During speech, vocal fold tissues must sustain the mechanical stresses required for vocalization. To do so the mechanical properties of the vocal folds must be maintained at appropriate levels. To properly regulate the mechanical properties of the vocal folds it is believed that vocal fold fibroblasts sense strains in the extracellular matrix (ECM) caused by mechanical stresses, and translate this information into an adaptive response such as an increase or decrease in ECM production. Our lab has undertaken research intended to improve our understanding of how cell behavior is affected by mechanical stimuli with the long term goal that this knowledge could be used to engineer replacement vocal fold tissue. We believe the future development of engineered vocal fold tissue could potentially benefit from cultivation within a bioreactor which closely mimics the mechanical environment of the native vocal fold.

We have previously developed a cyclic strain bioreactor and shown that cell seeded scaffolds can be conditioned with vocal-like cyclic strains to improve scaffold strength, stiffness and matrix distribution. Following on this work, we were interested in understanding what affect high frequency vibrations, a unique vocal fold stimuli, might have on cell behavior. To study the affect of vibration a bioreactor capable of conditioning cell seeded scaffolds at frequencies and amplitudes comparable to measured phonation values was developed. Using this system laryngeal fibroblasts were conditioned for 1 to 3 days. Following conditioning, matrix and matrix related gene expression, profibrotic cytokine production, and matrix protein accumulation were assessed with DNA microarray, ELISA, and immunofluorescent assays respectively. Assays revealed that the expression of several key matrix and matrix related genes, the secretion of the profibrotic cytokine TGFâ1, as well as the accumulation of fibronectin and collagen type 1 were all significantly increased over static controls.

At present the exact sequence of mechanical and biochemical events that regulate the properties of the vocal fold are unknown. However, our results seem to indicate that mechanical stimuli play an important role in regulating these properties. Our results suggest that matrix and matrix related cell behaviors which could affect in vitro tissue assembly are sensitive to both vibration and cyclic strain. Furthermore our results suggest that the response to each stimuli may be distinct, and as a result, bioreactors capable of delivering stimuli combinations could be exploited to influence in-vitro tissue assembly. We have recently developed a dual mode bioreactor capable of delivering both high frequency vibration and strain stimuli to cell seeded scaffolds. Using this latest bioreactor, extended conditioning experiments are currently under way which will examine the affect of stimuli combinations on matrix deposition and scaffold material properties. The results of this research could form the basis for future projects designed to develop engineered vocal fold tissue using a strategy that optimally integrates diverse stimuli.

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