Integrating Gelatin and Silicone to Engineer a Self-Oscillating Vocal Fold Model

Introduction: : The physiological function of the vocal folds, comprising a pair of soft connective tissues, is characterized by their self-sustained oscillations and quasi-static tension [1]. Elevated laryngeal musculature loading can increase laryngeal tension [2] and viscoelasticity [3]. Acoustically, the intricate relationships between airflow dynamics and the mechanical properties of vocal fold tissues dictate phonation. Airflow originating from the lungs traverses the trachea and larynx, passing between the vocal folds. This air propagation generates pressure differentials that induce vocal fold oscillation [4]. Voice disorders are highly prevalent, with a lifetime incidence of hoarseness estimated at approximately 30%, affecting 3% to 9% of the general population. There is a lack of reliable treatments. Bioreactor systems offer a controlled platform to simulate the phonatory environment, enabling the evaluation of mechanical stimulation's impact on vocal fold tissue [5]. These systems utilize airflow-induced self-oscillation to replicate the vibratory conditions characteristic of the human larynx, thereby facilitating the study of cellular responses and vibratory characteristics under physiologically relevant conditions [6]. Engineered bioreactor systems can be used to investigate the effects of mechanical forces on the cellular behavior and structural integrity of vocal folds [7]. This work aimed to integrate synthetic rubber and gelatin hydrogel to make a vibrating vocal fold bioreactor

Materials and

Methods: : Our design consists of a vocal fold replica system incorporating a body with a tubing network, a Plexiglass base, and a pair of synthetic silicone rubber vocal folds (Fig. 1). The model was scaled down to a quarter of its original size (μ-EPI, 1:4 ratio). The setup includes a Senor air compressor (PC 1010), a 100 PSI pressure transducer, an IP54 stainless steel pressure gauge, and an MF5700 series airflow meter. To capture dynamic supraglottal pressure, a digital microphone (SLM-25 sound level meter) was placed approximately 80 mm above the vocal folds. The microphone data was processed using software to compute the fundamental phonation frequency. Reynolds number (Re) served as the scaling criterion, and we tuned the mechanical properties by adjusting the Ecoflex-to-thinner ratio with 0%, 50%, 100%, and 150% thinner concentrations.

To simulate tissue interaction, a hydrogel element was embedded in the upper section of the model to shield cells from direct airflow exposure. The hydrogel’s volume and position (\~200 μL) were optimized. A high-speed camera was positioned above the hydrogel to capture real-time motion during oscillation. Gelatin methacryloyl (GelMA), prepared in-house, was introduced at concentrations of 3%, 5%, and 7% w/v. In the final stage, a GelMA-based hydrogel capsule (100–200 μL), embedded with human vocal fold fibroblasts, was incorporated into the model. This allowed perfusion studies and observation of tissue-like phonatory motion. The full assembly and placement are illustrated in Fig. 1.

Results, Conclusions, and Discussions:: Following the validation of our synthetic model (not shown here), we included the hydrogel part (Gelatin and GelMA) to assess the phonatory function. Figure 2 illustrates the fundamental phonatory response of our synthetic vocal folds for different material . Fig. 2A, digital kymography (DKG) reveals temporal oscillation patterns along a fixed glottal line, while glottal opening frame waveforms (GOFW) quantify time-resolved opening areas for both vocal folds [8-10]. They are supposed to show the vibration cycles at their maximum amplitude [8-10]. Vocal fold trajectories (VFT) were extracted and fitted using polynomial models to represent oscillatory paths. Figure 2B demonstrates spatial registration of the VKG line on the glottal midline, with left and right trajectories overlaid in color to highlight phase synchronization and amplitude differences. The data show the performance of the model and its relevance to in vivo phonatory functions. Figure 2C presents superior views of the glottis and key snapshots across a vibratory cycle, emphasizing mucosal wave propagation. The extracted parameters collectively enable a detailed assessment of vocal fold symmetry and phonatory function. These data confirm the validity of our model for replicating phonatory response [9-10]. In the last step, we verified the performance of our system in encapsulating cells while under air-flow oscillations (Fig. 3). The cell viability shows higher cell viability around the central area. At the same time, the edges led to more toxicity, and the cells showed lower densities at Day 7 (Fig. 3D). The metabolic activity of the cells was found to be comparable to the case of static control, confirming the effectiveness of our model in cell encapsulation.

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