Poster Q7 - Compression Stiffening in Adipose Tissues

Introduction: Mechanical cues from the extracellular matrix (ECM) are crucial for controlling cell behaviors and regulating tissue functions. These cues include mechanical stresses, which directly deform cells and tissues, and mechanical properties such as stiffness, viscosity, and poroelasticity. While studies have predominantly examined the effects of individual and static mechanical cues, physiological and pathological conditions often involve multiple dynamic cues simultaneously. For instance, adipose tissues in the human body are constantly subjected to mechanical forces, whether during sitting, lying on the back or front, especially pronounced in individuals with obesity, where gravity significantly contributes to these forces. These forces, predominantly compressive, not only produce mechanical stresses deforming the cells but also dynamically alter tissue mechanical properties. However, the collective impact of these mechanical cues on adipose cells and tissue functions remains inadequately explored, partly due to a lack of comprehensive understanding of tissue mechanical characteristics and behaviors. Here, we investigate the mechanical response of adipose tissues to physiologically relevant compressions. We first reveal a relaxation of compressive stress upon compression, signifying mechanical adaptations and changes in tissue structure. Notably, we observe a significant increase in tissue rigidity during compressions that are comparable to forces experienced during sitting. Moreover, this compression-induced stiffening varies between adipose tissue types, with high-fat diet conditions showing greater stiffening. Overall, our study underscores the importance of considering dynamically combined rigidity and stress as mechanical cues to elucidate matrix effects on cellular activities and tissue functions.

Materials and

Methods: In this study we utilize white adipose tissue (WAT) from male mice as the model system. We examine the mechanical properties of two distinct adipose tissue types: epididymal adipose tissue (eWAT), representing visceral adipose tissue, and inguinal adipose tissue (iWAT), representing subcutaneous adipose tissue. Our experiments incorporate both control lean mice and high-fat diet mice, characterized by a doubling of body weight on average, to represent both healthy and obesity conditions. High-fat diet mice showed substantial increases in both eWAT and iWAT mass, with eWAT expanding sixfold and iWAT tenfold compared to those in control mice.
The mechanical properties of the adipose tissues are determined via rheometry using a stress-controlled Kinexus rheometer. The top geometry plate applies both normal stresses and shear oscillations, enabling axial compressions and shear modulus measurements (Fig. 1A). WAT samples are prepared by being cut into 8 mm size cylinders to fit the two parallel plates. To eliminate the slipping between the sample and the plates, we apply tissue glue on the top surface and use sandpaper on the bottom plate. All measurements are done at 37℃ with humidity. Normal compressions are applied from 0 to 60% step-by-step, with 10% each step. Subsequent compression is only applied when the stress from the previous compression plateaus. The compression direction is perpendicular to the original skin direction. All shear oscillations are with 1% shear strain in 1 Hz frequency.

Results, Conclusions, and Discussions: We first determine stress relaxation over time upon normal compressions (Fig. 1B and C), observing that tissues from high-fat diet mice exhibit higher normal stresses, particularly at larger compression levels. Notably, the maximal stress observed (~40 kPa) is equivalent to the stress experienced by individuals while sitting.
From the normal stress and corresponding compression levels, we next calculate the apparent Young’s moduli. After each compression, the Young’s modulus increases dramatically and then relaxes with time (Fig. 2A and B). The equilibrium Young’s modulus under different compression levels (Fig. 2C) suggests that upon 60% compression, high-fat diet tissues stiffen by ~10x, whereas controls only by ~5x.
Simultaneously, shear oscillation is applied to measure the shear moduli of the tissues. Initially, without compression, the four types of tissues exhibit similar elastic shear moduli (Fig. 3A). However, under normal compressions, their shear moduli begin to diverge. Similar to Young’s modulus, there is a dynamic relaxation response in each shear modulus under each compression (Fig. 3B and C). Notably, control lean mice show ~10x increase in shear modulus for both eWAT and iWAT, while high-fat diet mice show increases of ~20x for eWAT and ~35x for iWAT (Fig. 3D). Given that individuals with obesity experience elevated levels of compression on their adipose tissues, due to both increased whole-body weight and greater adipose tissue gravity, it is likely that their tissues exhibit higher stiffness.
Further analysis correlates the equilibrated shear modulus and Young’s modulus from all compressions (Fig. 4A), suggesting that eWAT is generally incompressible, but iWAT may undergo water or fat oil flow during large compressions. Additionally, the linear relationship between shear modulus and normal stress (Fig. 4B and C) indicates potential mechanisms for compressional stiffening.
In summary, our research demonstrates that adipose tissues undergo significant stiffening under physiological compression forces, with larger effects observed in obesity-mimicking conditions. These findings highlight the significance of incorporating both rigidity and stress as mechanical cues to accurately study the role of ECM in regulating cell and tissue functions. This is especially crucial in conditions like obesity where multiple mechanical cues are substantially altered.

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