Two‐photon microscopy of healthy, infarcted and stem‐cell treated regenerating heart

Abstract

Two-photon excitation autofluorescence (produced in myocytes) and second-harmonic generation (produced mainly by collagen) allow label-free visualization of these two important components of myocardium. Because of their different emission wavelengths, these two signals can be separated spectrally. Here, we examine two-photon microscopy images of healthy, infarcted and stem-cell treated rat hearts. We find that in infarcted heart, regions distant from the site of infarct are similar to healthy tissue in composition (mostly myocytes, very little collagen) and organization (densely packed myocytes), but infarct regions are characterized by sparse myocytes and high collagen content indicative of scar tissue formation. Stem cell treated hearts, in contrast, show regions of intertwined myocytes and collagen throughout the infarct, suggesting reduced tissue damage. Finally, these results offer interesting insights into our ongoing polarized light studies of cardiac tissue anisotropy, and reveal that both tissue composition and tissue micro-organization are reflected in polarization-measured linear retardance values.

Fast Measurement of Sarcomere Length and Cell Orientation in Langendorff-Perfused Hearts Using Remote Focusing Microscopy

“…1,2 Confocal (1-photon) and 2-photon imaging modalities have been used to study calcium [3][4][5][6] and voltage [7][8][9][10] transients at cellular and subcellular 11 resolutions in the intact organ. Physiologically important parameters, such as sarcomere spacing 12,13 and intracellular calcium wave front speeds, [14][15][16][17] can also be directly measured. Given the anisotropic nature of myocardial tissue organization, quantification of measurements with a spatial component (distance or velocity) assumes that the cell orientation relative to the image plane is known or can be ignored.…”

Review of the emerging role of optical polarimetry in characterization of pathological myocardium

“…Interestingly, a distinct feature of the regrown myocytes, by means of NLM, was the local reorganization of collagen fibers; the alignment in a coherent manner parallel to the myocytes, indicating the unique capability of myocytes to profoundly influence the microstructural properties of the collagen scar at multiple scales. 16,26,27,29,30 Overall, these remodeling's of the myocardium were reflected in the increasing trend of anisotropy (and linear retardance) toward healthy counterparts, demonstrating the capability of optical polarimetry to visualize the extent and tracking such regenerative treatments of the myocardium.…”

“…Interestingly, NLM also explored special zones of no signals (neither TPEF nor SHG) in the infarct core region, most likely corresponding to regions of necrotic myocytes, not replaced by collagen yet. 16,26,27 4 Polarimetric Characterization of Stem-Cell-Regenerated Myocardium…”

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

“…Multimodal nonlinear microscopy has been used to examine the efficiency of novel therapies, i.e., stem cell therapy. In a rat model structural differences of healthy, infarcted and stem-cell treated regenerating heart were investigated by SHG and 2PF [152]. Beside structural imaging of disease related tissue modifications the second major field of application for nonlinear imaging techniques is visualizing dynamic processes.…”