Three-dimensional morphological characterization of colorectal pits from label-free microscopy images.

The prognosis of colorectal cancer (CRC), one of the most prevalent pathologies worldwide, is linked to early detection. Kudo's pit pattern classification states morphological pit patterns of the Lieberkühn crypts by analyzing the superficial mucosa, predicting the histology of colorectal lesions. Its use as a highly accurate two-dimensional diagnostic criterion has increased, mostly involving expert endoscopists' judgment. The processing of autofluorescence images could allow the diagnostic, bypassing staining techniques and decreasing the biopsies, resources and times involved in the inspection. That criterion could be extended by data of the pit three-dimensional (3D) morphology. Thus, this work was aimed at obtaining 3D morphological information by quantifying geometrical and shape descriptors through software processing and analysis of widefield autofluorescence microscopy image stacks acquired by fresh colon tissue samples from a murine model of CRC. Statistical analyses included pits from control mice and from the second (2nd), fourth (4th), and eighth (8th) weeks of treatment. Statistically significant differences were found for almost all parameters between the pits from control and from the 4th treated week, stating that the major morphological changes begin after the 2nd week. In particular, pits from control or initial treatment time points were more tubular, straighter and less rough than the ones from later treatment points. Therefore, they may be more associated to normal or non-neoplastic crypt lumens than linked to adenomas or even cancer crypts. These preliminary outcomes could be considered an advance in 3D pit morphology characterization.

Comparison of morphological changes of corneal collagen fibers treated with collagen crosslinking agents using second harmonic generation images.

Corneal cross-linking (CXL) is a common surgical procedure used to modify corneal biomechanics and stabilize keratoconus progression which is still under discussion. Its side effects, which are mostly related to anatomical unpredictability and stromal exposure, are the reason for the search for new CXL agents. In this work we have quantitatively evaluated the porcine corneal stroma architecture treated with collagen crosslinking agents such as riboflavin solutions and açai extract, using second harmonic generation microscopy. Aimed at evaluating the morphological changes in the corneal stroma after collagen crosslinking under a CXL chemical agent, a tubeness filter based Hessian matrix to obtain a 3D fiber characterization of the SHG images was applied. The results showed a curling effect and shortening of the collagen fibers treated with açai as compared to the control. They also showed a higher degree of clustering of the collagen fibers with larger empty spaces when compared to the other two groups. We believe that studies such as these presented in this paper are a good direct nondestructive and free labeling evaluation technique that allows the observation of morphologic features of corneas treated with new CXL agents.