RESUMEN
The alveolar-capillary barrier is composed of epithelial and endothelial cells interacting across a fibrous extracelluar matrix (ECM). Although remodeling of the ECM occurs during several lung disorders, it is not known how fiber structure and mechanics influences cell injury during cyclic airway reopening as occurs during mechanical ventilation (atelectrauma). We have developed a novel in vitro platform that mimics the micro/nano-scale architecture of the alveolar microenvironment and have used this system to investigate how ECM microstructural properties influence epithelial cell injury during airway reopening. In addition to epithelial-endothelial interactions, our platform accounts for the fibrous topography of the basal membrane and allows for easy modulation of fiber size/diameter, density and stiffness. Results indicate that fiber stiffness and topography significantly influence epithelial/endothelial barrier function where increased fiber stiffness/density resulted in altered cytoskeletal structure, increased tight junction (TJ) formation and reduced barrier permeability. However, cells on rigid/dense fibers were also more susceptible to injury during airway reopening. These results indicate that changes in the mechanics and architecture of the lung microenvironment can significantly alter cell function and injury and demonstrate the importance of implementing in vitro models that more closely resemble the natural conditions of the lung microenvironment.
Asunto(s)
Barrera Alveolocapilar/fisiología , Matriz Extracelular/metabolismo , Técnicas In Vitro , Microtecnología , Atelectasia Pulmonar/etiología , Atelectasia Pulmonar/metabolismo , Actinas/metabolismo , Células Epiteliales Alveolares/metabolismo , Materiales Biomiméticos , Células Cultivadas , Citoesqueleto/metabolismo , Células Endoteliales/metabolismo , Humanos , Microtecnología/instrumentación , Microtecnología/métodos , Atelectasia Pulmonar/patología , Uniones EstrechasRESUMEN
Personalized cancer therapies drive the need for devices that rapidly and accurately segregate cancer cells from solid tumors. One potential sorting strategy is to segregate populations of cells based on their relative strength of adhesion. To investigate the effect of surface hydrophilicity and cell phenotype on adhesion, primary human breast skin fibroblasts and keratinocytes and MCF-7 breast cancer cells were seeded onto air and CF(4) plasma-treated nanofibers followed by exposure to three shear stresses (200, 275 and 350 dynes per cm(2)) 1 hour after inoculation. No difference in strength of adhesion was measured in either fibroblasts or keratinocytes on either plasma treated-surface: all exhibited >60% of the initial cell count after a 5 minute exposure to 350 dynes per cm(2) of shear stress. In contrast, a significant difference between relative strength of adhesion on air versus CF(4) plasma-treated surfaces was observed for MCF-7 cells: 26% and 6.6% of cells remained on the air and CF(4) plasma-treated surfaces, respectively. The ability to sort this cancer cell line from two non-cancerous primary human cells was evaluated by inoculating a mixture of all three cell types simultaneously onto CF(4) treated nanofibers followed by 1 hour of culture and exposure to 350 dynes per cm(2) shear stress. The majority of MCF-7 cells were removed (0.7% remained) while a majority of fibroblasts and keratinocytes remained adhered (74 and 57%). Post-sorted MCF-7 viability and morphology remained unchanged, preserving the possibility of post-separation and analysis. These data suggest that the plasma treatment of electrospun scaffolds provides a tool useful in sorting cancer cells from a mixed cell population based on adhesion strength.
Asunto(s)
Neoplasias de la Mama/patología , Adhesión Celular/fisiología , Fluorocarburos/química , Nanofibras/química , Neoplasias de la Mama/ultraestructura , Supervivencia Celular/fisiología , Femenino , Humanos , Células MCF-7 , Microfluídica , Microscopía Electrónica de Rastreo , Microscopía de Contraste de Fase , Propiedades de SuperficieRESUMEN
A compliant terpolymer made of hexylmethacrylate (HMA), methylmethacrylate (MMA), and methacrylic acid (MAA) intended for use in small diameter vascular graft applications has been developed. The mechanical properties and in vitro biostability of this terpolymer have been previously characterized. The goal of this investigation was to examine the interactions between endothelial cells and the new terpolymer and to evaluate endothelial cell function. Electrospinning was used to produce both oriented and random terpolymer fiber scaffolds. Smooth solution cast films and tissue culture polystyrene were used as negative and positive controls, respectively. Human blood outgrowth endothelial cells and human umbilical vein endothelial cells were incubated with the test and control samples and characterized with respect to initial cell attachment, proliferation, viability, and maintenance of the endothelial cell phenotype. It was found that the terpolymer is cytocompatible allowing endothelial cell growth, with random fibers being more effective in promoting enhanced cellular activities than oriented fibers. In addition, endothelial cells cultured on these substrates appeared to maintain their phenotype. The results from this study demonstrate that electrospun HMA:MMA:MAA terpolymer has the potential to be used successfully in fabricating small diameter blood vessel replacements.
Asunto(s)
Prótesis Vascular , Comunicación Celular/efectos de los fármacos , Células Endoteliales/citología , Células Endoteliales/efectos de los fármacos , Metacrilatos/farmacología , Polímeros/farmacología , Ingeniería de Tejidos/métodos , Adhesión Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Células Cultivadas , Células Endoteliales/metabolismo , Humanos , Microscopía Electrónica de Rastreo , Porosidad/efectos de los fármacos , Factor de von Willebrand/metabolismoRESUMEN
OBJECTIVES: This laboratory study was designed to investigate the effect of controlled nanoporosity on the wear resistance of polymeric composites reinforced with silica gel powders and to determine the mechanisms controlling the abrasive wear properties of these unique nanostructured materials. METHODS: Silica gels were prepared by hydrolysis and condensation of tetraethylorthosilicate (TEOS) using four different catalysts to modify the porous structure of the resulting polysilicate silanation, an organic monomer (TEGDMA) containing various initiators was introduced into the gel powders to form a paste. The various pastes were then polymerized inside a glass mold. A pin-on-disk apparatus was then used to record the specimen length and number of revolutions. Abrasive wear rates were determined by regression analysis and statistical differences were determined by analysis of variance and multiple comparisons. BET was used to characterize the filler pore structure and scanning electron microscopy was used used to visually examine the abraded surfaces. RESULTS: Significant differences (p < 0.05) in the wear rates of the experimental composites were noted. Within the range of filler porosities examined, wear resistance was found to be linearly dependent (R2 = 0.983) on filler pore volume. The wear rates decreased with increasing filler porosity. HCl-catalyzed gels having low porosity produced composites having relatively limited abrasion resistance. In contrast, high porosity HF-catalyzed gels produced more wear-resistant composites. The abrasive wear resistance of these nanocomposites was not significantly affected by the level of silane coupling used in these experiments. SEM evaluation suggested that better wear resistance was associated with fine-scale plastic deformation of the wear surface and the absence of filler particle pullout. SIGNIFICANCE: Porous particles prepared via sol-gel show some promise as fillers that improve the wear resistance of photopolymerized resins. The wear resistance of the fillers appears to be directly related to nanoporous structure of the gel particles. Unlike conventional dental composites, these materials rely primarily on nanomechanical coupling for improved wear resistance. This new principle should benefit subsequent investigations.