| BackgroundHeterotopic ossification(HO)is defined as the formation of bone tissue in extraskeletal tissues such as muscle and soft tissue,and is a complication with a high incidence in the healing process after trauma.There are two ways of heterotopic ossification:intramembranous ossification and endochondral ossification.The current mainstream view is endochondral ossification,but the underlying mechanism is still unclear.Endochondral ossification includes four processes:recruitment of inflammatory factors,aggregation of mesenchymal cells,cartilage formation and endochondral ossification.It is known that the occurrence of inflammation plays an initiating role in HO.Macrophage migration inhibitory factor(Macrophage migration inhibitory factor,MIF),which can inhibit the random migration of macrophages,is an important cytokine that regulates inflammatory responses,and has been shown to play an important role in the development and maturation of the skeletal system.Studies have shown that Wnt signaling is closely related to cell proliferation,differentiation and apoptosis through the classic pathway dependent on β-catenin,and plays an important role in the process of differentiation of mesenchymal stem cells into osteoblasts.At the same time,MIF is also considered as an inducer of oxidative stress.Does MIF participate in the osteogenesis and development of heterotopic ossification?Does the Wnt/β-catenin signaling pathway play an important role in this process?The above scientific issues are worthy of further research and exploration.ObjectiveTo explore the effect of MIF on osteogenic differentiation and traumatic HO formation of tendon-derived stem cells(TDSCs)in vitro and its possible underlying mechanism.Materials and MethodsPurified mouse TDSCs were isolated in vitro,and at the same time of osteogenesis induction,different concentrations of MIF were added to observe the difference in their osteogenic differentiation ability.While adding exogenous MIF,TDSCs were treated with the MIF-specific antagonist 4-Iodo-6-phenylpyrimidine(4-IPP)or the Wnt/β-catenin pathway inhibitor IWR-1,respectively,and the osteogenic differentiation ability was observed,difference.At the same time,a hypoxic microenvironment was constructed in vitro to make TDSCs in a state of oxidative stress,and the effect of MIF on the osteogenic differentiation of TDSCs in a state of oxidative stress was evaluated.In animals,the mouse Achilles tendon amputation model was used to observe the effects of MIF combined with 4-IPP and IWR-1 or not on the formation of traumatic heterotopic ossification.ResultsIn vitro studies have shown that MIF increases the expression of cartilage biomarkers SOX9 and ossification indicators(ALP,OCN,and RUNX2)by activating the Wnt/β-catenin signaling pathway,and promotes the osteogenic differentiation of TDSCs.Treatment of Wnt/β-catenin signaling blockade with 4-IPP or IWR-1 inhibited MIF-induced osteogenic differentiation.At the same time,when simulating the hypoxic microenvironment of soft tissue injury in vitro,the addition of MIF can increase the level of intracellular ROS and weaken the activity of antioxidant enzymes,thereby aggravating the oxidative stress state of TDSCs and promoting the osteogenic differentiation of TDSCs.Inhibition of MIF can reverse the above results.In vivo studies have shown that MIF can significantly promote the formation of traumatic heterotopic ossification after Achilles tendon rupture.ConclusionsThis study confirmed that MIF has a positive regulatory effect on the osteogenic differentiation of TDSCs,and promotes the pathogenesis of mouse tendon traumatic HO in vivo,which may be related to the activation of Wnt/β-catenin pathway by MIF,and secondly,MIF can increase ROS level,aggravate oxidative stress,and make TDSCs differentiate towards osteogenesis.These findings may provide new potential therapeutic targets for the prevention of HO,and may provide a new direction for the treatment of HO. |
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