Northwestern University Feinberg School of Medicine

Evangelos Kiskinis Lab

Our Work

The lack of easy accessibility to the cells of the nervous system has hampered progress towards the discovery of degenerative mechanisms as well as more effective treatments for neurological diseases. The groundbreaking technology of reprogramming, which allows for the generation of patient specific induced pluripotent stem cells (iPSCs) has created an unprecedented opportunity for a new approach towards developing cellular models of human disease1,2. Pluripotent stem cells, which renew indefinite, retain the unique ability to generate every cell in the human body including the myriad neuronal subtypes that make up the nervous system (Figure 1).

We employ this approach to generate cortical excitatory and inhibitory neurons as well as spinal motor neurons and astrocytes from individual patients. We use gene-editing techniques to introduce or fix mutations and develop specific reporter cell lines. We study the neurons we make by typical methods including immunocytochemistry (Figure 2), biochemistry, global genomic analysis and live cell imaging (Video 1). We also examine the physiological properties of neurons by using multi-electrode array (MEA) systems that allow for population recordings of large quantities of neurons (Video 2), as well as the Optopatch3,4, which allows for targeted recordings of neuronal subtypes of choice in an all optical fashion.

Figure 1 (click image to view a larger size)

Figure 1


Defining the overlap in degenerative mechanisms of genetic types of ALS

As much as 15% of ALS patients suffer from familial forms of the disease, with genetic studies demonstrating that ALS can be caused by mutations in multiple genes that encode proteins involved in diverse cellular functions. The degree of pathogenic overlap in genetically distinct ALS classes remains unclear. To address this issue we are generating patient-specific iPSCs from genetic cases, using gene-editing approaches to fix the mutations and generate isogenic controls and are analyzing the phenotypic alterations in diseased motor neurons5,6.


Developing in vitro models of epileptic channelopathies

Dravet Syndrome (DS) or Severe Myoclonic Epilepsy Infancy (SMEI) is a neurodevelopmental disorder beginning in infancy and characterized by severe epileptic seizures. In most cases it is caused by de novo, loss of function mutations in SCN1A, which encodes the a-subunit of the neuronal voltage-gated sodium channel Nav1.1. We are interested in developing cellular models of DS and other channelopathies in order to define the mechanisms that lead to their dysfunction and epileptic disease.


Investigating the role of DNA methylation in human neurons

We are seeking to understand the potential role of DNA methylation in the development and function of human neurons. To do this we are generating neuronal subtypes from stem cell lines that do not express the enzymes that are involved in methylating/demethylating DNA and are assessing their ability to function, respond to stress and adapt to physiological stimuli.

Movie 1
Monitoring neuronal viability and cellular dynamics by the introduction of relevant reporters and live cell imaging.


Figure 2

Left - Neurilized stem cell culture stained for PAX6 (green), OCT4 (red) and DNA (blue). Right - Spinal motor neurons generated from an ALS patient, stained for MAP2 (green), ISL1 (red) and TDP43 (purple).


Movie 2
Monitoring electrical activity of neurons using multi-electrode systems.


References

1. Han, S. S., Williams, L. A. & Eggan, K. C. Constructing and deconstructing stem cell models of neurological disease. Neuron70, 626-644, doi:10.1016/j.neuron.2011.05.003 (2011).
2. Kiskinis, E. & Eggan, K. Progress toward the clinical application of patient-specific pluripotent stem cells. The Journal of clinical investigation120, 51-59, doi:10.1172/jci40553 (2010).
3. Hochbaum, D. R. et al. All-optical electrophysiology in mammalian neurons using engineered microbial rhodopsins. Nat Methods11, 825-833, doi:nmeth.3000 [pii] 10.1038/nmeth.3000 [doi].
4. Kralj, J. M., Douglass, A. D., Hochbaum, D. R., Maclaurin, D. & Cohen, A. E. Optical recording of action potentials in mammalian neurons using a microbial rhodopsin. Nat Methods9, 90-95, doi:nmeth.1782 [pii] 10.1038/nmeth.1782 [doi].
5. Kiskinis, E. et al. Pathways Disrupted in Human ALS Motor Neurons Identified through Genetic Correction of Mutant SOD1. Cell stem cell, doi:10.1016/j.stem.2014.03.004 (2014).
6. Wainger, B. J. et al. Intrinsic membrane hyperexcitability of amyotrophic lateral sclerosis patient-derived motor neurons. Cell reports7, 1-11, doi:10.1016/j.celrep.2014.03.019 (2014).