The Mechanotransduction In Myoblast Cultured In Vitro In PLGA-collage Scaffolds

Functional malocclusion is common, and its treatment method is functional orthopedics,namely a complicated biological reaction of maxillomandible to the mechanical forces brought from masticatory muscles by the aid of functional appliances. The functional remodeling of masticatory muscles happens initially in a new mechanical environment, which is the key of functional orthopedics and plays an important role in acquiring good orthodontic treatment results and retention. So it is necessary to orthodontics to study the remodeling reaction of muscle in a variety of mechanical environments and this will provide theoretic guidance to the clinical treatment of functional malocclusion.It is unclear how such mechanical stress induces masticatory muscles remodeling. As the myoblast is highly responsive to mechanical stimuli,it plays the important role in the masticatory muscles remodeling processes.However,it is not definitively determined the initial responses which involved in mechanotransdution mechanism in the myoblast.Therefore,we studied the proliferation and the mechanism after mechanical stimulation of cell cultured in vitro in a PLGA-collagen scaffold,in order to better understand the mechanotransdution process.Methods: Skeletal muscle cultures were prepared from maxillofacial skeletal muscle from 1-2 days neonatal SD rats as described by Blau's method. Three-dimensional PLGA-collagen scaffolds were constructed using solvent casting and salt leaching and air/jet electrospinning processes. A cyclic strain loading apparatus - "Forcel" four point bending device was used for mechanically loading. Strain ranged from 1000μstrain to 4000μstrain. The flow cytometry method was used to examine the proliferation of cells cultured in a 3D PLGA-collagen scaffold after loading tension stress. Examined the expression of integrinβ1 and ILK mRNA after stress by RT-PCR,the phosphorylation levels of paxillin protein by Western Blotting. The difference of expression between cells cultured in a 3D PLGA-collagen scaffold and cells cultured on 2D surface at gene and proteins levels were compared.Result:1. PLGA-collagen scaffolds constructed using air/jet electrospinning processes have good microstructure and biological safety.2. 2000μstrain tension loading is a normal range of mechanical strain of myoblast when cultured in a PLGA-collagen scaffold. It is unfavourable for proliferation of cells to endure a tension loading last more than 24h.3. Myoblasts can express integrinβ1 and ILK.Tension stress could activate integrinβ1 and ILK in the cells cultured both in a 3D scaffold and on a 2D surface,but there were difference between them.4. The stimulation of tension strain (1000μstrain,2000μstrain)could activate paxillin immediately,and 2000μstrain tension strain promote the phosphorylation levels of paxillin more than 1000μstrain tension strain. 5. It was concluded from this study that myoblast cells cultured in 3D systems experience greater mechanotransduction patterns when compared to 2D surfaces, the importance of local architecture conditions on cell initial responses to mechanical stimuli was emphasized.Conclusion: Integrinβ1 , ILK and paxillin take part in mechanotransduction of myoblasts cultured in a 3D PLGA-collagen scaffold with stimulation of physiologic magnitude tension stress.And it was concluded from this study that the local architecture conditions plays an importance role on cell initial responses to mechanical stimuli.