Abstract
The use of second-harmonic generation (SHG) microscopy in biomedical research is rapidly increasing. This is due in large part to the wide spread interest of using this imaging technique to examine the role of fibrillar collagen organization in diseases such as cancer. The co-examination of SHG images and traditional bright-field (BF) images of hematoxylin and eosin (H&E) stained tissue as a gold standard clinical validation is usually required. However, image registration of these two modalities has been mostly done by manually selecting corresponding landmarks which is labor intensive and error prone. We designed, implemented, and validated the first image intensity-based registration method capable of automatically aligning SHG images and BF images. In our algorithmic approach, a feature extractor is used to pre-process the BF image to block the content features not visible in SHG images and the output image is then aligned with the SHG image by maximizing the common image features. An alignment matrix maximizing the image mutual information is found by evolutionary optimization and the optimization is facilitated using a hierarchical multiresolution framework. The automatic registration results were compared to traditional manual registration to assess the performance of the algorithm. The proposed algorithm has been successfully used in several biomedical studies such as pancreatic and kidney cancer studies and shown great efficacy.
Highlights
Alterations of stromal organization are commonly observed in the development of tumors such as breast cancer and pancreatic ductal adenocarcinoma [1,2,3,4,5]
Due to the nonlinear property of Second harmonic generation (SHG), only collagen fibers in the tissue section can be observed in SHG image, while all components stained with hematoxylin and eosin (H&E) dye are visualized in the BF image
Collagen fibers are hard to be directly extracted in BF images, they mostly exist in the extracellular matrix (ECM) which can be separated in H&E stained images by segmenting out the regions containing the cells
Summary
Alterations of stromal organization are commonly observed in the development of tumors such as breast cancer and pancreatic ductal adenocarcinoma [1,2,3,4,5]. It has been recognized that the extracellular matrix (ECM) in the tumor microenvironment can regulate tumor cell behavior and tissue tension homeostasis [6,7]. As the major stromal component, constitutes the scaffold of tumor microenvironment and affects tumor development such as infiltration, angiogenesis, invasion, and migration. Second harmonic generation (SHG) imaging has been recognized as a powerful tool for revealing material non-linear optical response [8,9,10]. SHG imaging has been used to examine collagen molecular and structural reorganizations in diseases such as cancer, fibrosis, and connective tissue disorders as it is highly sensitive to the collagen structure alternations that occur in these diseases. SHG imaging has been used to examine collagen molecular and structural reorganizations in diseases such as cancer, fibrosis, and connective tissue disorders as it is highly sensitive to the collagen structure alternations that occur in these diseases. [11,12,13,14,15]
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