PURPOSE: Cellular proliferation is accompanied by cellular apoptosis. In the healing digital flexor tendon, molecular events concerning cellular apoptosis have not been investigated. This study aimed to investigate the relationship between cellular apoptosis and proliferation in early tendon healing. METHODS: The flexor digitorum profundus tendons of 50 long toes in 25 chickens were transected and were repaired surgically. On postoperative days 3, 7, 14, 21, and 28, we subjected tendons to in situ terminal deoxynucleotide transferase dUTP nick end labeling (TUNEL) assay to detect apoptotic cells, immunofluorescence staining with antibodies to proliferating cell nuclear antigen to assess proliferation, and Bcl-2, an anti-apoptotic protein, to assess responses suppressive to apoptosis. The positively labeled tenocytes were counted microscopically and compared statistically. We also stained sections with hematoxylin and eosin to observe their healing status. An additional 12 tendons (6 chickens) served as day 0 controls. RESULTS: Compared with tendons at day 0, the healing tendons had notably greater cellularity in both epitenon and endotenon areas. The total number of cells and number of TUNEL-positive cells peaked at day 3. At days 7 to 21, the number of proliferating cell nuclear antigen-positive cells peaked. At days 7 and 14, the cells positively stained with Bcl-2 peaked. At days 14 to 28, the total number of cells and TUNEL-positive cells decreased significantly compared with those at days 3 and 7, yet the numbers remained greater than those on day 0. CONCLUSIONS: Apoptosis in the healing tendons peaks at day 3, followed about 10 days later by the peak proliferation period. Because Bcl-2 serves to inhibit apoptosis, a later increase in Bcl-2-positive cells indicates that tendon apoptosis is inhibited. These findings indicate that tenocyte apoptosis is accelerated within several days after injury, followed by increases in cellular proliferation and activation of molecular events to inhibit apoptosis in 2 to 4 weeks. Copyright 2010 American Society for Surgery of the Hand. Published by Elsevier Inc. All rights reserved.
PURPOSE: Cellular proliferation is accompanied by cellular apoptosis. In the healing digital flexor tendon, molecular events concerning cellular apoptosis have not been investigated. This study aimed to investigate the relationship between cellular apoptosis and proliferation in early tendon healing. METHODS: The flexor digitorum profundus tendons of 50 long toes in 25 chickens were transected and were repaired surgically. On postoperative days 3, 7, 14, 21, and 28, we subjected tendons to in situ terminal deoxynucleotide transferase dUTP nick end labeling (TUNEL) assay to detect apoptotic cells, immunofluorescence staining with antibodies to proliferating cell nuclear antigen to assess proliferation, and Bcl-2, an anti-apoptotic protein, to assess responses suppressive to apoptosis. The positively labeled tenocytes were counted microscopically and compared statistically. We also stained sections with hematoxylin and eosin to observe their healing status. An additional 12 tendons (6 chickens) served as day 0 controls. RESULTS: Compared with tendons at day 0, the healing tendons had notably greater cellularity in both epitenon and endotenon areas. The total number of cells and number of TUNEL-positive cells peaked at day 3. At days 7 to 21, the number of proliferating cell nuclear antigen-positive cells peaked. At days 7 and 14, the cells positively stained with Bcl-2 peaked. At days 14 to 28, the total number of cells and TUNEL-positive cells decreased significantly compared with those at days 3 and 7, yet the numbers remained greater than those on day 0. CONCLUSIONS: Apoptosis in the healing tendons peaks at day 3, followed about 10 days later by the peak proliferation period. Because Bcl-2 serves to inhibit apoptosis, a later increase in Bcl-2-positive cells indicates that tendon apoptosis is inhibited. These findings indicate that tenocyte apoptosis is accelerated within several days after injury, followed by increases in cellular proliferation and activation of molecular events to inhibit apoptosis in 2 to 4 weeks. Copyright 2010 American Society for Surgery of the Hand. Published by Elsevier Inc. All rights reserved.