Recent Advances in the Field of 3D Modelling for Neurodegeneration

In the last few years, we have witnessed a tremendous advancement in cell culturing methods. The scene has been dominated by the optimization of 3D in vitro models either in the form of dissociated neuronal cultures or of the so-called organoids. The attempt of introducing new methods responds to the difficulty of capturing the multifactorial nature of diseases such as Alzheimer’s (AD) and Parkinson’s (PD) diseases in conventional models. There is certainly the need for developing more relevant pre-clinical set-ups and their use in association to stem cell technologies might provide previously unexplored modelling opportunities. The laboratories of professors Tanzi and Kim, first described in the context of AD, how the 3D environment allows recapitulating key features of AD pathology on-a-dish [1, 2]. Compared to standard 2D cultures, stem cell derived-neurons carrying familial AD mutations (APP and PS1), differentiated in 3D, show accumulation of critical amounts of amyloid-β peptide (Aβ) and hyperphosphorylated tau protein. In the context of PD, we show that 3D cultures of dopaminergic neurons carrying the LRRK2-G2019S mutation progressively degenerate in association with mitochondrial defects [3]. In 2D cultures, similar degeneration was previously observed only with the use of stress factors. An explanation for this different behavior might lie in the fact that cells embedded in matrix surrogates experience a more physiological equilibration and transport of soluble factors compared to 2D-maintained cells [4, 5]. The more confined environment favors the synthesis of extracellular matrix proteins and facilitates cell-cell interaction. Interestingly, gene and protein expression profiles have shown to significantly differ when comparing 2D and 3D cultures [6, 7]. Cells cultured in 2D show approximately 30% of differentially expressed genes compared to cells in vivo [8]. This is not surprising considering that the 2D environment provides cells with mechanical constraints and favors artificial geometries. Because of this fine mechanical and biochemical interplay between neurons and their surrounding matrix, disease-specific subtle phenotypes might be then missed when studied only in 2D.

A further step forward in disease-modelling optimization has been the use of cerebral organoids, first described in 2013 [9]. These self-organizing brain tissues with mature cortical neuronal subtypes might represent a good starting point to study AD, but provided also the hippocampal area of the brain can be better developed in the future. In the context of PD, we and others have generated organoids resembling the midbrain area of the human brain [10, 11].

The next challenge in this exciting field will be the improvement of quality control measures and standards. This in particular with the brain organoids as major variability has been described during their generation. In addition, the use of stem cell 3D models for the study of neurodegenerative mechanisms has been often criticized because of the thought that neurons do not reach a sufficient degree of maturation. However, 3D conditions have been shown to promote neuronal maturity and to increase adult tau isoforms levels [2]. That said, the field will need to introduce the aging component in the model to further refine the methods and capture faithfully more traits of neurodegenerative mechanisms. Although the technology currently presents several limitations, 3D is certainly a powerful approach which represents a step closer to the in vivo situation and can fuel new drug discovery options.

[1] Choi SH, Kim YH, Hebisch M, Sliwinski C, Lee S, D'Avanzo C, Chen H, Hooli B, Asselin C, Muffat J, Klee JB, Zhang C, Wainger BJ, Peitz M, Kovacs DM, Woolf CJ, Wagner SL, Tanzi RE, Kim DY (2014) A three-dimensional human neural cell culture model of Alzheimer's disease. Nature 515, 274-278.
[2] Kim YH, Choi SH, D'Avanzo C, Hebisch M, Sliwinski C, Bylykbashi E, Washicosky KJ, Klee JB, Brustle O, Tanzi RE, Kim DY (2015) A 3D human neural cell culture system for modeling Alzheimer's disease. Nat Protoc 10, 985-1006.
[3] Bolognin S, Fossépré M, Qing X, Jarazo J, Ščančar J, Moreno EL, Nickels SL, Wasner K, Ouzren N, Walter J, Grünewald A, Glaab E, Salamanca L, Fleming RMT, Antony PMA, Schwamborn JC (2018) 3D cultures of Parkinson's disease‐specific dopaminergic neurons for high content phenotyping and drug testing. Adv Sci (Weinh), doi: 10.1002/advs.201800927.
[4] Baker BM, Chen CS (2012) Deconstructing the third dimension: how 3D culture microenvironments alter cellular cues. J Cell Sci 125, 3015-3024.
[5] Ramanujan S, Pluen A, McKee TD, Brown EB, Boucher Y, Jain RK (2002) Diffusion and convection in collagen gels: implications for transport in the tumor interstitium. Biophys J 83, 1650-1660.
[6] Nelson CM, Bissell MJ (2005) Modeling dynamic reciprocity: engineering three-dimensional culture models of breast architecture, function, and neoplastic transformation. Semin Cancer Biol 15, 342-352.
[7] Baharvand H, Hashemi SM, Kazemi Ashtiani S, Farrokhi A (2006) Differentiation of human embryonic stem cells into hepatocytes in 2D and 3D culture systems in vitro. Int J Dev Biol 50, 645-652.
[8] Birgersdotter A, Sandberg R, Ernberg I (2005) Gene expression perturbation in vitro--a growing case for three-dimensional (3D) culture systems. Semin Cancer Biol 15, 405-412.
[9] Lancaster MA, Knoblich JA (2014) Generation of cerebral organoids from human pluripotent stem cells. Nat Protoc 9, 2329-2340.
[10] Monzel AS, Smits LM, Hemmer K, Hachi S, Moreno EL, van Wuellen T, Jarazo J, Walter J, Bruggemann I, Boussaad I, Berger E, Fleming RMT, Bolognin S, Schwamborn JC (2017) Derivation of human midbrain-specific organoids from neuroepithelial stem cells. Stem Cell Reports 8, 1144-1154.
[11] Jo J, Xiao Y, Sun AX, Cukuroglu E, Tran HD, Goke J, Tan ZY, Saw TY, Tan CP, Lokman H, Lee Y, Kim D, Ko HS, Kim SO, Park JH, Cho NJ, Hyde TM, Kleinman JE, Shin JH, Weinberger DR, Tan EK, Je HS, Ng HH (2016) Midbrain-like organoids from human pluripotent stem cells contain functional dopaminergic and neuromelanin-producing neurons. Cell Stem Cell 19, 248-257.

Last comment on 5 January 2019 by Doo Yeon Kim, Ph.D.


We would like to congratulate Drs. Bolognin, Antony, and Schwamborn on their excellent publication regarding a 3D human neural cell culture model of PD. We are very excited to see that the 3D Matrigel-based cell culture system accelerates pathogenic cascades in hiPSC-derived PD neurons as we have shown with AD neurons (Choi et al., 2014). In our previous study, we showed that the 3D Matrigel cell culture system accelerated accumulation/aggregation of pathogenic amyloid beta species and also increased the expression of adult neural markers including adult 4-repeat tau isoform, which is crucial for developing robust tau pathology.  It is quite interesting that the 3D Matrigel cell culture system also accelerated mitochondrial deficits and neuronal death in human PD iPSC-derived neurons. It is possible that similar mechanisms may play a role. Although α-synuclein aggregation was not detected in Dr. Bolognin’s 3D PD cellular model, we think it will be intriguing to check whether soluble pathogenic proteins, including extracellular alpha-synuclein, selectively accumulate in 3D Matrigel with PD iPSC-derived neurons. 


We also agree with Dr. Bolognin that these new 3D human cellular models provide a new research tool to study the pathogenic mechanism of neurodegeneration in a human brain-like environment. However, many challenges lie ahead for comprehensively recapitulating brain conditions during human neurodegeneration: 1) mimicking aging aspects to 3D neural cell culture models; 2) precisely reconstituting brain structures that are damaged during neurodegeneration, as Dr. Bolognin addressed; 3) generating sporadic disease models; 4) adding neuroinflammatory elements. In an attempt to more precisely mimic the disease conditions, our lab, in collaboration with Dr. Cho’s lab at UNCC, recently developed 3D triculture models that mimic neuroinflammation in AD by adding human microglia cells into our 3D neuron-astrocyte AD models (Park et al., 2018). We think it will be interesting to find whether adding neuroinflammatory elements to 3D PD cellular models can further accelerate pathogenic cascades in PD iPSC-derived neurons. 


Finally, we would like to briefly comment on implications of the 3D cell culture models for drug discovery. As pointed by Dr. Bolognin, there are many challenges to adapt self-assembling 3D organoid models into high-throughput drug screening system. However, ours and Dr. Bolognin’s 3D culture models can be easily adapted to large-scale drug testing since they do not need complex self-organization of neural stem cells. Although there are many limitations in current 3D neurodegeneration models, we strongly believe that some of the 3D models are already useful enough for drug screening. 


-Doo Yeon Kim, Joseph Park, Mehdi Jorfi, Rudolph E. Tanzi



Choi, S.H., Kim, Y.H., Hebisch, M., Sliwinski, C., Lee, S., D'Avanzo, C., Chen, H., Hooli, B., Asselin, C., Muffat, J., et al. (2014). A three-dimensional human neural cell culture model of Alzheimer’s disease. Nature 515, 274–278.

Park, J., Wetzel, I., Marriott, I., Dréau, D., D'Avanzo, C., Kim, D.Y., Tanzi, R.E., and Cho, H. (2018). A 3D human triculture system modeling neurodegeneration and neuroinflammation in Alzheimer's disease. Nat Neurosci 21, 941–951.