Tendinopathy is a tendon pathology often resulting from a failed healing response to tendon injury. Activated protein C (APC) is a natural anti-coagulant with anti-inflammatory and wound healing promoting functions, which are mainly mediated by its receptors, endothelial protein C receptor (EPCR) and protease activated receptors (PARs). This study aimed to determine whether APC stimulates tenocyte healing and if so, to assess the involvement of the receptors.

Mouse-tail tenocytes were isolated from 3-week-old wild type (WT), PAR- 1 knockout (KO) and PAR-2 KO mice. The expression of EPCR, PAR-1 and −2 and the effect of APC on tenocytes tendon healing and the underlying mechanisms were investigated by Reverse transcription real time PCR, western blot, 3- (4,5-dimethylthiazol-2-yl)-2,5- diphenyltetrazolium bromide (MTT) assay, zymography, and scratch wound healing/ migration assay.

When compared to WT cells, PAR-1 KO tenocytes showed increased cell proliferation (3.3-fold, p<0.0001), migration (2.7-fold, p<0.0001) and wound healing (3-fold, p<0.0001), whereas PAR-2 KO cells displayed decreased cell proliferation (0.6-fold, p<0.05) and no change in cell migration or wound healing. APC at 1 μg/ml stimulated WT and PAR-1 KO tenocyte proliferation (~1.3, respectively, p<0.05) and wound healing (~1.3-fold, respectively, p<0.05), and additionally promoted PAR1-KO cell migration (1.4-fold, p<0.0001). APC only increased the migration (2-fold, p<0.05) of PAR-2 KO tenocytes. The activation of AKT, extracellular signal-regulated kinase (ERK)-2, and glycogen synthase kinase (GSK)-β3, the intracellular molecules that are associated with cell survival/growth, and matrix metalloproteinase (MMP)-2 that is related to cell migration and wound healing, were increased in all three cell lines in response to APC treatment.

These findings show that PAR-1 and PAR-2 act differentially in tenocyte proliferation/migration/wound healing. APC likely promotes tenocyte proliferation/ wound healing via PAR-2, not PAR-1.