A new nondestructive cytometric assay based on resazurin metabolism and an organ culture model for the assessment of corneal viability

Perrot, S.; Dutertre-Catella, H.; Martin, Chantal; Warnet, Jean‐Michel; Rat, Patrice

doi:10.1002/cyto.a.10067

Cited by 29 publications

(19 citation statements)

References 39 publications

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“…For example, the cell culture model does not consider the stratified character of the conjunctival barrier, drug diffusion, conjunctival blood supply, mucin production and composition and tear fluid. Therefore, in vitro studies cannot exactly predict the properties of pharmaceuticals during in vivo use [15–16,22]. However the present study is in line to previous studies employing cell culture models for in vitro ocular toxicological studies in order to understand mechanisms of some external eye diseases [6,14–16]…”

Section: Discussionsupporting

confidence: 71%

Effect of different artificial tears against desiccation in cultured human epithelial cells

et al. 2012

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SummaryBackgroundA large number of artificial tears is widely used to treat dry eye symptoms. To test the efficacy of these drugs independent of individual parameters in vitro models are required. As described previously, we employed a reproducible in vitro cell culture system to evaluate the desiccation protection capability of some artificial tears. In the present paper data is presented of another set of pharmaceutical agents.Material/MethodsConjunctival epithelial cell line Chang 1-5c-4 (series 1) and the corneal cell line 2.040 pRSV-T (series 2) were cultured under standard conditions. Confluent cells were wetted for 20 min with artificial tears (Arufil® Uno, Arufil®, Lacrimal®, Lacophthal® sine, Siccaprotect®, Tears Again®, Vidisept® EDO, Vistil®, Wet Comod®) or PBS as a control. After exposure to a constant air flow for 0, 15, 30 and 45 minutes respectively, cells were incubated with the vital dye alamarBlue. Subsequently, absorption of the oxidised form of the dye was assessed using an ELISA-Reader.ResultsCell best survival rates in series 1 after 15 min were found for Lacrimal® (0.89), Wet Comod® (0.84) compared to PBS (0.66) and in series 2 for Vidisept® EDO (0.57) and Lacrimal® (0.56) compared to PBS (0.01). After 45 min highest survival was seen in series 1 for Lacrimal® (0.46) and Lacophthal® sine (0.36) compared to PBS (0.33) and in series 2 for Lacrimal® (−0.06) and Arufil (−0.16) compared to PBS (−0.23).ConclusionsBoth cell lines tested showed different susceptibility towards desiccation and the artificial tears showed differences in preventing cells from desiccation.

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Section: Discussionsupporting

confidence: 71%

Effect of different artificial tears against desiccation in cultured human epithelial cells

et al. 2012

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“…in 2003 [31] measured the emission spectra of a resazurin solution 40 μM and a resorufin solution 40 μM, both solutions prepared in methanol. They excited at 570 nm and obtained a maximum emission at 590 for resorufin and two different peaks for resazurin with a maximum at 630 nm.…”

Section: Discussionmentioning

confidence: 99%

A First Approach to Evaluate the Cell Dose in Highly Porous Scaffolds By Using a Nondestructive Metabolic Method

Divieto

Sassi

2015

Future Sci. OA

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Background:In cell-based therapies, in vitro studies on biomimetic cell–scaffold constructs can facilitate the determination of the cell dose, a key factor in guaranteeing the effectiveness of the treatment. However, highly porous scaffolds do not allow a nondestructive evaluation of the cell number. Our objective was to develop a nondestructive method for human mesenchymal stem cells dose evaluation in a highly porous scaffold for bone regeneration.Materials & measurement method:Proliferation trend of human mesenchymal stem cells on Biocoral® scaffolds was measured by a resazurin-based assay here optimized for 3D cultures. The method allows to noninvasively follow the cell proliferation on biocorals over 3 weeks with very high reproducibility.Conclusion:This reliable method could be a powerful tool in cell-based therapies for cell dose determination.

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“…The resazurin assay measures intrinsic cellular metabolic activity, which reduces resazurin and changes its color as a measurable indicator of the number of viable cells that are present in a test sample (34,47). Resazurin-based assays are commonly used for drug susceptibility analysis of prokaryotic (29) and eukaryotic (20,34,41,46) cells. Much like resazurin, the tetrazolium salt 2,3-bis(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide (XTT) is reduced by mitochondrial and cytoplasmic redox enzymes to a colored formazan compound with a direct correlation with cell proliferation.…”

mentioning

confidence: 99%

A Rapid, High-Throughput Viability Assay forBlastocystisspp. Reveals Metronidazole Resistance and Extensive Subtype-Dependent Variations in Drug Susceptibilities

Mirza

Teo

Upcroft

et al. 2011

Antimicrob Agents Chemother

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Blastocystis is an emerging protistan parasite of controversial pathogenesis. Although metronidazole (Mz) is standard therapy for Blastocystis infections, there have been accumulating reports of treatment failure, suggesting the existence of drug-resistant isolates. Furthermore, very little is known about Blastocystis susceptibility to standard antimicrobials. In the present study, we established resazurin and XTT viability microassays for Blastocystis spp. belonging to subtypes 4 and 7, both of which have been suggested to represent pathogenic zoonotic subtypes. The optimized resazurin assay was used to screen a total of 19 compounds against both subtypes. Interestingly, subtype 7 parasites were resistant to Mz, a 1-position-substituted 5-nitroimidazole (5-NI), while subtype 4 parasites were sensitive. Some cross-resistance was observed to tinidazole, another 1-position 5-NI. Conversely, subtype 4 parasites were resistant to emetine, while subtype 7 parasites were sensitive. Position 2 5-NIs were effective against both subtypes, as were ornidazole, nitazoxanide, furazolidone, mefloquine, quinicrine, quinine, cotrimoxazole (trimethoprim-sulfamethoxazole), and iodoacetamide. Both subtypes were resistant to chloroquine, doxycycline, paromomycin, ampicillin, and pyrimethamine. This is the first study to report extensive variations in drug sensitivities among two clinically important subtypes. Our study highlights the need to reevaluate established treatment regimens for Blastocystis infections and offers clear new treatment options for Mz treatment failures.Blastocystis is an emerging enteric protistan parasite with zoonotic potential (39,57,58). It is one of the most common parasites colonizing the human gut, with prevalences ranging between 10% of the population in developed countries and 50% in developing countries (58). It frequently infects immunocompromised individuals (27,40,59) and has a high prevalence in impoverished children (35) and HIV/AIDS (27) and cancer (59) patients. Individuals infected with Blastocystis present with common intestinal symptoms, such as abdominal pain, vomiting, and bloating, as well as mucous and watery diarrhea (58). Blastocystis infections are commonly associated with dermatological disorders (25, 67) and irritable bowel syndrome (54).Although metronidazole (Mz) treatment is considered firstline therapy for Blastocystis infections, therapeutic intervention is equivocal because of the large number of asymptomatic carriers and frequent reports of treatment failure (3,23,37,53,55). The confusion concerning the status of Blastocystis as a pathogen is primarily due to limitations of diagnostic techniques, purported subtype-dependent variations in parasite virulence, and variable host responses (55). The variation in treatment response suggests the presence of metronidazoleresistant (Mz r ) subtypes of the parasite, but there are currently no in vitro or in vivo data to support this hypothesis. Despite these controversies, interest in the parasite has increased in recent years, as sig...

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A new nondestructive cytometric assay based on resazurin metabolism and an organ culture model for the assessment of corneal viability

Cited by 29 publications

References 39 publications

Effect of different artificial tears against desiccation in cultured human epithelial cells

Effect of different artificial tears against desiccation in cultured human epithelial cells

A First Approach to Evaluate the Cell Dose in Highly Porous Scaffolds By Using a Nondestructive Metabolic Method

A Rapid, High-Throughput Viability Assay forBlastocystisspp. Reveals Metronidazole Resistance and Extensive Subtype-Dependent Variations in Drug Susceptibilities

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