Tissue engineering the human auricle by auricular chondrocyte-mesenchymal stem cell co-implantation.

Children suffering from microtia have few options for auricular reconstruction. Tissue engineering approaches attempt to replicate the complex anatomy and structure of the ear with autologous cartilage but have been limited by access to clinically accessible cell sources. Here we present a full-scale, patient-based human ear generated by implantation of human auricular chondrocytes and human mesenchymal stem cells in a 1:1 ratio. Additional disc construct surrogates were generated with 1:0, 1:1, and 0:1 combinations of auricular chondrocytes and mesenchymal stem cells. After 3 months in vivo, monocellular auricular chondrocyte discs and 1:1 disc and ear constructs displayed bundled collagen fibers in a perichondrial layer, rich proteoglycan deposition, and elastin fiber network formation similar to native human auricular cartilage, with the protein composition and mechanical stiffness of native tissue. Full ear constructs with a 1:1 cell combination maintained gross ear structure and developed a cartilaginous appearance following implantation. These studies demonstrate the successful engineering of a patient-specific human auricle using exclusively human cell sources without extensive in vitro tissue culture prior to implantation, a critical step towards the clinical application of tissue engineering for auricular reconstruction.

Anthropometric growth study of the ear in a Chinese population.

BACKGROUND: A large number of anthropometric studies of the auricle have been reported in different nations, but little data were available in the Chinese population. The aim of this study was to analyze growth changes in the ear by measuring the width and length of ears in a Chinese population. METHODS: A total of 480 participants were enrolled and classified into 1-, 3-, 5-, 7-, 9-, 12-, 14-, and 18-year groups (half were boys and half were girls in each group). Ear length, ear width, body weight, and body length were measured and recorded; ear index was calculated according to ear length and ear width. The growth of auricle and differences between genders were analyzed. Growth of ear in relation to body height and weight and the degree of emphasis on the length and width of the auricle were also analyzed. RESULTS: Ear length and width increased with age. Ear length achieved its mature size in both 14-year-old males and females. Ear width reached its mature size in males at 7 years and in females at 5 years. Different trends of ear index were shown between males and females. People in this population paid more attention to the length than the width of the auricle. CONCLUSIONS: The data indicated that ear development followed increase in age. There were gender and ethnic difference in the development of ear. These results may have potential implications for the diagnosis of congenital malformations, syndromes, and planning of ear reconstruction surgery.

Development of the head, pinnae, and acoustical cues to sound location in a precocial species, the guinea pig (Cavia porcellus).

The morphology of the head and pinna shape the spatial and frequency dependence of sound propagation that give rise to the acoustic cues to sound source location. During early development, the physical dimensions of the head and pinna increase rapidly. Thus, the binaural (interaural time and level differences, ITD and ILD) and monaural (spectral shape) cues are also hypothesized to change rapidly. Complex interactions between the size and shape of the head and pinna limit the accuracy of simple acoustical models (e.g. spherical) and necessitate empirical measurements. Here, we measured the cues to location in the developing guinea pig, a precocial species commonly used for studies of the auditory system. We measured directional transfer functions (DTFs) and the dimensions of the head and pinna in guinea pigs from birth (P0) through adulthood. Dimensions of the head and pinna increased by 87% and 48%, respectively, reaching adult values by ∼8 weeks (P56). The monaural acoustic gain produced by the head and pinna increased with frequency and age, with maximum gains at higher frequencies (>8 kHz) reaching values of 10-21 dB for all ages. The center frequency of monaural spectral notches also decreased with age, from higher frequencies (∼17 kHz) at P0 to lower frequencies (∼12 kHz) in adults. In all animals, ILDs and ITDs were dependent on both frequency and spatial location. Over development, the maximum ILD magnitude increased from ∼15 dB at P0 to ∼30 dB in adults (at frequencies >8 kHz), while the maximum low frequency ITDs increased from ∼185 µs at P0 to ∼300 µs in adults. These results demonstrate that the changes in the acoustical cues are directly related to changes in head and pinna morphology.

The importance of costal cartilage framework stabilization in microtia reconstruction: anthropometric comparison based on 216 cases.

This study explored anthropometric changes in the reconstructed auricle and the contralateral normal ear in a series of 216 microtia patients using different stabilization methods. Our main personal modifications concerning the preparation of the framework were the following: 1. The individualized framework grafting was based on patients with different ages and different degrees of the strength and thickness of the rib cartilage. 2. The framework was stabilized as a "C" shape by using a piece of cartilage or suture to reinforce the two end points of the "C". In group A (the thickness of cartilage was more than 5 mm), a block of residual cartilage fixed by wire was added between the tragus and the base frame of the inferior crus by the modified method but was not applied in the original method. In group B (the thickness of cartilage was less than 5 mm), a 4-0 braided suture was used to reinforce the two structures but was not used in the original method. No significant differences were found in the height or width measurements of the cartilage framework and the contralateral normal side in either group at the time of implantation. At the follow-up, the height and width measurements were obviously increased in both groups operated on by the original method compared with the initial implanted or contralateral normal measurements. There were no significant differences in the height or width measurements by the modified method in either group. The authors' techniques produced acceptable results and generated some useful parameters for the growth study of the reconstructed auricle and the contralateral normal ear. The modifications in framework stabilization allow a harmonious outline of the reconstructed auricle to be attained, which is almost symmetrical to the contralateral normal auricle.

Anthropometric measurements in 126 microtia reconstructions.

BACKGROUND: Due to its complex, three-dimensional morphology, auricular reconstruction remains one of the most challenging procedures in reconstructive surgery. A subject that remains controversial, however, is the question of the growth potential of the cartilaginous framework. This study explored the anthropometric changes of the reconstructed auricle and the contralateral normal ear in a series of Asian patients. METHODS: The records of 126 unilateral microtia patients in three age groups who underwent autogenous costal cartilage auricular reconstruction between 2007 and 2010 were reviewed. The average age was 14 years, and the average follow-up was 2.5 years. The auricular height was measured as the distance from the supra-auricle to the subauricle, and the width was measured as the distance from preauricle to the postauricle. Measurements of the height and width of the reconstructed auricle and the contralateral normal side were taken at implantation and at the final follow-up. Comparisons between the three age groups were performed using a paired Student t test to examine the mean auricular heights and widths for significant interval changes in size. RESULTS: The measurements showed average width increases of 1.24 mm (3.68%) and 1.35 mm (3.91%) in the reconstructed auricles of children and adolescents, respectively, but we did not find obvious changes in the adult group. No significant differences were found in the height measurement of the reconstructed auricle in the three groups. The mature size of the normal ear was reached by age 12, with slowing as patients entered adolescence and adulthood. Comparison of the reconstructed auricle and the contralateral normal ear revealed a very similar growth rate in the adult group. There were average height decreases of 0.77 mm (1.3%) and 1.3 mm (2.09%) in the reconstructed auricles of children and adolescents compared with the normal side. The results showed an average width increase of 1.13 mm (3.15%) in the adolescent group but not in the child or adult groups. CONCLUSIONS: The results of this study have generated some useful parameters for the study of growth of the reconstructed auricle and contralateral normal ear. This information serves to clarify the issue of auricular framework fabrication in terms of proper size, especially for Asian patients. Further investigation and analysis are necessary to provide further proof of the graft change.

High-fidelity tissue engineering of patient-specific auricles for reconstruction of pediatric microtia and other auricular deformities.

INTRODUCTION: Autologous techniques for the reconstruction of pediatric microtia often result in suboptimal aesthetic outcomes and morbidity at the costal cartilage donor site. We therefore sought to combine digital photogrammetry with CAD/CAM techniques to develop collagen type I hydrogel scaffolds and their respective molds that would precisely mimic the normal anatomy of the patient-specific external ear as well as recapitulate the complex biomechanical properties of native auricular elastic cartilage while avoiding the morbidity of traditional autologous reconstructions. METHODS: Three-dimensional structures of normal pediatric ears were digitized and converted to virtual solids for mold design. Image-based synthetic reconstructions of these ears were fabricated from collagen type I hydrogels. Half were seeded with bovine auricular chondrocytes. Cellular and acellular constructs were implanted subcutaneously in the dorsa of nude rats and harvested after 1 and 3 months. RESULTS: Gross inspection revealed that acellular implants had significantly decreased in size by 1 month. Cellular constructs retained their contour/projection from the animals' dorsa, even after 3 months. Post-harvest weight of cellular constructs was significantly greater than that of acellular constructs after 1 and 3 months. Safranin O-staining revealed that cellular constructs demonstrated evidence of a self-assembled perichondrial layer and copious neocartilage deposition. Verhoeff staining of 1 month cellular constructs revealed de novo elastic cartilage deposition, which was even more extensive and robust after 3 months. The equilibrium modulus and hydraulic permeability of cellular constructs were not significantly different from native bovine auricular cartilage after 3 months. CONCLUSIONS: We have developed high-fidelity, biocompatible, patient-specific tissue-engineered constructs for auricular reconstruction which largely mimic the native auricle both biomechanically and histologically, even after an extended period of implantation. This strategy holds immense potential for durable patient-specific tissue-engineered anatomically proper auricular reconstructions in the future.

Differentiation of adipose-derived stem cells into ear auricle cartilage in rabbits.

BACKGROUND: Adipose-derived stem cells have been reported as a novel candidate for the repair of cartilage injuries in vivo. METHODS: In order to assess their differentiation ability, adipose-derived stem cells isolated from rabbit fat tissue were injected into the midportion of a surgically created rabbit ear auricle cartilage defect. After several months, the auricles were resected, histopathologically assessed and compared with a control group. RESULTS: Histopathological examination of auricles removed three, four and five months after injection showed islands of new cartilage formation at the site of the surgically induced defect. Six months after injection, we observed well-formed, mature cartilaginous plates that completely filled the defect in the native cartilage. In the control group, there was no significant growth of new cartilage. CONCLUSION: The results of this study suggest the great potential of adipose-derived stem cells to repair damaged cartilage tissue in vivo.

Presurgical evaluation of retroauricular subcutaneous tissue thickness in BAHA surgery.

OBJECTIVE: A growing population of implanted patients benefit from the Bone-Anchored Hearing Aid (BAHA). An important step in the surgery is subcutaneous tissue reduction. The proper preparation of the implant site depends on relevant tissue removal, which is combined with incision, surgical technique, and preliminary retroauricular tissue thickness. STUDY DESIGN: Prospective analysis. SETTING: Authors attempt to measure the retroauricular subcutaneous tissue thickness and try to trace the correlation between retroauricular subcutaneous tissue thickness with objective parameters such as sex, height, weight, and age. PATIENTS: Group of hundred randomly selected patients were enrolled in the study. INTERVENTION(S): Standard ultrasound with liner transducer probe was applied to measure the tissue thickness. MAIN OUTCOME MEASURES: Sex, weight, height, and age were correlated with retroauricular tissue thickness; results also were calculated for body mass index. RESULTS: The retroauricular subcutaneous tissue thickness ranged between 2.0 and 11.0 mm with the mean value of 5.25 mm. Statistical validation revealed strong correlation with weight and body mass index. Statistical difference was found between retroauricular subcutaneous tissue thickness and sex, confirming women to have thinner subcutaneous tissue than men. The age of the patient also was predictive but without the linear correlation. Conducted measurements revealed no association between retroauricular subcutaneous tissue thickness and height of the patients enrolled in the study. CONCLUSION: Our results provide a predictive value during preoperative counseling of patients and planning the surgery. Basic information obtained during the examination can help choose the optimal implantation technique. The ultrasound examination applied in the study turned to be an easy and reliable method to assess the retroauricular tissue thickness before the surgery.

Concurrent development of the head and pinnae and the acoustical cues to sound location in a precocious species, the chinchilla (Chinchilla lanigera).

Sounds are filtered in a spatial- and frequency-dependent manner by the head and pinna giving rise to the acoustical cues to sound source location. These spectral and temporal transformations are dependent on the physical dimensions of the head and pinna. Therefore, the magnitudes of binaural sound location cues-the interaural time (ITD) and level (ILD) differences-are hypothesized to systematically increase while the lower frequency limit of substantial ILD production is expected to decrease due to the increase in head and pinna size during development. The frequency ranges of the monaural spectral notch cues to source elevation are also expected to decrease. This hypothesis was tested here by measuring directional transfer functions (DTFs), the directional components of head-related transfer functions, and the linear dimensions of the head and pinnae for chinchillas from birth through adulthood. Dimensions of the head and pinna increased by factors of 1.8 and 2.42, respectively, reaching adult values by ~6 weeks. From the DTFs, the ITDs, ILDs, and spectral shape cues were computed. Maximum ITDs increased by a factor of 1.75, from ~160 µs at birth (P0-1, first postnatal day) to 280 µs in adults. ILDs depended on source location and frequency exhibiting a shift in the frequency range of substantial ILD (>10 dB) from higher to lower frequencies with increasing head and pinnae size. Similar trends were observed for the spectral notch frequencies which ranged from 14.7-33.4 kHz at P0-1 to 5.3-19.1 kHz in adults. The development of the spectral notch cues, the spatial- and frequency-dependent distributions of DTF amplitude gain, acoustic directionality, maximum gain, and the acoustic axis were systematically related to the dimensions of the head and pinnae. The dimension of the head and pinnae in the chinchilla as well as the acoustical properties associated with them are mature by ~6 weeks.

Preweaning behaviors, developmental landmarks, and acrylamide and glycidamide levels after pre- and postnatal acrylamide treatment in rats.

At high levels of exposure, acrylamide monomer (AA) is a known neurotoxicant (LoPachin, 2004 [23]). The effects of lower levels of exposure, such as those experienced via a typical human diet, have not been widely investigated. Data at these levels are particularly relevant given the widespread human exposure through carbohydrate-containing foods cooked at high temperatures. Additionally, daily AA intake is estimated to be higher for infants and children. Earlier, we described behavioral alterations in preweaning rats resulting from developmental AA treatment (0.5-10.0mg/kg/day) (Garey et al., 2005 [14]). In the present study, the effects of lower doses were measured as well as serum AA and glycidimide (GA) levels in dams, fetuses, and young pups. Pregnant Fischer 344 dams (n=48-58/treatment group) were gavaged with 0.0, 0.1, 0.3, 1.0, or 5.0mg AA/kg/day beginning on gestational day 6 and ending on the day of parturition. Beginning on postnatal day 1 (PND 1) and continuing through PND 21, all pups/litter were gavaged with the same dose as their dam. There were no AA treatment effects on offspring fur development, pinnae detachment, or eye opening. Offspring body weight was somewhat decreased by 5.0mg/kg/day, particularly in males. However, righting reflex (PNDs 4-7), slant board (i.e., negative geotaxis) (PNDs 8-10), forelimb hang (PNDs 12-16), and rotarod behavior (PNDs 21-22) were not significantly altered by AA treatment. Male and female offspring of the 5.0mg/kg/day group were 30-49% less active in the open field at PNDs 19-20 (p<0.05). Serum AA levels of GD20 dams and their fetuses were comparable, indicating the ability of AA to cross the placental barrier. AA levels of pups were not affected by age (PND 1 and 22) or sex. In all rats, serum AA and GA levels exhibited a dose-response relationship. These data extend those of our previous study (Garey et al., 2005 [14]) and demonstrate that overt preweaning neurobehavioral effects are apparent in rats exposed to acrylamide pre- and postnatally, but only at the highest doses tested.

Is the large size of the pinna of the ear of the European hare (Lepus europaeus) due to its role in thermoregulation or in anterior capital shock absorption?

Large pinnae are characteristic of the Leporids, and the pinna is known to have a thermoregulatory role. Another role has been hypothesized for the pinna of Lepus spp., as a part of a suspensory system for the greater portion of the head, absorbing shock that might otherwise interfere with vision during high-speed locomotion. We compared the lengths of the pinnae of adult European hares Lepus europaeus from the source population in the cooler climate of England with those of the introduced population in the warmer climate of Australia, and we compared the lengths of the pinnae of hares that had grown in cooler weather with those that had grown in warmer weather. There were no significant differences between each of the comparisons, indicating that the size of the pinna is not determined by thermoregulatory requirements at rest. We compared the growth in length of the pinnae and the legs with growth in body mass, and growth in the mass of the pinnae with the masses of the head and the eyeballs, and found support for the suspension hypothesis. We suggest that the rapid growth of the pinna is because visual acuity is a function of absolute eye size, not relative eye size, yet juvenile hares are subject to the same predator pressure as adult hares, and equally need to maximize visual acuity while running at high speeds in dim light. We believe that the large size of the pinna is determined by its role in anterior capital suspension, not in thermoregulation.

Postnatal development of sound pressure transformations by the head and pinnae of the cat: Binaural characteristics.

There are three acoustical cues to sound location: Interaural time differences (ITDs), interaural level differences (ILDs), and monaural spectral shape cues. During development, the increasing interaural distance and pinnae size associated with a growing head and pinnae result in localization cues that change continuously until maturation is complete. Here the authors report measurements of both the physical dimensions of the head and pinnae, as well as acoustical measurements of the binaural localization cues of cats aged 1.3 weeks to adulthood. For a given source location, ILD magnitude tended to increase with both frequency and age. Moreover, the range of significant ILD production (approximately 10 dB) shifted with age from higher to lower frequencies. ITD magnitude increased with age. Partial correlation analyses revealed that increasing pinnae size accounted for approximately 31% of the variance in the development of ILDs while increasing head size accounted for virtually none. On the other hand, increases in both the head and pinnae sizes contributed to the development of the ITD cues accounting for approximately 71% and approximately 25% of the variance, respectively. ILD and ITD cues in cats reach maturity by approximately 16 and approximately 22 weeks, respectively, which match the time period over which the pinnae and head dimensions reach maturity.

Postnatal development of sound pressure transformations by the head and pinnae of the cat: monaural characteristics.

Although there have been many anatomical, physiological, and psychophysical studies of auditory development in cat, there have been no comparable studies of the development of the sound pressured transformations by the cat head and pinnae. Because the physical dimensions of the head and pinnae determine the spectral and temporal transformations of sound, as head and pinnae size increase during development, the magnitude and frequency ranges of these transformations are hypothesized to systematically change. This hypothesis was tested by measuring directional transfer functions (DTFs), the directional components of head-related transfer functions, and the linear dimensions of the head and pinnae in cats from the onset of hearing ( approximately 1.5 weeks) through adulthood. Head and pinnae dimensions increased by factors of approximately 2 and approximately 2.5, respectively, reaching adult values by approximately 23 and approximately 16 weeks, respectively. The development of the spectral notch cues to source location, the spatial- and frequency-dependent distributions of DTF amplitude gain (acoustic directionality), maximum gain, and the acoustic axis, and the resonance frequency and associated gain of the ear canal and concha were systematically related to the dimensions of the head and pinnae. These monaural acoustical properties of the head and pinnae in the cat are mature by 16 weeks.