Application and prospects of high-throughput screening for in vitro neurogenesis

Abstract

Over the past few decades, high-throughput screening (HTS) has made great contributions to new drug discovery. HTS technology is equipped with higher throughput, minimized platforms, more automated and computerized operating systems, more efficient and sensitive detection devices, and rapid data processing systems. At the same time, in vitro neurogenesis is gradually becoming important in establishing models to investigate the mechanisms of neural disease or deve lopmental processes. However, challenges remain in generating more mature and functional neurons with specific subtypes and in establishing robust and standardized three-dimensional (3D) in vitro models with neural cells cultured in 3D matrices or organoids representing specific brain regions. Here, we review the applications of HTS technologies on in vitro neurogenesis, especially aiming at identifying the essential genes, chemical small molecules and adaptive microenvironments that hold great prospects for generating functional neurons or more reproductive and homogeneous 3D organoids. We also discuss the developmental tendency of HTS technology, e.g., so-called next-generation screening, which utilizes 3D organoid-based screening combined with microfluidic devices to narrow the gap between in vitro models and in vivo situations both physiologically and pathologically.

Keywords: Cell differentiation; Cellular microenvironments; High-throughput screening; Neurogenesis; Stem cells; Three-dimensional cell culture.

Figure 1

Main protocols and applications of in vitro neurogenesis. Neurons can be generated in vitro through the differentiation of induced pluripotent stem cells or the transdifferentiation of somatic cells with the aid of transcription factors, growth factors or small molecules[51,131]. With the combination of 3D biomaterials, induced neurons can be applied in various fields, including modeling, drug screening, neural tissue engineering and regenerative medicine[213,232,276]. iPSCs: Induced pluripotent stem cells.

Figure 2

Procedures for 3D microenvironment screening. The mixture of neural cells and 3D biomaterials is printed through a microarrayer and subsequently spotted on the substrate of microarrays or micropillars. Then, culture medium is added to the microarray, and the micropillar is stamped into the microwell containing culture medium, including various combinations of growth factors and chemicals.

Figure 3

Summary of high-throughput screening platforms and their functions. Cell-free, cell-based and organoid-based HTS platforms can screen out target small molecules, genes, biomaterials and microenvironments to promote the survival, proliferation, differentiation and maturation of pluripotent stem cells, induce the transdifferentiation of somatic cells to other cell types, and even generate specific organoids for further applications, including disease modeling, development modeling, drug screening, tissue engineering and regenerative medicine. High-throughput screening (HTS) platforms include multiwell plates, microarrays and microfluidic chips, indicating the great potential in HTS in future studies. HTS: High-throughput screening; ECM: Extracellular matrix.