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Title: | 透過聲穿孔作用輔助藥物遞送並以調節腫瘤微環境硬度的方式增加放射敏感性 Sonoporation Assisted Tumor Microenvironment Stiffness Modulation for Enhanced Radiosensitivity in Radiation Therapy |
Authors: | 裴煜 YU PEI |
Advisor: | 李百祺 Pai-Chi Li |
Keyword: | 放射治療,腫瘤微環境,細胞外基質硬度,放射增敏劑,三維細胞培養系統,剪切波彈性影像,順鉑,LOX抑制劑,聲穿孔作用, Radiotherapy,tumor microenvironment,extracellular matrix stiffness,radiosensitizer,three-dimensional cell culture system,shear wave elasticity imaging,cisplatin,LOX inhibitor,sonoporation, |
Publication Year : | 2024 |
Degree: | 碩士 |
Abstract: | 放射治療(RT)作為臨床上治療癌症最主要的方式,如何最大化放射線對腫瘤組織的損傷並同時將對正常組織的副作用降到最低是目前主要研究的課題。近年來研究發現腫瘤微環境也是影響腫瘤行為和對治療的抗性的關鍵因素之一,細胞外基質(ECM)的重塑和過度沉積被證實會導致放射抗性以及耐藥性。細胞外基質的重塑和沉積在臨床上能夠以硬度的變化被觀察到,但腫瘤的硬度變化對治療效果的影響仍無法量化。臨床上廣泛使用化學藥物作為放射增敏劑,透過前處理造成DNA損傷來增強放射治療對癌細胞的效果,但由於化療藥物多半是全身給藥,所以其濃度和對正常細胞的損傷仍須探討。而微氣泡結合超音波造成的聲穿孔作用被發現可以增加細胞膜的通透性進而提升藥物和放射的敏感性及靶向性。近年來也發現DNA損傷和氧化應激可能對細胞外基質的重塑也有一定的影響,各種因素所導致的硬度變化對癌症治療的影響仍有待研究。為了瞭解硬度對治療的影響,首先最重要的是如何準確地測量硬度大小,本研究使用了毫米級三維細胞培養系統,並透過高頻單探頭超音波的剪切波彈性影像平台實現對毫米級體外模型的量測。固定探頭與邊界的距離預測得出樣品的剪切波速度。利用剪切波彈性影像的非侵入性在不損壞樣本的情況下對模型進行長期觀測,並進一步探討其細胞死亡率和DNA損傷等生物特性。實驗上使用臨床中與硬度有顯著關聯性的肝癌細胞Huh7,並透過改變膠原蛋白的濃度來模擬三種不同的起始硬度,藥物治療的部分嘗試了細胞毒性藥物(順鉑, cisplatin) 和細胞外基質靶向藥物(LOX抑制劑, BAPN )在成膠前進行24小時的前處理,聲穿孔處理也是在成膠前24小時作用,而放射治療的部分則是在成膠後24小時照射16Gy的劑量,最後培養96小時探討治療後硬度和生物特性的改變。放射治療在較軟的環境中造成的DNA損傷程度較高,同樣的現象也在順鉑或LOX抑制劑前處理結合放射治療的樣品中被發現,透過比較樣品的硬度變化可以推測DNA損傷可能促進了細胞外基質的重塑。藥物結合放射治療的部分,在順鉑前處理結合放射治療的聯合治療結果可以發現在越軟的起始環境下進行順鉑前處理,放射治療誘導細胞死亡的效果越好,達到放射增敏的效果,而LOX抑制劑在越硬的起始環境下的影響越大,抑制了細胞外基質重塑使DNA修復的速度降低,並且在較硬的初始環境下結合放射治療可以達到與在中等硬度做單純放射治療相似的效果。最後聲穿孔處理的部分,聲穿孔處理後的硬度皆有明顯的下降,藥物治療可以達到更高劑量的效果,結合放射治療後的效果也都有明顯的提升。本研究透過三維細胞培養系統結合剪切波彈性影像可以直接觀測細胞外基質硬度與生物特性的變化,觀察造成DNA損傷的細胞毒性藥物和細胞外基質靶向藥物結合放射治療和聲穿孔處理後初始硬度和隨時間硬度的變化,成功量化硬度對癌症治療的影響,了解腫瘤微環境硬度和放射/化學治療的相互作用,並成功開發了針對細胞外基質靶向的放射治療方式,有助於未來開發更有效的癌症治療方法。 Radiotherapy (RT) is currently the primary clinical strategy for cancer treatment. Maximizing the damage to tumor tissues with minimal side effects is the main challenge of current research. The tumor microenvironment plays a crucial role in tumor behavior and resistance to treatment. Chemotherapeutic agents are widely used in clinical practice to enhance the efficacy of RT. However, since most chemo-drugs are administered systemically, their concentration and damage to normal cells need further investigation. Sonoporation has been found to enhance the sensitivity to drugs and radiation. It has been discovered that DNA damage and oxidative stress may also have certain effects on the extracellular matrix (ECM) remodeling. The impact of stiffness changes caused by various factors on cancer treatment is still under investigation. This study developed a millimeter-scale 3D cell culture system and used the Shear Wave Elasticity Imaging (SWEI) to measure the in vitro model. Specifically, the SWEI is based on the speed of the reflected shear waves measured by a single-element ultrasound transducer. The non-invasive nature of SWEI allowed us to perform long-term observations of the 3D cell culture system without damaging the samples and further explore their biological characteristics through flow cytometry. Three different initial stiffness levels were simulated by changing the concentration of collagen. The changes in stiffness and biological characteristics were examined after 96 hours of cultivation. The study found that RT caused higher levels of DNA damage in softer environments. It was suggested that DNA damage might promote ECM remodeling. The results of the combined treatment with cisplatin pretreatment and RT show that the induction of cell death by RT is more effective in softer initial stiffness and achieves radiosensitization. On the other hand, the effect of BAPN was more significant in harder starting environments, inhibiting ECM remodeling and reducing the rate of DNA repair. Combining RT with BAPN in a stiffer initial environment produced similar effects to those of single RT in medium stiffness. Finally, in the study of sonoporation treatment, there was a significant decrease in stiffness after sonoporation. Drug treatments achieved higher dose effects, and the combination with RT showed a high enhancement. The study successfully quantified stiffness''s impact on cancer treatment using a 3D cell culture system combined with SWEI. The study developed a targeted RT approach for the ECM, contributing to developing more effective cancer treatments and understanding the interplay between tumor stiffness and radio/chemotherapy. |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/91641 |
DOI: | 10.6342/NTU202400215 |
Fulltext Rights: | 同意授權(限校園內公開) |
Appears in Collections: | 生醫電子與資訊學研究所 |
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