Kirkeby Lab

Creating a human fetal brain in the dish with microfluidics (the MiSTR model)

This project is an interdisciplinary collaboration joining forces from the fields of bioengineering and stem cell biology with the aim of producing a novel model of early human neural tube patterning. Through microfluidic engineering techniques, we expose hESCs to morphogenic gradients in vitro, thereby building a microenvironment in which the stem cells are patterned into structures resembling the early stages of the rostral-to-caudal regionalised human neural tube. With this technique, we produce an anatomically relevant 3D in vitro model of the developing human neural tube corresponding to around weeks 2-10 of fetal development – read more here.

Mapping human neural subtype identities through single cell RNAseq

Using the MiSTR model described above, we apply single cell RNA sequencing to map the complexity of human neural subtypes present during early and late neural specification. By mapping cells at a single cell level at several different time points, we identify the unique molecular signatures of different human neural subtypes and aim to dissect the trajectories underlying the regionalisation and subregionalisation of human neural progenitors and neurons during embryo development.

Identifying secreted factors from different human brain regions through proteomics

For this project, we use our MISTR model to search for human-specific patterns of developmental growth factors through unbiased mass-spec (MS) proteomic analysis of the secreted proteins produced from different neural regions during development. Novel candidates identified from this approach are explored for their biological function in directing neural cells towards specific neuronal subtypes and for inducing neuronal subtype maturation.

Studying the functions of long non-coding RNAs in human neural cells

Long non-coding RNAs (lncRNAs) are highly abundant in the human genome, and for the vast majority of these their functions are unknown. Here, we apply CRISPR gene editing techniques in human pluripotent cells (CRISPR knockout, CRISPR activation and CRISPR inhibition) to investigate the functions of novel lncRNAs during human neural specification and differentiation. We aim to uncover both general functions in neural differentiation as well as more specific functions in regional subtype specification.

Generating human hypothalamic neurons for studying neuronal control of appetite and metabolism

The hypothalamus is important for regulating appetite, metabolism and sleep, and hypothalamic neuronal dysfunction is coupled to obesity, type 2 diabetes and narcolepsy. As such, access to functional human hypothalamic neurons is of great interest to pharma, biotech and academia for use in disease modelling, drug screening and cell replacement therapies. However, protocols for generation of subtype-specific hypothalamic neurons are still suboptimal, partly due to a lack of knowledge on the developmental trajectories giving rise to each specific neuronal subtype of the hypothalamus. In this project, we generate protocols for subtype-specific differentiation of hESCs into different subregions of the hypothalamus, and we investigate the neuronal lineages through single cell RNA sequencing. We focus in particular on hypothalamic neuronal subtypes which are important for the regulation of appetite, metabolism and sleep

Generating human hypocretin (HCRT) neurons for treatment of narcolepsy

Narcolepsy is a chronic, neurological disease affecting approximately 20-50 out of 100,000 individuals. The disease onset is typically during adolescence or early adulthood, and symptoms include excessive daytime sleepiness, cataplexy, hypnagogic hallucinations and sleep paralysis. The most common form of the disease, Type 1 narcolepsy, is caused by the selective loss of HCRT neurons in the hypothalamus of the patients. In this project, we generate protocols for production of HCRT neurons from hESCs, and we investigate in animal models the potential of these neurons to be used as a transplantation therapy for narcolepsy, by regenerating the HCRT sleep circuit

Generating human basal forebrain cholinergic neurons for treatment of dementia

Neurodegenerative disorders such as Alzheimer’s Disease, Parkinson’s Disease and Lewy body disease are associated with the loss of basal forebrain cholinergic neurons, which can lead to memory loss and dementia. In this project, we develop protocols for generating cultures of human basal forebrain cholinergic neurons from stem cells and through animal models, we assess the potential of these cells to be used as a transplantation cell therapy for treatment of cognitive deficits. This project is part of the EU H2020 consortium NSC-Reconstruct.

Studying neuroinflammation in in vitro stem cell models of Parkinson’s Disease

Recent studies in the field of Parkinson’s Disease has put increased focus on the role of neuroinflammation in the pathogenesis of the disease. In this project, we study the factors involved in neuroinflammation in Parkinson’s Disease through the use of complex in vitro models containing human neurons, astrocytes and microglia. This work is funded as a Michael J Fox Foundation/ASAP Research consortium

STEM-PD: developing a stem cell therapy for treatment of Parkinson’s Disease

In our lab at Lund University, together with the lab of Malin Parmar, we work in a long-standing collaboration to develop a cell therapy for treatment of Parkinson’s Disease. In this project, we produce human dopaminergic progenitor cells from human embryonic stem cells for transplantation therapy to replace the dopamine neurons which are lost in the brains of Parkinson’s Disease patients. This work has resulted in the development of a clinical stem cell product (STEM-PD), which is anticipated to go into clinical trial in Parkinson’s Disease patients in late 2021/early 2022.  

Funding

The projects above have received funding from the following sources:

• Novo Nordisk Foundation, Hallas Møller Emerging Investigator fellowship
• Independent Research Fund Denmark, Sapere Aude Research Leader fellowship
• EU Horizon 2020 Research Consortium NSC-Reconstruct
• Michael J Fox Foundation/ASAP, Research consortium
• Lundbeck Foundation, Collaborative project & Postdoc Fellowship
• Knut and Alice Wallenberg Foundation, WCMM Fellowship

2022

Elabi OF, Pass R, Sormonta I, Nolbrant S, Drummond N, Kirkeby A, Kunath T, Parmar M, Lane EL. Human Embryonic Stem Cell-Derived Dopaminergic Grafts Alleviate L-DOPA Induced Dyskinesia. J Parkinsons Dis. 2022 Apr 22. https://doi.org/10.3233/jpd-212920

Isaksson M, Karlsson C, Laurell T, Kirkeby A, and Heusel M.MSLibrarian: Optimized Predicted Spectral Libraries for Data-Independent Acquisition Proteomics. Journal of Proteome Research, Jan 2022, https://doi.org/10.1021/acs.jproteome.1c00796

2021

Tomishima M and Kirkeby A. Bringing advanced therapies for PD to the clinic: The scientist’s perspective. Journal of Parkinson’s Disease, June 2021,  2021;11(s2):S135-S140. https://doi:10.3233/JPD-212685

Brambach M, Ernst A, Nolbrant S, Drouin-Oellet J, Kirkeby A, Parmar M, Olariu V. Neural tube patterning: From a minimal model for rostro-caudal pattening to an integrated model. iScience, May 2021, https://doi.org/10.1016/j.isci.2021.102559

2020

Rifes P, Isaksson M, Rathore GS, Aldrin—Kirk P, Møller OM, Barzaghi G, Lee J, Egerod KL, Rausch DM, Parmar M, Pers TH, Laurell T and Kirkeby A. Modeling neural tube development by differentiation of human embryonic stem cells in a microfluidic WNT gradient. Nature Biotechnology, May 2020, DOI: 10.1038/s41587-020-0525-0

Tiklova K, Nolbrant S, Björklund Å, Fiorenzano A, Heuer A, Gillberg L, Hoban D, Cardoso T, Adler A, Lundén-Miguel H, Volakakis N, Kirkeby A, Perlmann T, Sharma Y, Birtele M and Parmar M. Single Cell Transcriptomics Identifies Stem Cell-Derived Graft Composition in a Model of Parkinson’s Disease. Nature Communications, May 2020, https://doi.org/10.1038/s41467-020-16225-5

Korshunova I, Rhein S, García-González D, Stölting I, Pfisterer U, Barta A, Dmytriyeva O, Kirkeby A, Schwaninger M and Khodosevich K. Genetic modification increases the survival and the neuroregenerative properties of transplanted neural stem cells. JCI Insight, Jan 2020, doi: 10.1172/jci.insight.126268,

2019

Kirkeby, A., Barker, R.A. (2019). Parkinson disease and growth factors - is GDNF good enough?. Nature Reviews Neurology, doi: 10.1038/s41582-019-0180-6.

2018

Cardoso, T., Adler, A.F., Mattsson, B., Hoban, D.B., Nolbrant, S., Wahlestedt, J.N., Kirkeby, A., Grealish, S., Bjorklund, A., and Parmar, M. (2018). Target‐specific forebrain projections and appropriate synaptic inputs of hESC‐derived dopamine neurons grafted to the midbrain of parkinsonian rats. Journal of Comparative Neurology 526, 2133-2146,doi: 10.1002/cne.24500

Porrello, E.R. and Kirkeby, A. (2018). A symphony of stem cells in Vienna – looking to the future. Development 145, dev163501,doi: 10.1242/dev.163501

Abbot, S., Agbanyo, F., Ahlfors, J.E., Baghbaderani, B.A., Bartido, S., Bharti, K., Burke, C., Carlsson, B., Cavagnaro, J., Creasey, A., DiGiusto, D., Francissen, K., Gaffney, A., Goldring, C., Gorba, T., Griffiths, E., Hanatani, T., Hayakawa, T., Heki, T., Hoogendoorn, K., Kawamata, S., Kimura, H., Kirkeby, A., Knezevic, I., Lebkowski, J., Lin, S., Lin-Gibson, S., Lubiniecki, A., O'Shea, O., Pera, M., Petricciani, J., Pigeau, G., Ratcliffe, A., Sato, Y., Schumann, G.G., Shingleton, W., Stacey Chair, G., Sullivan, S., Svendsen, C.N., Trouvin, J.H., Vandeputte, J., Yuan, B.Z., and Zoon, K. (2018). Report of the international conference on manufacturing and testing of pluripotent stem cells. Biologicals, doi:10.1016/j.biologicals.2018.08.004.

Mondal, T., Juvvuna, P.K., Kirkeby, A., Mitra, S., Kosalai, S.T., Traxler, L., Hertwig, F., Wernig-Zorc, S., Miranda, C., Deland, L., Volland, R., Bartenhagen, C., Bartsch, D., Bandaru, S., Engesser, A., Subhash, S., Martinsson, T., Caren, H., Akyurek, L.M., Kurian, L., Kanduri, M., Huarte, M., Kogner, P., Fischer, M., and Kanduri, C. (2018). Sense-Antisense lncRNA Pair Encoded by Locus 6p22.3 Determines Neuroblastoma Susceptibility via the USP36-CHD7-SOX9 Regulatory Axis. Cancer Cell 33, 417-434.e417, doi:10.1016/j.ccell.2018.01.020.

2017

Lehnen D, Barral S, Cardoso T, Grealish S, Heuer A, Smiyakin A, Kirkeby A, Kollet J, Cremer H, Parmar M, Bosio A and Knöbel S. (2017). IAP-Based Cell Sorting Results in Homogeneous Transplantable Dopaminergic Precursor Cells Derived from Human Pluripotent Stem Cells, Stem Cell Reports 9, 1207-1220 doi:10.1016/j.stemcr.2017.08.016.

Nolbrant, S., Heuer, A., Parmar, M., and Kirkeby, A. (2017). Generation of high-purity human ventral midbrain dopaminergic progenitors for in vitro maturation and intracerebral transplantation. Nature Protocols 12, 1962-1979, doi:10.1038/nprot.2017.078.

Moraghebi, R., Kirkeby, A., Chaves, P., Ronn, R.E., Sitnicka, E., Parmar, M., Larsson, M., Herbst, A., and Woods, N.B. (2017). Term amniotic fluid: an unexploited reserve of mesenchymal stromal cells for reprogramming and potential cell therapy applications. Stem Cell Research and Therapy 8, 190, doi:10.1186/s13287-017-0582-6.

Kirkeby, A., Nolbrant, S., Tiklova, K., Heuer, A., Kee, N., Cardoso, T., Ottosson, D.R., Lelos, M.J., Rifes, P., Dunnett, S.B., Grealish, S., Perlmann, T., and Parmar, M. (2017). Predictive Markers Guide Differentiation to Improve Graft Outcome in Clinical Translation of hESC-Based Therapy for Parkinson's Disease. Cell Stem Cell 20, 135-148, doi:10.1016/j.stem.2016.09.004.

Kee, N., Volakakis, N., Kirkeby, A., Dahl, L., Storvall, H., Nolbrant, S., Lahti, L., Bjorklund, A.K., Gillberg, L., Joodmardi, E., Sandberg, R., Parmar, M., and Perlmann, T. (2017). Single-Cell Analysis Reveals a Close Relationship between Differentiating Dopamine and Subthalamic Nucleus Neuronal Lineages. Cell Stem Cell 20, 29-40, doi:10.1016/j.stem.2016.10.003.

Kirkeby, A., Parmar, M., and Barker, R.A. (2017). Strategies for bringing stem cell-derived dopamine neurons to the clinic: A European approach (STEM-PD). Progress in Brain Research, book series: Functional Neural Transplantation – IV, Elsevier, doi: 10.1016/bs.pbr.2016.11.011.

2016

Barker, R.A., Parmar, M., Kirkeby, A., Bjorklund, A., Thompson, L., and Brundin, P. (2016). Are Stem Cell-Based Therapies for Parkinson's Disease Ready for the Clinic in 2016? Journal of Parkinson’s Disease 6, 57-63, doi:10.3233/jpd-160798.

Heuer, A., Kirkeby, A., Pfisterer, U., Jonsson, M.E., and Parmar, M. (2016). hESC-derived neural progenitors prevent xenograft rejection through neonatal desensitisation. Experimental Neurology 282, 78-85, doi:10.1016/j.expneurol.2016.05.027.

Kirkeby, A., Perlmann, T., and Pereira, C.F. (2016). The stem cell niche finds its true north. Development 143, 2877-2881, doi:10.1242/dev.140095.

2015

Grealish, S., Heuer, A., Cardoso, T., Kirkeby, A., Jonsson, M., Johansson, J., Bjorklund, A., Jakobsson, J., and Parmar, M. (2015). Monosynaptic Tracing using Modified Rabies Virus Reveals Early and Extensive Circuit Integration of Human Embryonic Stem Cell-Derived Neurons. Stem Cell Reports 4, 975-983, doi:10.1016/j.stemcr.2015.04.011.

Jönsson, M.E., Nelander Wahlestedt, J., Åkerblom, M., Kirkeby, A., Malmevik, J., Brattaas, P.L., Jakobsson, J., and Parmar, M. (2015). Comprehensive analysis of microRNA expression in regionalized human neural progenitor cells reveals microRNA-10 as a caudalizing factor. Development 142, 3166-3177, doi:10.1242/dev.122747.

2014

Grealish, S., Diguet, E., Kirkeby, A., Mattsson, B., Heuer, A., Bramoulle, Y., Van Camp, N., Perrier, A.L., Hantraye, P., Bjorklund, A., and Parmar, M. (2014). Human ESC-derived dopamine neurons show similar preclinical efficacy and potency to fetal neurons when grafted in a rat model of Parkinson's disease. Cell Stem Cell 15, 653-665, doi:10.1016/j.stem.2014.09.017.

2013

Kirkeby, A., Nelander, J., and Parmar, M. (2013). Generating regionalized neuronal cells from pluripotency, a step-by-step protocol. Frontiers in Cellular Neuroscience 6, 64, doi:10.3389/fncel.2012.00064

2012

Kirkeby, A., Grealish, S., Wolf, D.A., Nelander, J., Wood, J., Lundblad, M., Lindvall, O., and Parmar, M. (2012). Generation of Regionally Specified Neural Progenitors and Functional Neurons from Human Embryonic Stem Cells under Defined Conditions. Cell Reports 1, 703-714, doi:10.1016/j.celrep.2012.04.009.

Kirkeby, Agnete & Malin Parmar (2012) Building authentic midbrain dopaminergic neurons from stem cells – lessons from development. (Review). Translational Neuroscience, 3(4), 314-319, doi: 10.2478/s13380-012-0041-x.

2011

Pfisterer, U.*, Kirkeby, A.*, Torper, O.*, Wood, J., Nelander, J., Dufour, A., Bjorklund, A., Lindvall, O., Jakobsson, J., and Parmar, M. (2011). Direct conversion of human fibroblasts to dopaminergic neurons. Proceedings of the National Academy of Sciences 108, 10343-10348, doi:10.1073/pnas.1105135108. *contributed equally

Kirkeby, A., Parmar, M., Jakobsson, J. (2011). Using endogenous microRNA expression patterns to visualize neural differentiation of human pluripotent cells. Human embryonic and induced pluripotent stem cells – lineage-specific differentiation protocols, Springer Protocols, doi:1007/978-1-61779-267-0_34.

2010

Sachdeva, R., Jonsson, M.E., Nelander, J., Kirkeby, A., Guibentif, C., Gentner, B., Naldini, L., Bjorklund, A., Parmar, M., and Jakobsson, J. (2010). Tracking differentiating neural progenitors in pluripotent cultures using microRNA-regulated lentiviral vectors. Proceedings of the National Academy of Sciences 107, 11602-11607, doi:10.1073/pnas.1006568107.

2008

Lapchak, P.A., Kirkeby, A., Zivin, J.A., and Sager, T.N. (2008). Therapeutic window for non-erythropoietic carbamylated erythropoietin to improve motor function following multiple infarct ischemic strokes in New Zealand white rabbits. Brain Research 1238, 208-14, doi:10.1016/j.brainres.2008.08.017.

Kirkeby, A., Torup, L., Bochsen, L., Kjalke, M., Abel, K., Theilgaard-Monch, K., Johansson, P.I., Bjorn, S.E., Gerwien, J., and Leist, M. (2008). High-dose erythropoietin alters platelet reactivity and bleeding time in rodents in contrast to the neuroprotective variant carbamylerythropoietin (CEPO). Thrombosis and Haemostasis 99, 720-8, doi:10.1160/TH07-03-0208.

Leist, M., Bremer, S., Brundin, P., Hescheler, J., Kirkeby, A., Krause, K.H., Poerzgen, P., Puceat, M., Schmidt, M., Schrattenholz, A., Zak, N.B., and Hentze, H. (2008). The biological and ethical basis of the use of human embryonic stem cells for in vitro test systems or cell therapy.Altex 25, 163-190.

2007

Montero, M., Poulsen, F.R., Noraberg, J., Kirkeby, A., van Beek, J., Leist, M., and Zimmer, J. (2007). Comparison of neuroprotective effects of erythropoietin (EPO) and carbamylerythropoietin (CEPO) against ischemia-like oxygen-glucose deprivation (OGD) and NMDA excitotoxicity in mouse hippocampal slice cultures. Experimental Neurology 204, 106-117, doi:10.1016/j.expneurol.2006.09.026.

Kirkeby, A., van Beek, J., Nielsen, J., Leist, M., and Helboe, L. (2007). Functional and immunochemical characterisation of different antibodies against the erythropoietin receptor. Journal of Neuroscience Methods 164, 50-58, doi:10.1016/j.jneumeth.2007.03.026.