非線性光學顯微術觀察生物組織熱致變化

Observation of Heat Propagation and Heat Induced Denaturation in Biological Tissues by Non-linear Optical Microscopy

廖建盛(Chien-Sheng Liao)

指導教授 ： 朱士維

國立臺灣大學/理學院/物理研究所/碩士(2010年)

https://doi.org/10.6342/NTU.2010.00038

摘要

熱在生物組織的重要性可由逐漸受重視的熱療法來說明。例如，透過加熱角膜引起的蛋白質收縮提供了非侵入式矯正遠視和散光的方法。分子的熱反應研究提供了對於熱療法的最佳化及減少不需要的组織傷害。另外在手術期間，提供非侵入式和高空間解析度的溫度影像也是相當重要的。在我們的研究中，我們利用由倍頻顯微術(SHGM)觀察人體的最豐富的蛋白質：第一類型膠原蛋白的熱致變化，以及利用雙光子激發螢光作葉子內部的溫度影像和對葉綠素的熱相變化的探討。
因為二倍頻與第一型膠原蛋白的三股螺旋的分子排列有很大的關係，這個非線性信號提供了在熱致變化時的分子結構的改變。在倍頻顯微術的觀察下，我們在第一型膠原蛋白熱致變化過程中發現了新的變化：de-crimp。在de-crimp過程中，由於第一型膠原蛋白纖維間的鍵結的斷裂造成纖維的自然捲曲樣式消失了，而倍頻信號強度未有改變，表示三股螺線分子仍然維持原有結構。在更高温，我們觀察收縮量與倍頻的強度變化並將熱致變化分為三個階段的變化。
溫度影像上，由於有許多的研究著重在葉子的螢光會因為本身溫度而有強度的改變，因此我們使用葉子當作試驗生物組織的溫度影像的樣本。另外，由於我們使用雙光子激發螢光，這個非線性的激發方式提供了細胞等級解析度的三維成像可能，這是目前常用的熱影像方法所不能達到的。此外透過螢光變化的熱影像提供了估計熱變化中的相變化的能力。

關鍵字

非線性光學顯微術 ； 熱力學 ； 生醫影像

並列摘要

Recently, thermal therapies have been applied to treatment of diseases increasingly. For examples, by heating cornea the induced shrinkage of collagen provides precise way to correct the hyperopia and astigmatism. Better understanding of molecular response to heat is not only of fundamental importance, but also necessary to optimize the heating strategies to reduce unwanted tissue damage. Moreover, it is helpful for surgical need to visualize distribution of temperature in the biological tissue by optical method. In our study, we observe heat induced denaturation of type 1 collagen which is the most abundant protein in human body by second harmonic generation microscopy (SHGM) and demonstrate a way to visualize distribution of temperature in the cell of plant by two photon excited microscopy.
Considering that SHG is related to the molecular packing of the triple helix in collagen fibers, this nonlinear signal provides an insight of molecular dynamics during thermal denaturation. With the aid of SHG microscopy, we found a new step in collagen thermal denaturation process, de-crimp. During the de-crimp step, the characteristic crimp pattern of collagen fascicles disappeared due to the breakage of interconnecting bonds between collagen fibrils, while SHG intensity remained unchanged, suggesting the intactness of the triple helical molecules. At higher temperature, shrinkage is observed with strongly reduced SHG intensity, indicating denaturation at the molecular level. Referring to thermal imaging heat-induced change of chlorophyll fluorescence in intact leaves has been studied actively as a result fresh leaf is a suitable biological tissue for demonstration of thermal visualization by quantifying the intensity of fluorescence. Additionally, with the advantages of non-linear optical microscopy, this technique provides optical sectioning ability to visualize temperature distribution in three dimensions in sub-μm resolution, which is not achieved by any conventional method.

並列關鍵字

Non-linear optical microscopy ； Thermal dynamics ； Medical imaging

參考文獻

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