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

Mechanical properties of porcine cartilage after uniform RF heating.

Background and objectivesThermally 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 methodsThe 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.ResultsThe 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.ConclusionsAlthough 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

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

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

Mechanical properties of porcine cartilage after uniform RF heating.

Background and objectivesThermally 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 methodsThe 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.ResultsThe 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.ConclusionsAlthough 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

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

ImportanceElectromechanical 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.ObjectivesTo 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 specimensIn 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 measuresThe 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.ResultsTreated 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 relevanceElectromechanical 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 evidenceNA

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

ImportanceElectromechanical 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.ObjectivesTo 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 specimensIn 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 measuresThe 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.ResultsTreated 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 relevanceElectromechanical 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 evidenceNA

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

Background/objectivesSimilar 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 methodsThe 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.ResultsShape 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.ConclusionsLCR 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