Achilles tendon transection was performed on 40 rats (all on the same day) and the animals were randomized to 4 different groups; Fresh/No preconditioning, Fridge, Frozen and Preconditioning high (Table (Table2).2). Sample collection was performed 12 days post-surgery (Fig. 4) and followed by mechanical testing (Fig. 5). Before the samples were subjected to a pull-to-failure test, the refrigerated samples were stored for 24 h at 8 °C and the frozen samples were stored for 7 days at − 18 °C. This test was therefore performed on different days for each group. The fresh, refrigerated, and frozen samples were all tested by a pull-to-failure test without preconditioning.

Study design for mechanical measurements.

Precond. preconditioning; × means that the procedure was executed.

Experimental setup. (A) On day 0, a transversally skin incision was performed on the right hind limb and the tendon complex was exposed. The plantaris tendon was removed and the Achilles tendon was transected. (B) Animals were euthanized and tissue samples were collected 12 days post-surgery. (C) Samples were fixed in a metal clamp in the material-testing machine. Mechanical testing was performed according to each group.

Representative overview of the type of mechanical test and preconditioning level performed in each experiment. Graphs are from a fresh sample. (A, B) represents the preconditioning graph, where 5 cycles of 2 N (A) or 5 N (B) were applied. (C) Pull-to-failure test. (D) Creep test with a constant load of 8 N for 300 s. Graphs were obtained from the mechanical testing (100R; DDL INC., EDEN PRAIRIE, MN) software MtestW, version5.1.0 (ADMET).

In the preconditioned high group, samples were fresh and subjected to 5 cycles of 5 N of preconditioning before the pull-to-failure test. This group was thereafter compared to the fresh samples with no preconditioning.

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