The effects of primary cilia-mediated mechanical stimulation on nestin+-BMSCs during bone-tendon healing.

Abstract

Introduction: Mechanical stimulation has been proven to promote bone-tendon interface (BTI) healing, but the mechanism remains unclear.

Objective: 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.

Methods: 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.

Results: 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.

Conclusion: 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.