Excessive mechanical stretch stress induces tendon cell apoptosis via mechanosensitive ion channel protein Piezo1

To investigate the role and mechanism of Piezo1, a mechanically-sensitive ion channel protein, in excessive mechanical stretch induced apoptosis of tendon cells.

Tendon cells of 8-week-old male C57 mice（n=20, body mass: 22~26 g）were isolated and cultured, and a cell model of mechanical stretch induced apoptosis of tendon cells was established using the Flexcell system. Tendon cells were divided into the control group, the 20% elongation group, the 20% elongation+Yoda1 group, the 20% elongation+GsMTx4 group, the 20% elongation+Piezo1 knockdown lentivirus group（Lv-Piezo1）and 20% elongation+control lentivirus group（Lv-Ctrl）. Mitochondrial membrane potential assay, flow cytometry assay, Western blot assay and calcium fluorescence probe were used to detect the role of Piezo1 in tendon cell apoptosis. Tendon cells were divided into the 20% elongation group, the 20% elongation+siRNA-mediated Calpain2 knockdown group（si-Calpain2）, and the 20% elongation+control siRNA group（si-Ctrl）. Western blot assay was used to detect the activation of Piezo1 downstream signal Calpain2/BAX/Caspase3 axis.

The level of apoptosis with excessive mechanical tensile stress was higher in the 20% elongation group than those in the control group（P<0.05）, and Yoda1 promoted mechanical tensile stress induced apoptosis in tendon cells（P<0.05）, while GsMTx4 and Lv-Piezo1 showed the opposite effect（P<0.05）. The expression of Calpain2, BAX and cleaved-Caspase3 in tendon cells was enhanced by excessive mechanical stretch stress（P<0.05）. The 20% elongation+si-Calpain2 group had lower expression levels of BAX and cleaved-Caspase3 than those in the 20% elongation and the 20% elongation+control siRNA group（P<0.05）. Knockdown of Calpain2 inhibited the expression of BAX and cleaved-Caspase3（P<0.05）and attenuated apoptosis in tendon cells（P<0.05）.

Excessive mechanical stretch stress can induce apoptosis of tendon cells via the activation of Piezo1 and the downstream Calpain2/BAX/Caspase3 pathways. Piezo1 holds promise as a potential therapeutic target for tendinopathy.