Atomic Force Microscopy of Tissue Sections Is a Useful Complementary Tool in Biomedical Morphological Studies
The aim of the study was to demonstrate a good diagnostic potential of atomic force microscopy (AFM) in tracking morphological changes in the extracellular matrix (ECM) of connective tissue due to pathological processes. Here we summarize our experience in AFM application in a number of biomedical studies on the connective tissue disease, both for the research and clinical purposes.
Materials and Methods. Depending on theproject application (experimental or clinical), the tissue specimens were harvested either from animals, or from patients in the course of their surgical treatment, or post mortem.
AFM images of fixed tissue slices on glass slides were acquired with a Solver P47 AFM instrument (NT-MDT, Russia), in the semi-contact mode. For mechanical properties mapping, the images were acquired on air in the PeakForce Quantitative Nanomechanical Mapping mode (PeakForce QNM®), using a MultiMode 8 atomic force microscope (Bruker, USA). The regions of interest for scanning were selected in accordance with the histological assignments for the same sample, based on the view of a sample in the built-in optical microscope of the AFM instrument setup. To quantify the changes in the ECM morphology visualized by AFM imaging, we applied flicker-noise spectroscopy parameterization.
Results. AFM has been shown to reveal visible deviations from the normal morphology of the ECM in diseased tissues. We found that AFM and related techniques are capable of tracking disease-related changes at different levels of collagen organization in the ECM. At the microscale, AFM may detect loosening and disorganization of collagen fibers (e.g., in a dysplastic process), or the opposite process of their packing into tight parallel bundles in a fibrotic process. AFM may also monitor the ratio between collagen and non-fibrous material of the ECM, for example, in inflammatory and neoplastic processes. At the level of collagen fibrils, AFM may reveal early signs of the matrix destruction and remodeling not visible at the microscopic level. The flicker-noise spectroscopy parameters provide quantification of the morphological changes visualized by AFM. The PeakForce QNM® and nanoindentation studies give a further insight into the state of ECM via tracking changes in the local mechanical and adhesive properties.
All our AFM studies appeared in a good agreement with the histological findings and generally had a superior sensitivity to pathology-related ECM rearrangements.
Conclusion. AFM may serve as a valuable complementary diagnostic tool for tracking pathological changes in the connective tissue.
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