In Vitro Model of Human Skeletal Muscle Tissues with Contractility Fabricated by Immortalized Human Myogenic Cells

Abstract

In the development process for drugs used to treat skeletal muscle, cell‐based contractile force assays have been considered as a useful in vitro test. Immortalized human myogenic cells are promising as cell sources for reproducible and well‐characterized in vitro models. In this study, it is investigated whether immortalized human myogenic cells, Hu5/KD3, have suitable contractile ability and the potential to be used as cell sources for contractile force assays. Muscle tissues are fabricated using Hu5/KD3 cells on the microfabricated devices used to measure contractile force. The tissues generate a tetanic force of ≈30 µN in response to the electrical pulse stimulation (EPS). Gene expression analysis of the myosin heavy chain (MYH) isoform indicates that the tissues mostly consisted of muscle fibers expressing MYH7 or/and MYH8. The addition of dexamethasone or lovastatin decreases the contractile force of the tissues, indicating that the tissues have the potential to evaluate drug candidates designed to treat muscle atrophy or statin‐induced myopathy. It is also demonstrated that the contractile force of tissues increased when EPS is applied as an artificial exercise. These results indicate that the Hu5/KD3 tissues can be employed for contractile force assays and would be useful for in vitro human skeletal muscle models.

Compartmentalized three-dimensional human neuromuscular tissue models fabricated on a well-plate-format microdevice

“…Furthermore, in this study, we used immortalized human myogenic cells, Hu5/KD3. In our previous study, we fabricated engineered Hu5/KD3 skeletal muscle tissues and showed that they generated a larger contractile force than that of the tissues engineered from other cell types, including C2C12, primary cells, and reprogrammed stem cells (28,29). This result suggests that the maturation of Hu5/KD3 tissues was more advanced than those of C2C12 (8,11,14) or reprogrammed stem cells (15) used in other studies.…”

“…The copyright holder for this preprint this version posted January 8, 2021. ; https://doi.org/10.1101/2021.01.07.424253 doi: bioRxiv preprint 5 for a contractile force measurement of the developed engineered SkM tissues (27)(28)(29)…”

“…1). We integrated the microtunnels for axons into the microdevices for a contractile force measurement of the developed engineered SkM tissues (27)(28)(29)…”

“…Subsequently, we observed contraction of the SkM tissues for 10 days.2.5 Evaluation of the contractility of SkM tissues cocultured with MNsThe displacement of the tip of the micropost was observed under an upright microscope (BX53, OLYMPUS, Japan) equipped with ORCA-Spark (Hamamatsu Photonics, Hamamatsu, Japan). The contractile force was calculated using the displacement of the tip using the reported equation(29).For the measurement of contraction of the cocultured SkM tissues induced by spontaneous firing of MNs, the displacement of the tip of the micropost was observed within 5 min after removing the tissues from the 37 °C incubator to avoid the loss of MN activities due to the temperature decrease. To activate the MNs, glutamate was added at 400 µM into the MN chamber.…”

Development of a human neuromuscular tissue-on-a-chip model on a 24-well-plate-format compartmentalized microfluidic device

“…The copyright holder for this preprint this version posted January 8, 2021. ; https://doi.org/10.1101/2021.01.07.424253 doi: bioRxiv preprint for a contractile force measurement of the developed engineered SkM tissues (27)(28)(29). The developed microdevices that fit a well of a 24-well plate have two chambers connected by the microtunnels for axons.…”