Effects of electromechanical reshaping on mechanical behavior of exvivo bovine tendon.

BACKGROUND: Electromechanical reshaping is a novel, minimally invasive means to induce mechanical changes in connective tissues, and has the potential to be utilized in lieu of current orthopedic therapies that involve tendons and ligaments. Electromechanical reshaping delivers an electrical current to tissues while under mechanical deformation, causing in situ redox changes that produce reliably controlled and spatially limited mechanical and structural changes. In this study, we examine the feasibility of altering Young's modulus and inducing a shape deformation using an ex vivo bovine Achilles tendon model. METHODS: Tendon was mechanically deformed in two different modes: (1) elongation to assess for tensile modulus and (2) compression to assess for compressive modulus. Electromechanical reshaping was applied to tendon specimens via flat plate platinum electrodes (6 V, 3 min) while simultaneously under mechanical strain for 15 min. FINDINGS: In elongation mode, post-electromechanical reshaping samples demonstrated a significant decrease in Young's modulus compared to pretreatment samples (66.02 and 45.12 MPa, respectively, p < 0.0049). In compression mode, posttreatment samples illustrated a significant shape change, with an increase in diameter (10.62 to 11.36 mm, p < 0.05) and decrease in thickness (4.13 to 3.62 mm, p < 0.05). INTERPRETATION: Results demonstrated a tissue softening effect without lengthening deformation during elongation, and a shortening effect without compromising compressive stiffness during compression. Electromechanical reshaping's reliable, low-cost, and efficacious methodology in inducing mechanical and structural connective tissue modifications illustrates a potential for future alternative orthopedic applications. Future studies will optimize and refine electromechanical reshaping to address clinically relevant geometries and methods such as needle techniques.

In vivo needle-based electromechanical reshaping of pinnae: New Zealand White rabbit model.

IMPORTANCE: Electromechanical reshaping (EMR) is a low-cost, needle-based, and simple means to shape cartilage tissue without the use of scalpels, sutures, or heat that can potentially be used in an outpatient setting to perform otoplasty. OBJECTIVES: To demonstrate that EMR can alter the shape of intact pinnae in an in vivo animal model and to show that the amount of shape change and the limited cell injury are proportional to the dosimetry. DESIGN, SETTING, AND SPECIMENS: In an academic research setting, intact ears of 18 New Zealand white rabbits underwent EMR using 6 different dosimetry parameters (4 V for 5 minutes, 4 V for 4 minutes, 5 V for 3 minutes, 5 V for 4 minutes, 6 V for 2 minutes, and 6 V for 3 minutes). A custom acrylic jig with 2 rows of platinum needle electrodes was used to bend ears at the middle of the pinna and to perform EMR. Treatment was repeated twice per pinna, in proximal and distal locations. Control pinnae were not subjected to current application when being bent and perforated within the jig. Pinnae were splinted for 3 months along the region of the bend using soft silicon sheeting and a cotton bolster. MAIN OUTCOMES AND MEASURES: The ears were harvested the day after splints were removed and before euthanasia. Photographs of ears were obtained, and bend angles were measured. Tissue was sectioned for histologic examination and confocal microscopy to assess changes to microscopic structure and cellular viability. RESULTS: Treated pinnae were bent more and retained shape better than control pinnae. The mean (SD) bend angles in the 7 dosimetry groups were 55° (35°) for the control, 60° (15°) for 4 V for 4 minutes, 118° (15°) for 4 V for 5 minutes, 88° (26°) for 5 V for 3 minutes, 80° (17°) for 5 V for 4 minutes, 117° (21°) for 6 V for 2 minutes, and 125° (18°) for 6 V for 3 minutes. Shape change was proportional to electrical charge transfer, which increased with voltage and application time. Hematoxylin-eosin staining of the pinnae identified localized areas of cell injury and fibrosis in the cartilage and in the surrounding soft tissue where the needle electrodes were inserted. This circumferential zone of injury (range, 1.5-2.5 mm) corresponded to dead cells on cell viability assay, and the diameter of this region increased with total electrical charge transfer to a maximum of 2.5 mm at 6 V for 3 minutes. CONCLUSIONS AND RELEVANCE: Electromechanical reshaping produced shape change in intact pinnae of rabbits in this expanded in vivo study. A short application of 4 to 6 V can achieve adequate reshaping of the pinnae. Tissue injury around the electrodes increases with the amount of total current transferred into the tissue and is modest in spatial distribution. This study is a critical step toward evaluation of EMR in clinical trials. LEVEL OF EVIDENCE: NA.

Preliminary investigations on therapy thresholds for laser dosimetry, cryogen spray cooling duration, and treatment cycles for laser cartilage reshaping in the New Zealand white rabbit auricle.

Previous studies have demonstrated the feasibility of laser irradiation (λ = 1.45 µm) in tandem with cryogen spray cooling (CSC) to reshape rabbit auricular cartilage using a total energy density of 14 J/cm(2). The aim of this study was to further explore and identify the dosimetry parameter space for laser output energy, CSC duration, and treatment cycles required to achieve shape change while limiting skin and cartilage injury. Ten New Zealand white rabbits were treated with the 1.45 µm diode laser combined with cryogen spray cooling (Candela Smoothbeam™, Candela Co., Wayland, MA, USA). The ear's central portion was bent around a cylindrical jig and irradiated in consecutive spots of 6 mm diameter (13 or 14 J/cm(2) per spot) along three rows encompassing the bend. CSC was delivered during irradiation in cycles consisting of 25-35 ms. At thin and thick portions of the ear, 4-7 and 6-10 treatment cycles were delivered, respectively. After surgery, ears were examined and splinted for 6 weeks. Treatment parameters resulting in acceptable (grades 1 and 2) and unacceptable (grade 3) skin injuries for thick and thin regions were identified, and shape change was observed. Confocal and histological analysis of cartilage tissue revealed several outcomes correlating to laser dosimetry, CSC duration, and treatment cycles. These outcomes included expansion of cartilage layers (thickening), partial cartilage injuries, and full-thickness cartilage injuries. We determined therapy thresholds for laser output energy, cryogen spray cooling duration, and treatment cycles in the rabbit auricular model. These parameters are a starting point for future clinical procedures aimed at correcting external ear deformities.

In vivo laser cartilage reshaping with carbon dioxide spray cooling in a rabbit ear model: a pilot study.

BACKGROUND/OBJECTIVES: Similar to conventional cryogen spray cooling, carbon dioxide (CO2) spray may be used in combination with laser cartilage reshaping (LCR) to produce cartilage shape change while minimizing cutaneous thermal injury. Recent ex vivo evaluation of LCR with CO2 cooling in a rabbit model has identified a promising initial parameter space for in vivo safety and efficacy evaluation. This pilot study aimed to evaluate shape change and cutaneous injury following LCR with CO2 cooling in 5 live rabbits. STUDY DESIGN/MATERIALS AND METHODS: The midportion of live rabbit ears were irradiated with a 1.45 µm wavelength diode laser (12 J/cm(2)) with simultaneous CO2 spray cooling (85 millisecond duration, 4 alternating heating/cooling cycles per site, 5 to 6 irradiation sites per row for 3 rows per ear). Experimental and control ears (no LCR) were splinted in the flexed position for 30 days following exposure. A total of 5 ears each were allocated to the experimental and control groups. RESULTS: Shape change was observed in all irradiated ears (mean 70 ± 3°), which was statistically different from control (mean 37 ± 11°, P = 0.009). No significant thermal cutaneous injury was observed, with preservation of the full thickness of skin, microvasculature, and adnexal structures. Confocal microscopy and histology demonstrated an intact and viable chondrocyte population surrounding irradiated sites. CONCLUSIONS: LCR with CO2 spray cooling can produce clinically significant shape change in the rabbit auricle while minimizing thermal cutaneous and cartilaginous injury and frostbite. This pilot study lends support for the potential use of CO2 spray as an adjunct to existing thermal-based cartilage reshaping modalities. An in vivo systematic evaluation of optimal laser dosimetry and cooling parameters is required.

Mechanical properties of porcine cartilage after uniform RF heating.

BACKGROUND AND OBJECTIVES: Thermally mediated modalities of cartilage reshaping utilize localized heating of cartilage combined with mechanical deformation to achieve new geometries. We sought to determine the steady state elastic modulus of thermally modified cartilage without deformation, as this provides a constraint in mechanical models of the shape change process. STUDY DESIGN/MATERIALS AND METHODS: The main objective of this study was to characterize the steady state elastic modulus of porcine septal cartilage after uniform heating with radiofrequency (RF) to peak temperatures of 50 ± 5, 65 ± 5, and 85 ± 5°C. The cartilage was divided into three equally sized regions, designated as anterior, middle and posterior. Each region was then sectioned into two specimens with the proximal component serving as a paired control. RESULTS: The data confirm that there is high baseline variability in control steady state elastic moduli between animals. Also, the control values confirm a decreasing steady state elastic modulus from anterior to posterior. There is no statistical significance (P > 0.05) found between the elastic moduli of control and treated samples. CONCLUSIONS: Although shape change and retention have been fairly well characterized, little is known about the specific relation between steady state elastic modulus of cartilage and maximum treatment temperature. We determined that the difference of steady state elastic modulus between control and treated porcine septal samples was not statistically significant after uniform heating with RF to peak temperatures of 50 ± 5, 65 ± 5, and 85 ± 5°C. Ultimately, the results of this study do not pertain to the regions of heated cartilage that are shaped to hold a new form; however, it does show that the regions that are not mechanically deformed do return to the original pre-treatment elastic modulus. This is still useful information that may be used in finite element models to predict changes in internal stress distributions of cartilage after laser reshaping.

Changes in the tangent modulus of rabbit septal and auricular cartilage following electromechanical reshaping.

Transforming decades' old methodology, electromechanical reshaping (EMR) may someday replace traditionally destructive surgical techniques with a less invasive means of cartilage reshaping for reconstructive and esthetic facial surgery. Electromechanical reshaping is essentially accomplished through the application of voltage to a mechanically deformed cartilage specimen. While the capacity of the method for effective reshaping has been consistently shown, its associated effects on cartilage mechanical properties are not fully comprehended. To begin to explore the mechanical effect of EMR on cartilage, the tangent moduli of EMR-treated rabbit septal and auricular cartilage were calculated and compared to matched control values. Between the two main EMR parameters, voltage and application time, the former was varied from 2-8 V and the latter held constant at 2 min for septal cartilage, 3 min for auricular cartilage. Flat platinum electrodes were used to apply voltage, maintaining the flatness of the specimens for more precise mechanical testing through a uniaxial tension test of constant strain rate 0.01 mm/s. Above 2 V, both septal and auricular cartilage demonstrated a slight reduction in stiffness, quantified by the tangent modulus. A thermal effect was observed above 5 V, a newly identified EMR application threshold to avoid the dangers associated with thermoforming cartilage. Optimizing EMR application parameters and understanding various side effects bridge the gap between EMR laboratory research and clinical use, and the knowledge acquired through this mechanical study may be one additional support for that bridge.

Survival of chondrocytes in rabbit septal cartilage after electromechanical reshaping.

Electromechanical reshaping (EMR) has been recently described as an alternative method for reshaping facial cartilage without the need for incisions or sutures. This study focuses on determining the short- and long-term viability of chondrocytes following EMR in cartilage grafts maintained in tissue culture. Flat rabbit nasal septal cartilage specimens were bent into semi-cylindrical shapes by an aluminum jig while a constant electric voltage was applied across the concave and convex surfaces. After EMR, specimens were maintained in culture media for 64 days. Over this time period, specimens were serially biopsied and then stained with a fluorescent live-dead assay system and imaged using laser scanning confocal microscopy. In addition, the fraction of viable chondrocytes was measured, correlated with voltage, voltage application time, electric field configuration, and examined serially. The fraction of viable chondrocytes decreased with voltage and application time. High local electric field intensity and proximity to the positive electrode also focally reduced chondrocyte viability. The density of viable chondrocytes decreased over time and reached a steady state after 2-4 weeks. Viable cells were concentrated within the central region of the specimen. Approximately 20% of original chondrocytes remained viable after reshaping with optimal voltage and application time parameters and compared favorably with conventional surgical shape change techniques such as morselization.

Viability of human septal cartilage after 1.45 microm diode laser irradiation.

BACKGROUND AND OBJECTIVES: Chondrocyte viability following laser irradiation and reshaping has not been established for human nasal septal cartilage. Knowledge of the relationship between thermal injury and laser dosimetry is needed in order to optimize septal laser cartilage reshaping. The objective of this study was to determine the depth and width of thermal injury in human septal cartilage following laser irradiation. STUDY DESIGN/MATERIALS AND METHODS: Excess fresh nasal septal cartilage sections from rhinoplasty or septoplasty operations were irradiated using a 1.45 microm diode laser 1.25-3.6 W (2.8 mm spot diameter) with 1 second fixed exposure time, and then at exposure times of 1-4 seconds for a fixed power of 1.25 W. An infrared camera recorded surface temperature profiles during irradiation, and the temperature data were incorporated into a rate process model to numerically estimate thermal damage. Calcein AM and ethidium homodimer-1 fluorescent dyes combined with confocal laser microscopy (CLM) were used to measure thermal damage. RESULTS: CLM demonstrated clear demarcation between dead and living cells following irradiation. The extent of non-viable chondrocyte distributions increased with power and exposure time. The maximum depths of injury were 1,012 and 1,372 microm after 3.6 W 1 second and 1.25 W 4 seconds irradiation respectively. The damage predictions made by the rate process model underestimated thermal injury when compared with CLM measurements. CONCLUSIONS: The assay system identified regions of non-viable chondrocytes in human septal cartilage and defined how thermal injury varies with dosimetry when using a 1.45 microm diode laser.

Analysis of Nd:YAG laser-mediated thermal damage in rabbit nasal septal cartilage.

BACKGROUND AND OBJECTIVES: Laser cartilage reshaping (LCR) involves the use of photo-thermal heating to reshape cartilage. Its clinical relevance depends on the ability to minimize thermal injury in irradiated regions. The present study seeks to understand the safety of LCR by determining shape change and resultant tissue viability as a function of laser dosimetry. STUDY DESIGN/MATERIALS AND METHODS: Rabbit nasal septal cartilage were irradiated using a Nd:YAG laser (lambda = 1.32 microm, 5.4 mm spot diameter) with different exposure times of 4, 6, 8, 10, 12, and 16 seconds and powers of 4, 6, and 8 W. Temperature on the cartilage surface in the laser-irradiated region was collected using infrared thermography, this data was then used to predict tissue damage via a rate process model. A Live/Dead viability assay combined with fluorescent confocal microscopy was used to measure the amount of thermal damage generated in the irradiated specimens. RESULTS: Considerable thermal injury occurred at and below the laser-reshaping parameters that produced clinically relevant shape change using the present Nd:YAG laser. Confocal microscopy identified dead cells spanning the entire cross-sectional thickness of the cartilage specimen (about 500 microm thick) at laser power density and exposure times above 4 W and 6 seconds; damage increased with time and irradiance. The damage predictions made by the rate process model compared favorably with measured data. CONCLUSIONS: These results demonstrate that significant thermal damage is concurrent with clinically relevant shape change. This contradicts previous notions that there is a privileged laser dosimetry parameter where clinically relevant shape change and tissue viability coexist.

Shape retention in porcine and rabbit nasal septal cartilage using saline bath immersion and Nd:YAG laser irradiation.

BACKGROUND AND OBJECTIVES: The process of altering the shape of cartilage using heat has been referred to as thermoforming, and presents certain clinical benefits in reconstructive surgical procedures within the head and neck. Thermoforming allows cartilage in the upper airway and face to be reshaped without the use of classic surgical maneuvers such as carving, morselizing, or suturing. The goal of this study was to determine the dependence of cartilage shape change on both temperature and laser dosimetry using two thermoforming methods: saline bath immersion and laser irradiation. STUDY DESIGN/MATERIALS AND METHODS: Ex-vivo rabbit and porcine nasal septal cartilages were mechanically deformed and reshaped using the two thermoforming methods. With saline bath immersion using rabbit cartilage, each specimen was deformed by securing it to a small copper tube (outer diameter 8 mm) using dental bands. For porcine cartilage immersed in a saline bath, each sample was mechanically deformed between two pieces of wire mesh attached to a semicircular acrylic block. With both porcine and rabbit cartilage, the specimen and apparatus were then immersed in a hot saline bath for time intervals varying from 20 and 320 seconds and at constant temperatures between 62 and 74 degrees C. In laser reshaping, the cartilage specimens were mechanically deformed on a jig and consecutively irradiated with an Nd:YAG laser (lambda = 1.32 microm) in several spots for 6-16 seconds and irradiances of 10.2-40.7 W/cm2 per spot. After either saline bath heating or irradiation, cartilage specimens were immersed in room temperature saline for 15 minutes, then upon removal from the jig the length between the ends of each specimen was measured in order to calculate the resulting bend angle. RESULTS: The transition zone for cartilage reshaping was defined as where a significant increase in bend angle was observed between consecutive times of immersion/irradiation at the same temperature/irradiance. For the saline bath experiments, the transition zone was observed between 59-68 degrees C and 62-68 degrees C for porcine and rabbit cartilage, respectively. Similar transition zones occurred with laser irradiation below irradiances of 20.4 W/cm2 for both porcine and rabbit cartilage. In addition, the dosimetry pairs in the transition zones produce peak temperatures below the thresholds determined from the saline bath immersion studies. CONCLUSIONS: The critical transition temperature region was determined by the sharp increase in bend angle at consecutive times of immersion at the same temperature. This range was determined to be 59-68 degrees C and 62-68 degrees C for porcine and rabbit cartilage, respectively. Similar transition zones for dosimetry occurred below 20.4 W/cm2 during cartilage irradiation in both species.