Engineered myogenic microtissues derived from human skeletal myoblasts offer unique opportunities for varying skeletal muscle tissue engineering applications, such as in vitro drug-testing and disease modelling. However, more complex models require the incorporation of vascular structures, which remains to be challenging. In this study, myogenic spheroids were generated using a high-throughput, non-adhesive micropatterned surface. Since monoculture spheroids containing human skeletal myoblasts were unable to remain their integrity, co-culture spheroids combining human skeletal myoblasts and human adipose-derived stem cells were created. When using the optimal ratio, uniform and viable spheroids with enhanced myogenic properties were achieved. Applying a pre-vascularization strategy, through addition of endothelial cells, resulted in the formation of spheroids containing capillary-like networks, lumina and collagen in the extracellular matrix, whilst retaining myogenicity. Moreover, sprouting of endothelial cells from the spheroids when encapsulated in fibrin was allowed. The possibility of spheroids, from different maturation stages, to assemble into a more large construct was proven by doublet fusion experiments. The relevance of using three-dimensional microtissues with tissue-specific microarchitecture and increased complexity, together with the high-throughput generation approach, makes the generated spheroids a suitable tool for in vitro drug-testing and human disease modeling.

Reconstruction of bone tissue defects is a problematic area of the modern world. Temporary “platforms” of various materials for improving cell adhesion and proliferation have been extensively researched in recent decades. β-tricalcium phosphate (β-TCP) is a suitable biocompatible, biodegradable material used for bone regeneration. The creation of scaffolds with specifically designed surface structures will enable bone engineering applications that require navigated cell proliferation on a substrate with pre-set geometric limits. In this study, an innovative laser-based technique for surface modification was applied to improve the morphological properties of the surface of β-TCP pellets for proper cell surface environment creation. The obtained topographies with diverse processing parameters were compared. Homogenous microgroove structures, less than 100 µm, without the onset of melting and crack formation, were produced. The contribution from the accumulation effect of a diverse number of laser pulses (N = 1–100) on the final structure dimensions was examined. The microstructured scaffolds were investigated by confocal laser scanning microscopy (CLSM), scanning electron microscopy (SEM), energy-dispersive X-ray spectroscopy (EDX), Fourier-transform infrared spectroscopy (FTIR), and X-ray diffraction (XRD) analyses. We studied the effect of the patterned surface of the material on the mouse calvaria osteoblast (MC3T3) cells’ viability and cytotoxicity from 1 to 7 days. The results indicated that cell behavior was affected by microscale dimensions of the surface.