Neutral Red Uptake, Cellular Growth and Lysosomal Function: In Vitro Effects of 24 Metals

1993 ◽  
Vol 21 (4) ◽  
pp. 501-507 ◽  
Author(s):  
Guillermo Repetto ◽  
Pilar Sanz
Keyword(s):  
Neuroblastoma Cells ◽  
Neutral Red ◽  
Cellular Growth ◽  
Metal Compounds ◽  
Lysosomal Activity ◽  
Neutral Red Assay ◽  
Neutral Red Uptake ◽  
Group B ◽  
Relative Uptake

Specific toxic interference in lysosomal activity was evaluated by the relative uptake of neutral red by lysosomes. In this adaptation of the standard neutral red cytotoxicity assay, the results are expressed relative to cell culture protein content to avoid misinterpretation due to the influence on cell proliferation of the chemicals tested. Neuro-2a mouse neuroblastoma cells were exposed in vitro to 24 metal compounds and the lysosomal activity was quantified. Five different patterns of alterations were identified. Group A (ZnCl2, NaAsO2, Na2HAsO4, CdCl2, SnCl2, HgCl2, HgCH3Cl and TlCOOCH3) produced an inhibition of the relative uptake of the dye, in marked contrast to the greater inhibition shown by the neutral red uptake assay. Group B (MgCl2, A1C13, KMnO4) NiCl2, BaCl2 and Pb[NO3]2) showed less inhibition with strong parallelism with the neutral red assay. In Group C (CrCl3, Na2Cr2O7, FeCl3, CoCl2, CuCl2 and Sn[C2H5]4), low-dose stimulation, and inhibition at higher concentrations were found. Group D (LiCl and CrCl2) stimulated uptake, and Group E (MnCl2 and Sr[NO3]2) produced no significant modifications. The relative uptake of neutral red can be a convenient tool for the study of specific toxic alterations of lysosomes.

The EYTEX™ System and the Neutral Red Uptake Inhibition Assay in Cultured Hep G2 Cells as Alternative Methods for In Vivo Eye Irritation Following the EEC Protocol

1990 ◽  
Vol 17 (4) ◽  
pp. 325-333
Author(s):  
Paul J. Dierickx ◽  
Virginia C. Gordon
Keyword(s):  
Neutral Red ◽  
Alternative Methods ◽  
Inhibition Assay ◽  
Hep G2 ◽  
Hep G2 Cells ◽  
Eye Irritation ◽  
Uptake Inhibition ◽  
In Vitro Methods ◽  
Neutral Red Uptake

The neutral red uptake inhibition assay and the EYTEX™ system were investigated as alternative methods for the assessment of eye irritation, determined according to the EEC protocol. The 17 test chemicals used were mainly organic solvents. The xenobiotics were applied to Hep G2 cells for 24 hours at different concentrations. Neutral red uptake inhibition was then measured. The results are expressed as the NI50 value, which is the concentration of test compound required to induce a 50% reduction in neutral red uptake. The same chemicals were also tested as coded samples by the EYTEX™ test according to the manufacturer's directions. A nearly identical quantitative correlation was found for both in vitro methods with corneal opacity scores: r = 0.84 for EYTEX™ scores and r = 0.83 for log NI50, expressed in μg/ml. Whilst these correlations are certainly not perfect, it is clear that both in vitro methods can be used as valuable prescreening methods.

Stromal cell and human prostatic epithelial cell in-vitro co-coltures: Growth and morphology

1996 ◽  
Vol 63 (1_suppl) ◽  
pp. 65-68
Author(s):  
S. De Angeli ◽  
A. Fandella ◽  
C. Gatto ◽  
S. Buoro ◽  
C. Favretti ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Stromal Cells ◽  
Marked Reduction ◽  
Stromal Cell ◽  
Optical Microscope ◽  
Neutral Red ◽  
Cellular Growth ◽  
Murine Fibroblasts ◽  
Prostatic Epithelial Cell

A study was carried out on the effect of stroma-epithelium interaction on cellular growth and morphology in co-coltures of U285 prostatic epithelial cells with human prostatic and esophageal stromal cells and with murine fibroblasts of the 3T3-J2 line. The proliferation rate was determined by growth tests of neutral red and kenacid blue. Morphological observations were made under optical microscope on the same cultures used for the growth tests. Results highlighted a marked reduction in cellular growth in the co-cultures compared to control cultures, as well as the tendency of the stromal and epithelial cells to re-organise themselves in pseudo-acinous structures.

Quantitative In Vitro Assessment of Phototoxicity by a Fibroblast-Neutral Red Assay

1992 ◽  
Vol 98 (5) ◽  
pp. 725-729 ◽  
Author(s):  
Richard M Lasarow ◽  
R Rivkah Isseroff ◽  
Edward C Gomez
Keyword(s):  
Neutral Red ◽  
Neutral Red Assay ◽  
In Vitro Assessment

scholarly journals Biocompatibility Evaluation of Electrospun Scaffolds of Poly (L-Lactide) with Pure and Grafted Hydroxyapatite

2017 ◽  
Vol 58 (4) ◽  
Author(s):  
José Manuel Cornejo-Bravo ◽  
Luis Jesús Villarreal-Gómez ◽  
Ricardo Vera-Graziano ◽  
María Raquel Vega-Ríos ◽  
José Luis Pineda-Camacho ◽  
...  
Keyword(s):  
Bacterial Adhesion ◽  
Neutral Red ◽  
C2c12 Cells ◽  
Surrounding Tissue ◽  
Neutral Red Assay ◽  
Electrospun Scaffolds ◽  
Muscle Tissues ◽  
Hematoxylin Eosin

<p>The objective of this work was to evaluate the biocompatibility of scaffolds of poly(<em>L</em>-lactide) with pure and grafted hydroxyapatite, at various concentrations of reinforcement. The biocompatibility tests were carried out <em>in vivo </em>in Wistar rats by implanting the material into the subcutaneous and muscle tissues from 1 to 14 weeks and evaluating the surrounding tissue stained with hematoxylin-eosin. For <em>in vitro </em>assays, MTT and neutral red assay were used to evaluate any cytotoxicity in Mioblast Muscle C2C12 Cells (ATCC® CRL-1772™) and Bovine Coronary Artery Endothelial Cells (BCAEC); <em>Escherichia coli </em>and <em>Staphylococcus aureus </em>were used to evaluate bacterial adhesion. All variants of scaffolds provoked a mild inflammatory response, without showing necrosis. No evidence of cytotoxicity was presented in cell viability tests and good bacterial cell adhesion was visualized for all of the materials studied.</p>

scholarly journals 413. Differences in Inflammatory Mechanisms in Pseudomonas aeruginosa and Staphyloccoccus auereus Infections in Cystic Fibrosis

2019 ◽  
Vol 6 (Supplement_2) ◽  
pp. S209-S209
Author(s):  
Melissa S Phuong ◽  
Rafael E Hernandez ◽  
Subash Sad
Keyword(s):  
Cystic Fibrosis ◽  
Pseudomonas Aeruginosa ◽  
Cell Death ◽  
Cytokine Production ◽  
Neutral Red ◽  
Chronic Infections ◽  
Neutral Red Assay ◽  
The One ◽  
The University

Abstract Background Chronic bacterial lung infections are the primary cause of morbidity and mortality in cystic fibrosis (CF). The most common CF pathogens, Pseudomonas aeruginosa (P. aeruginosa) or Staphylococcus aureus (S. aureus), are common commensal or environmental organisms that adapt to the CF lung. We sought to investigate whether adaptation from early lung colonizer to chronic pathogen alters the bacterial effects on host inflammation. Methods P. aeruginosa (n = 25) and S. aureus (n = 25) isolates from CF patients with early and chronic infections were acquired from Seattle Children’s CF. Environmental (n = 8) and clinical, non-CF P. aeruginosa (n = 8) isolates were obtained from the University of Ottawa. P. aeruginosa reference strain PA14 and PA14 transposon mutants for T3SS and flagellin were used to observe the relationship between cell death and cytokine production. We infected THP-1-derived macrophages (PMA differentiated) in vitro for 3 hours with various MOIs. We subsequently measured cell death of THP-1-derived macrophages using neutral red assay and cytokine production using ELISAs. Results Infections with PA14 mutants and non-CF P. aeruginosa isolates demonstrated that rapid cell death of THP-1-derived macrophages caused a reduction in cytokine production relative to strains that did not cause as much cell death. At 10 MOI, early P. aeruginosa isolates from CF patients induced more THP-1-derived macrophage cell death compared with chronic isolates (P < 0.0001). Chronic P. aeruginosa isolates induced greater production of TNF, IL-8, and IL-6 (P < 0.01, P < 0.0001, and P < 0.0001, respectively) compared with early strains. No difference in IL-1β production was observed. When controlling for cell death between the two groups by using heat-killed bacteria, the only difference maintained was in TNF production (P < 0.01). Between early and chronic S. aureus isolates, the one difference observed was greater IL-8 production among early isolates (P < 0.01). Conclusion Chronic P. aeruginosa isolates from CF patients induce less cell death but more TNF, IL-8, and IL-6 production compared with early isolates. This suggests that P. aeruginosa producing chronic infections induce inflammatory signals that may contribute to increased morbidity among CF patients. Disclosures All authors: No reported disclosures.

scholarly journals Differential Effects of the Flavonolignans Silybin, Silychristin and 2,3-Dehydrosilybin on Mesocestoides vogae Larvae (Cestoda) under Hypoxic and Aerobic In Vitro Conditions

Molecules ◽  
2018 ◽  
Vol 23 (11) ◽  
pp. 2999 ◽  
Author(s):  
Gabriela Hrčková ◽  
Terézia Kubašková ◽  
Oldřich Benada ◽  
Olga Kofroňová ◽  
Lenka Tumová ◽  
...  
Keyword(s):  
Metabolic Activity ◽  
Neutral Lipids ◽  
Neutral Red ◽  
Suitable Model ◽  
Glucose Content ◽  
Morphological Alterations ◽  
Mitochondrial Functions ◽  
Neutral Red Uptake ◽  
Mesocestoides Vogae

Mesocestoides vogae larvae represent a suitable model for evaluating the larvicidal potential of various compounds. In this study we investigated the in vitro effects of three natural flavonolignans—silybin (SB), 2,3-dehydrosilybin (DHSB) and silychristin (SCH)—on M. vogae larvae at concentrations of 5 and 50 μM under aerobic and hypoxic conditions for 72 h. With both kinds of treatment, the viability and motility of larvae remained unchanged, metabolic activity, neutral red uptake and concentrations of neutral lipids were reduced, in contrast with a significantly elevated glucose content. Incubation conditions modified the effects of individual FLs depending on their concentration. Under both sets of conditions, SB and SCH suppressed metabolic activity, the concentration of glucose, lipids and partially motility more at 50 μM, but neutral red uptake was elevated. DHSB exerted larvicidal activity and affected motility and neutral lipid concentrations differently depending on the cultivation conditions, whereas it decreased glucose concentration. DHSB at the 50 μM concentration caused irreversible morphological alterations along with damage to the microvillus surface of larvae, which was accompanied by unregulated neutral red uptake. In conclusion, SB and SCH suppressed mitochondrial functions and energy stores, inducing a physiological misbalance, whereas DHSB exhibited a direct larvicidal effect due to damage to the tegument and complete disruption of larval physiology and metabolism.

The neutral red assay can be used to evaluate cell viability during autophagy or in an acidic microenvironment in vitro

2020 ◽  
pp. 1-9
Author(s):  
Jorge G. Gomez-Gutierrez ◽  
Neal Bhutiani ◽  
Molly W. McNally ◽  
Phillip Chuong ◽  
Wenyuan Yin ◽  
...  
Keyword(s):  
Cell Viability ◽  
Neutral Red ◽  
Acidic Microenvironment ◽  
Neutral Red Assay

Aquatic Pollutants Tested in vitro with Early Passage Fish Cells

1987 ◽  
Vol 15 (2) ◽  
pp. 116-122
Author(s):  
Harvey Babich ◽  
Ellen Borenfreund
Keyword(s):  
Neutral Red ◽  
Fibroblast Cells ◽  
Bluegill Sunfish ◽  
Epithelioid Cells ◽  
Fibroblastic Cell ◽  
Early Passage ◽  
Neutral Red Assay ◽  
Aquatic Pollutants ◽  
Fish Cells

Epithelioid cells derived from fin tissues of bluegill sunfish fingerlings were used at early passage with the neutral red assay to assess the relative cytotoxicities of organochlorine pesticides. These fin cells were able to differentiate between the cytotoxicities of various test agents, although they were less sensitive than were the BF-2 cells, an established fibroblastic cell-line derived from the caudal trunk of bluegill fry. Both the fin epithelioid and the BF-2 fibroblast cells lacked significant xenobiotic metabolising capacity, based on determinations of 7-ethoxycoumarin O-deethylase as the indicator of monooxygenase activities. Such enzymatic activity could not be induced by exposure of the cultures to Arochlor 1254.

The Neutral Red Uptake Assay: Comments on the Results of the Istituto Superiore di Sanità in the EC/HO Validation Study

1998 ◽  
Vol 26 (1) ◽  
pp. 61-68
Author(s):  
Annalaura Stammati ◽  
Franco Zampaglioni ◽  
Cristiana Zanetti
Keyword(s):  
Validation Study ◽  
Rank Correlation ◽  
Neutral Red ◽  
Home Office ◽  
Uptake Assay ◽  
Neutral Red Uptake ◽  
British Home ◽  
Chemical Groups

The neutral red uptake (NRU) assay was included, among others, in a validation study sponsored by the European Commission/British Home Office (EC/HO) study, for its reliability as an in vitro alternative to the Draize eye irritancy test. The test was performed in parallel by four laboratories (Istituto Superiore di Sanità [ISS], Microbiological Associates, Hatano Research Institute and Kurabo Industries) on 60 selected chemicals. The results obtained by the ISS are reported in this paper. A poor rank correlation was obtained between the in vivo endpoint and the ISS in vitro results for the full set of chemicals and for the subsets, with the exception of surfactants, by an independent statistics group. The same unsatisfactory results were obtained by the ISS group when the rank correlation was calculated for compounds divided into chemical groups. The performance of the NRU assay, as an alternative to the Draize eye irritancy test, is discussed.

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