scholarly journals First Description of the Generation of Human Schwann Cells Transfected With CRISPR Cas9 and a Model of Metformin Mitochondrial Metabolism in Metachromatic Leukodystrophy

2021 ◽  
Author(s):  
Nayibe Tatiana Sanchez Alvarez ◽  
Paula Katherine Bautista-Niño ◽  
Juanita Trejos-Suárez ◽  
Norma Cecilia Serrano-Díaz
Keyword(s):  
Cell Death ◽  
Cell Viability ◽  
Schwann Cells ◽  
Metachromatic Leukodystrophy ◽  
In Vitro Study ◽  
Basal Respiration ◽  
Ros Production ◽  
Transfected Cells ◽  
Significant Difference

Abstract Background: Metachromatic leukodystrophy (MLD) is a neurological lysosomal deposit disease that has an impact on public health despite its low incidence in the population. Existing treatments are expensive and inefficient. Few reports in the literature on pathophysiological events related to enzyme deficiency and subsequent accumulation of sulfatides; therefore, the use of metformin as an alternative treatment was evaluated in vitro to counteract the effects. Methodology: An experimental in vitro study that sought to determine the effect of the use of metformin on the accumulation of sulfates in glycolysis and mitochondrial function in an in vitro model of metachromatic leukodystrophy. Human Schwann cells (CSH) transfected with CRISPR Cas9 and without transfection were treated with different concentrations of sulfatides and metformin. Cell viability was evaluated by MTT and SYTOX Green; mitochondrial and glycolytic function by Seahorse XFe24, determination of reactive oxygen species (ROS) and cell death. Results: In the MTT trials, we found that treatment with different concentrations of sulfates did not affect cell viability. Transfected CSH showed higher cell death and ROS production when exposed to 100 µM sulfatides with a statistically significant difference (p <0.001), compared to nontransfected CSH cells. Sulfatides at concentrations of 10 to 100 µM affect mitochondrial bioenergetics as concentrations increase in transfected cells, in nontransfected cells they respond metabolically to exposure; Furthermore, transfected cells show a decrease in basal respiration and maximum respiration after being exposed to a concentration of 100 µM of sulphates; however, in double treatment of these cells with both sulfates and Metformin, respiration also decreases. Maximum and normal mitochondrial respiratory capacity. Conclusion: This research describes for the first time the generation of transfected CSH and the bioenergetic and mitochondrial effect of sulfates in Schwann cells, treatment with 500 µM of Metformin restores metabolic activity of these cells and decreases ROS production, as well as prevention of cell death.

scholarly journals A Preliminary Research Study on the Cytotoxicity of Expired and Non-expired Composite Resins: In Vitro Study

2020 ◽  
pp. 227-238
Author(s):  
Magrur Kazak DDS, PhD ◽  
Nazmiye Donmez DDS, PhD ◽  
Fatemeh Bahadori PhD ◽  
Vildan Betul Yenigun PhD ◽  
Abdurrahim Kocyigit MD, PhD
Keyword(s):  
Cell Viability ◽  
Composite Resin ◽  
In Vitro Study ◽  
Negative Control Group ◽  
Negative Control ◽  
Composite Resins ◽  
Control Group ◽  
Significant Difference ◽  
Preliminary Research

Objective: Studies have focused on use of non-expired composites. Unfortunately some clinicians still use expired composite resins without considering their effects. The objective of this in vitro preliminary research was to investigate cytotoxicity of expired(6-months) and non-expired composite resins. Materials and methods: Expired (E) and non-expired (NE) samples of one bulk-fill (Tetric N-Ceram Bulk-fill [TNB], Ivoclar Vivadent), two nano-hybrid (Tetric N-Ceram [TN], Ivoclar Vivadent; Clearfil Majesty ES-2 [CM], Kuraray) composite resins were tested on L929 fibroblast cells. Medium covering cells was removed then plastic rings (2-mm height) were filled with non-polymerized composite resins, placed in direct contact with cells and polymerized with LED light curing unit (LCU). Three samples were prepared for each group. After polymerization, removed medium was added to the cells. Cells that were left without medium (WOM) and cells that were exposed to LCU were used as positive control groups. Cells without any treatment were used as negative control group (C). Cells were incubated with tested materials for 7-days to evaluate cytotoxicity. Cell viability was calculated by sulforhodamine B test as a percentage (%). One-way ANOVA and post-hoc Tukey tests were used for statistical analyses (p<0.05). Results: Comparison between E and NE groups of same composite resins did not result in statistically significant differences (p>0.05), except between TN NE and TN E (p<0.05). TN E group was significantly more cytotoxic than TN NE group. When NE composite resin groups were compared to each other, statistically significant difference was only obtained between TNB NE and TN NE (p<0.05). Among all tested groups, TN NE group showed the least cytotoxic profile. No statistically significant differences were determined when E composite resin groups were compared to each other (p>0.05). All experimental groups compared with C group showed statistically significant cytotoxicity (p<0.05). A statistically significant difference existed between LCU and C groups (p<0.05). Conclusions: In clinical practice, expired composite resins should never be used. Although a correlation was found between expiration dates of nano-hybrid composite resins and cell viability, opposite data were obtained for bulk-fill composite resin. Researches are still required to evaluate biocompatibility of bulk-fill composite resins at various thicknesses with current LCUs.

scholarly journals Establishing Irreversible Electroporation Electric Field Potential Threshold in A Suspension In Vitro Model for Cardiac and Neuronal Cells

2021 ◽  
Vol 10 (22) ◽  
pp. 5443
Author(s):  
Sahar Avazzadeh ◽  
Barry O’Brien ◽  
Ken Coffey ◽  
Martin O’Halloran ◽  
David Keane ◽  
...  
Keyword(s):  
Cell Death ◽  
Electric Field ◽  
Cell Viability ◽  
Cardiac Fibroblasts ◽  
Irreversible Electroporation ◽  
Field Potential ◽  
Optimal Threshold ◽  
Delayed Cell Death ◽  
Significant Difference

Aims: Irreversible electroporation is an ablation technique being adapted for the treatment of atrial fibrillation. Currently, there are many differences reported in the in vitro and pre-clinical literature for the effective voltage threshold for ablation. The aim of this study is a direct comparison of different cell types within the cardiovascular system and identification of optimal voltage thresholds for selective cell ablation. Methods: Monophasic voltage pulses were delivered in a cuvette suspension model. Cell viability and live–dead measurements of three different neuronal lines, cardiomyocytes, and cardiac fibroblasts were assessed under different voltage conditions. The immediate effects of voltage and the evolution of cell death was measured at three different time points post ablation. Results: All neuronal and atrial cardiomyocyte lines showed cell viability of less than 20% at an electric field of 1000 V/cm when at least 30 pulses were applied with no significant difference amongst them. In contrast, cardiac fibroblasts showed an optimal threshold at 1250 V/cm with a minimum of 50 pulses. Cell death overtime showed an immediate or delayed cell death with a proportion of cell membranes re-sealing after three hours but no significant difference was observed between treatments after 24 h. Conclusions: The present data suggest that understanding the optimal threshold of irreversible electroporation is vital for achieving a safe ablation modality without any side-effect in nearby cells. Moreover, the evolution of cell death post electroporation is key to obtaining a full understanding of the effects of IRE and selection of an optimal ablation threshold.

scholarly journals Preeclampsia with Intrauterine Growth Restriction Generates Morphological Changes in Endothelial Cells Associated with Mitochondrial Swelling—An In Vitro Study

2019 ◽  
Vol 8 (11) ◽  
pp. 1994 ◽  
Author(s):  
Dorota Formanowicz ◽  
Agnieszka Malińska ◽  
Marcin Nowicki ◽  
Katarzyna Kowalska ◽  
Karolina Gruca-Stryjak ◽  
...  
Keyword(s):  
Endothelial Cell ◽  
Cell Viability ◽  
Pregnant Women ◽  
Intrauterine Growth Restriction ◽  
Morphological Changes ◽  
In Vitro Study ◽  
Growth Restriction ◽  
Ros Production ◽  
Intrauterine Growth

Pregnancy complicated by preeclampsia (PE) and intrauterine growth restriction (IUGR) promotes endothelial cell (EC) dysfunction. Our in vitro study aimed to evaluate the endothelial cell morphology after acute and chronic exposition to medium supplemented with serum taken from healthy pregnant women and women with IUGR and IUGR with PE. In the same condition, ECs viability, proliferation, reactive oxygen species (ROS) production, and serum concentration of vascular endothelial growth factor (VEGF) were also measured. Pregnant women with IUGR and IUGR with PE-delivered babies with reduced body mass and were characterized in elevated blood pressure, urine protein loss, and reduced level of VEGF. The 24 hours of exposition did not exert any morphological changes in ECs, except the reduction in cell viability, but prolonged exposition resulted in significant morphological changes concerning mostly the swelling of mitochondria with accompanying ROS production, cell autophagy, reduced cell viability, and proliferation only in complicated pregnancies. In conclusion, the sera taken from women with IUGR and IUGR with PE show a detrimental effect on ECs, reducing their viability, proliferation, and generating oxidative stress due to dysfunctional mitochondria. This multidirectional effect might have an adverse impact on the cardiovascular system in women with IUGR and PE.

Comparative Study to Evaluate the Surface Detail Reproduction of Dental Stone after Immersion in Various Different Disinfectant Solutions, Under Stereomicroscope 10 X Magnification –An In-Vitro Study

2017 ◽  
Vol 9 (3) ◽  
Author(s):  
Rathika Rai ◽  
M. A. Easwaran ◽  
K. T. Dhivya
Keyword(s):  
Sodium Hypochlorite ◽  
In Vitro Study ◽  
Control Group ◽  
Immersion Method ◽  
Significant Difference ◽  
Surface Irregularities ◽  
Disinfectant Solution ◽  
Surface Detail ◽  
Group B

Aim: To evaluate the surface detail reproduction of dental stone this is immersed in different disinfectant solution and studied under stereomicroscope. Methodology: Total number of 30 specimens of dental stone (Type III) were made with measurements of 1.5cm diameter and 1cm height .This samples are divided in to 3 groups group A,B,C. were A is immersed in Distilled water which was taken as control group ;B is immersed in 2% Glutaraldehyde and C is immersed in 5%sodium hypochlorite. Each specimen were immersed in the disinfectant solution for 15 minutes and dried under room temperature for 24 hrs. After 24 hrs each specimens are studied under stereomicroscope for surface details. Result: The results showed no significant difference in the surface irregularities and porosities for a group 1 and group 2 except group 3 which showed significant increase in the porosities, surface irregularities and erosions after disinfection with 5% NaHOCl by immersion method. Conclusion: The surface detail reproduction capacity of die stone was adversely affected when 5% Sodium hypochlorite was used as disinfectant solution when compare d to control group and 2% Glutaraldehyde

Stew in its Own Juice: Protein Homeostasis Machinery Inhibition Reduces Cell Viability in Multiple Myeloma Cell Lines

2019 ◽  
Vol 19 (2) ◽  
pp. 112-119 ◽  
Author(s):  
Mariana B. de Oliveira ◽  
Luiz F.G. Sanson ◽  
Angela I.P. Eugenio ◽  
Rebecca S.S. Barbosa-Dantas ◽  
Gisele W.B. Colleoni
Keyword(s):  
Multiple Myeloma ◽  
Cell Death ◽  
Cell Viability ◽  
Cell Lines ◽  
Treatment Options ◽  
Compensatory Mechanism ◽  
Protein Homeostasis ◽  
Protein Response ◽  
Rpmi 8226

Introduction:Multiple myeloma (MM) cells accumulate in the bone marrow and produce enormous quantities of immunoglobulins, causing endoplasmatic reticulum stress and activation of protein handling machinery, such as heat shock protein response, autophagy and unfolded protein response (UPR).Methods:We evaluated cell lines viability after treatment with bortezomib (B) in combination with HSP70 (VER-15508) and autophagy (SBI-0206965) or UPR (STF- 083010) inhibitors.Results:For RPMI-8226, after 72 hours of treatment with B+VER+STF or B+VER+SBI, we observed 15% of viable cells, but treatment with B alone was better (90% of cell death). For U266, treatment with B+VER+STF or with B+VER+SBI for 72 hours resulted in 20% of cell viability and both treatments were better than treatment with B alone (40% of cell death). After both triplet combinations, RPMI-8226 and U266 presented the overexpression of XBP-1 UPR protein, suggesting that it is acting as a compensatory mechanism, in an attempt of the cell to handle the otherwise lethal large amount of immunoglobulin overload.Conclusion:Our in vitro results provide additional evidence that combinations of protein homeostasis inhibitors might be explored as treatment options for MM.

scholarly journals Human Neuronal Cell Lines as An In Vitro Toxicological Tool for the Evaluation of Novel Psychoactive Substances

2021 ◽  
Vol 22 (13) ◽  
pp. 6785
Author(s):  
Valeria Sogos ◽  
Paola Caria ◽  
Clara Porcedda ◽  
Rafaela Mostallino ◽  
Franca Piras ◽  
...  
Keyword(s):  
Cell Death ◽  
Neuronal Cell ◽  
In Vitro Study ◽  
Dependent Manner ◽  
Novel Psychoactive Substances ◽  
Psychoactive Substances ◽  
Concentration Dependent Manner ◽  
Mechanisms Of Toxicity ◽  
Neuronal Cell Lines

Novel psychoactive substances (NPS) are synthetic substances belonging to diverse groups, designed to mimic the effects of scheduled drugs, resulting in altered toxicity and potency. Up to now, information available on the pharmacology and toxicology of these new substances is very limited, posing a considerable challenge for prevention and treatment. The present in vitro study investigated the possible mechanisms of toxicity of two emerging NPS (i) 4′-methyl-alpha-pyrrolidinoexanophenone (3,4-MDPHP), a synthetic cathinone, and (ii) 2-chloro-4,5-methylenedioxymethamphetamine (2-Cl-4,5-MDMA), a phenethylamine. In addition, to apply our model to the class of synthetic opioids, we evaluated the toxicity of fentanyl, as a reference compound for this group of frequently abused substances. To this aim, the in vitro toxic effects of these three compounds were evaluated in dopaminergic-differentiated SH-SY5Y cells. Following 24 h of exposure, all compounds induced a loss of viability, and oxidative stress in a concentration-dependent manner. 2-Cl-4,5-MDMA activates apoptotic processes, while 3,4-MDPHP elicits cell death by necrosis. Fentanyl triggers cell death through both mechanisms. Increased expression levels of pro-apoptotic Bax and caspase 3 activity were observed following 2-Cl-4,5-MDMA and fentanyl, but not 3,4-MDPHP exposure, confirming the different modes of cell death.

scholarly journals Marginal and Internal Fit of Ceramic Prostheses Fabricated from Different Chairside CAD/CAM Systems: An In Vitro Study

2021 ◽  
Vol 11 (2) ◽  
pp. 857
Author(s):  
Keunbada Son ◽  
Kyu-Bok Lee
Keyword(s):  
Computer Aided Design ◽  
In Vitro Study ◽  
Vitro Study ◽  
Marginal Gap ◽  
Significant Difference ◽  
Cad Cam ◽  
Ceramic Crowns ◽  
Internal Fit ◽  
Cam Systems

The purpose of this in vitro study was to evaluate marginal and internal fits of ceramic crowns fabricated with chairside computer-aided design and manufacturing (CAD/CAM) systems. An experimental model based on ISO 12836:2015 was digitally scanned with different intraoral scanners (Omnicam (CEREC), EZIS PO (DDS), and CS3500 (Carestream)). Ceramic crowns were fabricated using the CAD/CAM process recommended by each system (CEREC, EZIS, and Carestream systems; N = 15). The 3-dimensional (3D) marginal and internal fit of each ceramic crown was measured using a 3D inspection software (Geomagic control X). Differences among the systems and various measurements were evaluated using the Kruskal–Wallis test. Statistically significant differences were validated using pairwise comparisons (α = 0.05). Occlusal gaps in the CEREC, EZIS, and Carestream groups were 113.0, 161.3, and 438.2 µm, respectively (p < 0.001). The axial gaps were 83.4, 78.0, and 107.9 µm, respectively. The marginal gaps were 77.8, 99.3, and 60.6 µm, respectively, and the whole gaps were 85.9, 107.3, and 214.0 µm, respectively. Significant differences were observed with the EZIS system compared with the other two systems in terms of the marginal gap sizes. The CEREC system showed no significant differences among the four measured regions. However, the EZIS and Carestream systems did show a statistically significant difference (p < 0.05). All three systems were judged to be capable of fabricating clinically acceptable prostheses, because the marginal gap, which is the most important factor in the marginal fit of prostheses, was recorded to be below 100 µm in all three systems.

scholarly journals Pyrrolizidine Alkaloids Induce Cell Death in Human HepaRG Cells in a Structure-Dependent Manner

2020 ◽  
Vol 22 (1) ◽  
pp. 202
Author(s):  
Josephin Glück ◽  
Julia Waizenegger ◽  
Albert Braeuning ◽  
Stefanie Hessel-Pras
Keyword(s):  
Cell Death ◽  
Cell Viability ◽  
Pyrrolizidine Alkaloids ◽  
Defense Mechanism ◽  
Hepatoma Cell ◽  
Hepatoma Cell Line ◽  
Dependent Manner ◽  
Human Hepatoma Cell

Pyrrolizidine alkaloids (PAs) are a group of secondary metabolites produced in various plant species as a defense mechanism against herbivores. PAs consist of a necine base, which is esterified with one or two necine acids. Humans are exposed to PAs by consumption of contaminated food. PA intoxication in humans causes acute and chronic hepatotoxicity. It is considered that enzymatic PA toxification in hepatocytes is structure-dependent. In this study, we aimed to elucidate the induction of PA-induced cell death associated with apoptosis activation. Therefore, 22 structurally different PAs were analyzed concerning the disturbance of cell viability in the metabolically competent human hepatoma cell line HepaRG. The chosen PAs represent the main necine base structures and the different esterification types. Open-chained and cyclic heliotridine- and retronecine-type diesters induced strong cytotoxic effects, while treatment of HepaRG with monoesters did not affect cell viability. For more detailed investigation of apoptosis induction, comprising caspase activation and gene expression analysis, 14 PA representatives were selected. The proapoptotic effects were in line with the potency observed in cell viability studies. In vitro data point towards a strong structure–activity relationship whose effectiveness needs to be investigated in vivo and can then be the basis for a structure-associated risk assessment.

scholarly journals In vitro study of human osteoblast proliferation and morphology on orthodontic mini-implants

2015 ◽  
Vol 85 (6) ◽  
pp. 920-926 ◽  
Author(s):  
Ricardo Carvalho Bueno ◽  
Roberta Tarkany Basting
Keyword(s):  
Chemical Composition ◽  
In Vitro Study ◽  
Chemical Components ◽  
Control Group ◽  
Osteoblast Proliferation ◽  
Mini Implants ◽  
Significant Difference ◽  
Implant Anchorage ◽  
Positive Effect

ABSTRACT Objective:  To evaluate the proliferation and morphology of human osteoblasts cultured on two brands of mini-implants after 24, 48, and 72 hours, in addition to the chemical composition found on their surface. Materials and Methods:  Two brands of mini-implant (Morelli and Neodent) were evaluated; polystyrene was used as a control group (n  =  3). Osteoblasts were cultured on the surface of sterilized mini-implants in a CO2 incubator at different time periods (24, 48, and 72 hours). Osteoblast proliferation was quantified by scanning electron microscopy using up to 5000× magnification, and cell morphology was analyzed by a single observer. For the chemical analysis, spectroscopy X-ray fluorescence was used to identify and quantify chemical components on the surface of the mini-implants. Results:  Two-way ANOVA showed no significant interaction between the factors studied (P  =  0.686). A Tukey test revealed no significant difference in osteoblast proliferation between the mini-implants at all studied periods; however, a difference in cell proliferation was detected between the Neodent and the control group (P  =  .025). For all groups, time had a direct and positive effect on osteoblast proliferation (P &lt; .001). The significant elements present in both brands of mini-implants were titanium, aluminum, vanadium, and iron. Conclusions:  Osteoblast proliferation was present on the mini-implants studied, which increased over time; however, no significant difference between brands was observed. No difference was seen between the mini-implants evaluated in terms of chemical composition. Cell adhesion after 72 hours suggests that areas of bone remodeling can be achieved, thus initiating the process of mini-implant anchorage.

8-Formylophiopogonanone B induces ROS-mediated apoptosis in nasopharyngeal carcinoma CNE-1 cells

2021 ◽  
Author(s):  
Ya-jing Zhang ◽  
Zhen-lin Mu ◽  
Ping Deng ◽  
Yi-dan Liang ◽  
Li-chuan Wu ◽  
...  
Keyword(s):  
Nasopharyngeal Carcinoma ◽  
Cell Viability ◽  
Tumor Cells ◽  
High Efficiency ◽  
Pharmacological Action ◽  
Migration And Invasion ◽  
Ros Production ◽  
Biological Functions ◽  
Intracellular Ros

Abstract Cancer is one of the leading causes of death in the world. It is very important to find drugs with high efficiency, low toxicity, and low side effects for the treatment of cancer. Flavonoids and their derivatives with broad biological functions have been recognized as anti-tumor chemicals. 8-Formylophiopogonanone B (8-FOB), a naturally existed homoisoflavonoids with rarely known biological functions, needs pharmacological evaluation. In order to explore the possible anti-tumor action of 8-FOB, we used six types of tumor cells to evaluate in vitro effects of this agent on cell viability and tested the effects on clone formation ability, scratching wound-healing, and apoptosis. In an attempt to elucidate the mechanism of pharmacological action, we examined 8-FOB-induced intracellular oxidative stress and -disrupted mitochondrial function. Results suggested that 8-FOB could suppress tumor cell viability, inhibit cell migration and invasion, induce apoptosis, and elicit intracellular ROS production. Among these six types of tumor cells, the nasopharyngeal carcinoma CNE-1 cells were the most sensitive cancer cells to 8-FOB treatment. Intracellular ROS production played a pivotal role in the anti-tumor action of 8-FOB. Our present study is the first to document that 8-FOB has anti-tumor activity in vitro and increases intracellular ROS production, which might be responsible for its anti-tumor action. The anti-tumor pharmacological effect of 8-FOB is worthy of further investigation.

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