In order to faithfully model the human spinal cord, multiple cell types need to be present: motor neurons, astrocytes, oligodendrocytes, and microglia. Motor neurons are the cells that transmit impulses to our muscles, allowing us to move; astrocytes surround our neurons and help keep them healthy, removing waste and providing metabolic support; oligodendrocytes produce myelin (which is like insulation for our neurons), protecting our neurons and allowing signals to travel faster along them; microglia are our brain and spinal cord’s resident immune cells, helping to protect us. All of these cells have been shown to not function properly in ALS.
These cells can be generated from induced pluripotent stem cells (iPSCs), which start life as skin or blood cells taken from patients, which are then reverted to a stem cell state. This means they retain the genetic background of the patient (vital for disease modelling) and can theoretically be differentiated into any cell type in the body. The astrocytes in our model are directly induced from skin cells called fibroblasts. All of these cells can then be embedded in a substrate and printed into 3D structures, in a process that looks a bit like using writing icing on a cake.
iPSC-derived motor neurons grown in a 3D plate. This image shows iPSC-derived motor neurons which have been stained for the enzyme choline acetyltransferase (ChAT), which makes the neurotransmitter needed for motor neurons to communicate with each other. This image is made of around 100 images stacked on top of each other. The stacking of images is necessary as 3D cultures are much thicker than 2D cultures.
The EU Joint Programme – Neurodegenerative Disease Research (JPND) ORGTHERAPY project aims to generate a 3D-printed iPSC-derived model of the spinal cord to study C9orf72 amyotrophic lateral sclerosis/frontotemporal dementia (ALS/FTD). These are two diseases that can be caused by mutations in the C9orf72 gene. It is still unknown exactly why some patients with these mutations end up with ALS, FTD, or both. If we can improve the way we model these diseases, by using human-derived cells in a 3D system, it could result in data that translates well into potential treatments for these diseases, as well as further our understanding about what causes them.
iPSC-derived motor neurons and iAstrocytes 3D printed into gelatine scaffolds. The four panels show cell nucleus staining (blue), neuron staining (green) and astrocyte staining (red) of cells that were embedded in gelatine and then 3D printed into structures. These structures can then be cut and stained to produce images such as these (you can read more about this technology here).
Collaborating between different universities, there are multiple disciplines that are being combined in order to develop this model, with experts in 3D printing of cells, microfluidic devices and iPSC-derived cells all working together. Working as a team, particularly between different disciplines, is extremely important for science and allows us to advance new and ground-breaking techniques that will ultimately help patients.
Visitors from abroad! Collaboration with University of Eastern Finland
In November 2022, Mireia Gomez Budia – a PhD student from the University of Eastern Finland – came to our lab to help train Marianne in how to make iPSC-derived microglia, as part of the JPND project. The trip was a success, with Mireia providing lots of valuable expertise in culturing microglia, which are our brain’s resident immune cells. These cells have a lot of involvement in ALS, meaning it’s vital that our models of ALS include them. It is hoped that in 2023 we will be able to combine these iPSC-derived microglia into our 3D model, which has already included iPSC-derived motor neurons and induced astrocytes from Marianne’s trip to Sweden in 2021.