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Objective: Shilajit is a pale-brown to blackish-brown organic mineral substance available from Himalayan rocks. We demonstrated that in type I obese humans, shilajit supplementation significantly upregulated extracellular matrix (ECM)-related genes in the skeletal muscle. Such an effect was highly synergistic with exercise. The present study (clinicaltrials.gov NCT02762032) aimed to evaluate the effects of shilajit supplementation on skin gene expression profile and microperfusion in healthy adult females. Methods: The study design comprised six total study visits including a baseline visit (V1) and a final 14-week visit (V6) following oral shilajit supplementation (125 or 250 mg bid). A skin biopsy of the left inner upper arm of each subject was collected at visit 2 and visit 6 for gene expression profiling using Affymetrix Clariom™ D Assay. Skin perfusion was determined by MATLAB processing of dermascopic images. Transcriptome data were normalized and subjected to statistical analysis. The differentially regulated genes were subjected to Ingenuity Pathway Analysis (IPA®). The expression of the differentially regulated genes identified by IPA® were verified using real-time polymerase chain reaction (RT-PCR). Results: Supplementation with shilajit for 14 weeks was not associated with any reported adverse effect within this period. At a higher dose (250 mg bid), shilajit improved skin perfusion when compared to baseline or the placebo. Pathway analysis identified shilajit-inducible genes relevant to endothelial cell migration, growth of blood vessels, and ECM which were validated by quantitative real-time polymerase chain reaction (RT-PCR) analysis. Conclusions: This work provides maiden evidence demonstrating that oral shilajit supplementation in adult healthy women induced genes relevant to endothelial cell migration and growth of blood vessels. Shilajit supplementation improved skin microperfusion.
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Matriz Extracelular/efectos de los fármacos , Microvasos/efectos de los fármacos , Minerales , Resinas de Plantas , Piel , Transcriptoma/efectos de los fármacos , Administración Oral , Adulto , Matriz Extracelular/metabolismo , Femenino , Humanos , Minerales/administración & dosificación , Minerales/farmacología , Resinas de Plantas/administración & dosificación , Resinas de Plantas/farmacología , Piel/irrigación sanguínea , Piel/efectos de los fármacosRESUMEN
Hyperglycemia (HG) induces genome-wide cytosine demethylation. Our previous work recognized miR-200b as a critical angiomiR, which must be transiently downregulated to initiate wound angiogenesis. Under HG, miR-200b downregulation is not responsive to injury. Here, we demonstrate that HG may drive vasculopathy by epigenetic modification of a miR promoter. In human microvascular endothelial cells (HMECs), HG also lowered DNA methyltransferases (DNMT-1 and DNMT-3A) and compromised endothelial function as manifested by diminished endothelial nitric oxide (eNOS), lowered LDL uptake, impaired Matrigel tube formation, lower NO production, and compromised VE-cadherin expression. Bisulfite-sequencing documented HG-induced miR-200b promoter hypomethylation in HMECs and diabetic wound-site endothelial cells. In HMECs, HG compromised endothelial function. Methyl donor S-adenosyl-L-methionine (SAM) corrected miR-200b promoter hypomethylaton and rescued endothelial function. In vivo, wound-site administration of SAM to diabetic mice improved wound perfusion by limiting the pathogenic rise of miR-200b. Quantitative stable isotope labeling by amino acids in cell culture (SILAC) proteomics and ingenuity pathway analysis identified HG-induced proteins and principal clusters in HMECs sensitive to the genetic inhibition of miR-200b. This work presents the first evidence of the miR-200b promoter methylation as a critical determinant of diabetic wound angiogenesis.
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Angiopatías Diabéticas/genética , Epigénesis Genética , MicroARNs/genética , Animales , Línea Celular , Metilación de ADN , ADN Metiltransferasa 3A , Diabetes Mellitus Experimental , Angiopatías Diabéticas/metabolismo , Angiopatías Diabéticas/patología , Modelos Animales de Enfermedad , Células Endoteliales/metabolismo , Epigénesis Genética/efectos de los fármacos , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Hiperglucemia/genética , Ratones , Ratones Transgénicos , Neovascularización Patológica/genética , Neovascularización Patológica/metabolismo , Regiones Promotoras Genéticas , Selenometionina/análogos & derivados , Selenometionina/farmacologíaRESUMEN
Unlike the epidermis, which regenerates continually, hair follicles anchored in the subcutis periodically regenerate by spontaneous repetitive cycles of growth (anagen), degeneration (catagen), and rest (telogen). The loss of hair follicles in response to injuries or pathologies such as alopecia endangers certain inherent functions of the skin. Thus, it is of interest to understand mechanisms underlying follicular regeneration in adults. In this work, a phytochemical rich in the natural vitamin E tocotrienol (TRF) served as a productive tool to unveil a novel epidermal pathway of hair follicular regeneration. Topical TRF application markedly induced epidermal hair follicle development akin to that during fetal skin development. This was observed in the skin of healthy as well as diabetic mice, which are known to be resistant to anagen hair cycling. TRF suppressed epidermal E-cadherin followed by 4-fold induction of ß-catenin and its nuclear translocation. Nuclear ß-catenin interacted with Tcf3. Such sequestration of Tcf3 from its otherwise known function to repress pluripotent factors induced the plasticity factors Oct4, Sox9, Klf4, c-Myc, and Nanog. Pharmacological inhibition of ß-catenin arrested anagen hair cycling by TRF. This work reports epidermal E-cadherin/ß-catenin as a novel pathway capable of inducing developmental folliculogenesis in the adult skin.
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Cadherinas/genética , Folículo Piloso/efectos de los fármacos , Fitoquímicos/farmacología , Regeneración/efectos de los fármacos , Tocotrienoles/farmacología , beta Catenina/genética , Animales , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Cadherinas/antagonistas & inhibidores , Cadherinas/metabolismo , Núcleo Celular/efectos de los fármacos , Núcleo Celular/metabolismo , Regulación de la Expresión Génica , Folículo Piloso/crecimiento & desarrollo , Folículo Piloso/metabolismo , Factor 4 Similar a Kruppel , Factores de Transcripción de Tipo Kruppel/genética , Factores de Transcripción de Tipo Kruppel/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Proteína Homeótica Nanog/genética , Proteína Homeótica Nanog/metabolismo , Factor 3 de Transcripción de Unión a Octámeros/genética , Factor 3 de Transcripción de Unión a Octámeros/metabolismo , Transporte de Proteínas/efectos de los fármacos , Proteínas Proto-Oncogénicas c-myc/genética , Proteínas Proto-Oncogénicas c-myc/metabolismo , Regeneración/genética , Factor de Transcripción SOX9/genética , Factor de Transcripción SOX9/metabolismo , Transducción de Señal , beta Catenina/agonistas , beta Catenina/metabolismoRESUMEN
Peripheral vasculopathies cause severe wound hypoxia inducing the hypoxamiR miR-210. High level of miR-210, persisting in wound-edge tissue as ischemic memory, suppresses oxidative metabolism and inhibits cell proliferation necessary for healing. In wound-edge tissue of chronic wound patients, elevated miR-210 was tightly associated with inhibition of epidermal cell proliferation as evident by lowered Ki67 immunoreactivity. To inhibit miR-210 in murine ischemic wound-edge tissue, we report the formulation of antihypoxamiR functionalized gramicidin lipid nanoparticles (AFGLN). A single intradermal delivery of AFGLN encapsulating LNA-conjugated anti-hypoximiR-210 (AFGLNmiR-210) lowered miR-210 level in the ischemic wound-edge tissue. In repTOP™mitoIRE mice, AFGLNmiR-210 rescued keratinocyte proliferation as visualized by in vivo imaging system (IVIS). 31P NMR studies showed elevated ATP content at the ischemic wound-edge tissue following AFGLNmiR-210 treatment indicating recovering bioenergetics necessary for healing. Consistently, AFGLNmiR-210 improved ischemic wound closure. The nanoparticle based approach reported herein is effective for miR-directed wound therapeutics warranting further translational development.
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Antibacterianos/administración & dosificación , Gramicidina/administración & dosificación , Nanopartículas , Cicatrización de Heridas , Animales , Humanos , Isquemia/metabolismo , Queratinocitos , Lípidos , Ratones , MicroARNsRESUMEN
This work sought to develop a robust and clinically relevant swine model of critical limb ischemia (CLI) involving the onset of ischemic muscle necrosis. CLI carries about 25-40% risk of major amputation with 20% annual mortality. Currently, there is no specific treatment that targets the ischemic myopathy characteristic of CLI. Current swine models of CLI, with tolerable side-effects, fail to achieve sustained ischemia followed by a necrotic myopathic endpoint. Such limitation in experimental model hinders development of effective interventions. CLI was induced unilaterally by ligation-excision of one inch of the common femoral artery (CFA) via infra-inguinal minimal incision in female Yorkshire pigs (n = 5). X-ray arteriography was done pre- and post-CFA transection to validate successful induction of severe ischemia. Weekly assessment of the sequalae of ischemia on limb perfusion, and degree of ischemic myopathy was conducted for 1 month using X-ray arteriography, laser speckle imaging, CTA angiography, femoral artery duplex, high resolution ultrasound and histopathological analysis. The non-invasive tissue analysis of the elastography images showed specific and characteristic pattern of increased muscle stiffness indicative of the fibrotic and necrotic outcome expected with associated total muscle ischemia. The prominent onset of skeletal muscle necrosis was evident upon direct inspection of the affected tissues. Ischemic myopathic changes associated with inflammatory infiltrates and deficient blood vessels were objectively validated. A translational model of severe hindlimb ischemia causing ischemic myopathy was successfully established adopting an approach that enables long-term survival studies in compliance with regulatory requirements pertaining to animal welfare.
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Enfermedades Musculares , Rabdomiólisis , Porcinos , Femenino , Animales , Isquemia Crónica que Amenaza las Extremidades , Miembro Posterior/irrigación sanguínea , Rabdomiólisis/complicaciones , Enfermedades Musculares/patología , Isquemia/patología , Necrosis/patología , Músculo Esquelético/patología , Modelos Animales de EnfermedadRESUMEN
The α-tocotrienol (TCT) form of natural vitamin E is more potent than the better known α-tocopherol against stroke. Angiographic studies of canine stroke have revealed beneficial cerebrovascular effects of TCT. This work seeks to understand the molecular basis of such effect. In mice, TCT supplementation improved perfusion at the stroke-affected site by inducing miR-1224. miRNA profiling of a laser-capture-microdissected stroke-affected brain site identified miR-1224 as the only vascular miR induced. Lentiviral knockdown of miR-1224 significantly blunted the otherwise beneficial effects of TCT on stroke outcomes. Studies on primary brain microvascular endothelial cells revealed direct angiogenic properties of miR-1224. In mice not treated with TCT, advance stereotaxic delivery of an miR-1224 mimic to the stroke site markedly improved stroke outcomes. Mechanistic studies identified Serpine1 as a target of miR-1224. Downregulation of Serpine1 augmented the angiogenic response of the miR-1224 mimic in the brain endothelial cells. The inhibition of Serpine1, by dietary TCT and pharmacologically, increased cerebrovascular blood flow at the stroke-affected site and protected against stroke. This work assigns Serpine1, otherwise known to be of critical significance in stroke, a cerebrovascular function that worsens stroke outcomes. miR-1224-dependent inhibition of Serpine1 can be achieved by dietary TCT as well as by the small-molecule inhibitor TM5441.
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Sweating and heat buildup at the skin-liner interface is a major challenge for persons with limb loss. Liners made of heat-non-conducting materials may cause sweating of the residual limb and may result in liners slipping off the skin surface especially on a warm day or during high activity, causing skin breakdown and affecting limb health. To address this, we evaluated the efficacy of the vented liner-socket system (VS, Össur) compared to Seal-In silicone liner and non-vented socket (nVS, Össur) in reducing relative humidity (RH) during increased sweat. Nine individuals with limb loss using nVS were randomized to VS or nVS and asked for activity in a 20-min treadmill walk. RH was significantly attenuated (p = 0.0002) and perceived sweating, as reported by prosthesis users, improved (p = 0.028) with VS, patient-reported comprehensive lower limb amputee socket survey (CLASS) outcomes to determine the suspension, stability, and comfort were not significantly different between VS and nVS. There are limited rigorous scientific studies that clearly provide evidence-based guidelines to the prosthetist in the selection of liners from numerous available options. The present study is innovative in clearly establishing objective measures for assessing humidity and temperatures at the skin-liner interface while performing activity. As shown by the measured data and perceived sweat scores provided by the subjects based on their daily experience, this study provided clear evidence establishing relative humidity at the skin-liner interface is reduced with the use of a vented liner-socket system when compared to a similar non-vented system.
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Amputados , Miembros Artificiales , Humanos , Muñones de Amputación , Tibia , Amputación Quirúrgica , Extremidad Inferior/cirugía , Diseño de PrótesisRESUMEN
INTRODUCTION: Between 5% and 20% of all combat-related casualties are attributed to burn wounds. A decrease in the mortality rate of burns by about 36% can be achieved with early treatment, but this is contingent upon accurate characterization of the burn. Precise burn injury classification is recognized as a crucial aspect of the medical artificial intelligence (AI) field. An autonomous AI system designed to analyze multiple characteristics of burns using modalities including ultrasound and RGB images is described. MATERIALS AND METHODS: A two-part dataset is created for the training and validation of the AI: in vivo B-mode ultrasound scans collected from porcine subjects (10,085 frames), and RGB images manually collected from web sources (338 images). The framework in use leverages an explanation system to corroborate and integrate burn expert's knowledge, suggesting new features and ensuring the validity of the model. Through the utilization of this framework, it is discovered that B-mode ultrasound classifiers can be enhanced by supplying textural features. More specifically, it is confirmed that statistical texture features extracted from ultrasound frames can increase the accuracy of the burn depth classifier. RESULTS: The system, with all included features selected using explainable AI, is capable of classifying burn depth with accuracy and F1 average above 80%. Additionally, the segmentation module has been found capable of segmenting with a mean global accuracy greater than 84%, and a mean intersection-over-union score over 0.74. CONCLUSIONS: This work demonstrates the feasibility of accurate and automated burn characterization for AI and indicates that these systems can be improved with additional features when a human expert is combined with explainable AI. This is demonstrated on real data (human for segmentation and porcine for depth classification) and establishes the groundwork for further deep-learning thrusts in the area of burn analysis.
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Inteligencia Artificial , Quemaduras , Humanos , Porcinos , Animales , UltrasonografíaRESUMEN
Tissue injury to skin diminishes miR-200b in dermal fibroblasts. Fibroblasts are widely reported to directly reprogram into endothelial-like cells and we hypothesized that miR-200b inhibition may cause such changes. We transfected human dermal fibroblasts with anti-miR-200b oligonucleotide, then using single cell RNA sequencing, identified emergence of a vasculogenic subset with a distinct fibroblast transcriptome and demonstrated blood vessel forming function in vivo. Anti-miR-200b delivery to murine injury sites likewise enhanced tissue perfusion, wound closure, and vasculogenic fibroblast contribution to perfused vessels in a FLI1 dependent manner. Vasculogenic fibroblast subset emergence was blunted in delayed healing wounds of diabetic animals but, topical tissue nanotransfection of a single anti-miR-200b oligonucleotide was sufficient to restore FLI1 expression, vasculogenic fibroblast emergence, tissue perfusion, and wound healing. Augmenting a physiologic tissue injury adaptive response mechanism that produces a vasculogenic fibroblast state change opens new avenues for therapeutic tissue vascularization of ischemic wounds.
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Fibroblastos , Piel , Cicatrización de Heridas , Animales , Humanos , Ratones , Antagomirs/farmacología , Antagomirs/uso terapéutico , Fibroblastos/metabolismo , Fibroblastos/fisiología , Oligonucleótidos/farmacología , Piel/metabolismo , Cicatrización de Heridas/genética , Cicatrización de Heridas/fisiologíaRESUMEN
Non-invasive, repeated interrogation of the same wound is necessary to understand the tissue repair continuum. In this work, we sought to test the significance of non-invasive high-frequency high-resolution ultrasound technology for such interrogation. High-frequency high-resolution ultrasound imaging was employed to investigate wound healing under fetal and adult conditions. Quantitative tissue cellularity and elastic strain was obtained for visualization of unresolved inflammation using Vevo strain software. Hemodynamic properties of the blood flow in the artery supplying the wound-site were studied using color Doppler flow imaging. Non-invasive monitoring of fetal and adult wound healing provided unprecedented biomechanical and functional insight. Fetal wounds showed highly accelerated closure with transient perturbation of wound tissue cellularity. Fetal hemodynamics was unique in that sharp fall in arterial pulse pressure (APP) which was rapidly restored within 48h post-wounding. In adults, APP transiently increased post-wounding before returning to the pre-wounding levels by d10 post-wounding. The pattern of change in the elasticity of wound-edge tissue of diabetics was strikingly different. Severe strain acquired during the early inflammatory phase persisted with a slower recovery of elasticity compared to that of the non-diabetic group. Wound bed of adult diabetic mice (db/db) showed persistent hypercellularity compared to littermate controls (db/+) indicative of prolonged inflammation. Normal skin strain of db/+ and db/db were asynchronous. In db/db, severe strain acquired during the early inflammatory phase persisted with a slower recovery of elasticity compared to that of non-diabetics. This study showcases a versatile clinically relevant imaging platform suitable for real-time analyses of functional wound healing.
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Diagnóstico por Imagen/métodos , Ultrasonografía/métodos , Animales , Fenómenos Biomecánicos , Femenino , Hemodinámica/fisiología , Imagenología Tridimensional/métodos , Ratones , Embarazo , Cicatrización de Heridas/fisiologíaRESUMEN
BACKGROUND: Basilar artery occlusion (BAO) is a subset of posterior circulation stroke that carries a mortality as high as 90%. The current clinical standard to diagnose ischemic stroke include computerized tomography (CT), CT angiography and perfusion and magnetic resonance imaging (MRI). Large animal pre-clinical models to accurately reflect the clinical disease as well as methods to assess stroke burden and evaluate treatments are lacking. METHODS: We describe a canine model of large vessel occlusion (LVO) stroke in the posterior circulation, and developed a laser speckle imaging (LSI) protocol to monitor perfusion changes in real time. We then utilized high b-value DWI (b=1800s/mm2) MRI to increase detection sensitivity. We also evaluated the ability of magnetic resonance angiography (MRA) to assess arterial occlusion and correlate with DSA. Finally, we verified infarct size from apparent diffusion coefficient (ADC) mapping with histology. Results: Administration of thromboembolism occluded the basilar artery as tracked by DSA (n=7). LSI correlated with DSA, demonstrating a reduction in perfusion after stroke onset that persisted throughout the experiment, allowing us to monitor perfusion in real time. DWI with an optimized b-value for dogs illustrated the stroke volume and allowed us to derive ADC and magnetic resonance angiography (MRA) images. The MRA performed at the end of the experiment correlated with DSA performed after occlusion. Finally, stroke burden on MRI correlated with histology. CONCLUSIONS: Our studies demonstrate real time perfusion imaging using LSI of a canine thromboembolic LVO model of posterior circulation stroke, which utilizes multimodal imaging important in the diagnosis and treatment of ischemic stroke.
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Infarto Cerebral/diagnóstico por imagen , Rayos Láser , Imagen por Resonancia Magnética , Perfusión , Accidente Cerebrovascular/diagnóstico por imagen , Animales , Arteriopatías Oclusivas/complicaciones , Arteriopatías Oclusivas/diagnóstico por imagen , Arteria Basilar/diagnóstico por imagen , Arteria Basilar/patología , Arteria Basilar/fisiopatología , Infarto Cerebral/diagnóstico , Infarto Cerebral/patología , Infarto Cerebral/fisiopatología , Angiografía por Tomografía Computarizada , Imagen de Difusión por Resonancia Magnética , Modelos Animales de Enfermedad , Perros , Angiografía por Resonancia Magnética , Accidente Cerebrovascular/diagnóstico , Accidente Cerebrovascular/fisiopatología , Volumen SistólicoRESUMEN
Epithelial to mesenchymal transition (EMT) and wound vascularization are two critical interrelated processes that enable cutaneous wound healing. Zinc finger E-box binding homeobox 1 (ZEB1), primarily studied in the context of tumor biology, is a potent EMT activator. ZEB1 is also known to contribute to endothelial cell survival as well as stimulate tumor angiogenesis. The role of ZEB1 in cutaneous wounds was assessed using Zeb1+/- mice, as Zeb1-/- mice are not viable. Quantitative stable isotope labeling by amino acids in cell culture (SILAC) proteomics was used to elucidate the effect of elevated ZEB1, as noted during hyperglycemia. Under different glycemic conditions, ZEB1 binding to E-cadherin promoter was investigated using chromatin immunoprecipitation. Cutaneous wounding resulted in loss of epithelial marker E-cadherin with concomitant gain of ZEB1. The dominant proteins downregulated after ZEB1 overexpression functionally represented adherens junction pathway. Zeb1+/- mice exhibited compromised wound closure complicated by defective EMT and poor wound angiogenesis. Under hyperglycemic conditions, ZEB1 lost its ability to bind E-cadherin promoter. Keratinocyte E-cadherin, thus upregulated, resisted EMT required for wound healing. Diabetic wound healing was improved in ZEB+/- as well as in db/db mice subjected to ZEB1 knockdown. This work recognizes ZEB1 as a key regulator of cutaneous wound healing that is of particular relevance to diabetic wound complication.
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Transición Epitelial-Mesenquimal/fisiología , Neovascularización Fisiológica/fisiología , Cicatrización de Heridas/fisiología , Homeobox 1 de Unión a la E-Box con Dedos de Zinc/metabolismo , Adulto , Animales , Glucemia , Cadherinas/genética , Cadherinas/metabolismo , Células Cultivadas , Diabetes Mellitus/metabolismo , Células Endoteliales/metabolismo , Femenino , Humanos , Queratinocitos/metabolismo , Masculino , Ratones , Ratones Noqueados , Persona de Mediana Edad , Regiones Promotoras Genéticas , Regulación hacia Arriba , Homeobox 1 de Unión a la E-Box con Dedos de Zinc/genéticaRESUMEN
Temperature distribution is a crucial factor in determining the outcome of laser phototherapy in cancer treatment. Magnetic resonance imaging (MRI) is an ideal method for 3-D noninvasive temperature measurement. A 7.1-T MRI was used to determine laser-induced high thermal gradient temperature distribution of target tissue with high spatial resolution. Using a proton density phase shift method, thermal mapping is validated for in vivo thermal measurement with light-absorbing enhancement dye. Tissue-simulating phantom gels, biological tissues, and tumor-bearing animals were used in the experiments. An 805-nm laser was used to irradiate the samples, with laser power in the range of 1 to 3 W. A clear temperature distribution matrix within the target and surrounding tissue was obtained with a specially developed processing algorithm. The temperature mapping showed that the selective laser photothermal effect could result in temperature elevation in a range of 10 to 45 degrees C. The temperature resolution of the measurement was about 0.37 degrees C with 0.4-mm spatial resolution. The results of this study provide in vivo thermal information and future reference for optimizing laser dosage and dye concentration in cancer treatment.
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Neoplasias de la Mama/patología , Neoplasias de la Mama/terapia , Hipertermia Inducida/métodos , Terapia por Láser/métodos , Imagen por Resonancia Magnética/métodos , Cirugía Asistida por Computador/métodos , Animales , Femenino , Ratas , Ratas Wistar , Resultado del TratamientoRESUMEN
Tissue surface temperature distribution on the treatment site can serve as an indicator for the effectiveness of a photothermal therapy. In this study, both infrared thermography and theoretical simulation were used to determine the surface temperature distribution during laser irradiation of both gel phantom and animal tumors. Selective photothermal interaction was attempted by using intratumoral indocyanine green enhancement and irradiation via a near-infrared laser. An immunoadjuvant was also used to enhance immunological responses during tumor treatment. Monte Carlo method for tissue absorption of light and finite difference method for heat diffusion in tissue were used to simulate the temperature distribution during the selective laser photothermal interaction. An infrared camera was used to capture the thermal images during the laser treatment and the surface temperature was determined. Our findings show that the theoretical and experimental results are in good agreement and that the surface temperature of irradiated tissue can be controlled with appropriate dye and adjuvant enhancement. These results can be used to control the laser tumor treatment parameters and to optimize the treatment outcome. More importantly, when used with immunotherapy as a precursor of immunological responses, the selective photothermal treatment can be guided by the tissue temperature profiles both in the tumor and on the surface.
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Terapia por Láser , Neoplasias Mamarias Animales/cirugía , Temperatura Cutánea , Animales , Pollos , Femenino , Ratones , Ratones Endogámicos BALB C , Método de Montecarlo , Fantasmas de Imagen , Termografía/métodos , Resultado del TratamientoRESUMEN
Cutaneous microvasculopathy complicates wound healing. Functional assessment of gated individual dermal microvessels is therefore of outstanding interest. Functional performance of laser speckle contrast imaging (LSCI) systems is compromised by motion artefacts. To address such weakness, post-processing of stacked images is reported. We report the first post-processing of binary raw data from a high-resolution LSCI camera. Sharp images of low-flowing microvessels were enabled by introducing inverse variance in conjunction with speckle contrast in Matlab-based program code. Extended moving window averaging enhanced signal-to-noise ratio. Functional quantitative study of blood flow kinetics was performed on single gated microvessels using a free hand tool. Based on detection of flow in low-flow microvessels, a new sharp contrast image was derived. Thus, this work presents the first distinct image with quantitative microperfusion data from gated human foot microvasculature. This versatile platform is applicable to study a wide range of tissue systems including fine vascular network in murine brain without craniotomy as well as that in the murine dorsal skin. Importantly, the algorithm reported herein is hardware agnostic and is capable of post-processing binary raw data from any camera source to improve the sensitivity of functional flow data above and beyond standard limits of the optical system.
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Encéfalo/irrigación sanguínea , Pie/irrigación sanguínea , Hemorreología , Rayos Láser , Microcirculación , Algoritmos , Encéfalo/diagnóstico por imagen , Pie/diagnóstico por imagen , Humanos , Aumento de la Imagen , Procesamiento de Imagen Asistido por Computador , Masculino , Relación Señal-RuidoRESUMEN
Objective: (1) Develop a standardized approach to quantitatively measure residual limb skin health. (2) Report reference residual limb skin health values in people with transtibial and transfemoral amputation. Approach: Residual limb health outcomes in individuals with transtibial (n = 5) and transfemoral (n = 5) amputation were compared to able-limb controls (n = 4) using noninvasive imaging (hyperspectral imaging and laser speckle flowmetry) and probe-based approaches (laser doppler flowmetry, transcutaneous oxygen, transepidermal water loss, surface electrical capacitance). Results: A standardized methodology that employs noninvasive imaging and probe-based approaches to measure residual limb skin health are described. Compared to able-limb controls, individuals with transtibial and transfemoral amputation have significantly lower transcutaneous oxygen tension, higher transepidermal water loss, and higher surface electrical capacitance in the residual limb. Innovation: Residual limb health as a critical component of prosthesis rehabilitation for individuals with lower limb amputation is understudied in part due to a lack of clinical measures. Here, we present a standardized approach to measure residual limb health in people with transtibial and transfemoral amputation. Conclusion: Technology advances in noninvasive imaging and probe-based measures are leveraged to develop a standardized approach to quantitatively measure residual limb health in individuals with lower limb loss. Compared to able-limb controls, resting residual limb physiology in people that have had transfemoral or transtibial amputation is characterized by lower transcutaneous oxygen tension and poorer skin barrier function.
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Selective photothermal interaction using dye enhancement has proven to be effective in minimizing surrounding tissue damage and delivering energy to target tissue. During laser irradiation, the process of photon absorption and thermal energy diffusion in the target tissue and its surrounding tissue are crucial. Such information allows the selection of proper operating parameters such as dye concentrations, laser power, and exposure time for optimal therapeutic effect. Combining the Monte Carlo method for energy absorption and the finite difference method for heat diffusion, the temperature distributions in target tissue and surrounding tissue in dye enhanced laser photothermal interaction are obtained. Different tissue configurations and dye enhancement are used in the simulation, and different incident beam sizes are also used to determine optimum beam sizes for various tissue configurations. Our results show that the algorithm developed in this study could predict the thermal outcome of laser irradiation. Our simulation indicates that with appropriate absorption enhancement of the target tissue, the temperature in the target tissue and in the surrounding tissue can be effectively controlled. This method can be used for optimization of lesion treatment using laser photothermal interactions. It may also provide guidance for laser immunotherapy in cancer treatment, since the immunological responses are believed to be related to tissue temperature changes.
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Diagnóstico por Computador/métodos , Hipertermia Inducida/métodos , Terapia por Láser , Modelos Biológicos , Neoplasias/fisiopatología , Neoplasias/terapia , Animales , Simulación por Computador , Transferencia de Energía/efectos de la radiación , Humanos , Método de Montecarlo , TemperaturaRESUMEN
BACKGROUND: Wound assessment relies on visual evaluation by physicians. Such assessment is largely subjective and presents the opportunity to explore the use of emergent technologies. METHODS: Emergent and powerful noninvasive imaging technologies applicable to assess burn and chronic wounds are reviewed. RESULTS: The need to estimate wound depth is critical in both chronic wound and burn injury settings. Harmonic ultrasound technology is powerful to study wound depth. It addresses the limitations of optical imaging with limited depth of penetration. What if a wound appears epithelialized by visual inspection, which shows no discharge yet is covered by repaired skin that lacks barrier function? In this case although the wound is closed as defined by current standards, it remains functionally open, presenting the risk of infection and other postclosure complications. Thus, assessment of skin barrier function is valuable in the context of assessing wound closure. Options for the study of tissue vascularization are many. If noncontact and noninvasive criteria are of importance, laser speckle imaging is powerful. Fluorescence imaging is standard in several clinical settings and is likely to serve the wound clinics well as long as indocyanine green injection is not of concern. A major advantage of harmonic ultrasound imaging of wound depth is that the same system is capable of providing information on blood flow dynamics in arterial perforators. CONCLUSION: With many productive imaging platforms to choose from, wound care is about to be transformed by technology that would help assess wound severity.
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Piel/diagnóstico por imagen , Heridas y Lesiones/diagnóstico por imagen , Quemaduras/diagnóstico por imagen , Quemaduras/patología , Quemaduras/fisiopatología , Enfermedad Crónica , Humanos , Piel/lesiones , Piel/patología , Piel/fisiopatología , Cicatrización de Heridas/fisiología , Heridas y Lesiones/patología , Heridas y Lesiones/fisiopatologíaRESUMEN
This work represents the first study employing non-invasive high-resolution harmonic ultrasound imaging to longitudinally characterize skin wound healing. Burn wounds (day 0-42), on the dorsum of a domestic Yorkshire white pig were studied non-invasively using tandem digital planimetry, laser speckle imaging and dual mode (B and Doppler) ultrasound imaging. Wound depth, as measured by B-mode imaging, progressively increased until day 21 and decreased thereafter. Initially, blood flow at the wound edge increased up to day 14 and subsequently regressed to baseline levels by day 21, when the wound was more than 90% closed. Coinciding with regression of blood flow at the wound edge, there was an increase in blood flow in the wound bed. This was observed to regress by day 42. Such changes in wound angiogenesis were corroborated histologically. Gated Doppler imaging quantitated the pulse pressure of the primary feeder artery supplying the wound site. This pulse pressure markedly increased with a bimodal pattern following wounding connecting it to the induction of wound angiogenesis. Finally, ultrasound elastography measured tissue stiffness and visualized growth of new tissue over time. These studies have elegantly captured the physiological sequence of events during the process of wound healing, much of which is anticipated based on certain dynamics in play, to provide the framework for future studies on molecular mechanisms driving these processes. We conclude that the tandem use of non-invasive imaging technologies has the power to provide unprecedented insight into the dynamics of the healing skin tissue.
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Ultrasonografía , Cicatrización de Heridas , Animales , Biomarcadores , Biopsia , Diagnóstico por Imagen/métodos , Diagnóstico por Imagen de Elasticidad , Modelos Animales , Flujo Sanguíneo Regional , Porcinos , Ultrasonografía/métodosRESUMEN
BACKGROUND: Dicer endonuclease, critical for maturation of miRNAs, is depleted in certain forms of cardiomyopathy which results in differential expression of certain microRNAs. We sought to elucidate the mechanisms underlying the rapid loss of cardiac function following cardiac-specific Dicer depletion in adult mice. RESULTS: Conditional Dicer deletion in the adult murine myocardium demonstrated compromised heart function, mitochondrial dysfunction and oxidant stress. Elevated miR-15b was observed as an early response to Dicer depletion and was found to silence Pim-1 kinase, a protein responsible for maintaining mitochondrial integrity and function. Anti-miRNA based suppression of induced miRNA-15b rescued the function of Dicer-depleted adult heart and attenuated hypertrophy. CONCLUSIONS: Anti-miRNA based suppression of inducible miRNA-15b can prevent rapid loss of cardiac function in a Dicer-depleted adult heart and can be a key approach worthy of therapeutic consideration.