Immunohistochemical (IHC) technique was utilized to detect TGF-β and VEGF in tissue slices by labeling them with HRP that could react with a specific substrate to give a brown color in tissue.25 Briefly, tissue slices (4 µm thickness) were immunohistochemically stained with the primary rabbit antibodies to TGF-β and VEGF (Catalog #: ab229856 and ab53465, respectively; Abcam®, Cambridge, UK). Image quantitation was performed via image analysis software (ImageJ, 1.48a, NIH, USA), as optical density (OD) of brown-stained (positive) cells across six different areas for each rat section.

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