Two-dimensional (2D) human skeletal muscle fiber cultures are ill-equipped to support the contractile properties of maturing muscle fibers. This limits their application to the study of adult human neuromuscular junction (NMJ) development, a process requiring maturation of muscle fibers in the presence of motor neuron endplates. Here we describe a three-dimensional (3D) co-culture method whereby human muscle progenitors mixed with human pluripotent stem cell-derived motor neurons self-organize to form functional NMJ connections. Functional connectivity between motor neuron endplates and muscle fibers is confirmed with calcium imaging and electrophysiological recordings. Notably, we only observed epsilon acetylcholine receptor subunit protein upregulation and activity in 3D co-cultures. Further, 3D co-culture treatments with myasthenia gravis patient sera shows the ease of studying human disease with the system. Hence, this work offers a simple method to model and evaluate adult human NMJ de novo development or disease in culture.

A highly elastic hybrid hydrogel of methacryloyl‐substituted recombinant human tropoelastin (MeTro) and graphene oxide (GO) nanoparticles are developed. The synergistic effect of these two materials significantly enhances both ultimate strain (250%), reversible rotation (9700°), and the fracture energy (38.8 ± 0.8 J m−2) in the hybrid network. Furthermore, improved electrical signal propagation and subsequent contraction of the muscles connected by hybrid hydrogels are observed in ex vivo tests.