Recognition of serous ovarian tumors in human samples by multimodal nonlinear optical microscopy

Abstract

We used a multimodal nonlinear optics microscopy, specifically two-photon excited fluorescence (TPEF), second and third harmonic generation (SHG∕THG) microscopies, to observe pathological conditions of ovarian tissues obtained from human samples. We show that strong TPEF + SHG + THG signals can be obtained in fixed samples stained with hematoxylin and eosin (H&E) stored for a very long time, and that H&E staining enhanced the THG signal. We then used the multimodal TPEF-SHG-THG microscopies in a stored file of H&E stained samples of human ovarian cancer to obtain complementary information about the epithelium∕stromal interface, such as the transformation of epithelium surface (THG) and the overall fibrillary tissue architecture (SHG). This multicontrast nonlinear optics microscopy is able to not only differentiate between cancerous and healthy tissue, but can also distinguish between normal, benign, borderline, and malignant specimens according to their collagen disposition and compression levels within the extracellular matrix. The dimensions of the layers of epithelia can also be measured precisely and automatically. Our data demonstrate that optical techniques can detect pathological changes associated with ovarian cancer.

Second harmonic generation microscopy analysis of extracellular matrix changes in human idiopathic pulmonary fibrosis

“…SHG directly probes the structure of collagen and has been used to describe ECM alterations in several diseases, such as cancers, fibroses, and connective tissue disorders. [14][15][16][17][18][19][20][21][22][23][24][25][26][27] Multiphoton microscopy of elastin has also been used for several applications, including imaging skin and cardiovascular tissues, often in conjunction with SHG and coherent anti-Stokes Raman scattering. [28][29][30] SHG and TPEF microscopy has not yet been used extensively for lung tissues and has been limited to mouse models.…”

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

“…[43,124] The structure of epithelium of the head and neck [125], the colon [126] and intestine [127] can be visualized, but also the layered structure of teeth [18] and the collagen of the vocal folds [128,129] has been imaged. 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].…”

“…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].…”

Multimodal label-free microscopy

“…Fortunately, several reviews already adequately cover this¯eld. 45,77 In addition to these combinations which have been well established in the past years, more speci¯c use has also been made of multimodal nonlinear responses, such as combining several harmonics of auto°uorescence, 78 characterizing cell types such as cancerous cells through their nonlinear responses, 79 di®erentiating several tissue types in mucosa 80 or performing in vivo imaging of complete small organisms. 50,81,82 Due to the coherent state of the harmonic generation, polarization-resolved measurements are also possible, and work well in particular for SHG, where the non-symmetric requirements makes it very sensitive to the polarization state of the excitation beam.…”