A rat excised larynx model of vocal fold scar

Abstract

Purpose: To develop and evaluate a rat excised larynx model for the measurement of acoustic, aerodynamic, and vocal fold vibratory changes resulting from vocal fold scar.

Method: Twenty-four 4-month-old male Sprague-Dawley rats were assigned to 1 of 4 experimental groups: chronic vocal fold scar, chronic vocal fold scar treated with 100-ng basic fibroblast growth factor (bFGF), chronic vocal fold scar treated with saline (sham treatment), and unscarred untreated control. Following tissue harvest, histological and immunohistochemical data were collected to confirm extracellular matrix alteration in the chronic scar group; acoustic, aerodynamic, and high-speed digital imaging data were collected using an excised larynx setup in all groups. Phonation threshold pressure (P(th)), glottal resistance (R(g)), glottal efficiency (E(g)), vibratory amplitude, and vibratory area were used as dependent variables.

Results: Chronically scarred vocal folds were characterized by elevated collagen Types I and III and reduced hyaluronic acid abundance. Phonation was achieved, and data were collected from all control and bFGF-treated larynges; however, phonation was not achieved with 3 of 6 chronically scarred and 1 of 6 saline-treated larynges. Compared with control, the chronic scar group was characterized by elevated P(th), reduced E(g), and intralarynx vibratory amplitude and area asymmetry. The bFGF group was characterized by P(th) below control-group levels, E(g) comparable with control, and vocal fold vibratory amplitude and area symmetry comparable with control. The sham group was characterized by P(th) comparable with control, E(g) superior to control, and vocal fold vibratory amplitude and area symmetry comparable with control.

Conclusions: The excised larynx model reported here demonstrated robust deterioration across phonatory indices under the scar condition and sensitivity to treatment-induced change under the bFGF condition. The improvement observed under the sham condition may reflect unanticipated therapeutic benefit or artifact. This model holds promise as a tool for the functional characterization of biomechanical tissue changes resulting from vocal fold scar and the evaluation of experimental therapies.

Figure 1

Scale photographs of a rat excised larynx. A: Superior view, relative to human fingers. B: Magnified anterior view. C: Magnified superior view.

Figure 2

Schematic of the excised larynx setup used in this experiment.

Figure 3

A: Photograph of a mounted rat excised larynx. B: HSDI of the glottis during excised larynx phonation, taken from a larynx with unilateral chronic scar.

Figure 4

Representative histological and immunohistochemical images of chronically scarred and control vocal folds. A: Alcian blue stain for HA. HA is stained blue and its presence was confirmed using a hyaluronidase digestion control (see text). B: Masson’s trichrome stain. Collagen is stained blue. C: Collagen I immunostain. Collagen I is green; cell nuclei are blue. D: Collagen III immunostain. Collagen III is red; cell nuclei are blue.

Figure 5

P_th data across experimental groups. The p-value reflects a one-way ANOVA, which was performed on natural logarithm transformed data.

Figure 6

Mean R_g and E_g data across experimental groups. The p-values reflect LDA based comparisons of the control condition against the other experimental conditions, which were performed on natural logarithm transformed data. Additional statistically significant comparisons are reported in the text.

Figure 7

Vibratory amplitude and area data from two larynges representative of the control and scar groups, collected from sequential experimental runs at P_s values below 4 kPa.

Figure 8

Vibratory amplitude and area data measured at P_th and 1.96 kPa. Data are expressed as a ratio of left versus right vocal fold, with a value of 1 representing symmetry. The p-values reflect the ANOVA fixed effect for experimental group. ANOVAs were performed on natural logarithm transformed data.