Deep tissue injury (DTI) is a severe kind of pressure ulcers formed by sustained deformation of muscle tissues over bony prominences. As a major clinical issue, DTI affects people with physical disabilities, and is obviously related to the load-bearing capacity of muscle cells in various in vivo conditions. Oxidative stress, either induced by reperfusion immediately following tissue unloading or in chronic inflammatory conditions, may affect the cellular capacity against subsequent mechanical damages. It is important to understand the multiscale mechanobiological mechanism of DTI etiology and develop preventive approach to lower the risk of DTI.

Firstly, I measured the compressive damage threshold of C2C12 mouse myoblasts with or without pretreatment of hydrogen peroxide (H₂O₂) as an oxidative agent to understand how changes in the oxidative environment may contribute to the development of DTI. Spherical indentation was applied onto a layer of agarose gel covering a monolayer of myoblasts. The spatial profile of the measured percentage cell damage was correlated with the radially varying stress field determined by finite element (FE) analysis to estimate the compressive stress damage threshold. Results supported that chronic exposure to high-dosage oxidative stress could compromise the capability of muscle cells to withstand compressive damages, while short exposure to low-dosage oxidative stress could enhance such capability.

Secondly, I experimentally studied the effects of extrinsic H₂O₂ on the actin cytoskeletal geometry in C2C12 myoblasts. A confocal-based cell specific FE model was built to study the effects of stress fiber density, fiber cross-sectional area, tensile pre-strain in the fibers, elastic moduli of stress fibers, actin cortex, actin cytoskeleton (including both actin stress fibers and actin cortex), nucleus and cytoplasm on a myoblast’s compressive resistance. The results supported that a decrease in actin cytoskeletal elastic modulus could increase the average tensile strain on the actin cortex-membrane structure compromising the compressive resistance of a myoblast. The extent of cell damage could be estimated by a Weibull distribution function.

Thirdly, tissue compression can potentially compromise lymphatic transport and cause accumulation of metabolic biowastes, which may cause further cell damage under continuous mechanical loading. In vitro, biowastes collected from cells damaged under prolonged low compressive stress were toxic and could compromise compressive resistance of normal myoblasts. In silico, I used COMSOL to simulate the compressive stress distribution and the diffusion of biowastes in a semi-3D buttock FE model showing biowastes release would cause earlier propagation of tissue damage. This study highlighted the importance of biowastes in the development of DTI to clinical pressure ulcers under prolonged skeletal compression.

Fourthly, I found cyclic mechanical stimulation on monolayer of myoblasts could enhance viability of muscle cells, the compressive stress damage threshold of muscle cells and cell plasma membrane resealing ratio under prolonged compression. A platen indenter was applied to generate 20% strain in 0.5% agarose gel covering monolayer of myoblasts. The frequency and amplitude of the additional cyclic strain imposed on the gel was 1 Hz and 0.2% respectively for 2 hours. I propose this approach as a potential clinical measure to reduce the risk of DTI.

In the end, the limitations of this study were discussed and some future research directions were proposed.

深層組織受損是一種嚴重的壓瘡疾病，是由於骨突出而引發的肌肉組織形 變。作為一個重要臨床問題，深層組織受損困擾著有生理殘疾的人群，很明顯 也與不同體內環境中肌肉細胞的承受載荷能力有關。組織長期受壓後撤銷壓力 所帶來快速倒灌注或者長期炎症反應會引起氧化逆境，這種逆境可能會影響肌 肉細胞抵抗後續機械損傷的能力。研究深層組織受損發病機理的多尺度力生物 學機理和開發預防降低深層組織受損發病的風險是重要的。

首先，我測量了 C2C12 小鼠成肌肉細胞受氧化劑過氧化氫前處理的情況下 壓應力損傷閾值。借此來理解過氧化環境的變化在深層組織受損過程中起的作 用。球形壓頭壓入一層瓊脂糖水凝膠，膠底部培養了單層成肌肉細胞。細胞死 亡百分比的空間分佈與有限元分析計算得到的應力場做對應之後估計得壓應力 損傷閾值。結果表明慢性高劑量的氧化逆境會降低肌肉細胞對壓力損傷的抵抗 能力，然而短時間低計量的氧化逆境則可以增強抵抗能力。

其次，我利用細胞實驗研究外源性過氧化氫對成肌肉細胞細胞骨架幾何結 構的影響。利用共聚焦顯微鏡建立的細胞特異性有限元模型研究以下參數對成 肌肉細胞的抗壓應力的能力：應力纖維密度，應力纖維橫截面積，纖維中預拉 應變，應力纖維、肌動蛋白皮層、激動蛋白細胞骨架（同時包括肌動蛋白應力 纖維和肌動蛋白皮層）、細胞核、細胞質的彈性模量。結果顯示肌動蛋白細胞骨 架的彈性模量下降會減弱成肌肉細胞抵抗壓應力的能力。細胞損傷的程度可以 由韋伯分佈函數來估計。

第三，組織受壓之後可能阻礙淋巴傳輸系統導致代謝廢物的累計，從而在 持續的壓應力作用下造成進一步的細胞受損。細胞實驗表明，細胞在承受長期 低壓應力後損傷所釋放出來的生物廢料具有生物毒性，同時也會降低正常肌細 胞抵抗壓應力能力。通過仿真模擬，我利用 COMSOL 多物理場仿真軟體建立半三 維臀部有限元模型來仿真臀部的壓應力分佈以及生物廢料的自由擴散。結果顯 示生物廢料的釋放會造成組織受損更早期的擴散。此項研究突出了生物廢料在 長期骨骼的壓力作用下深層組織受損發展成臨床壓瘡中的重要性。

第四，我發現對單層成肌肉細胞的週期性力刺激可以增加在長期壓力下肌 肉細胞存活率，壓應力損傷閾值和細胞膜修復率。一個平板壓頭作用在覆蓋在 單層成肌肉細胞上的瓊脂糖水凝膠上產生 20%應變。額外施加在水凝膠上的週期 性應變的頻率和幅值分別為 1 赫茲和 0.2%，作用時間為 2 小時。我提出這個方 法作為潛在的可以降低深層組織受損發病風險的臨床手段。

最後，我做出總結，同時討論本論文中的局限性，也提出了一些未來研究 的方向。