Generation of human motor units with functional neuromuscular junctions in microfluidic devices
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Generation of human motor units with functional neuromuscular junctions in microfluidic devices. / Dittlau, Katarina Stoklund; Krasnow, Emily N.; Fumagalli, Laura; Vandoorne, Tijs; Baatsen, Pieter; Kerstens, Axelle; Giacomazzi, Giorgia; Pavie, Benjamin; Rossaert, Elisabeth; Beckers, Jimmy; Sampaolesi, Maurilio; Van Damme, Philip; Van Den Bosch, Ludo.
In: Journal of Visualized Experiments, Vol. 2021, No. 175, e62959, 2021.Research output: Contribution to journal › Journal article › Research › peer-review
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TY - JOUR
T1 - Generation of human motor units with functional neuromuscular junctions in microfluidic devices
AU - Dittlau, Katarina Stoklund
AU - Krasnow, Emily N.
AU - Fumagalli, Laura
AU - Vandoorne, Tijs
AU - Baatsen, Pieter
AU - Kerstens, Axelle
AU - Giacomazzi, Giorgia
AU - Pavie, Benjamin
AU - Rossaert, Elisabeth
AU - Beckers, Jimmy
AU - Sampaolesi, Maurilio
AU - Van Damme, Philip
AU - Van Den Bosch, Ludo
N1 - Publisher Copyright: © 2021 JoVE Creative Commons Attribution-NonCommercial-NoDerivs 3.0 Unported License.
PY - 2021
Y1 - 2021
N2 - Neuromuscular junctions (NMJs) are specialized synapses between the axon of the lower motor neuron and the muscle facilitating the engagement of muscle contraction. In motor neuron disorders, such as amyotrophic lateral sclerosis (ALS) and spinal muscular atrophy (SMA), NMJs degenerate, resulting in muscle atrophy and progressive paralysis. The underlying mechanism of NMJ degeneration is unknown, largely due to the lack of translatable research models. This study aimed to create a versatile and reproducible in vitro model of a human motor unit with functional NMJs. Therefore, human induced pluripotent stem cell (hiPSC)-derived motor neurons and human primary mesoangioblast (MAB)-derived myotubes were co-cultured in commercially available microfluidic devices. The use of fluidically isolated micro-compartments allows for the maintenance of cell-specific microenvironments while permitting cell-to-cell contact through microgrooves. By applying a chemotactic and volumetric gradient, the growth of motor neuron-neurites through the microgrooves promoting myotube interaction and the formation of NMJs were stimulated. These NMJs were identified immunocytochemically through co-localization of motor neuron presynaptic marker synaptophysin (SYP) and postsynaptic acetylcholine receptor (AChR) marker α-bungarotoxin (Btx) on myotubes and characterized morphologically using scanning electron microscopy (SEM). The functionality of the NMJs was confirmed by measuring calcium responses in myotubes upon depolarization of the motor neurons. The motor unit generated using standard microfluidic devices and stem cell technology can aid future research focusing on NMJs in health and disease.
AB - Neuromuscular junctions (NMJs) are specialized synapses between the axon of the lower motor neuron and the muscle facilitating the engagement of muscle contraction. In motor neuron disorders, such as amyotrophic lateral sclerosis (ALS) and spinal muscular atrophy (SMA), NMJs degenerate, resulting in muscle atrophy and progressive paralysis. The underlying mechanism of NMJ degeneration is unknown, largely due to the lack of translatable research models. This study aimed to create a versatile and reproducible in vitro model of a human motor unit with functional NMJs. Therefore, human induced pluripotent stem cell (hiPSC)-derived motor neurons and human primary mesoangioblast (MAB)-derived myotubes were co-cultured in commercially available microfluidic devices. The use of fluidically isolated micro-compartments allows for the maintenance of cell-specific microenvironments while permitting cell-to-cell contact through microgrooves. By applying a chemotactic and volumetric gradient, the growth of motor neuron-neurites through the microgrooves promoting myotube interaction and the formation of NMJs were stimulated. These NMJs were identified immunocytochemically through co-localization of motor neuron presynaptic marker synaptophysin (SYP) and postsynaptic acetylcholine receptor (AChR) marker α-bungarotoxin (Btx) on myotubes and characterized morphologically using scanning electron microscopy (SEM). The functionality of the NMJs was confirmed by measuring calcium responses in myotubes upon depolarization of the motor neurons. The motor unit generated using standard microfluidic devices and stem cell technology can aid future research focusing on NMJs in health and disease.
U2 - 10.3791/62959
DO - 10.3791/62959
M3 - Journal article
C2 - 34570099
AN - SCOPUS:85116594836
VL - 2021
JO - Journal of Visualized Experiments
JF - Journal of Visualized Experiments
SN - 1940-087X
IS - 175
M1 - e62959
ER -
ID: 368623153