Optical Biomarkers of Serous and Mucinous Human Ovarian Tumor Assessed with Nonlinear Optics Microscopies

Abstract

BackgroundNonlinear optical (NLO) microscopy techniques have potential to improve the early detection of epithelial ovarian cancer. In this study we showed that multimodal NLO microscopies, including two-photon excitation fluorescence (TPEF), second-harmonic generation (SHG), third-harmonic generation (THG) and fluorescence lifetime imaging microscopy (FLIM) can detect morphological and metabolic changes associated with ovarian cancer progression.Methodology/Principal FindingsWe obtained strong TPEF + SHG + THG signals from fixed samples stained with Hematoxylin & Eosin (H&E) and robust FLIM signal from fixed unstained samples. Particularly, we imaged 34 ovarian biopsies from different patients (median age, 49 years) including 5 normal ovarian tissue, 18 serous tumors and 11 mucinous tumors with the multimodal NLO platform developed in our laboratory. We have been able to distinguish adenomas, borderline, and adenocarcinomas specimens. Using a complete set of scoring methods we found significant differences in the content, distribution and organization of collagen fibrils in the stroma as well as in the morphology and fluorescence lifetime from epithelial ovarian cells.Conclusions/SignificanceNLO microscopes provide complementary information about tissue microstructure, showing distinctive patterns for serous and mucinous ovarian tumors. The results provide a basis to interpret future NLO images of ovarian tissue and lay the foundation for future in vivo optical evaluation of premature ovarian lesions.

Quantitative comparison of 3D third harmonic generation and fluorescence microscopy images

“…THG microscopy is also establishing itself as an important tool for studying intact tissue. It has been successfully applied to image unstained samples such as insect embryos, plant seeds and intact mammalian tissue [3], epithelial tissues [7][8][9], zebrafish embryos [10], zebrafish nervous system [4], and mouse brain [5]. THG is a nonlinear optical process that depends on the third-order susceptibility c (3) of the tissue and the phase-matching conditions.…”

“…Like other multi-photon techniques [10][11][12][13][14][15][16][17][18], THG not only enables the recording of label-free images of unfixed 3D volumes of tissue [1][2][3][4][5][6][7][8][9][10] free from spatial distortion artifacts inherent to histopathology, but also potentially allows feedback on the nature of the tissue, i. e. whether it is healthy or tumorous, to the surgeon during surgery, as the relative speed of the imaging modalities approaches 'real' time, and no preparation steps of the tissue are required [5,6,[19][20][21][22][23]. THG was shown recently to yield label-free images of ex-vivo human tumor tissue of histopathological quality, in real-time [6].…”

Increased metabolic activity detected by FLIM in human breast cancer cells with desmoplastic reaction: a pilot study

“…The SPC Image software (Becker & Hickl) was used to analyze the fluorescence lifetime of each metabolic molecules (NAD(P)H and FAD), as described previously by our group17 , with applied binning of 5 x 5. Data are expressed by median values and interquartile range.…”

Accumulating advantages, reducing limitations: Multimodal nonlinear imaging in biomedical sciences – The synergy of multiple contrast mechanisms

“…Particularly the differences in tissue architecture and the cellular appearance occurring during carcinogenesis are detectable for colonic cancer [126] and head and neck squamous cell carcinoma [102,125,130,131]. The same is true for visualizing the distinct morphology and characteristic structures of the inner organs and disease related modifications, e.g., for kidney tumors [114], pancreas and pancreatic cancer [132,133], bladder [134], ovary and ovarian tumors [135][136][137], and lung carcinoma [138]. In addition the structure of the musculoskeletal system has been visualized, in particular the structure of muscle [109] and tendon [34,35], but also of bones and cartilage [59,139].…”