Utility of cell viability assays for use with ex vivo vocal fold epithelial tissue

Abstract

Objective Ex vivo models are routinely used to investigate the barrier function of the vocal fold epithelium. However, there are limited reports on assays that can be used to investigate the effect of clinically-relevant challenges on vocal fold epithelial tissue viability. Our objective was to determine the utility of two assays routinely used in cell culture – a cellular metabolic activity assay and a cell membrane integrity assay - to investigate the viability of ex vivo porcine vocal fold epithelium. Study Design Prospective, ex vivo animal study Methods Porcine vocal folds were exposed to acrolein, hydrochloric acid, or hydrogen peroxide challenge. An untreated, sham challenge was included as a control. Assays including metabolic activity, cell membrane integrity, and histology were used to determine whether challenges reduced epithelial viability as compared to sham. Results Cell membrane integrity and metabolic activity assays detected reductions in viability following hydrochloric acid and hydrogen peroxide challenges but not acrolein challenge as compared to sham. No challenge produced significant changes in epithelial appearance as evidenced by light microscopy. Conclusion Metabolic activity and cell membrane integrity assays are valuable tools that can be used to evaluate the viability of ex vivo vocal fold epithelial tissue following clinically-relevant challenges. As viability is reduced, the ability of epithelial tissue to maintain its barrier function is compromised. Accurate assessment of viability may provide us clues into understanding mechanisms underlying vocal fold epithelial injury and disease.

Role of voice rest following laser resection of vocal fold lesions: A randomized controlled trial

“…A canine model has shown that voice rest precipitates re‐epithelialization offering protection to the lamina propria . Gene expression and protein levels of markers found in the breakdown of tissue such as matrix metalloproteinase‐1 also have been shown to be upregulated in a rabbit model of phonation‐induced dynamic tissue stress . These results suggest that the mechanical stress involved in speaking, especially following surgery, could alter and negatively influence postoperative wound healing.…”

“…4 Gene expression and protein levels of markers found in the breakdown of tissue such as matrix metalloproteinase-1 also have been shown to be upregulated in a rabbit model of phonation-induced dynamic tissue stress. 5,6 These results suggest that the mechanical stress involved in speaking, especially following surgery, could alter and negatively influence postoperative wound healing. Conversely, low levels of vocal loading have been shown to decrease inflammatory markers found in laryngeal secretions, as well as vocal fold fibroblasts, suggesting that active phonation could have an antiinflammatory effect and promote wound healing.…”

Acute Acrolein Exposure Induces Impairment of Vocal Fold Epithelial Barrier Function

“…Vocal fold epithelial metabolic activity was assessed using a modified MTT assay following a published protocol [ 34 ]. Epithelia samples were dissected from 7 larynges (fourteen vocal folds), punched (4 mm in diameter), and weighed.…”

Fish Scales Produce Cortisol upon Stimulation with ACTH

“…To examine tissue viability during the incubation, the Thiazolyl Blue Tetrazolium Bromide (MTT) assay was performed on whole scales as previously described for animal tissues [ 26 , 27 ], with a few modifications in order to perform the assay on a 96-well plate. In brief, after the incubation described above was completed, 10–15 mg of scales from a subset of samples were incubated for 2 h in 96-well plates in 200 ul MTT (2 mg ml −1 ; Sigma-Aldrich, M2128, Taufkirchen, Germany) dissolved in the incubation medium on a rotating platform.…”