乳腺组织的拉曼光谱线性模型研究

本研究甄选10个代表乳腺组织拉曼活性成分的基谱,在国内首次构建一种乳腺组织拉曼谱的线性回归模型。用2 000多个正常和非正常乳腺组织拉曼谱对该模型进行统计检验,模型显著性F检验的置信度全部为1,多元决定系数的平均值为0.95,表明线性模型假设成立、拟合效果良好。10个基谱代表脂肪、细胞质、细胞间质、DNA、血液、β-胡萝卜素、胆固醇等的拉曼谱,基谱的归一化拟合系数间接反映出这些成分的相对含量。用该模型拟合正常和肿瘤乳腺组织的拉曼宏观谱,病变前后细胞质和DNA的拟合系数增大、脂肪拟合系数减小,这反映出它们相对含量的增减,与已知的病理学结果一致。该研究有助于理解乳腺肿瘤组织的生化变化,并可能为分析该变化提供了有效手段。     

人体肿瘤组织的拉曼光谱相关系数成像

运用逐点扫描(Scanning)采谱技术,测量了人体乳腺肿瘤组织的阵列拉曼光谱集(Mapping);利用自编的软件,分析和计算了该谱集与参考谱DNA、细胞质等拉曼谱的相关系数;并将其映射为灰度值,构建了点阵的相关系数成像。结果表明,相关系数成像较好地反映了参考谱所代表的生化成分的分布;DNA谱与组织中其他成分谱的相关性很小,因此DNA谱的相关系数成像能够直接地反映成像区域DNA的分布,可信度尤其高。该研究结果为进一步研究乳腺肿瘤的拉曼诊断方法奠定了一定基础。     

乳腺组织形态基元共焦显微拉曼光谱的研究

利用共焦显微拉曼方法,测定了乳腺组织切片的Mapping拉曼谱,光谱采样体积达到μm尺度.通过对这些共焦拉曼谱与商业提纯肌动蛋白、胶原等生化物质做出的谱的分析比较,分离出了来自细胞质、细胞间质、脂肪以及乳腺沉积物这些组织形态基元的拉曼谱;用K阶簇分析方法,还获得了细胞核基元谱;基元谱与提纯化学物质谱的相关分析印证了组织形态基元谱的来源.还分析总结了这些组织形态基元谱的特征.这些工作为深入理解组织形态与其拉曼谱的关系以及乳腺组织形态拉曼模型的建立奠定了基础.     

乳腺癌病人单个细胞的Raman光谱

给出了乳腺癌病人正常乳腺细胞与癌细胞的拉曼光谱。从实验谱线中得到：癌变细胞的拉曼谱整体变弱, 就相对强度而言, DNA的2个磷酸根骨架峰782,1 084 cm-1和脱氧核糖-磷酸振动峰1 155及1 262 cm-1拉曼谱线明显的减少; 表征A型(DNA)构像的特征峰812 cm-1及谱线979,668 cm-1消失, 并有新峰1 175 cm-1出现, 905 cm-1的谱线增强并有6 cm-1的红移, 这说明DNA的磷酸根骨架有一定的断裂; 从而导致癌细胞的分裂繁殖失去有效的控制。在癌变组织细胞的拉曼谱中还发现了很强的一类与钙硬化密切相关的特征峰960 cm-1; 通过这些特征拉曼谱线的变化, 为癌症的诊断和治疗提供了有力的实验依据。     

基于基底细胞癌的太赫兹波光谱分析

获得了基底细胞癌变组织的太赫兹(THz)光谱和图像,并进行了组织病理学的分析,采用太赫兹时域光谱技术,对石蜡包埋的基底细胞癌组织进行透射式实验研究,获得了透过率和吸收系数的光谱,并利用癌变组织和健康组织区分度明显的1.3THz的吸收系数进行成像得到了与宏观组织较符合的THz图像.从组织病理学的角度阐述了太赫兹谱线及图像的成因,结合生物大分子的THz光谱,分别讨论了细胞密度、DNA的含量、水化作用对太赫兹光谱和成像的影响.实验结果表明:癌变组织与健康组织相比对THz吸收较多.太赫兹光谱和成像技术有望用于术前基底细胞癌的轮廓诊断.     

肺正常组织与癌变组织的表面增强拉曼光谱

采用共焦显微拉曼光谱进行肺正常组织与癌变组织的表面增强拉曼光谱测试,拉曼位移测试范围为300～1700cm-1 。根据得到的肺正常和癌变组织的表面增强拉曼光谱,对比分析了两者的特点与差异。选取蛋白质主链构象中构象不灵敏的CH₂弯曲振动在1454cm-1的谱线为内标,发现相比肺正常组织,癌变组织蛋白质主链中酰胺Ⅲ和酰胺Ⅰ谱带的有序构象明显减少,对α螺旋和无规则卷曲的损伤比较严重,β回折几乎消失,而骨架C—N,C—C伸缩振动却出现了与前2个谱带截然相反的状况,有序构象显著增加,蛋白质侧链构象变化与主链相比则相对复杂;DNA的骨架磷酸基团、脱氧核糖、磷酸-脱氧核糖和碱基的含量总体呈增加趋势;RNA的情况与DNA类似;磷脂的链内纵向有序性参数明显下降。     

乳腺肿瘤周边组织的拉曼光谱研究

使用REMSHAW显微共焦拉曼光谱仪测量了40例手术切除乳腺肿瘤周边(肿块边约5 mm)组织的拉曼光谱.通过对浸润性导管癌、乳腺增生、纤维腺瘤等乳腺肿瘤周边组织的拉曼光谱分析,发现不同乳腺肿瘤周边组织的拉曼光谱有显著差异.这些差异在肿瘤诊断中,可作为乳腺癌和其他乳腺肿瘤拉曼光谱的特征,为乳腺疾病诊断提供依据.乳腺肿瘤周边组织的拉曼光谱对乳腺肿瘤诊断具有重要的价值.     

菲涅尔区域的乳腺三维超声成像技术*

本文提出了一种基于菲涅尔区域的乳腺三维超声成像技术,可实现病变组织早期的定性诊断与定量监测。主要研究内容包括：采用透射方式,以慢度作为反演参量,将正演与反演的透射时间差作为迭代判据,基于菲涅尔区域法实现乳腺组织的二维反演成像;根据柱面传感阵列的分布特点,将乳腺三维超声成像问题转为二维成像问题,进而实现乳腺三维超声成像;计算三维病变成像声速及尺寸误差,并分析了菲涅尔区域法的成像分辨率以及入射频率对成像分辨率的影响。本文研究结果证明基于柱面传感阵列的菲涅尔区域法可实现乳腺三维超声成像并有效实现病变早期监测与诊断。     

光敏剂CPD4对肿瘤组织DNA损伤的拉曼光谱研究

观测了小牛胸腺DNA、腹水型S_(180)肿瘤组织DNA以及光敏剂CPD_4对肿瘤组织DNA损伤的拉曼光谱。肿瘤组织DNA是分别从注射了光敏剂和生理盐水的荷瘤小鼠的肿瘤中提取的。实验结果表明,肿瘤组织DNA的拉曼光谱与正常组织DNA的有所差异,最明显的是脱氧核糖的特征谱线在正常组织DNA拉曼光谱中出现而在肿瘤组织DNA中却几乎消失或变为很弱的小峰。这些差异可作为肿瘤早期诊断的参考。实验结果还表明,光敏剂对肿瘤组织DNA各碱基产生不同程度的破坏,在DNA修复酶的作用下,可使脱氧核糖和鸟嘌呤得到修复。     

基于拉曼光谱技术的甲状腺疾病检测的研究

基于光学成像与光谱技术的无损检测是生物医学光学交叉领域研究的重要发展方向。其中拉曼光谱技术可获得检测对象的生化成分的“指纹信息”,被广泛应用于面向生物分子,细胞以及生物组织的检测诊断研究。甲状腺疾病尤其肿瘤的临床检测往往涉及多方法和技术手段的结合,且存在一定的诊断难度,因此发展新的检测技术方法具有重要的意义。首先综述了拉曼光谱技术在甲状腺细胞系的单细胞拉曼光谱检测与分析,然后介绍甲状腺病理组织和甲状腺正常组织的拉曼光谱鉴别诊断(特别介绍了本研究小组开展以银纳米粒子为增强基底的甲状腺离体组织SERS光谱研究情况),以及拉曼光谱技术在甲状腺激素等方面的研究概况。最后简要探讨了拉曼光谱技术在该领域的研究应用前景和发展方向。     

Comparison of molecular images as defined by Raman spectra between normal mucosa and squamous cell carcinoma in the oral cavity

Raman spectroscopy has been effectively applied to clinically differentiate normal and cancerous mucosal tissues. Micro‐Raman spectroscopy provides a tool to better understand the molecular basis for the Raman clinical signal. The objective of the current study was to utilize micro‐Raman spectroscopy to define the molecular/spectral differences between normal and abnormal squamous cell carcinoma (SCC) in oral mucosa (in vitro). Understanding this may help in identifying unique spectra or may be useful for in vivo application of this technology. Micro‐Raman (confocal) spectroscopy was used to obtain molecular images of normal and SCC cells of human oral mucosa. Four fresh flashed‐frozen tumor and four matched normal tongue specimens were studied. The spectra covered a wavenumber range from 300 to 4000 cm⁻¹ with a spectral resolution of 8 cm⁻¹ and a spatial resolution of 1.0 µm. The cells were located within thin sections of tongue mucosa biopsies. The excitation wavelength of 515 nm was used. We were able to obtain Raman images with rich information about the spectroscopic and structural features within the cytoplasm, cell membrane, and cell nuclei. Significant spectral differences were observed between the Raman images of normal and malignant squamous cells. The heterogeneity of tumor cells within the abnormal tissue was also demonstrated. Spectral differences demonstrated between both tissue types have provided important information regarding the origins of specific signals within the cells of each tissue type. In our search for specific spectral biomarkers, we believe that a cell surface protein, greatly upregulated in SCC cells, was discovered at 1583 cm⁻¹. Copyright © 2010 John Wiley & Sons, Ltd.     

傅里叶变换红外光谱用于胆囊癌细胞及其细胞核结构的基础研究

应用傅里叶变换红外光谱(FTIR)对体外培养胆囊癌细胞、细胞核进行检测,对照胆囊癌组织光谱特征,研究胆囊癌细胞株及细胞核的光谱表现;为红外光谱法诊断胆囊癌奠定细胞学基础。应用美国Nicolet(尼高力)公司5700-Ⅱ型红外光谱仪,组织标本放置于OMNIC采样器表面进行检测,记录红外光谱;体外培养胆囊癌细胞株(GBC-SD)及提取的细胞核涂于欧米采样器表面进行红外检测,记录红外光谱,得到时间轴上体外培养细胞株及细胞核的光谱图;选择特征性光谱与胆囊癌组织光谱进行比对。结果发现胆囊癌细胞株红外光谱特征与相应癌组织光谱特征存在异同。从而可得出结论：将体外培养肿瘤细胞株及提取的肿瘤细胞核,进行FTIR测定是行之有效的细胞红外光谱检测手段,能得到体外培养细胞的红外谱图;胆囊癌组织所表现的红外光谱特征具有胆囊癌细胞本身的红外光谱特征,同时也具有组织自身的复杂特点;FTIR为应用于胆囊癌的定性诊断提供了更加丰富的细胞学基础。     

Research on the Raman spectral character and diagnostic value of squamous cell carcinoma of oral mucosa

Early diagnosis of oral carcinomas is essential for successful treatment. The purpose of the present study is to apply near‐infrared Raman spectroscopy to detect oral squamous cell carcinoma (SCC) and leukoplakia (OLK), in order to establish the diagnostic model of the Raman spectra of oral diseases. We collected Raman spectra of normal, OLK and SCC by near‐infrared Fourier transform Raman spectroscopy. The biochemical variations between different lesions were analyzed by the characteristic bands in the subtracted mean spectra. Gaussian radial basis function support vector machines (SVM) were used to classify spectra and establish the diagnostic models. Major differences were observed in the range between 800 and 1800 cm⁻¹. Compared with normal mucosa, high contents of protein, DNA and lipid in SCC and OLK were observed, but the difference between OLK and normal tissue was not as much as that between normal and SCC. SVM displayed a powerful ability in the classifying of normal and SCC, and the specificity, sensitivity and accuracy were 100, 97.56 and 98.75%, respectively. In discriminating between the OLK and normal groups, the three parameters were 85, 68 and 72.5%. The algorithm showed good ability in grouping and modeling of OLK and SCC, and the three parameters were 95, 97.43 and 96.25%. Combined with SVM, near‐infrared Raman spectroscopy can detect biochemical variations in oral normal mucosa, OLK and SCC, and establish diagnostic models accurately. Copyright © 2009 John Wiley & Sons, Ltd.     

内吞金纳米粒子的鼻咽癌细胞SERS光谱

采用内吞方法将金纳米粒子引入细胞内,测试分析单个活性CNE-1鼻咽癌细胞的常规拉曼光谱和SERS光谱,并对其进行初步谱峰归属。CNE-1细胞的常规拉曼光谱有6个主要的拉曼峰:718,1001,1123,1336,1446和1660cm-1;沉积于细胞内的金纳米粒子强烈地增强了细胞内生化物质拉曼信号,在内吞金纳米粒子的CNE-1细胞的拉曼光谱中出现了20多个SERS拉曼信号,主要拉曼峰的强度明显高于常规拉曼信号。DNA骨架振动(1026,1097,1336和1585cm-1)证明金纳米粒子通过内吞作用而进入细胞核内。结果表明,基于胶体金SERS技术可能为活性鼻咽癌细胞内生化物质的探测提供一种高灵敏的方法。     

Raman spectroscopic analysis of breast cancer tissues: identifying differences between normal,invasive ductal carcinoma and ductal carcinoma in situ of the breast tissue

A relatively non‐destructive method employing Raman spectroscopy for the analysis of histopathological specimens is described. Raman spectroscopy has allowed qualitative analysis of the same specimen used for histopathological evaluation. Breast cancer tissues have been analysed to demonstrate the feasibility of the chemical changes taking place in the biological tissue, which can be identified precisely, and the results are reproducible. Raman analysis of tissue sections provides distinct spectra that can be used to distinguish between the nuclear grades of ductal carcinoma in situ (DCIS) and invasive ductal carcinoma (IDC) of the breast. Sixty cases of breast carcinoma including DCIS and IDC and seven cases of normal breast tissues were studied employing the Raman spectroscopic technique. This study reports for the first time spectral differences between DCIS grades. It is concluded that Raman spectroscopy can objectively distinguish between DCIS and IDC grades and is non‐destructive and reproducible. It should become possible in future to use Raman spectroscopy as an informative and quantitative method suitable for classification of grades and diagnosis of breast carcinoma. Copyright © 2007 John Wiley & Sons, Ltd.     

Comparison of connective tissue invaded by Lewis lung carcinoma to healthy connective tissue by means of micro‐Raman spectroscopy

We have measured the micro‐Raman spectra of mouse tissues invaded by Lewis lung carcinoma (LLC). We have also carried out categorical principal component analysis (CATPCA) on the acquired spectra. The results indicate that the tumor tissues can be well discriminated from normal tissues by the first two principal components extracted from the spectra. Furthermore, we have found that the concentrations of nucleic acids and lipids/fatty acids in the tumor are considerably higher than those in the normal tissue, whereas the collagen concentration is lower. These differences can be detected and characterized by Raman images using the 788 cm⁻¹ DNA/RNA band and the 1301 cm⁻¹ lipid/fatty acid band. Copyright © 2009 John Wiley & Sons, Ltd.