The effects of primary cilia-mediated mechanical stimulation on nestin+-BMSCs during bone-tendon healing
Huabin Chen, Han Xiao, Bing Wu, Xin Shi, Changbiao Guan, Jianzhong Hu, Tao Zhang, Hongbin Lü
Abstract
• Mechanical stimulation (MS) enhances the bone-tendon interface (BTI) healing. • Nestin + -BMSCs participate the BTI healing and responsed to injury and MS. • MS facilitates the proliferation, migration and differentiation of nestin + -BMSCs. • Primary cilia play an important role in the response of nestin + -BMSCs to MS. • MS affects the biological function of nestin + -BMSCs through actin/Hippo/YAP signaling pathway. Mechanical stimulation has been proven to promote bone-tendon interface (BTI) healing, but the mechanism remains unclear. To investigate the effects of mechanical stimulation on the biological behavior of nestin + -bone mesenchymal stem cells (BMSCs) during the BTI healing, and to reveal the mechanisms of mechanical stimulation affecting BTI healing by primary cilia on the nestin + -BMSCs. Transgenic tracing mice (nestin cre ERT2 :: IFT88 fl/fl /ROSA26 YFP) with primary cilia on nestin + -BMSCs conditioned knocked out were constructed, and the littermates (nestin cre ERT2 :: ROSA26 YFP) with normal cilia on nestin + -BMSCs were the control. After establishing mouse supraspinatus insertion injury models, samples were collected at week-2 (n = 5 per group), 4 and 8 (n = 15 per group, respectively). In vivo, the repair efficiency was evaluated by histology, imaging, biomechanics, and the migration of nestin + -BMSCs, detected by immunofluorescence staining. In vitro, nestin + BMSCs were sorted and stimulated by tensile force to study the mechanisms of primary cilium-mediated mechanosensitive basis. Mechanical stimulation (MS) accelerated the recruitment of nestin + -BMSCs and promoted osteogenic and chondrogenic capacity. Histological, imaging and biomechanical results showed that the BTI healing quality of the IFT88 +/+ , MS group was better than that of the other groups. After the conditionally knockout IFT88 in nestin + -BMSCs, the repair ability of the BTI was obviously deteriorated, even though mechanical stimulation did not increase significantly (IFT88 -/- , MS group). In vitro results showed the tensile loading enhanced the proliferation, migration and osteogenic or chondrogenic gene expression of nestin + -BMSCs with normal cilia. On the other hand, osteogenesis and chondrogenic expression were significantly decreased after inhibiting actin- Hippo/YAP pathway components. The primary cilia mediated mechanical stimulation regulated osteogenic and chondrogenic differentiation potential of nestin + -BMSCs through the actin- Hippo/YAP pathway, and then promoted the BTI healing process.