Increased apoptosis at the late stage of tendon healing

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Abstract
The mechanism for the clearance of excess healing fibroblasts at the end of tendon healing has not been reported despite the importance of maintaining tissue homeostasis. This study investigated the role of apoptosis in cell turnover in a rat central 1/3 patellar tendon donor site injury model. At days 4, 7, 14, 28, months 2 and 6, the rats were killed. Patellar tendons without injury served as control. Apoptotic cells were determined by an in situ terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end-labeling (TUNEL) assay and anti-active caspase-3 antibodies, while proliferating cells were determined by anti-proliferating cell nuclear antigen antibodies. The total fibroblast-like cell density in the center of the wound increased from day 4 and thereafter steadily returned to normal. In situ TUNEL assay showed few positive staining cells in the wound at days 4 and 7. The percentages of TUNEL-positive fibroblast-like cells showing morphological characteristics of apoptosis increased sharply and reached the maximum on day 28 (median %: 31.38%). No fibroblast-like cell was stained at month 6 and the healed tissue was similar to that in a normal uninjured tendon. A similar trend was observed with active caspase-3 immunohistochemistry. In conclusion, an increase in apoptosis at the end of tendon healing coincided with a decrease in cellularity.