Healing of Tympanic Membrane Perforations: an Experimental Study

Most acute tympanic membrane perforations heal spontaneously without any residual complication, whereas a fraction transforms into a chronic perforation. The current treatment of a chronic perforation is surgical repair, which is a costly, time consuming and uncomfortable procedure particularly for the pediatric patients. The objective of this thesis is to evaluate the underlying mechanisms of the reparative process in acute and chronic perforations. This is done by monitoring the structural and mechanical properties of the tympanic membrane in various situations during healing. Furthermore, an aim is to test the efficacy of stem cells as a healing adjuvant both in acute and chronic perforations. On otomicroscopic examination the reduction of the laser perforation size was evident between day four and five after myringotomy. Histological analysis revealed thickening of the tympanic membrane at and around the myringotomy site at two and four weeks post-myringotomy in light microscopic sections. Transmission electron microscopic sections revealed a five-fold thickness increase in the lamina propria. Proliferation of fibroblasts and accumulation of large amounts of extra-cellular substances suggests an attempt of the body’s reparative mechanism to revert the damage. The lamina propria is thickened still at six months after myringotomy. The fibers of the lamina propria are loosely packed with seemingly unorganized orientation. These findings suggest that the healing process is not yet finished after half a year. A striking histological feature was the larger thickening increase in the lamina propria of the tympanic membranes that were treated with stem cells, as compared to the untreated. Growth factors or other secreted substances from the stem cells might have influenced the proliferation of fibroblasts and the production of extra-cellular substances to contribute to this accentuated thickness. An in vitro model in the rat was developed and adjusted for moiré interferometry measurements in order to assess the stiffness of normal and healed myringotomized tympanic membranes. Moiré interferometry is an optical, non-contacting technique by which the shape and pressure-produced displacement of an object can be measured. This method provides a full-field, 3-dimensional, overall stiffness change along with local stiffness variation of different portions of the tympanic membrane. The stiffness of myringotomized and healed tympanic membranes is almost normal already at two weeks after myringotomy. A huge over-production of fibers during the proliferative phase of the healing seem to limit the damage caused by the myringotomy and thus provide extra strength to this area of reorganization. Long-term measurement of the myringotomized and healed tympanic membranes showed slightly reduced strength as compared to normal. Acute perforations treated with mouse embryonic stem cells did not show any enhanced healing as compared to untreated. Teratoma, which is a known complication to embryonic stem cell treatment, was not detected in any of the ears after a follow-up time of six months. The use of human mesenchymal stem cell treatment on chronic perforations showed better healing as compared to the untreated. Thus, it appears to be of great importance to continue to study this type of stem cells in the treatment of chronic tympanic membrane perforations.