(C-90) Alignment of Mechanical and Anatomical Data in Ex Vivo Myocardium

Introduction:: The mechanical deviations of heart tissue are of great importance. The structural reorganization of the myocardium that comes with hypertrophy and fibrosis produces a pathologic mechanical behavior. This has been studied in both bulk bi-axial analysis and with gross regional tissue analysis. While effective at characterizing tissue blocks, these studies do not bridge the mechanical change at the tissue level to the micromechanical behavior of the cellular and subcellular components. In order to appropriately examine that bridge between subcellular and tissue mechanics, this work successfully registered mechanical data obtained from atomic force microscopy (AFM) to images obtained from two-photon (TPEF) and second harmonic generation (SHG) microscopy.

Materials and Methods:: AFM analysis was conducted on an Asylum MFP-3D BIO using pyramidal tips (r=7nm). Scans were conducted using contact mode in a droplet of 1X PBS at a high sensitivity (trigger point < 500 mV). TPEF/SHG images were excited at 850 and collected with a 414/46 bandpass filter in the forward. Alignment was conducted using a lab-built coverslip assembled by perforating PDMS microfilm (< 20 um). The film is detectable on both imaging systems and allowed for linear alignment of the images.

Results, Conclusions, and Discussions:: Results: Alignment was successful and matched the predicted region. Mechanical behavior as a function of local composition was observationally clear but only had a clear difference between ECM and cellular regions. Micron-level measurements were conducted in the AFM but did not produce contrast between subcellular regions such as collagen molecules and sarcomeres. Discussion: AFM mechanical analysis is difficult to retrieve in fluid and requires patience. The lack of contrast between micro components in the AFM images is a reflection of the sensitivity of the device and its ability to read the deformations caused by broader forces such as the cell surface and its proteins. The alignment of the system is very promising and allows for high level analysis without the need for costly materials or custom hybrid imaging apparatus.

Conclusion: The success of the project will lead to more analysis attempting high resolution data. This study can be a low cost and effective steppingstone for other work that aims to link the micromechanical behavior to both the bulk properties and the local anatomy.




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