Viability of Equine Chondrocytes After Exposure to Mepivacaine and Ropivacaine In Vitro

Silva, Gabriele Biavaschi; Côrte, Flávio Desessards De La; Brass, Karin Érica; Palma, Heloísa Einloft; Gallio, Miguel; Cantarelli, Camila; Bertolin, Kalyne; Krause, Alexandre; Wergutz, Julien; Krause, Luciana Maria Fontanari; Antoniazzi, Alfredo Q.

doi:10.1016/j.jevs.2019.02.019

Cited by 11 publications

(14 citation statements)

References 20 publications

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“…The MTT assay detects changes in cellular metabolism, namely the activity of mitochondrial reductases, whereas fluorescent labeling relies on membrane indicators of cellular viability. It is possible that early changes in cellular metabolism occur prior to changes in the plasma membrane that provide evidence of loss of viability 19 . The results of the two studies cited provide evidence that the MTT assay used in this study should be as sensitive as or more sensitive than fluorescent labeling and flow cytometry techniques when determining the in vitro chondrotoxic effects of bupivacaine 19,20 …”

Section: Discussionmentioning

confidence: 74%

“…It is possible that early changes in cellular metabolism occur prior to changes in the plasma membrane that provide evidence of loss of viability. 19 The results of the two studies cited provide evidence that the MTT assay used in this study should be as sensitive as or more sensitive than fluorescent labeling and flow cytometry techniques when determining the in vitro chondrotoxic effects of bupivacaine. 19,20 An unexpected result noted with LEB was that the short-term lack of toxicity was not replicated with longterm culture.…”

Section: Discussionmentioning

confidence: 75%

“…Fluorescent staining of viable and nonviable cells and cell counting are more direct means of determining viability compared with MTT assay. When researchers compared MTT results to fluorescent labeling and flow cytometry evaluating equine chondrocytes exposed to ropivacaine and mepivacaine for 30 minutes, 19 they found that the MTT assay detected greater loss of viability compared with fluorescent labeling and flow cytometry. This is in contrast to the results of another study 20 in which researchers found no major difference between MTT assay and flow cytometry for evaluating equine chondrocytes exposed to bupivacaine, lidocaine, and F I G U R E 2 Box plot illustrating clonogenic assay results.…”

Section: Discussionmentioning

confidence: 99%

“…Among other methods for assessing chondrocyte viability after a toxic insult, the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and clonogenic assays have been used routinely for cells grown in monolayer. [19][20][21][22] These tests were chosen for their cost and expedience. The MTT test is a metabolic test that tests cellular viability indirectly by detecting cellular metabolism of the MTT reagent to produce formazan crystals.…”

Section: Introductionmentioning

confidence: 99%

“…The objective of this study was to evaluate cell viability and cytotoxicity from low concentrations of SFB and from LEB in canine chondrocytes by using a monolayer culture system. Among other methods for assessing chondrocyte viability after a toxic insult, the 3‐(4,5‐dimethylthiazol‐2‐yl)‐2,5‐diphenyltetrazolium bromide (MTT) and clonogenic assays have been used routinely for cells grown in monolayer 19‐22 . These tests were chosen for their cost and expedience.…”

Section: Introductionmentioning

confidence: 99%

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Effect of bupivacaine concentration and formulation on canine chondrocyte viability in vitro

2021

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Objective: To quantitate bupivacaine concentration and formulation effects on chondrocyte viability in vitro. Study design: Controlled laboratory study. Sample population: Primary canine chondrocyte isolates. Methods: Cell passage 3 and 4 canine chondrocytes were exposed to 0.9% saline; canine chondrocyte growth medium; 0.4, 0.5, 0.6, 1.5, 2.5, 3.5, or 5 mg/ mL preservative-free standard formulation bupivacaine (SFB); or 13.3 or 6.65 mg/mL liposomal encapsulated bupivacaine (LEB) for 1 hour. Chondrocyte viability and clonogenicity were quantitated with 3-(4,5-dimethylthiazol-2-31 yl)-2,5-diphenyltetrazolium bromide (MTT) and clonogenic assays, respectively. Differences among concentrations and formulations were assessed with Kruskal-Wallis and Dwass-Steel-Critchlow-Fligner post hoc tests. Results: Growth medium had the highest cell viability based on MTT metabolism. Similarly, all LEB concentration groups had higher cell viability compared with SFB concentration cells treated with 3.5 or 5 mg/mL SFB (P < .03). Among SFB concentrations, cell viability was higher at 0.6 mg/mL compared with at 2.5 mg/mL or greater (P < .03). Cell clonogenicity was not significantly different between saline, culture medium, or 0.5 mg/mL SFB. Clonogenicity was lower with all tested LEB concentrations compared with saline or medium (P < .02). Conclusion: In vitro toxicity of SFB on canine chondrocytes is concentration dependent. Liposomal encapsulated bupivacaine may have time-dependent effects resulting in chondrotoxicity. Clinical significance: Clinically relevant concentrations of SFB after a single injection may not result in chondrotoxic effects in vitro. Liposomal encapsulated bupivacaine should not be used in the articular environment.

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Section: Discussionmentioning

confidence: 74%