Histological and immunohistochemical evaluations of the right knee joints.

<p>(<b>a</b>) Haematoxylin and Eosin stain of the right knee joint PFG samples. Yellow arrow points at the exposed area of cartilage, leading to subchondral bone (green arrow) of the control knee joints. Treated samples (BMSCs and ADSCs) reflected regenerated engineered cartilages (white arrows) covering the subchondral bone, though not smoothly packed like the native cartilage (black arrow). Scale represents 70 µm. (<b>b</b>) Safranin O stains of the right knee joint (patella) samples. The regenerated cartilage on both treated knee (BMSCs and ADSCs) stained positive (white arrow). They were homogenous to the histochemical properties of accumulated proteoglycans revealed via Safranin O, but not smooth as the native cartilage (black arrow). Yellow arrow points at the degenerated area of cartilage on the control; green arrow, the subchondral bone. Scale represents 70 µm. (<b>c</b>) Immunohistochemistry images of the right knee joint (PFG) samples. Collagen type II protein staining for both treated samples (BMSCs and ADSCs) demonstrated positive staining within the ground tissues and were visible throughout the matrixes, evidenced by the green fluorescence tagged with the secondary antibody (black arrow). The native articular cartilage served as positive control (black arrow). The absence of green fluorescence on the ground tissue of the engineered cartilages without the primary antibody (yellow arrow) served as the negative control. Scale represents 35 µm. (<b>d</b>) Immunohistochemistry images of the right knee joint (PFG) samples. SOX9 staining for BMSCs and ADSCs demonstrated positivity within the ground tissues and were visible throughout the matrixes by the green fluorescence tagged with the secondary antibody (black arrow). The native articular cartilage served as positive control (black arrow), while the absence of green fluorescence on the ground tissue of the engineered cartilages without the primary antibody (yellow arrow) served as the negative control. Scale represents 35 µm. (<b>e</b>) Immunohistochemistry images of the right knee joint (PFG) samples. Collagen type I staining for BMSCs and ADSCs demonstrated diminished positivity within the ground tissues. This occurred mainly as scattered clusters of the green fluorescence tagged with the secondary antibody (black arrow). The fibrous cartilage served as positive control (black arrow), while the absence of green fluorescence on the ground tissue of the engineered cartilages without the primary antibody (yellow arrow) served as the negative control. Scale represents 35 µm. <b>Note</b>: The authors wish to state that a superimposed histological image of Safranin O and PKH26 dye or the immuno signals with PKH26 dye would have made better proves of the presence of the injected cells, but owing to technical challenges with these processes and the dye stability, it was not accomplished.