2021 ◽  
Vol 26 (10) ◽  
pp. 2997
Author(s):  
Christine Quentin ◽  
Rūta Gerasimaitė ◽  
Alexandra Freidzon ◽  
Levon S. Atabekyan ◽  
Gražvydas Lukinavičius ◽  
...  
Keyword(s):  
Fluorescence Microscopy ◽  
Aqueous Solutions ◽  
Time Lapse ◽  
Drug Uptake ◽  
Live Cells ◽  
Direct Visualization ◽  
Cellular Environment ◽  
Time Lapse Imaging ◽  
The Impact

Amlodipine, a unique long-lasting calcium channel antagonist and antihypertensive drug, has weak fluorescence in aqueous solutions. In the current paper, we show that direct visualization of amlodipine in live cells is possible due to the enhanced emission in cellular environment. We examined the impact of pH, polarity and viscosity of the environment as well as protein binding on the spectral properties of amlodipine in vitro, and used quantum chemical calculations for assessing the mechanism of fluorescence quenching in aqueous solutions. The confocal fluorescence microscopy shows that the drug readily penetrates the plasma membrane and accumulates in the intracellular vesicles. Visible emission and photostability of amlodipine allow confocal time-lapse imaging and the drug uptake monitoring.

scholarly journals Cells Lacking PA200 Adapt to Mitochondrial Dysfunction by Enhancing Glycolysis via Distinct Opa1 Processing

2021 ◽  
Vol 22 (4) ◽  
pp. 1629
Author(s):  
Abdennour Douida ◽  
Frank Batista ◽  
Pal Boto ◽  
Zsolt Regdon ◽  
Agnieszka Robaszkiewicz ◽  
...  
Keyword(s):  
Atp Synthase ◽  
Neuroblastoma Cells ◽  
Selective Inhibition ◽  
Basal Respiration ◽  
Live Cells ◽  
Mitochondrial Morphology ◽  
Transcriptional Level ◽  
Genetic Ablation ◽  
The Impact

The conserved Blm10/PA200 proteins are proteasome activators. Previously, we identified PA200-enriched regions in the genome of SH-SY5Y neuroblastoma cells by chromatin immunoprecipitation (ChIP) and ChIP-seq analysis. We also found that selective mitochondrial inhibitors induced PA200 redistribution in the genome. Collectively, our data indicated that PA200 regulates cellular homeostasis at the transcriptional level. In the present study, our aim is to investigate the impact of stable PA200 depletion (shPA200) on the overall transcriptome of SH-SY5Y cells. RNA-seq data analysis reveals that the genetic ablation of PA200 leads to overall changes in the transcriptional landscape of SH-SY5Y neuroblastoma cells. PA200 activates and represses genes regulating metabolic processes, such as the glycolysis and mitochondrial function. Using metabolic assays in live cells, we showed that stable knockdown of PA200 does not change basal respiration. Spare respiratory capacity and proton leak however are slightly, yet significantly, reduced in PA200-deficient cells by 99.834% and 84.147%, respectively, compared to control. Glycolysis and glycolytic capacity show a 42.186% and 26.104% increase in shPA200 cells, respectively, compared to control. These data suggest a shift from oxidative phosphorylation to glycolysis especially when cells are exposed to oligomycin-induced stress. Furthermore, we observed a preserved long and compact tubular mitochondrial morphology after inhibition of ATP synthase by oligomycin, which might be associated with the glycolytic change of shPA200 cells. The present study also demonstrates that the proteolytic cleavage of Opa1 is affected, and that the level of OMA1 is significantly reduced in shPA200 cells upon oligomycin-induced mitochondrial insult. Together, these findings suggest a role for PA200 in the regulation of metabolic changes in response to selective inhibition of ATP synthase in an in vitro cellular model.

Multi-mode light microscopy of microtubule assembly dynamics and chromosome movement in vivo and In vitro

1996 ◽  
Vol 54 ◽  
pp. 730-731
Author(s):  
E. D. Salmon ◽  
J. C. Waters ◽  
C. Waterman-Storer
Keyword(s):  
Imaging System ◽  
Ccd Camera ◽  
Transmitted Light ◽  
Microtubule Assembly ◽  
Live Cells ◽  
Optical Efficiency ◽  
Multiple Band ◽  
Multi Mode

We have developed a multi-mode digital imaging system which acquires images with a cooled CCD camera (Figure 1). A multiple band pass dichromatic mirror and robotically controlled filter wheels provide wavelength selection for epi-fluorescence. Shutters select illumination either by epi-fluorescence or by transmitted light for phase contrast or DIC. Many of our experiments involve investigations of spindle assembly dynamics and chromosome movements in live cells or unfixed reconstituted preparations in vitro in which photodamage and phototoxicity are major concerns. As a consequence, a major factor in the design was optical efficiency: achieving the highest image quality with the least number of illumination photons. This principle applies to both epi-fluorescence and transmitted light imaging modes. In living cells and extracts, microtubules are visualized using X-rhodamine labeled tubulin. Photoactivation of C2CF-fluorescein labeled tubulin is used to locally mark microtubules in studies of microtubule dynamics and translocation. Chromosomes are labeled with DAPI or Hoechst DNA intercalating dyes.

The Predominant microRNAs in β-cell Clusters for Insulin Regulation and Diabetic Control

2020 ◽  
Vol 21 (7) ◽  
pp. 722-734
Author(s):  
Adele Soltani ◽  
Arefeh Jafarian ◽  
Abdolamir Allameh
Keyword(s):  
Mirna Expression ◽  
Expression Profiles ◽  
Expression Patterns ◽  
Diabetic Control ◽  
In Vitro Proliferation ◽  
Cell Functions ◽  
Mirna Expression Profiles ◽  
Multiple Processes ◽  
The Impact

micro (mi)-RNAs are vital regulators of multiple processes including insulin signaling pathways and glucose metabolism. Pancreatic β-cells function is dependent on some miRNAs and their target mRNA, which together form a complex regulative network. Several miRNAs are known to be directly involved in β-cells functions such as insulin expression and secretion. These small RNAs may also play significant roles in the fate of β-cells such as proliferation, differentiation, survival and apoptosis. Among the miRNAs, miR-7, miR-9, miR-375, miR-130 and miR-124 are of particular interest due to being highly expressed in these cells. Under diabetic conditions, although no specific miRNA profile has been noticed, the expression of some miRNAs and their target mRNAs are altered by posttranscriptional mechanisms, exerting diverse signs in the pathobiology of various diabetic complications. The aim of this review article is to discuss miRNAs involved in the process of stem cells differentiation into β-cells, resulting in enhanced β-cell functions with respect to diabetic disorders. This paper will also look into the impact of miRNA expression patterns on in vitro proliferation and differentiation of β-cells. The efficacy of the computational genomics and biochemical analysis to link the changes in miRNA expression profiles of stem cell-derived β-cells to therapeutically relevant outputs will be discussed as well.

The Impact of Antioxidants from the Diet on Breast Cancer Cells Monitored by Raman Microspectroscopy

2018 ◽  
Vol 16 (2) ◽  
pp. 127-137
Author(s):  
Paula Sofia Coutinho Medeiros ◽  
Ana Lúcia Marques Batista de Carvalho ◽  
Cristina Ruano ◽  
Juan Carlos Otero ◽  
Maria Paula Matos Marques
Keyword(s):  
Breast Cancer ◽  
Cancer Cells ◽  
Cell Line ◽  
Breast Cancer Cells ◽  
Triple Negative ◽  
Breast Adenocarcinoma ◽  
Coumaric Acid ◽  
Human Breast ◽  
The Impact

Background: The impact of the ubiquitous dietary phenolic compound p-coumaric acid on human breast cancer cells was assessed, through a multidisciplinary approach: Combined biological assays for cytotoxicity evaluation and biochemical profiling by Raman microspectroscopic analysis in cells. </P><P> Methods: Para-coumaric acid was shown to exert in vitro chemoprotective and antitumor activities, depending on the concentration and cell line probed: a significant anti-invasive ability was detected for the triple-negative MDA-MB-231 cells, while a high pro-oxidant effect was found for the estrogen- dependent MCF-7 cells. A striking cell selectivity was obtained, with a more noticeable outcome on the triple-negative MDA-MB-231 cell line. Results: The main impact on the cellular biochemical profile was verified to be on proteins and lipids, thus justifying the compound´s anti-invasive effect and chemoprotective ability. Conclusion: p-Coumaric acid was thus shown to be a promising chemoprotective/chemotherapeutic agent, particularly against the low prognosis triple-negative human breast adenocarcinoma.

VDAC1 Mediated Anticancer Activity of Gallic Acid in Human Lung Adenocarcinoma A549 Cells

2018 ◽  
Vol 18 (2) ◽  
pp. 255-262 ◽  
Author(s):  
Aikebaier Maimaiti ◽  
Amier Aili ◽  
Hureshitanmu Kuerban ◽  
Xuejun Li
Keyword(s):  
Western Blot ◽  
Cell Viability ◽  
Gallic Acid ◽  
Molecular Mechanisms ◽  
A549 Cells ◽  
Dependent Manner ◽  
Lung Carcinoma Cells ◽  
Induced Apoptosis ◽  
The Impact

Aims: Gallic acid (GA) is generally distributed in a variety of plants and foods, and possesses cell growth-inhibiting activities in cancer cell lines. In the present study, the impact of GA on cell viability, apoptosis induction and possible molecular mechanisms in cultured A549 lung carcinoma cells was investigated. Methods: In vitro experiments showed that treating A549 cells with various concentrations of GA inhibited cell viability and induced apoptosis in a dose-dependent manner. In order to understand the mechanism by which GA inhibits cell viability, comparative proteomic analysis was applied. The changed proteins were identified by Western blot and siRNA methods. Results: Two-dimensional electrophoresis revealed changes that occurred to the cells when treated with or without GA. Four up-regulated protein spots were clearly identified as malate dehydrogenase (MDH), voltagedependent, anion-selective channel protein 1(VDAC1), calreticulin (CRT) and brain acid soluble protein 1(BASP1). VDAC1 in A549 cells was reconfirmed by western blot. Transfection with VDAC1 siRNA significantly increased cell viability after the treatment of GA. Further investigation showed that GA down regulated PI3K/Akt signaling pathways. These data strongly suggest that up-regulation of VDAC1 by GA may play an important role in GA-induced, inhibitory effects on A549 cell viability.

Effects of Transport Medium Composition on in vitro Drug Permeation across Excised Pig Intestinal Tissue

2020 ◽  
Vol 10 ◽  
Author(s):  
Bianca Peterson ◽  
Henrico Heystek ◽  
Josias H. Hamman ◽  
Johan D. Steyn
Keyword(s):  
Medium Composition ◽  
Screening Tests ◽  
Intestinal Tissue ◽  
Predictive Values ◽  
Transport Medium ◽  
Intestinal Fluids ◽  
Lower Permeability ◽  
The Impact ◽  
Simulated Intestinal Fluids

Background:: Knowledge of the permeation characteristics of new chemical entities across biological membranes is essential to drug research and development. Transport medium composition may affect the absorption of compounds during in vitro drug transport testing. To preserve the predictive values of screening tests, the possible influence of transport media on the solubility of model drugs, and on the activities of tight junctions and efflux transporter proteins (e.g. P-glycoprotein) must be known. Objective:: The aim of this study was to compare the impact of different transport media on the bi-directional transport of standard compounds, selected from the four classes of the Biopharmaceutical Classification System (BCS), across excised pig intestinal tissue. Methods:: The Sweetana-Grass diffusion apparatus was used for the transport studies. Krebs-Ringer bicarbonate (KRB) buffer and simulated intestinal fluids in the fed (FeSSIF) and fasted (FaSSIF) states were used as the three transport media, while the chosen compounds were abacavir (BCS class 1), dapsone (BCS class 2), lamivudine (BCS class 3) and furosemide (BCS class 4). Results:: Abacavir exhibited lower permeability in both the simulated intestinal fluids than in the KRB buffer. Dapsone showed similar permeability in all media. Lamivudine exhibited lower permeability in FaSSIF than in the other two media. Furosemide exhibited improved transport with pronounced efflux in FaSSIF. Conclusion:: Different permeation behaviors were observed for the selected drugs in the respective media, which may have resulted from their different physico-chemical properties, as well as from the effects that dissimilar transport media components had on excised pig intestinal tissue.

The Relationship Between Highly Visual Social Media and Young People’s Mental Health: A Scoping Review (Preprint)

2020 ◽  
Author(s):  
Alanna McCrory
Keyword(s):  
Mental Health ◽  
Social Media ◽  
Young People ◽  
Scoping Review ◽  
Quantitative Methods ◽  
Grey Literature ◽  
Significant Proportion ◽  
Mental Wellbeing ◽  
Published Evidence ◽  
The Impact

UNSTRUCTURED Users of highly visual social media (HVSM), such as Snapchat and Instagram, share their messages through images, rather than relying on words. A significant proportion of people that use these platforms are adolescents. Previous research reveals mixed evidence regarding the impact of online social technologies on this age group’s mental wellbeing, but it is uncertain whether the psychological effects of visual content alone differ from text-driven social media. This scoping review maps existing literature that has published evidence about highly visual social media, specifically its psychological impact on young people. Nine electronic databases and grey literature from 2010 until March 2019 were reviewed for articles describing any aspect of visual social media, young people and their mental health. The screening process retrieved 239 articles. With the application of eligibility criteria, this figure was reduced to 25 articles for analysis. Results indicate a paucity of data that exclusively examines HVSM. The predominance of literature relies on quantitative methods to achieve its objectives. Many findings are inconsistent and lack the richness that qualitative data may provide to explore the reasons for theses mixed findings.

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