A matrigel-free method to generate matured human cerebral organoids using 3D-Printed microwell arrays

The current methods of generating human cerebral organoids rely excessively on the use of Matrigel or other external extracellular matrices(ECM)for cell micro-environmental modulation.Matrigel embedding is problematic for long-term culture and clinical applications due to high inconsistency and other limitations.In this study,we developed a novel microwell culture platform based on 3D printing.This platform,without using Matrigel or external signaling molecules(i.e.,SMAD and Wnt inhibitors),successfully generated matured human cerebral organoids with robust formation of high-level features(i.e.,wrinkling/folding,lumens,neuronal layers).The formation and timing were comparable or superior to the current Matrigel methods,yet with improved consistency.The effect of microwell geometries(curvature and resolution)and coating materials(i.e.,mPEG,Lipidure,BSA)was studied,showing that mPEG outperformed all other coating materials,while curved-bottom microwells outperformed flat-bottom ones.In addition,high-resolution printing outperformed low-resolution printing by creating faithful,isotropically-shaped microwells.The trend of these effects was consistent across all developmental characteristics,including EB formation efficiency and sphericity,organoid size,wrinkling index,lumen size and thickness,and neuronal layer thickness.Overall,the microwell device that was mPEG-coated,high-resolution printed,and bottom curved demonstrated the highest efficacy in promoting organoid development.This platform provided a promising strategy for generating uniform and mature human cerebral organoids as an alternative to Matrigel/ECM-embedding methods.

Advanced 4D-bioprinting technologies for brain tissue modeling and study

Although the process by which the cortical tissues of the brain fold has been the subject of considerable study and debate over the past few decades,a single mechanistic description of the phenomenon has yet to be fully accepted.Rather,two competing explanations of cortical folding have arisen in recent years;known as the axonal tension and the differential tangential expansion models.In the present review,these two models are introduced by analyzing the computational,theoretical,materials-based,and cell studies which have yielded them.Then Four-dimensional bioprinting is presented as a powerful technology which can not only be used to test both models of cortical folding de novo,but can also be used to explore the reciprocal effects that folding associated mechanical stresses may have on neural development.Therein,the fabrication of‘smart’tissue models which can accurately simulate the in vivo folding process and recapitulate physiologically relevant stresses are introduced.We also provide a general description of both cortical neurobiology as well as the cellular basis of cortical folding.Our discussion also entails an overview of both 3D and 4D bioprinting technologies,as well as a brief commentary on recent advancements in printed central nervous system tissue engineering.

Fundamental functional differences between gyri and sulci: implications for brain function, cognition, and behavior

Folding of the cerebral cortex is a prominent characteristic of mammalian brains.Alterations or deficits in cortical folding are strongly correlated with abnormal brain function,cognition,and behavior.Therefore,a precise mapping between the anatomy and function of the brain is critical to our understanding of the mechanisms of brain structural architecture in both health and diseases.Gyri and sulci,the standard nomenclature for cortical anatomy,serve as building blocks to make up complex folding patterns,providing a window to decipher cortical anatomy and its relation with brain functions.Huge efforts have been devoted to this research topic from a variety of disciplines including genetics,cell biology,anatomy,neuroimaging,and neurology,as well as involving computational approaches based on machine learning and artificial intelligence algorithms.However,despite increasing progress,our understanding of the functional anatomy of gyro-sulcal patterns is still in its infancy.In this review,we present the current state of this field and provide our perspectives of the methodologies and conclusions concerning functional differentiation between gyri and sulci,as well as the supporting information from genetic,cell biology,and brain structure research.In particular,we will further present a proposed framework for attempting to interpret the dynamic mechanisms of the functional interplay between gyri and sulci.Hopefully,this review will provide a comprehensive summary of anatomo-functional relationships in the cortical gyro-sulcal system together with a consideration of how these contribute to brain function,cognition,and behavior,as well as to mental disorders.