Expression of HSP70 in Human Skin After Cryolipolysis Treatment.

BACKGROUND: Cryolipolysis nonsurgically targets and reduces subcutaneous fat through controlled cooling of skin and underlying fatty tissue. Although skin changes after cryolipolysis treatment have been observed clinically, the mechanisms by which these occur are not well understood. OBJECTIVES: The aim of this study was to investigate the expression of heat shock protein 70 (HSP70) in the epidermal and dermal layers of human skin following cryolipolysis treatment. METHODS: Subjects (N = 11; average age, 41.8 years; average BMI, 29.59 kg/m2) were recruited to receive cryolipolysis treatment with a vacuum cooling cup applicator (-11°C/35 minutes) prior to abdominoplasty surgery. Treated and untreated abdominal tissue samples were harvested immediately after surgery (average follow-up, 15 days; range, 3 days to 5 weeks). Immunohistochemistry for HSP70 was performed on all samples. Slides were digitized and quantified in epidermal and dermal layers. RESULTS: There was higher epidermal and dermal HSP70 expression in cryolipolysis-treated pre-abdominoplasty samples vs untreated samples. There was a 1.32-fold increase of HSP70 expression in the epidermis (P < .05) and a 1.92-fold increase in the dermis (P < .04) compared with untreated samples. CONCLUSIONS: We found significant induction of HSP70 after cryolipolysis treatment in epidermal and dermal layers. HSP70 has potential therapeutic benefits and is recognized to have a role in skin protection and adaption after thermal stress. Although cryolipolysis is popular for subcutaneous fat reduction, cryolipolytic HSP induction in the skin may prove valuable for additional therapeutic applications, including skin wound healing, remodeling, rejuvenation, and photoprotection.

Molecular and Histological Evidence Detailing Clinically Observed Skin Improvement Following Cryolipolysis.

BACKGROUND: In addition to body contouring, there is anecdotal and clinical evidence of reduced laxity caused by skin tightening after cryolipolysis. However, it has not been established how cryolipolysis triggers dermal changes. OBJECTIVES: The aim of this study was to investigate the fundamental mechanisms behind clinically observed dermal changes by molecular and immunohistochemistry (IHC) analytical methods. METHODS: This feasibility study involved 7 subjects who received cryolipolysis treatment. Tissue samples were harvested from 3 days to 5 weeks after treatment. RNA-sequencing examined differential gene expression of major collagens. RNA in situ hybridization (RNA-ISH) investigated the distribution of 1 of the gene markers for collagen type I (COL1A1). IHC for procollagen type I, heat shock protein 47 (HSP47), transforming growth factor ß (TGF-ß), and tropoelastin was performed and quantified. RESULTS: Gene expression analysis highlighted a gradual upregulation of collagen mRNA genes. RNA-ISH confirmed upregulation of COL1A1 mRNA and showed a homogeneous distribution through the dermis. IHC showed increases in protein expression. Quantification revealed a 3.62-fold increase of procollagen type I (P < 0.0071), a 2.91-fold increase of TGF-ß (P < 0.041), a 1.54-fold increase of HSP47 (P < 0.007), and a 1.57-fold increase of tropoelastin (P < 0.39) compared with untreated areas. CONCLUSIONS: This study revealed significant induction of molecular and protein markers of type I collagen, which supports neocollagenesis and may play an essential role in clinically relevant skin improvement. A dermal remodeling process driven by increased TGF-ß and higher expression of HSP47 was observed. Overall, these data provide the first evidence of dermal remodeling and clarify the mechanism by which cryolipolysis may induce skin improvement.

Clinical Study Demonstrates that Electromagnetic Muscle Stimulation Does Not Cause Injury to Fat Cells.

BACKGROUND AND OBJECTIVES: A previous pre-clinical study on electromagnetic muscle stimulation (EMMS) suggested that fat cell apoptosis occurs following treatment in a porcine model. While EMMS can induce changes in muscle, the effect on fat tissue is not established. This clinical study sought to assess adipose tissue response to EMMS in comparison to cryolipolysis treatment. STUDY DESIGN/MATERIALS AND METHODS: Study subjects were recruited prior to abdominoplasty to receive body contouring treatments and subsequently to obtain tissue for histological analysis. Non-invasive abdominal treatments were delivered using a commercially available (n = 6) or prototype (n = 3) EMMS system or a cryolipolysis system (n = 2). Subjects received a single EMMS treatment (100% intensity for 30 minutes) or a single cryolipolysis treatment (-11°C for 35 minutes) to the abdomen. Superficial and deep (i.e., adjacent to muscle layer) subcutaneous adipose tissue was harvested at set timepoints post-treatment. The presence or absence of an inflammatory response was evaluated using standard hematoxylin and eosin (H&E) staining. As adipocytes that are destined to become apoptotic cannot be distinguished by traditional H&E staining during the early phases of injury, irreversible fat cell injury was assessed using perilipin immunofluorescence. RESULTS: Following H&E histological analysis at 3, 10, 11, and 17 days post-treatment, no EMMS-treated samples showed an inflammatory response in either the superficial or deep subcutaneous adipose tissue. For the cryolipolysis-treated adipose tissue, however, the H&E staining revealed a marked inflammatory response with an influx of neutrophils, lymphocytes, and macrophages at timepoints consistent with previous histological studies. Further, loss of perilipin staining provided clear visual evidence of irreversible fat cell injury in the cryolipolysis-treated adipose tissue. In contrast, the electromagnetic muscle stimulation-treated samples showed persistence of perilipin staining of adipose tissue indicating that all fat cells were viable. CONCLUSION: This study failed to demonstrate either fat cell injury or inflammatory response following EMMS treatment. While electromagnetic muscle stimulation may non-invasively induce muscle changes, this clinical study found no evidence of an impact injurious or otherwise on subcutaneous fat. © 2020 The Authors. Lasers in Surgery and Medicine published by Wiley Periodicals LLC.

Thermal and elastic response of subcutaneous tissue with different fibrous septa architectures to RF heating: numerical study.

BACKGROUND AND OBJECTIVE: Radiofrequency currents are commonly used in dermatology to treat cutaneous and subcutaneous tissues by heating. The subcutaneous morphology of tissue consists of a fine, collagenous and fibrous septa network enveloping clusters of adipocyte cells. The architecture of this network, namely density and orientation of septa, varies among patients and, furthermore, it correlates with cellulite grading. In this work we study the effect of two clinically relevant fibrous septa architectures on the thermal and elastic response of subcutaneous tissue to the same RF treatment; in particular, we evaluate the thermal damage and thermal stress induced to an intermediate- and a high-density fibrous septa network architecture that correspond to clinical morphologies of 2.5 and 0 cellulite grading, respectively. STUDY DESIGN/MATERIALS AND METHODS: We used the finite element method to assess the electric, thermal and elastic response of a two-dimensional model of skin, subcutaneous tissue and muscle subjected to a relatively long, constant, low-power RF treatment. The subcutaneous tissue is constituted by an interconnected architecture of fibrous septa and fat lobules obtained by processing micro-MRI sagittal images of hypodermis. As comparison criteria for the RF treatment of the two septa architectures, we calculated the accumulated thermal damage that corresponds to 63% loss in cell viability. RESULTS: Electric currents preferentially circulated through the fibrous septa in the subcutaneous tissue. However, the intensity of the electric field was higher within the fat because it is a poor electric conductor. The power absorption in the fibrous septa relative to that in the fat varied with septum orientation: it was higher in septa with vertical orientation and lower in septa with horizontal orientation. Overall, maximum values of electric field intensity, power absorption and temperature were similar for both fibrous septa architectures. However, the high-density septa architecture (cellulite grade 0) had a more uniform and broader spatial distribution of power absorption, resulting in a larger cross-sectional area of thermal damage (≈1.5 times more). Volumetric strains (expansion and contraction) were small and similar for both network architectures. During the first seconds of RF exposure, the fibrous septa were subjected to thermal expansion regardless of orientation. In the long term, the fibrous septa contracted due to the thermal expansion of fat. Skin and muscle were subjected to significantly higher Von Mises stresses (measure of yield) or distortion energy than the subcutaneous tissue. CONCLUSION: The distribution of electric currents within subcutaneous tissues depends on tissue morphology. The electric field is more intense in septum oriented along the skin to muscle (top to bottom) direction, creating lines or planes of preferential heating. It follows that the more septum available for preferential heating, the larger the extent of volumetric RF-heating and thermal damage to the subcutaneous tissue. Thermal load alone, imposed by long-exposure to heating up to 50 °C, results in small volumetric expansion and contraction in the subcutaneous tissue. The subcutaneous tissue is significantly less prone to non-reversible deformation by a thermal load than the skin and muscle.

Effect of fibrous septa in radiofrequency heating of cutaneous and subcutaneous tissues: computational study.

BACKGROUND AND OBJECTIVES: Radiofrequency (RF) energy exposure is a popular non-invasive method for generating heat within cutaneous and subcutaneous tissues. Subcutaneous fat consists of fine collagen fibrous septa meshed with clusters of adipocytes having distinct structural, electrical and thermal properties that affect the distribution and deposition of RF energy. The objectives of this work are to (i) determine the electric and thermal effects of the fibrous septa in the RF heating; (ii) investigate the RF heating of individual fat lobules enclosed by fibrous septa; and, (iii) discuss the clinical implications. METHODS AND RESULTS: We used the finite element method to model the two-dimensional, time-dependent, electro-thermal response of a three-layer tissue (skin, subcutaneous fat, and muscle). We considered two different configurations of subcutaneous fat tissue: a homogenous layer of fat only and a honeycomb-like layer of fat with septa. Architecture of the fibrous septa was anatomically accurate, constructed from sagittal images from human micro-MRI. For a large electrode applied to the skin surface, results show that the absorbed electric power density is greater in some septa than in the surrounding fat lobules, favoring the flux of electric current density. Fibers aligned parallel to the electric field have higher electric flux and, consequently, absorb more power. Heat transfer from the septa occurs over time during and after RF energy delivery. There is a greater temperature rise in fat with fibrous septa. CONCLUSIONS: The presence of septa affects the local distribution of the static electric field, facilitates the flux of electric current and enhances the bulk electric power absorption of the subcutaneous fat layer. Fibrous septa aligned with the local electric field have higher absorbed power density than septa oriented perpendicular to the electric field. Individual fat lobules gain heat instantly by local power absorption and, eventually, by diffusion from the surrounding septa.

Fractional Skin Harvesting: Autologous Skin Grafting without Donor-site Morbidity.

BACKGROUND: Conventional autologous skin grafts are associated with significant donor-site morbidity. This study was conducted to determine feasibility, safety, and efficacy of a new strategy for skin grafting based on harvesting small columns of full-thickness skin with minimal donor-site morbidity. METHODS: The swine model was used for this study. Hundreds of full-thickness columns of skin tissue (~700 µm diameter) were harvested using a custom-made harvesting device, and then applied directly to excisional skin wounds. Healing in donor and graft sites was evaluated over 3 months by digital photographic measurement of wound size and blinded, computer-aided evaluation of histological features and compared with control wounds that healed by secondary intention or with conventional split-thickness skin grafts (STSG). RESULTS: After harvesting hundreds of skin columns, the donor sites healed rapidly without scarring. These sites reepithelialized within days and were grossly and histologically indistinguishable from normal skin within 7 weeks. By contrast, STSG donor sites required 2 weeks for reepithelialization and retained scar-like characteristics in epidermal and dermal architecture throughout the experiment. Wounds grafted with skin columns resulted in accelerated reepithelialization compared with ungrafted wounds while avoiding the "fish-net" patterning caused by STSG. CONCLUSION: Full-thickness columns of skin can be harvested in large quantities with negligible long-term donor-site morbidity, and these columns can be applied directly to skin wounds to enhance wound healing.

Selective and localized radiofrequency heating of skin and fat by controlling surface distributions of the applied voltage: analytical study.

At low frequencies (hundreds of kHz to a few MHz), local energy absorption is proportional to the conductivity of tissue and the intensity of the internal electric field. At 1 MHz, the electric conductivity ratio between skin and fat is approximately 10; hence, skin would heat more provided the intensity of the electric field is similar in both tissues. It follows that selective and localized heat deposition is only feasible by varying electric fields locally. In this study, we vary local intensities of the internal electric field in skin, fat and muscle by altering its direction through modifying surface distributions of the applied voltage. In addition, we assess the long-term effects of these variations on tissue thermal transport. To this end, analytical solutions of the electric and bioheat equations were obtained using a regular perturbation method. For voltage distributions given by second- and eight-degree functions, the power absorption in fat is much greater than in skin by the electrode center while the opposite is true by the electrode edge. For a sinusoidal function, the absorption in fat varies laterally from greater to lower than in skin, and then this trend repeats from the center to the edge of the electrode. Consequently, zones of thermal confinement selectively develop in the fat layer. Generalizing these functions by parametrization, it is shown that radiofrequency (RF) heating of layered tissues can be selective and precisely localized by controlling the spatial decay, extent and repetition of the surface distribution of the applied voltage. The clinical relevance of our study is to provide a simple, non-invasive method to spatially control the heat deposition in layered tissues. By knowing and controlling the internal electric field, different therapeutic strategies can be developed and implemented.

Analytical solution of the Pennes equation for burn-depth determination from infrared thermographs.

A serious problem in emergency medicine is the correct evaluation of skin burn depth to make the appropriate choice of treatment. In clinical practice, there is no difficulty in classifying first- and third-degree burns correctly. However, differentiation between the IIa (superficial dermal) and IIb (deep dermal) wounds is problematic even for experienced practitioners. In this work, the use of surface skin temperature for the determination of the depth of second-degree burns is explored. An analytical solution of the 3D Pennes steady-state equation is obtained assuming that the ratio between burn depth and the burn size is small. The inverse problem is posed in a search space consisting of geometrical parameters associated with the burned region. This space is searched to minimize the error between the analytical and experimental skin surface temperatures. The technique is greatly improved by using local one-dimensionality to provide the shape of the burned region. The feasibility of using this technique and thermography to determine skin burn depth is discussed.

Particle motion in unsteady two-dimensional peristaltic flow with application to the ureter.

Particle motion in an unsteady peristaltic fluid flow is analyzed. The fluid is incompressible and Newtonian in a two-dimensional planar geometry. A perturbation method based on a small ratio of wave height to wavelength is used to obtain a closed-form solution for the fluid velocity field. This analytical solution is used in conjunction with an equation of motion for a small rigid sphere in nonuniform flow taking Stokes drag, virtual mass, Faxén, Basset, and gravity forces into account. Fluid streamlines and velocity profiles are calculated. Theoretical values for pumping rates are compared with available experimental data. An application to ureteral peristaltic flow is considered since fluid flow in the ureter is sometimes accompanied by particles such as stones or bacteriuria. Particle trajectories for parameters that correspond to calcium oxalates for calculosis and Escherichia coli type for bacteria are analyzed. The findings show that retrograde or reflux motion of the particles is possible and bacterial transport can occur in the upper urinary tract when there is a partial occlusion of the wave. Dilute particle mixing is also investigated, and it is found that some of the particles participate in the formation of a recirculating bolus, and some of them are delayed in transit and eventually reach the walls. This can explain the failure of clearing residuals from the upper urinary tract calculi after successful extracorporeal shock wave lithotripsy. The results may also be relevant to the transport of other physiological fluids and industrial applications in which peristaltic pumping is used.