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BACKGROUND: In transposition flaps, thicker tissue and higher degrees of rotation are associated with increased pivotal restraint; however, limited experimental data exist quantifying the degree to which these affect flap biomechanics. The use of artificial skin models in conjunction with digital image correlation technology allows for investigation into biomechanical properties of skin flaps. OBJECTIVE: To quantify the effects of tissue thickness and rotational angles on pivotal restraint within transposition flaps using artificial skin models. METHODS: Ninety degree bilobed and trilobed flaps were used to close defects in artificial skin models of increasing thicknesses. Digital image correlation was used to quantify strain. Quantitative and qualitative differences in strain were assessed in increasing flap thicknesses and between flap designs. RESULTS: Increasing flap thickness was associated with decreasing strain. In the bilobed flap, increasing thickness was associated with displacement of the flap pivot point away from the distal flap edge. Comparatively, lower angles of rotation in the trilobed flap were not associated with migration of the flap pivot point. CONCLUSION: Increased pivotal restraint observed in higher degrees of rotation is due to migration of the flap pivot point. This model supports the common practice of decreasing flap angles to compensate for pivotal restraint.
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Piel Artificial , Colgajos Quirúrgicos , Fenómenos Biomecánicos , Ensayo de Materiales , Fotograbar , Rotación , Resistencia a la TracciónRESUMEN
Mast cells have functional plasticity affected by their tissue microenvironment, which greatly impacts their inflammatory responses. Because lactic acid (LA) is abundant in inflamed tissues and tumors, we investigated how it affects mast cell function. Using IgE-mediated activation as a model system, we found that LA suppressed inflammatory cytokine production and degranulation in mouse peritoneal mast cells, data that were confirmed with human skin mast cells. In mouse peritoneal mast cells, LA-mediated cytokine suppression was dependent on pH- and monocarboxylic transporter-1 expression. Additionally, LA reduced IgE-induced Syk, Btk, and ERK phosphorylation, key signals eliciting inflammation. In vivo, LA injection reduced IgE-mediated hypothermia in mice undergoing passive systemic anaphylaxis. Our data suggest that LA may serve as a feedback inhibitor that limits mast cell-mediated inflammation.
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Anafilaxia/prevención & control , Antiinflamatorios no Esteroideos/farmacología , Retroalimentación Fisiológica , Inmunoglobulina E/genética , Ácido Láctico/farmacología , Mastocitos/efectos de los fármacos , Agammaglobulinemia Tirosina Quinasa/genética , Agammaglobulinemia Tirosina Quinasa/inmunología , Anafilaxia/inducido químicamente , Anafilaxia/inmunología , Anafilaxia/patología , Animales , Dinitrofenoles/administración & dosificación , Dinitrofenoles/antagonistas & inhibidores , Femenino , Regulación de la Expresión Génica , Cetoprofeno/farmacología , Ácido Láctico/inmunología , Ácido Láctico/metabolismo , Mastocitos/inmunología , Mastocitos/patología , Ratones , Ratones Endogámicos C57BL , Proteína Quinasa 3 Activada por Mitógenos/genética , Proteína Quinasa 3 Activada por Mitógenos/inmunología , Transportadores de Ácidos Monocarboxílicos/genética , Transportadores de Ácidos Monocarboxílicos/inmunología , Cavidad Peritoneal/patología , Fosforilación/efectos de los fármacos , Cultivo Primario de Células , Albúmina Sérica/administración & dosificación , Albúmina Sérica/antagonistas & inhibidores , Transducción de Señal , Piel/efectos de los fármacos , Piel/inmunología , Piel/patología , Quinasa Syk/genética , Quinasa Syk/inmunología , Simportadores/genética , Simportadores/inmunologíaRESUMEN
Lactic acid (LA) is present in tumors, asthma, and wound healing, environments with elevated IL-33 and mast cell infiltration. Although IL-33 is a potent mast cell activator, how LA affects IL-33-mediated mast cell function is unknown. To investigate this, mouse bone marrow-derived mast cells were cultured with or without LA and activated with IL-33. LA reduced IL-33-mediated cytokine and chemokine production. Using inhibitors for monocarboxylate transporters (MCT) or replacing LA with sodium lactate revealed that LA effects are MCT-1- and pH-dependent. LA selectively altered IL-33 signaling, suppressing TGF-ß-activated kinase-1, JNK, ERK, and NF-κB phosphorylation, but not p38 phosphorylation. LA effects in other contexts have been linked to hypoxia-inducible factor (HIF)-1α, which was enhanced in bone marrow-derived mast cells treated with LA. Because HIF-1α has been shown to regulate the microRNA miR-155 in other systems, LA effects on miR-155-5p and miR-155-3p species were measured. In fact, LA selectively suppressed miR-155-5p in an HIF-1α-dependent manner. Moreover, overexpressing miR-155-5p, but not miR-155-3p, abolished LA effects on IL-33-induced cytokine production. These in vitro effects of reducing cytokines were consistent in vivo, because LA injected i.p. into C57BL/6 mice suppressed IL-33-induced plasma cytokine levels. Lastly, IL-33 effects on primary human mast cells were suppressed by LA in an MCT-dependent manner. Our data demonstrate that LA, present in inflammatory and malignant microenvironments, can alter mast cell behavior to suppress inflammation.
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Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Inflamación/prevención & control , Interleucina-33/inmunología , Ácido Láctico/farmacología , Mastocitos/efectos de los fármacos , Mastocitos/inmunología , MicroARNs/genética , Animales , Células Cultivadas , Relación Dosis-Respuesta a Droga , Femenino , Humanos , Inflamación/inmunología , Masculino , Mastocitos/metabolismo , Ratones , Ratones Endogámicos C57BL , MicroARNs/biosíntesis , Relación Estructura-ActividadRESUMEN
We present an insert-based approach to fabricate scalable and multiplexable microfluidic devices for 3D cell culture and integration with downstream detection modules. Laser-cut inserts with a layer of electrospun fibers are used as a scaffold for 3D cell culture, with the inserts being easily assembled in a 3D-printed fluidic device for flow-based studies. With this approach, the number and types of cells (on the inserts) in one fluidic device can be customized. Moreover, after an investigation (i.e., stimulation) under flowing conditions, the cell-laden inserts can be removed easily for subsequent studies including imaging and cell lysis. In this paper, we first discuss the fabrication of the device and characterization of the fibrous inserts. Two device designs containing two (channel width = 260 µm) and four (channel width = 180 µm) inserts, respectively, were used for different experiments in this study. Cell adhesion on the inserts with flowing media through the device was tested by culturing endothelial cells. Macrophages were cultured and stimulated under different conditions, the results of which indicate that the fibrous scaffolds under flow conditions result in dramatic effects on the amount and kinetics of TNF-α production (after LPS stimulation). Finally, we show that the cell module can be integrated with a downstream absorbance detection scheme. Overall, this technology represents a new and versatile way to culture cells in a more in vivo fashion for in vitro studies with online detection modules. Graphical abstract This paper describes an insert-based microfluidic device for 3D cell culture that can be easily scaled, multiplexed, and integrated with downstream analytical modules.
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Técnicas de Cultivo de Célula/instrumentación , Dispositivos Laboratorio en un Chip , Análisis de la Célula Individual/instrumentación , Animales , Bovinos , Adhesión Celular , Línea Celular , Células Endoteliales/citología , Diseño de Equipo , Macrófagos/citología , Ratones , Impresión Tridimensional , Células RAW 264.7RESUMEN
Degeneration of the nucleus pulposus (NP) is the primary cause of back pain in almost 80% of the world population. The current gold standard treatment for a degenerated NP is a spinal fusion surgery which is costly, temporary, and extremely invasive. Research has been moving towards minimally invasive methods to lessen the collateral damage created during surgery. The use of a tissue-engineered scaffold has the potential to promote a healthy and hydrated environment to regenerate the NP. Cryogels are unique polymeric scaffolds composed of a highly connected, macroporous structure, and are capable of maintaining stability under high deformations. For this study, cryogels have been developed using gelatin and poloxamer 407 (P407) at varying ratios to determine the ideal combination of stability, water retention, and pore size. For the application of NP regeneration, a gelatin-P407 cryogel should be both stable and a well hydrated carrier. The cryogels created varied from a 1:1 gelatin to P407 ratio to a 10:1 ratio. The inclusion of P407 in the cryogels resulted in a significant increase in hydrophilicity, ideal pore size for cell infiltration, mechanical stability over 28 days, and cell infiltration after just 21 days. This novel gelatin-P407 composite cryogel has the potential to be a practical alternative to the spinal fusion procedure, saving patients hundreds of thousands of dollars and, ideally, leading to improved patient outcome.
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Criogeles/química , Núcleo Pulposo/metabolismo , Poloxámero/química , Regeneración , Andamios del Tejido/química , Animales , Adhesión Celular , Peces/metabolismo , Gelatina/química , Humanos , Microscopía Electrónica de Rastreo , Polímeros/química , Porosidad , Fusión Vertebral , Ingeniería de Tejidos/métodos , Microtomografía por Rayos XRESUMEN
Organs-on-a-chip has emerged as a powerful tool for pharmacological and physiological studies. A key part in the construction of such a model is the ability to pattern or culture cells in a biomimetic fashion. Most of the reported cells-on-a-chip models integrate cells on a flat surface, which does not accurately represent the extracellular matrix that they experience in vivo. Electrospinning, a technique used to generate sub-micron diameter polymer fibers, has been used as an in vitro cell culture substrate and for tissue engineering applications. Electrospinning of fibers directly into a fully sealed fluidic channel using a conventional setup has not been possible due to issues of confining the fibers into a discrete network. In this work, a dynamic focusing method was developed, with this approach enabling direct deposition of electrospun fibers into a fully sealed fluidic channel, to act as a matrix for cell culture and subsequent studies under continuous flowing conditions. Scanning electron microscopy of electrospun polycaprolactone fibers shows that this method enables the formation of fibrous layers on the inner wall of a 3D-printed fluidic device (mean fiber size = 1.6 ± 0.6 µm and average pore size = 113 ± 19 µm(2)). Cells were able to be cultured in this 3D scaffold without the addition of adhesion proteins. Media was pumped through the channel at high flow rates (up to 400 µL min(-1)) during a dynamic cell culture process and both the fibers and the cells were found to be strongly adherent. A PDMS fluidic device was also prepared (from a 3D printed mold) and coated with polycaprolactone fibers. The PDMS device enables optical detection and confocal imaging of cultured cells on the fibers. Finally, macrophages were cultured in the devices to study how the fibrous scaffold can affect cell behavior. It was found that under lipopolysaccharide stimulation, macrophages cultured on PCL fibers inside of a channel secreted significantly more cytokines than those cultured on a thin layer of PCL in a channel or directly on the inner channel wall. Overall, this study represents a new approach for in vitro cell studies, where electrospinning can be used to easily and quickly create 3D scaffolds that can improve the culture conditions in microfluidic devices.
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Dispositivos Laboratorio en un Chip , Impresión Tridimensional , Andamios del Tejido , Animales , Técnicas de Cultivo de Célula , Células Cultivadas , Fibroblastos/citología , Humanos , Ratones , Poliésteres , Células RAW 264.7/citología , Ingeniería de TejidosRESUMEN
This study investigates the novel approach of placing a ventricular assist pump in the descending aorta in series configuration with the heart and compares it with the two traditional approaches of left-ventricle-to-ascending-aorta (LV-AA) and left-ventricle-to-descending-aorta (LV-DA) placement in parallel with the heart. Experiments were conducted by using the in-house simulator of the cardiovascular blood-flow loop (SCVL). The results indicate that the use of the LV-AA in-parallel configuration leads to a significant improvement in the systemic and pulmonic flow as the level of continuous flow is increased; however, this approach is considered highly invasive. The use of the LV-DA in-parallel configuration leads to an improvement in the systemic and pulmonic flow at lower levels of continuous flow but at higher levels of pump support leads to retrograde flow. In both in-parallel configurations, increasing the level of pump continuous flow leads to a decrease in pulsatility to a certain extent. The results of placing the pump in the descending aorta in series configuration show that the pressure drop upstream of the pump facilitates cardiac output as a result of afterload reduction. In addition, the pressure rise downstream of the pump may assist with renal perfusion. However, at the same time, the pressure drop generated at the proximal part of the descending aorta induces a slight drop in carotid perfusion, which would be autoregulated by the brain in a native cardiovascular system. The pulse wave analysis shows that placing the pump in the descending aorta leads to improved pulsatility in comparison with the traditional in-parallel configurations.
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Aorta Torácica/cirugía , Aorta/cirugía , Ventrículos Cardíacos/cirugía , Corazón Auxiliar , Simulación por Computador , Diseño de Equipo , Insuficiencia Cardíaca/cirugía , Hemodinámica , Humanos , Modelos CardiovascularesRESUMEN
STUDY DESIGN: A mechanical and chemical model of intervertebral disk (IVD) degeneration was developed by examining the enzymatic degradation of the nucleus pulposus (NP), and a gelatin-based restoration study was performed. OBJECTIVE: It was hypothesized that forced enzymatic degradation of the NP will mimic natural degeneration through the loss of disk height and that an injection of a gelatin solution will restore mechanical function. SUMMARY OF BACKGROUND DATA: Collagen and proteoglycans are essential for normal NP function. Their chemical destruction, combined with light mechanical loading, will mimic degeneration. Previous studies have determined that collagenase and matrix metalloproteinase-3 are directly implicated in IVD degradation; therefore, these enzymes were used in this model. MATERIALS AND METHODS: On the basis of preliminary testing, 0.5% collagenase, 1% collagenase, and 0.0025% metalloproteinase-3 in phosphate-buffered saline (PBS) were injected directly into the NP of various motion segments from a young bovine lumbar spine and subjected to light cyclic loading. To restore disk height and mechanical function, 20% gelatin in PBS at 70°C was injected into a degraded disk and subjected to the same loading conditions after an allotted hardening time. RESULTS: Mechanical testing showed statistically significant changes in disk height between control segments, 1% collagenase, and 0.5% collagenase. 0.5% collagenase had the most accurate appearance and loading pattern of degeneration upon disk transection postloading. A trend in restoration of disk function, given by the lessened loss of disk height upon loading, was observed with injection of gelatin after degradation with 0.5% collagenase. CONCLUSIONS: This study demonstrated the potential to create a degenerative model using enzymatic degradation of the NP and the possibility to restore function with an injectable therapy. Although gelatin is not a clinically viable option, it provides preliminary data for other injectable IVD therapies.
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Degeneración del Disco Intervertebral/metabolismo , Degeneración del Disco Intervertebral/fisiopatología , Disco Intervertebral/fisiopatología , Vértebras Lumbares/fisiología , Modelos Químicos , Vértebras Torácicas/fisiología , Animales , Fenómenos Biomecánicos/fisiología , Bovinos , Colágeno/metabolismo , Colágeno/farmacología , Colagenasas/metabolismo , Colagenasas/farmacología , Gelatina/farmacología , Disco Intervertebral/efectos de los fármacos , Disco Intervertebral/enzimología , Vértebras Lumbares/efectos de los fármacos , Vértebras Lumbares/enzimología , Metaloproteinasa 3 de la Matriz/metabolismo , Metaloproteinasa 3 de la Matriz/farmacología , Vértebras Torácicas/efectos de los fármacos , Vértebras Torácicas/enzimología , Soporte de Peso/fisiologíaRESUMEN
Tissue-engineered scaffolds have been identified as appropriate templates for bone regeneration, especially complex geometries seen in craniofacial defects. Here we describe the general fabrication and modification of hydrogels, cryogels, and electrospun scaffolds. These scaffolds offer a variety of templates for facilitating bone growth and regeneration in craniofacial applications.
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Regeneración Ósea , Ingeniería de Tejidos , Huesos , Criogeles , Andamios del TejidoRESUMEN
BACKGROUND/OBJECTIVES: Chronic pressure ulcers affect patient health, emotional state, and quality of life, causing considerable morbidity and mortality in addition to contributing to significant health care costs from lengthy hospitalizations to advanced home care and surgical care costs. The conventional treatment of these wounds can be slow due to their chronic inflammatory state and the senescence of local reparative cells. Platelet-rich plasma (PRP) therapy has been growing as a viable treatment alternative for a number of clinical applications and has potential benefit for use in chronic wounds. The sustained release of large quantities of autologous growth factors, cytokines, and other mediators found in PRP plus the favorable mononuclear cell profile of PRP may help us to stimulate wound healing and resolve chronic inflammation. METHODS: Three veterans with spinal cord injury (SCI), presenting with chronic stage IV pressure ulcers, were treated with a sustained release PRP therapy to stimulate wound healing. RESULTS: PRP treatment consistently resulted in the formation of granulation tissue and improved vascularity for each of the three patients treated, while reducing the overall ulcer area and volume. CONCLUSION: The controlled release of growth factors from PRP demonstrated a positive stimulatory effect on the healing rate of chronic pressure ulcers in individuals with SCI.
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Plasma Rico en Plaquetas , Úlcera por Presión/complicaciones , Úlcera por Presión/terapia , Traumatismos de la Médula Espinal/complicaciones , Cicatrización de Heridas , Adulto , Enfermedad Crónica , Humanos , Masculino , Persona de Mediana Edad , Cuadriplejía/complicacionesRESUMEN
BACKGROUND: Platelet-rich-plasma (PRP) is used to treat knee osteoarthritis; however, mechanistic evidence of PRP effectiveness for pain relief is limited. OBJECTIVE: To assess molecular biomarkers and mesenchymal stem cells (MSCs) in synovial fluid during PRP treatment of the osteoarthritic knee joint. DESIGN: Single blinded, randomized, placebo controlled pilot study. SETTING: Veterans Affairs Medical Center. PARTICIPANTS: Seventeen participants with mild to moderate knee osteoarthritis were randomized in a 2:1 placebo-controlled ratio, receiving PRP or saline (placebo) intra-articular injection into the knee joint. METHODS: Knee synovial fluid was analyzed before the respective injections and again 10 days following injection. Participants were followed up to 12 months completing visual analog scale (VAS) and Western Ontario and McMaster Universities Osteoarthritis Index (WOMAC) questionnaires at intervals over that period. MAIN OUTCOME MEASURES: The effects of PRP on synovial protein and MSC gene expression levels were measured by multiplex enzyme-linked immunosorbent assay and quantitative polymerase chain reaction. RESULTS: Novel biomarkers including levels of interleukin (IL)-5, IL-6, IL-10, and tumor necrosis factor-α were measured in synovial fluid 10 days after PRP treatment. Altered gene expression profiles in MSCs from patients treated with PRP were observed for matrix metalloproteinases and inflammatory markers (IL-6, IL-8, CCL2, TNF-α). A2M protease was significantly increased following PRP treatment (P = .005). WOMAC scores declined for up to 3 months from baseline levels and remained low at 6 and 12 months in the PRP group. In contrast, WOMAC scores for patients receiving the saline injection were relatively unchanged for up to 12 months. CONCLUSIONS: We report significant changes for the biomarker A2M (P = .005) as well as differences in expression of cellular markers and postulate that PRP modulates the local knee synovial environment by altering the inflammatory milieu, matrix degradation, and angiogenic growth factors. The PRP treatment group had less pain and stiffness and improved function scores.
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Osteoartritis de la Rodilla , Plasma Rico en Plaquetas , Humanos , Ácido Hialurónico , Inyecciones Intraarticulares , Articulación de la Rodilla , Osteoartritis de la Rodilla/tratamiento farmacológico , Resultado del TratamientoRESUMEN
Osteons are the repeating unit throughout cortical bone, consisting of canals filled with blood and nerve vessels surrounded by concentric lamella of hydroxyapatite-containing collagen fibers, providing mechanical strength. Creating a biodegradable scaffold that mimics the osteon structure is crucial for optimizing cellular infiltration and ultimately the replacement of the scaffold with native cortical bone. In this study, a modified air-gap electrospinning setup was exploited to continuously wrap highly aligned polycaprolactone polymer nanofibers around individual 1393 bioactive glass microfibers, resulting in a synthetic structure similar to osteons. By varying the parameters of the device, scaffolds with polymer fibers wrapped at angles between 5-20° to the glass fiber were chosen. The scaffold indicated increased cell migration by demonstrating unidirectional cell orientation along the fibers, similar to recent work regarding aligned nerve and muscle regeneration. The wrapping decreased the porosity from 90% to 80%, which was sufficient for glass conversion through ion exchange validated by inductively coupled plasma. Scaffold degradation was not cytotoxic. Encapsulating the glass with polymer nanofibers caused viscoelastic deformation during three-point bending, preventing typical brittle glass fracture, while maintaining cell migration. This scaffold design structurally mimics the osteon, with the intent to replace its material compositions for better regeneration.
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Alginate is a highly tailorable, biocompatible polymer whose properties can be tuned to mimic the properties of native nucleus pulposus (NP) tissue. Platelet-rich plasma (PRP) is a highly accessible, inexpensive, and readily available mix of pro-regenerative factors. By functionalizing alginate with PRP, a mechanically optimized, bioactive alginate NP analogue may stimulate NP cells to proliferate and accumulate matrix over a longer period of time than if the PRP were solely encapsulated within the hydrogel. In this study, PRP was chemically bound to alginate using carbodiimide chemistry and mechanically, physically, and cytologically compared to plain alginate as well as alginate containing free-floating lyophilized PRP. The alginates were mechanically and physically characterized; PRP-conjugated alginate had similar mechanical properties to controls and had the benefit of retained PRP proteins within the hydrogel. Human nucleus pulposus cells (hNPCs) were seeded within the modified alginates and cultured for 14 days. Quantification data of glycosaminoglycans suggests that PRP-incorporated alginate has the potential to increase ECM production within the characterized alginate constructs, and that PRP-functionalized alginate can retain protein within the hydrogel over time. This is the first study to functionalize the milieu of PRP proteins onto alginate and characterize the mechanical and physical properties of the modified alginates. This study also incorporates hNPCs into the characterized PRP-modified alginates to observe phenotypic maintenance when encapsulated within the in situ gelling constructs.
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Alginatos/química , Materiales Biocompatibles/química , Carbodiimidas/química , Hidrogeles/química , Plasma Rico en Plaquetas/química , Andamios del Tejido/química , Proliferación Celular , Células Cultivadas , Matriz Extracelular/metabolismo , Glicosaminoglicanos/metabolismo , Humanos , Hidrogeles/uso terapéutico , Inyecciones , Fenómenos Mecánicos , Núcleo Pulposo/citología , Ingeniería de TejidosRESUMEN
Hydrogels that can respond to multiple external stimuli represent the next generation of advanced functional biomaterials. Here, a series of multimodal hydrogels were synthesized that can contract and expand reversibly over several cycles while changing their mechanical properties in response to blue and red light, as well as heat (â¼50 °C). The light-responsive behavior was achieved through a photoredox-based mechanism consisting of photoinduced electron transfer from a zinc porphyrin photocatalyst in its excited state to oligoviologen-based macrocrosslinkers, both of which were integrated into the hydrogel polymer network during gel formation. Orthogonal thermoresponsive properties were also realized by introducing N-isopropyl acrylamide (NIPAM) monomer simultaneously with hydroxyethyl acrylate (HEA) in the pre-gel mixture to produce a statistical 60 : 40 HEA : NIPAM polymer network. The resultant hydrogel actuators - crosslinked with either a styrenated viologen dimer (2V4+-St) or hexamer (6V12+-St) - were exposed to red or blue light, or heat, for up to 5 h, and their rate of contraction, as well as the corresponding changes in their physical properties (i.e., stiffness, tensile strength, Young's modulus, etc.), were measured. The combined application of blue light and heat to the 6V12+-St-based hydrogels was also demonstrated, resulting in hydrogels with more than two-fold faster contraction kinetics and dramatically enhanced mechanical robustness when fully contracted. We envision that the reported materials and the corresponding methods of remotely manipulating the dynamic hydrogels may serve as a useful blueprint for future adaptive materials used in biomedical applications.
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The aim of this study is to evaluate a core-shell nanofiber as a useful matrix for tuning Rosuvastatin (RSV) release and osteogenic differentiation in vitro. Polyvinyl alcohol (PVA) and silk fibroin were used as the shell and the core, respectively. To obtain a linear and beadless core-shell structure and an optimal release profile, the shell/core flow rate ratio was varied (0, 0.4, 0.6, 0.8, and 1). Formation of continuous nanofibers with an obvious core/sheath structure was proved using scanning electron microscopy (SEM) and transmission electron microscopy (TEM). Differential scanning calorimetry indicated the presence of two distinct phase structures in the nanofibers. Also, RSV molecules were dispersed in an amorphous state in the nanofibers. The in vitro release profile of the core-shell structure exhibited a biphasic release profile and the amount of released RSV was controlled by adjusting the shell flow rate. Human adipose-derived stem cells cultured on the RSV loaded nanofibers were found to improve cell proliferation and assist osteogenic differentiation as revealed by Alizarin red staining and real-time RT-PCR.
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Tejido Adiposo/citología , Fibroínas/química , Nanofibras/química , Osteogénesis/efectos de los fármacos , Alcohol Polivinílico/química , Rosuvastatina Cálcica/farmacología , Células Madre/citología , Ingeniería de Tejidos/métodos , Adulto , Animales , Bombyx , Rastreo Diferencial de Calorimetría , Adhesión Celular/efectos de los fármacos , Adhesión Celular/genética , Diferenciación Celular/efectos de los fármacos , Diferenciación Celular/genética , Proliferación Celular/efectos de los fármacos , Proliferación Celular/genética , Liberación de Fármacos , Femenino , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Cinética , Persona de Mediana Edad , Nanofibras/ultraestructura , Osteogénesis/genética , Células Madre/efectos de los fármacos , Células Madre/metabolismo , HumectabilidadRESUMEN
Manuka honey is an ancient remedy to improve wound healing; however, an effective delivery system is needed to facilitate extended release of honey into wounds. We developed an electrospun dermal regeneration template consisting of a poly (ε-caprolactone) (PCL) scaffold embedded with 1%, 5%, 10%, or 20% manuka honey. In vitro studies demonstrated that honey PCL scaffolds were not toxic to macrophages, and they allowed for macrophage infiltration into the scaffolds. Vascular endothelial growth factor (VEGF), a marker of angiogenesis, was released by macrophages cultured with scaffolds and macrophage/scaffold conditioned media promoted endothelial cell tube formation in an angiogenesis assay. In a full thickness murine wound model, the scaffolds prevented rapid wound contraction. In vivo, cells infiltrated the scaffolds by post-wounding day 7, but the honey scaffolds did not affect collagen deposition at that time. In summary, preliminary studies investigating the effect of honey on tissue repair show that scaffolds prevent rapid wound contraction, allow for cell infiltration, and promote angiogenesis. © 2019 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater 107B:2620-2628, 2019.
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Miel , Macrófagos/metabolismo , Andamios del Tejido/química , Cicatrización de Heridas , Heridas y Lesiones/terapia , Animales , Femenino , Humanos , Macrófagos/patología , Ratones , Heridas y Lesiones/metabolismo , Heridas y Lesiones/patologíaRESUMEN
Honey is well-known for its wound healing capability and Manuka honey (MH) contains a unique Manuka factor, providing an additional antibacterial agent. Previously, there has not been a practical way to apply MH to a wound site, which renders treatment for an extended period extremely difficult. Tissue-engineered scaffolds offer an alternative treatment method to standard dressings by providing varying geometries to best treat the specific tissue. MH was incorporated into cryogels, hydrogels, and electrospun scaffolds to assess the effect of scaffold geometry on bacterial clearance and adhesion, as well as cellular adhesion. Electrospun scaffolds exhibited a faster release due to the nanoporous fibrous geometry which led to a larger partial bacterial clearance as compared to the more three-dimensional cryogels (CG) and hydrogels (HG). Similarly, the fast release of MH from the electrospun scaffolds resulted in reduced bacterial adhesion. Overall, the fast MH release of the electrospun scaffolds versus the extended release of the HG and CG scaffolds provides differences in cellular/bacterial adhesion and advantages for both short and long-term applications, respectively. This manuscript provides a comparison of the scaffold pore structures as well as bacterial and cellular properties, providing information regarding the relationship between varying scaffold geometry and MH efficacy.
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Significance: Historically, honey has been regarded as a potent agent in bacterial inhibition and wound healing. An increased prevalence of antibiotic resistant pathogens spurred an initial resurgence in honey's clinical popularity, with it quickly finding a place in wound care and regenerative medicine. However, this renewed usage demanded a need for improved delivery and overall research of its bioactive properties. This review provides an overview of the antibacterial properties and clinical use of honey. Recent Advances: The past and present clinical use of honey is noted, focusing specifically on burns and ulcers, as these are the most common applications of the natural agent. While honey is often used without modification clinically, there are also commercially available products ranging from dressings to gels, which are discussed. Critical Issues: Despite these products growing in popularity, the need for improved delivery and a structure to support wound healing could improve the treatment method. Future Directions: Tissue engineering scaffolds can provide an alternative method of honey delivery with research focusing primarily on electrospun scaffolds, hydrogels, and cryogels. Current studies on these scaffolds are discussed with respect to their advantages and potential for future clinical work. Overall, this review provides a comprehensive overview of the properties of honey, its current use in wound healing, and the potential for future incorporation into tissue-engineered scaffolds to provide an innovative wound healing agent.
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Volumetric muscle loss (VML) is a loss of over â¼10% of muscle mass that results in functional impairment. Although skeletal muscle possesses the ability to repair and regenerate itself following minor injuries, VML injuries are irrecoverable. Currently, there are no successful clinical therapies for the treatment of VML. Previous studies have treated VML defects with decellularized extracellular matrix (D-ECM) scaffolds derived from either pig urinary bladder or small intestinal submucosa. These therapies were unsuccessful due to the poor mechanical stability of D-ECM leading to quick degradation in vivo. To circumvent these issues, in this manuscript aligned nanofibers of D-ECM were created using electrospinning that mimicked native muscle architecture and provided topographical cues to primary satellite cells. Additionally, combining D-ECM with polycaprolactone (PCL) improved the tensile mechanical properties of the electrospun scaffold. In vitro testing shows that the electrospun scaffold with aligned nanofibers of PCL and D-ECM supports satellite cell growth, myogenic protein expression, and myokine production.
Asunto(s)
Matriz Extracelular/metabolismo , Músculo Esquelético/metabolismo , Mioblastos/metabolismo , Nanofibras/química , Andamios del Tejido/química , Animales , Anisotropía , Bovinos , Proliferación Celular , Células Cultivadas , ADN/análisis , Masculino , Poliésteres , Ratas , Ratas Endogámicas Lew , Regeneración , Estrés Mecánico , Resistencia a la Tracción , Ingeniería de Tejidos/métodosRESUMEN
Skeletal muscle is inept in regenerating after traumatic injuries due to significant loss of basal lamina and the resident satellite cells. To improve regeneration of skeletal muscle, we have developed biomimetic sponges composed of collagen, gelatin, and laminin (LM)-111 that were crosslinked with 1-ethyl-3-(3-dimethyl aminopropyl) carbodiimide (EDC). Collagen and LM-111 are crucial components of the muscle extracellular matrix and were chosen to impart bioactivity whereas gelatin and EDC were used to provide mechanical strength to the scaffold. Morphological and mechanical evaluation of the sponges showed porous structure, water-retention capacity and a compressive modulus of 590-808 kPa. The biomimetic sponges supported the infiltration and viability of C2 C12 myoblasts over 5 days of culture. The myoblasts produced higher levels of myokines such as VEGF, IL-6, and IGF-1 and showed higher expression of myogenic markers such as MyoD and myogenin on the biomimetic sponges. Biomimetic sponges implanted in a mouse model of volumetric muscle loss (VML) supported satellite, endothelial, and inflammatory cell infiltration but resulted in limited myofiber regeneration at 2 weeks post-injury. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 107A: 92-103, 2019.