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The neuronal pathological changes were also observed in the acquired digital images saved as uncompressed 24-bit RGB TIFF files in the software program AnalySIS (Olympus Soft Imaging System GmbH, Münster, Germany) performing quantitative analysis of neuronal damage in the karyopyknotic areas

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Images were analyzed using the “Adiposoft” plug-in software in ImageJ (NIH).[22] Briefly, images were converted to 8-bit images and scald to 0.366 microns per pixel (corresponding to 20× magnification on the Leica microscope)