RESUMEN
Deep tissue injury (DTI) is a life-threatening type of pressure ulcer which initiates subdermally with muscle necrosis at weight-bearing anatomical locations, where localized elevated tissue strains exist. Though it has been suggested that excessive sustained soft tissue strains might compromise cell viability, which then initiates the DTI, there is no experimental evidence to describe how specifically such a process might take place. Here, we experimentally test the hypothesis that macroscopic tissue deformations translated to cell-level deformations and in particular, to localized tensile strains in the plasma membrane (PM) of cells, increase the permeability of the PM which could disrupt vital transport processes. In order to determine whether PM permeability changes can occur due to static stretching of cells we measured the uptake of fluorescein isothiocyanate (FITC)-labeled Dextran (molecular weight = 4 kDa) by deformed vs. undeformed myoblasts, using a fluorescence-activated cell sorting (FACS) method. These PM permeability changes were then correlated with tensile strains in the PM which correspond to the levels of substrate tensile strain (STS) that were applied in the experiments. The PM strains were evaluated by means of confocal-microscopy-based cell-specific finite element (FE) modeling. The FACS studies demonstrated a statistically significant rise in the uptake of the FITC-labeled Dextran with increasing STS levels in the STS ≤ 12% domain, which thereby indicates a rise in the permeability of the PM of the myoblasts with the extent of the applied cellular deformation. The cell-specific FE modeling simulating the experiments further demonstrated that applying average PM tensile strains which exceed 3%, or, applying peak PM tensile strains over 9%, substantially increases the permeability of the PM of myoblasts to the Dextran. Moreover, the permeability of the PM grew rapidly with any further increase in PM strains, though there were no significant changes in the uptake above average and peak PM tensile strain values of 9 and 26%, respectively. These results provide an experimental basis for studying the theory that cell-level deformation-diffusion relationships may be involved in determining the tolerance of soft tissues to sustained mechanical loading, as relevant to the etiology of DTI.
Asunto(s)
Mioblastos/fisiología , Animales , Ingeniería Biomédica/instrumentación , Línea Celular , Permeabilidad de la Membrana Celular , Separación Celular , Dextranos , Análisis de Elementos Finitos , Citometría de Flujo , Fluoresceína-5-Isotiocianato/análogos & derivados , Colorantes Fluorescentes , Ratones , Microscopía Confocal , Modelos Biológicos , Úlcera por Presión/etiología , Úlcera por Presión/patología , Úlcera por Presión/fisiopatología , Estrés Mecánico , Resistencia a la TracciónRESUMEN
Many biological consequences of external mechanical loads applied to cells depend on localized cell deformations rather than on average whole-cell-body deformations. Such localized intracellular deformations are likely to depend, in turn, on the individual geometrical features of each cell, e.g., the local surface curvatures or the size of the nucleus, which always vary from one cell to another, even within the same culture. Our goal here was to characterize cell-to-cell variabilities in magnitudes and distribution patterns of localized tensile strains that develop in the plasma membrane (PM) and nuclear surface area (NSA) of compressed myoblasts, in order to identify resemblance or differences in mechanical performances across the cells. For this purpose, we utilized our previously developed confocal microscopy-based three-dimensional cell-specific finite element modeling methodology. Five different C2C12 undifferentiated cells belonging to the same culture were scanned confocally and modeled, and were then subjected to compression in the simulation setting. We calculated the average and peak tensile strains in the PM and NSA, the percentage of PM area subjected to tensile strains above certain thresholds and the coefficient of variation (COV) in average and peak strains. We found considerable COV values in tensile strains developing at the PM and NSA (up to ~35%) but small external compressive deformations induced greater variabilities in intracellular strains across cells compared to large deformations. Interestingly, the external deformations needed to cause localized PM or NSA strains exceeding each threshold were very close across the different cells. Better understanding of variabilities in mechanical performances of cells-either of the same type or of different types-is important for interpreting experimental data in any experiments involving delivery of mechanical loads to cells.
Asunto(s)
Análisis de Elementos Finitos , Fenómenos Mecánicos , Mioblastos/citología , Animales , Fenómenos Biomecánicos , Línea Celular , Membrana Celular/metabolismo , Núcleo Celular/metabolismo , Espacio Intracelular/metabolismo , Ratones , Resistencia a la TracciónRESUMEN
In the present study, we employ our recently developed confocal microscopy-based cell-specific finite element (FE) modeling method, which is suitable for large deformation analyses, to conduct inverse FE analyses aimed at determining the shear modulus of the cytoplasm of cultured skeletal myoblasts, G(cp), and its variation across a number of cells. We calibrate these cell-specific models against experimental data describing the force-deformation behavior of the same cell type, which were published by Peeters et al. (2005b) [J. Biomech.]. The G(cp) calculated for five different myoblasts were contained in the range of 0.8-2.4 kPa, with the median value being 1 kPa, the mean being 1.4 kPa, and the standard deviation being 0.7 kPa. The normalized sum of squared errors resulting from the fit between experimental and calculated force-deformation curves ranged between 0.12-0.73%, and Pearson correlations for all fits were greater than 0.99. Determining the mechanical properties of the cytoplasm through cell-specific FE will now allow calculation of cell stresses using cell-specific FE under various cell loading configurations, in support of experimental work in cellular mechanics.
Asunto(s)
Fuerza Compresiva , Citoplasma , Análisis de Elementos Finitos , Mioblastos Esqueléticos/citología , Resistencia al Corte , Estrés Mecánico , Animales , Calibración , Línea Celular , RatonesRESUMEN
This study introduces a new confocal microscopy-based three-dimensional cell-specific finite element (FE) modeling methodology for simulating cellular mechanics experiments involving large cell deformations. Three-dimensional FE models of undifferentiated skeletal muscle cells were developed by scanning C2C12 myoblasts using a confocal microscope, and then building FE model geometries from the z-stack images. Strain magnitudes and distributions in two cells were studied when the cells were subjected to compression and stretching, which are used in pressure ulcer and deep tissue injury research to induce large cell deformations. Localized plasma membrane and nuclear surface area (NSA) stretches were observed for both the cell compression and stretching simulation configurations. It was found that in order to induce large tensile strains (>5%) in the plasma membrane and NSA, one needs to apply more than approximately 15% of global cell deformation in cell compression tests, or more than approximately 3% of tensile strains in the elastic plate substrate in cell stretching experiments. Utilization of our modeling can substantially enrich experimental cellular mechanics studies in classic cell loading designs that typically involve large cell deformations, such as static and cyclic stretching, cell compression, micropipette aspiration, shear flow and hydrostatic pressure, by providing magnitudes and distributions of the localized cellular strains specific to each setup and cell type, which could then be associated with the applied stimuli.
Asunto(s)
Imagenología Tridimensional/métodos , Microscopía Confocal/métodos , Modelos Biológicos , Mioblastos/citología , Mioblastos/fisiología , Animales , Línea Celular , Tamaño de la Célula , Fuerza Compresiva/fisiología , Simulación por Computador , Módulo de Elasticidad/fisiología , Dureza/fisiología , Ratones , Estrés Mecánico , Resistencia a la Tracción/fisiologíaRESUMEN
The purposes of the present study were to (1) determine the internal plantar mechanical stresses in diabetic and healthy subjects during everyday activities, and (2) identify stress parameters potentially capable of distinguishing between diabetic and healthy subjects. A self-designed, portable, real-time and subject-specific foot load monitor which employs the Hertz contact theory was utilized to determine the internal dynamic plantar tissue stresses in 10 diabetic patients and 6 healthy subjects during free walking and outdoors stair climbing. Internal stress parameters and average stress-doses were evaluated, and the results obtained from the two groups were compared. Internal plantar stresses and averaged stress-doses during free walking and outdoors stairs climbing in the diabetic group were 2.5-5.5-fold higher than in the healthy group (p<0.001; stair climbing comparisons incorporated data from five diabetic patients). The interfacial pressures measured during free walking were slightly higher ( approximately 1.5-fold) in the diabetic group (p<0.05), but there was no significant difference between the two groups during stairs climbing. We conclude that during walking and stair climbing, internal plantar tissue stresses are considerably higher than foot-shoe interface pressures, and in diabetic patients, internal stresses substantially exceed the levels in healthy. The proposed method can be used for rating performances or design of footwear for protecting sub-dermal plantar tissues in patients who are at risk for developing foot ulcers. It may also be helpful in providing biofeedback to neuropathic diabetic patients.
Asunto(s)
Diabetes Mellitus/fisiopatología , Pie/fisiopatología , Adulto , Anciano , Fenómenos Biomecánicos , Femenino , Úlcera del Pie/fisiopatología , Humanos , Masculino , Persona de Mediana Edad , Monitoreo Fisiológico/instrumentación , Estrés Mecánico , Caminata/fisiologíaRESUMEN
Vertebral compression fractures are a potentially severe injury, which is characteristic to osteoporotic elderly. Despite being a significant healthcare problem, the etiology of compression fractures is not fully understood, and there are no biomechanical models in the literature that describe the development of these fractures based on cancellous bone failure accumulation. The objective of this study was therefore to develop a computational model of tissue-level failure accumulation in vertebral cancellous bone, which eventually leads to compression fractures. The model predicts the accumulated percentage of broken trabeculae delta in a vertebral region of interest (ROI) over 60 years, by employing Euler's theory for elastic buckling. The accumulated failure delta is calculated as function of the daily activity characteristics and rate of annual bone loss (RABL) with aging. An RABL of unity represents the normal bone loss attributed to aging per se, whereas RABL>1 is assumed to represent pathological bone metabolism such as osteoporosis. Simulations were conducted for a range of RABLs, to determine the effect of changes in bone metabolism on the accumulation of bone failure. Results showed that bone failure rapidly increased with RABL. Generally, trabecular failure was shown to become more severe for RABL>4. Total failure was exhibited at RABL=7.5 for the central ROI, and at RABL=8.5 for the sub-endplate ROI. We concluded that vertebral compression fractures advance monotonically between the age of 50-55 years and 70 years, and may accelerate thereafter if RABL is high (~8). Additionally, the model identified weight lifting as the action that most dramatically accelerated the destruction of osteoporotic spinal cancellous bone. The present biomechanical model is useful for understanding the etiology of compression fractures, and potentially, depending on further experimental characterization of RABL, for considering the effects of medications that influence bone metabolism on patient prognosis.