Effects of vibration on human vocal fold extracellular matrix composition and the resultant tissue viscoelastic properties are difficult to study in vivo. Therefore, an in vitro bioreactor, simulating the in vivo physiological environment, was explored. A stress-controlled commercial rheometer was used to administer shear vibrations to living tissues at stresses and frequencies corresponding to male phonation, while simultaneously measuring tissue viscoelastic properties. Tissue environment was evaluated and adjustments made in order to sustain cell life for short term experimentation up to $6\mathrm{h}$ . Cell nutrient medium evaporation, osmolality, pH, and cell viability of cells cultured in three-dimensional synthetic scaffolds were quantified under comparably challenging environments to the rheometer bioreactor for 4 or $6\mathrm{h}$ . The functionality of the rheometer bioreactor was demonstrated by applying three vibration regimes to cell-seeded three-dimensional substrates for $2\mathrm{h}$ . Resulting strain was quantified throughout the test period. Rheologic data and cell viability are reported for each condition, and future improvements are discussed.

To mimic in vivo vibration of vocal fold cells, we studied the controllability and range of frequency, acceleration, duration, and shear stress in a new bioreactor attachment. The custom multiwell disc appliance fits into a commercially built rheometer, together termed a torsional rheometer bioreactor (TRB). Previous attachments to the TRB were capable of 50–100 Hz vibrations at relatively high strains but were limited to single-sample experiments. The TRB-multiwell disc system accommodates 20 samples in partially fluid-filled wells in an aseptic environment delivering three different acceleration conditions to different samples simultaneously. Frequency and amplitude used to calculate acceleration along with duration and shear stress were controllable and quantifiable using a combination of built-in rheometer sensors, manufacturer software, and smooth particle hydrodynamics (SPH) simulations. Computed shear stresses at the well bottom using SPH in two and three dimensions were verified with analytical approximations. Results demonstrate capabilities of the TRB-multiwell disc system that, when combined with computational modeling, provide quantifiable vibration parameters covering frequencies 0.01–250 Hz, accelerations of 0.02–300 m/s², and shear stresses of 0.01–1.4 Pa. It is well-suited for studying cell function underlying vocal fold lamina propria homeostasis, inflammation, and wound healing under differential vibration conditions.

Engineered vocal fold tissues will be grown under conditions of shear strain, the magnitude of which is determined by known strains during human vocal fold oscillation. A cup and plate system is used with the Bohlin CVO 120 rheometer to apply a shear strain of approximately 0.4 radians at frequencies of 20–60 Hz. Baseline viscoelastic measurements are recorded for the synthetic scaffolding material into which different cell lines will be injected and grown to form their own matrix. The methodology allows duration and repetitions of strain exposure to seeded scaffolding to be varied and for subsequent viscoelastic measurements to be compared. [Work supported by NIDCD.]