Alginate‐Laminin Hydrogel Supports Long‐Term Neuronal Activity in 3D Human Induced Pluripotent Stem Cell‐Derived Neuronal Networks

Abstract

For 3D neural cultures durable hydrogels are required, which persist over a long differentiation period and thus enable the maturation of neuronal networks (NN). Here, 3D models based on human induced pluripotent stem cell‐derived neural progenitor cells that are embedded in hydrogels of either pure alginate or alginate functionalized with the extracellular matrix protein laminin 111 (L111) are established. This study analyzes material characteristics such as porosity, L111 distribution and shear viscosity, cell compatibility of hydrogels by measuring viability and cytotoxicity, and neural function by monitoring cell migration, differentiation as well as NN formation and activity on multielectrode arrays. The addition of L111 increases neural migration and enhances differentiation into neurons and astrocytes as well as synaptogenesis in alginate hydrogels. NN formed in hydrogels are electrically active for up to 206 d and L111‐supplementation further increases electrical activity, network maturation, and synchronicity compared to 2D controls and NN grown in pure alginate hydrogels. L111 addition to alginate gels further accelerates recovery of electrical activity after blockage of sodium channels with tetrodotoxin. In conclusion, NN grown in alginate‐L111 hydrogel blends are promising models for future long‐term applications in disease modeling, drug or chemical evaluation.

Long-term in vitro culture of 3D brain tissue model based on chitosan thermogel

“…iPSC-derived cells have a huge potential to model human diseases and to uncover disease mechanisms and novel therapeutic targets [78]. Current 3D culture techniques are based on various approaches, including self-organizing organoids [79], neural tissue [80], and hydrogel based artificial ECMs [81,82]. The literature reports several examples of 3D brain-like tissue models that rely on the use of the polysaccharide alginate with iPSCs, showing its ability to sustain differentiation into dopaminergic/glutamatergic neurons in long-term cultures representing an alternative model for applications in disease modeling (Parkinson's disease) and drug or chemical evaluation for mechanistic studies [80,83].…”

“…Current 3D culture techniques are based on various approaches, including self-organizing organoids [79], neural tissue [80], and hydrogel based artificial ECMs [81,82]. The literature reports several examples of 3D brain-like tissue models that rely on the use of the polysaccharide alginate with iPSCs, showing its ability to sustain differentiation into dopaminergic/glutamatergic neurons in long-term cultures representing an alternative model for applications in disease modeling (Parkinson's disease) and drug or chemical evaluation for mechanistic studies [80,83]. Additionally, several studies have been focused on hyaluronic acid (HA) or HA-chondroitinbased hydrogels in their efforts to develop 3D neural models.…”

“…These studies have aimed to identify the essential chemical cues required for creating a neuronal matrix that can trap the cell-produced ECM and neurotrophic factors, remodel the matrix, and support neurite outgrowth [84]. Human neurons and glial cells have also been successfully cultured in Matrigel and PEG-heparin-based hydrogels [80,84]. However, some of these biomaterials require a biofunctionalization to support neuronal cell adhesion and growth, and they often have limitations, including high batch-to-batch variability and low stability over time.…”

“…Long-term cultures hold the potential to expand the utility of in vitro neuronal networks in disease modeling and in drug screening [83,86]. To date, only a few studies have documented long durations of in vitro cultures [16,80,87]. However, it is important to underline that typically hydrogels, in long term cultures, demonstrated to be able to support the 3D culture of neuronal cell clusters/spheroids.…”

Electroconductive Collagen‐Carbon Nanodots Nanocomposite Elicits Neurite Outgrowth, Supports Neurogenic Differentiation and Accelerates Electrophysiological Maturation of Neural Progenitor Spheroids

“…To gain a deeper understanding of the underlying mechanisms that contribute to such enhanced electrophysiological activity, we investigated by IF and Western blotting the expression of synapsin (i.e., a crucial phosphoprotein known to promote the establishment and maintenance of synaptic connections by actively regulating the release of neurotransmitters) and that of NMDAr, which are primarily involved in the transmission of excitatory information. [97][98][99][100] Figure 7A shows that NP spheroids embedded in CE_CND1 exhibit higher levels of synapsin, namely 0.27 versus 0.16 and 0.18 for the CE and CE_rCND1 groups, respectively (Figure 7C,D). On the other hand, NMDA receptor levels are comparable for the conditions tested (Figure 7B,C,E).…”