Naama Shoham

Mechanotransduction in adipocytes

Obesity is widely recognized as a major public health problem due to its strong association with a number of serious chronic diseases including hyperlipidemia, hypertension, type II diabetes and coronary atherosclerotic heart disease. During the development of obesity, the positive energy balance involves recruitment of new adipocytes from preadipocytes in adipose tissue, which have proliferated and differentiated. Given that cells in adipose tissues are physiologically exposed to compound mechanical loading: tensile, compressive and shear strains/stresses, which are caused by bodyweight loads as well as by weight-bearing, it is important to determine whether the adipose conversion process is influenced by mechanical stimulations. In this article we provide a comprehensive review of the experimental studies addressing mechanotransduction in adipocytes, as well as of mathematical and computational models that are useful for studying mechanotransduction in adipocytes or for quantifying the responsiveness of adipocytes to different types of mechanical loading. The new understanding that adipogenesis is influenced by mechanical stimulations has the potential to open new and important research paths, driven by mechanotransduction, to explore mechanisms as well as treatment approaches in obesity and related conditions.

Static mechanical stretching accelerates lipid production in 3T3-L1 adipocytes by activating the MEK signaling pathway.

Understanding mechanotransduction in adipocytes is important for research of obesity and related diseases. We cultured 3T3-L1 preadipocytes on elastic substrata and applied static tensile strains of 12% to the substrata while inducing differentiation. Using an image processing method, we monitored lipid production for a period of 3-4 wk. The ratio of %-lipid area per field of view (FOV) in the stretched over nonstretched cultures was significantly greater than unity (P < 0.05), reaching ∼1.8 on average starting from experimental day ∼10. The superior coverage of the FOV by lipids in the stretched cultures was due to significantly greater sizes of lipid droplets (LDs) with respect to nonstretched cultures, starting from experimental day ∼10 (P < 0.05), and due to significantly more LDs per cell between days ∼10 and ∼17 (P < 0.05). The statically stretched cells also differentiated significantly faster than the nonstretched cells within the first ∼10 days (P < 0.05). Adding peroxisome proliferator-activated receptor-γ (PPARγ) antagonist did not change these trends, as the %-lipid area per FOV in the stretched cultures that received this treatment was still significantly greater than in the nonstretched cultures without the PPARγ antagonist (14.44 ± 1.96% vs. 10.21 ± 3%; P < 0.05). Hence, the accelerated adipogenesis in the stretched cultures was not mediated through PPARγ. Nonetheless, inhibiting the MEK/MAPK signaling pathway reduced the extent of adipogenesis in the stretched cultures (13.53 ± 5.63%), bringing it to the baseline level of the nonstretched cultures without the MEK inhibitor (10.21 ± 3.07%). Our results hence demonstrate that differentiation of adipocytes can be enhanced by sustained stretching, which activates the MEK signaling pathway.

Adipocyte stiffness increases with accumulation of lipid droplets.

Adipogenesis and increase in fat tissue mass are mechanosensitive processes and hence should be influenced by the mechanical properties of adipocytes. We evaluated subcellular effective stiffnesses of adipocytes using atomic force microscopy (AFM) and interferometric phase microscopy (IPM), and we verified the empirical results using finite element (FE) simulations. In the AFM studies, we found that the mean ratio of stiffnesses of the lipid droplets (LDs) over the nucleus was 0.83 ± 0.14, from which we further evaluated the ratios of LDs over cytoplasm stiffness, as being in the range of 2.5 to 8.3. These stiffness ratios, indicating that LDs are stiffer than cytoplasm, were verified by means of FE modeling, which simulated the AFM experiments, and provided good agreement between empirical and model-predicted structural behavior. In the IPM studies, we found that LDs mechanically distort their intracellular environment, which again indicated that LDs are mechanically stiffer than the surrounding cytoplasm. Combining these empirical and simulation data together, we provide in this study evidence that adipocytes stiffen with differentiation as a result of accumulation of LDs. Our results are relevant to research of adipose-related diseases, particularly overweight and obesity, from a mechanobiology and cellular mechanics perspectives.

The mechanics of hyaluronic acid/adipic acid dihydrazide hydrogel: Towards developing a vessel for delivery of preadipocytes to native tissues

Promising treatment approaches in repairing tissue defects include implementation of regenerative medicine strategies, particularly delivery of preadipocytes to sites where adipose tissue damage needs to be repaired or where fat needs to be generated. In this study, we suggest that the injectable hyaluronic acid/adipic acid dihydrazide (HA/ADH) hydrogel may be an adipose-tissue-like material in terms of biological compatibility as well as mechanical behavior. First, we show that the hydrogel enables and supports growth, proliferation and differentiation of 3T3-L1 preadipocytes. Second, given that adipose tissue is a weight-bearing biological structure, we investigate the large deformation mechanical behavior of the hydrogel with and without embedded preadipocytes, by performing confined and unconfined compression tests and then calibrating a strain energy density (SED) function to the results. Four test groups were examined: (1) Hydrogel specimens right after the preparation without cells, (2) and (3) 3-days-cultured hydrogel specimens with and without cells, respectively, and (4) 6-days-cultured hydrogel specimens with cells. A one-term Ogden SED was found to adequately describe the hyperelastic behavior of the hydrogel specimens in all experimental groups. Importantly, we found that the mechanical properties of the hydrogel, when subjected to compression, are in good agreement with those of native adipose tissue, with the better fit occurring 3-6 days after preparation of the hydrogel. Third, computational finite element studies of the mechanical (stress-strain) behavior of the HA/ADH hydrogel when containing mature adipocytes indicated that the stiffnesses of the constructs were mildly affected by the presence of the adipocytes. Hence, we conclude that injectable HA/ADH hydrogel may serve as a vessel for protecting preadipocytes during, and at a short-term after delivery to native tissues, e.g. in research towards regenerative medicine in tissue reconstructions.

Deformations, mechanical strains and stresses across the different hierarchical scales in weight-bearing soft tissues

Sustained internal tissue loads (deformations, mechanical strains and stresses) which develop during immobile weight-bearing postures such as while in bed or in a chair were identified as a fundamental cause for the onset and progression of pressure ulcers (PUs), particularly of the deep tissue injury (DTI) type. The sustained loading may compromise tissue viability either directly, by geometrically distorting cells, or indirectly, by distorting the vasculature or lymphatic networks or, at the micro-scale, by distorting cellular organelles involved in regulating transport, e.g. the plasma membrane, since transport-control-mechanisms are essential for adequate biological function of cells. In this article we provide a comprehensive, rigorous review of the up-to-date published computational-modeling-work as well as relevant experimental studies concerning tissue deformations, strains and stresses across the different hierarchical scales: tissue-scale [cm], meso-scale [mm] and cell-scale [μm]. Viability of tissues exposed to sustained loading should be investigated in all dimensional scales, from the macro to micro, in order to provide complete understanding of the etiology of PUs and DTIs and in particular, for identifying individuals for whom and conditions at which the susceptibility to these injuries might be greater. Emerging relevant bioengineering methods of computer simulation such as multiscale and multiphysics modeling will undoubtedly contribute to the aetiological research in this field in the near future.

The influence of mechanical stretching on mitosis, growth, and adipose conversion in adipocyte cultures

The mechanotransduction of adipocytes is not well characterized in the literature. In this study, we employ stochastic modeling fitted to experiments for characterizing the influence of mechanical stretching delivered to adipocyte monolayers on the probabilities of commitment to the adipocyte lineage, mitosis, and growth after mitosis in 3T3-L1 adipocytes. We found that the probability of a cell to become committed to the adipocyte lineage in a single division when cultured on an elastic substrate was 0.025, which was indistinguishable between cultures that were radially stretched (to 12% strain) and control cultures. The probability of undergoing mitosis however was different between the groups, being 0.4 in the stretched cultures and 0.6 in the controls. The probability of growing after mitosis was affected by the stretching as well and was 0.9 and 0.8 in the stretched and control groups, respectively. We conclude that static stretching of the substrate of adipocyte cultures influences the mitotic potential of the cells as well as the growth potential post-mitosis. The present work provides better understanding of the mechanotransduction of adipocytes and in particular quantify how stretching influences the likelihood of cell proliferation and differentiation and, consequently, adipogenesis in the adipocyte cultures.

Simulating single cell experiments in mechanical testing of adipocytes

This study introduces new three-dimensional finite element cell modeling for simulating the structural, large deformation behavior of maturing adipocytes, based on empirically acquired geometrical properties of cultured adipocyte cells. We created models of adipocyte differentiation and maturation, which represented four stages along that process. The modeling focused on two specific and commonly used experimental setups, one involving compression of individual adipocytes and the other stretching of adipocytes. Both are physiological loading regimes for fat tissues and cells in vivo, and both are often employed for testing cell responses to deformations in the context of obesity and pressure ulcer research. In both simulation types, and in all the cell models, external loads induced localized effective Lagrange strains in the plasma membrane that reached maximum values over the lipid droplets (LDs). We also observed that the effective stresses (averaged across the entire cell volume in each model case) increased with cell maturation and varied between cells with different structure and dimensions. This result points to an increase in the effective cell stiffness with maturation, which would have been expected, since the volume of the stiffer LDs increases as adipocytes mature. Overall, the mechanical behavior of an individual cell is influenced not only by the external mechanical loads that are exerted, but also by the cell structure and dimensions, and is fundamental to any interpretation of cell mechanics experiments, and particularly for testing adipocytes.

Effects of accumulation of lipid droplets on load transfer between and within adipocytes

Adipogenesis, a process of cell proliferation followed by the accumulation of lipid droplets (LDs), is accompanied by morphological changes in adipocytes, leading to a gradual rise in the structural stiffness of these cells. The increase in cellular structural stiffness can potentially influence the localized deformations of adjacent adipocytes in weight-bearing fat tissues, which, based on previous work, may accelerate intracytoplasmatic lipid production to form even larger and more tightly packed intracellular LDs. This process is based on mechanotransduction phenomena which are hypothesized (again, following empirical studies), to play a critical role in “en mass” adipocyte hypertrophy, and hence are important to characterize through computational modeling. Accordingly, we examined here how maturing adipocytes may affect localized loads acting on adjacent immature cells, using a set of finite element models of adipocytes embedded in an extracellular matrix. The peak strain energy density at the plasma membrane (PM) of the adipocytes, when constructs were externally loaded, was found to depend on the levels of lipid accumulation in the neighboring cells if the external compressive and shear deformations were large enough (

Contoured Foam Cushions Cannot Provide Long-term Protection Against Pressure-Ulcers for Individuals with a Spinal Cord Injury: Modeling Studies.

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