機械和電刺激對肌肉生成有正向作用。然而，如何在設備上結合機械和電刺激以有效地觀察細胞或組織，這仍然是一個挑戰。因此，我們設計一種生物裝置，能夠同時機械和電刺激細胞。在本研究中，我們以小鼠骨骼成肌細胞C2C12作為研究對象，研究其在經過機械/電刺激後的分化行為。雖然刺激有助於分化，但MTT生物活性測試顯示電刺激或循環拉伸刺激的強度過強會導致生物活性下降。在細胞排列實驗中發現直流電刺激並不影響C2C12排列方向，而拉伸量至少9%以上的循環拉伸刺激即能促使C2C12沿著與拉伸方向垂直的方向進行高度排列。而複合刺激實驗中其中細胞排列取決於拉伸方向，並不受電場的方向所影響。在肌管分化的實驗中， qPCR結果證實拉伸刺激或電刺激皆能夠增加分化標誌基因的表達，肌球重鏈蛋白(MHC)免疫染色實驗指出兩種刺激都會有效地增加肌管的數量及提升分化率，且拉伸刺激有助於控制肌管的排列。與單一刺激相比，複合刺激不僅保有維持肌管排列的效果，還能使分化標誌基因更加上調控或維持高水準，這些結果都指出拉伸/電複合刺激對於肌肉組織工程極具潛力。;Mechanical and electrical stimulations have been applied to pormote myogenesis. However, how to simultaneously apply these two stimuli to treat cells on a device with easy observation is still a challenge. Therefore, we designed a multi-well device which can stimulate cells mechanically and electrically. Mouse skeletal C2C12 myoblasts were examined of their differentiation under mechanical and electrical stimulations. The MTT assay showed that high levels of electrical and cyclic stretching stimulations resulted in decreasing cell viability. Regading cell morphology, Cyclic stretching equal or higher 9% of strain promoted C2C12 cells to align perpendicular to the stretching direction. However, electrical stimulation did not affect cell alignment. When these two stimulations were simultaneously performed, cell alignment only depended on the stretching direction. Quantative PCR (qPCR) results showed that both stretching and electrical stimulations enhanced the expression of differentiation marker genes. The immunostaining of myosin heavy chain (MHC) protein also indicated that both stimulations effectively increased myotubes, and myotubes were aligned under stretching stimulation. Compared with sole electrical or stretching stimulation, the combination of electrical and stretching stimulations not only aligned myotubes, but also maintained or even upregulated myo-differentiation genes. These results all indicate that the combination application of electrical and stretching stimulations is a potential strategy for muscle tissue engineering.
