The development of techniques to create and use multiphase microstructured hydrogels (granular hydrogels or microgels) has enabled the generation of cultures with more biologically relevant architecture and use of structured hydrogels is especially pertinent to the development of new types of central nervous system (CNS) culture models and therapies. We review material choice and the customisation of hydrogel structure, as well as the use of hydrogels in developmental models. Combining the use of structured hydrogel techniques with developmentally relevant tissue culture approaches will enable the generation of more relevant models and treatments to repair damaged CNS tissue architecture.

Significance: We developed a label-free and noninvasive single-cell Raman microspectroscopy (SCRM)-based platform to identify neural cell lineages derived from clinically relevant human induced pluripotent stem cells (hiPSCs). Through large-scale Raman spectral analysis, we can distinguish hiPSCs and hiPSC-derived neural cells using their intrinsic biochemical profile. We identified glycogen as a Raman biomarker for neuronal differentiation and validated the results using conventional glycogen detection assays. The parameters obtained from SCRM were processed by a novel machine learning method based on t-distributed stochastic neighbor embedding (t-SNE)-enhanced ensemble stacking, enabling highly accurate and robust cell classification. The platform and the proposed biomarker should also be applicable to other cell types and can shed light on developmental biology and glycogen metabolism disorders.

Primary rodent neurons and immortalised cell lines have overwhelmingly been used for in vitro studies of traumatic injury to peripheral and central neurons, but have some limitations of physiological accuracy. Motor neurons (MN) derived from human induced pluripotent stem cells (iPSCs) enable the generation of cell models with features relevant to human physiology. To facilitate this, it is desirable that MN protocols both rapidly and efficiently differentiate human iPSCs into electrophysiologically active MNs. In this study, we present a simple, rapid protocol for differentiation of human iPSCs into functional spinal (lower) MNs, involving only adherent culture and use of small molecules for directed differentiation, with the ultimate aim of rapid production of electrophysiologically functional cells for short-term neural injury experiments. We show successful differentiation in two unrelated iPSC lines, by quantifying neural-specific marker expression, and by evaluating cell functionality at different maturation stages by calcium imaging and patch clamping. Differentiated neurons were shown to be electrophysiologically altered by uniaxial mechanical deformation. Spontaneous network activity decreased with applied stretch, indicating aberrant network connectivity. These results demonstrate the feasibility of this rapid, simple protocol for differentiating iPSC-derived MNs, suitable for in vitro neural injury studies focussing on electrophysiological alterations caused by mechanical deformation or trauma.

Corneal disease is one of the most important causes of blindness worldwide. Currently, the dominating treatment of corneal blindness is corneal transplantation. However, the main source of cornea for transplantation is based on donations which is far from enough to meet the requirement (less than 1:70 of cases). The severe shortage of donor cornea promotes the studies of effective corneal alternatives. However, many problems remain and can't be solved in current researches, such as original geometry reconstruction and ocular optical function restoring. 3D bioprinting can be a promising approach for corneal substitution. The advantages of this technology in corneal regeneration enable personalized corneal implant and single or multi-layer corneal equivalents with controllable structure and designed refractive ability. In this review, the progress, applications and limitations of most influential works among current keratoprosthesis and tissue-engineering cornea researches are discussed. Then the applications of 3D bioprinting in manufacturing multi-layered structures and surface are mentioned. Further, the potential, advantages in current research of 3D bioprinting single or multi-layer corneal equivalents and alternatives are discussed. Finally, an insight into the technical challenges and prospective facing the future research of 3D bioprinting corneal alternatives in vivo and in vitro is provided.