Induced Pluripotent Stem Cells for treatment of cardiovascular and respiratory diseases

Speaker:             Ulrich Martin

Department:     Cell biology

Location:            Erasmus MC Rotterdam

Date:                    15-5-2017

Author:               Katja Slangewal

The endogenous heart regeneration after a myocardial infarct is far from sufficient in mammals. Actually, less than 50% of the cardiomyocytes in a mammal heart are replaced during the entire life span. These facts immediately show the importance of the development of stem cell based regenerative treatments. This is the main area of research for Ulrich Martin from the Hannover Medical School, centre for Regenerative Medicine. During his talk, he did not only talk about the use of induced pluripotent stem cells (iPSCs) as therapeutic for heart repair, but also about the use of patient specific iPSCs in context for cystic fibrosis (a severe monogenic disease).

It has become more and more clear that quite often a general type of disease differs a lot from patient to patient. Emphasizing the need of patient specific treatments. Martin and his team work on patient specific iPSCs for disease modelling and the search to new drugs. The production of iPSCs (figure 1) becomes more and more efficient. The reprogramming step has become routine and it is even possible to use big tanks for the production of large numbers of iPSCs. This in contrast to a few years ago, when the production of iPSCs was tedious, time consuming, intensive and about low numbers of cells. The automatization and upscaling of stem cell production however was needed for high throughput and industrialization. For the first time in history, even big pharma are interested in stem cells.

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Figure 1: iPSCs are formed by taking somatic cells (for instant adult fibroblast cells) and adding reprogramming factors (KLF4, SOX2, c-Myc, Nanog, Oct-3/4 and LIN-28). After culturing the iPSCs and if desired adding mutations, the iPSCs can be differentiated in various tissues.

Now back to one of the applications Martin and his team focus on: treatment of Cystic Fibrosis. Cystic fibrosis is a severe and quite common disease, 1:2000 new-borns is diagnosed with CF. The disease is almost always caused by a single point mutation in the CFTR gene: F508del-CFTR. This mutation leads to a shorter and dysfunctional protein. Martin and his team want to investigate this mutation in iPSCs.

The workflow of Martin and his team goes as follows: first they generate CF specific iPSCs from the peripheral blood of CF patients. For his research, he uses patients with varieties in severity of CF. Next, he wants to pick the interesting clones (in which CFTR is expressed but not functional). Since the known antibodies for CFTR are not reliable, the lab uses a CFTR construct labelled with tdtomato. To screen for functionality, he uses eYFP labelling of the cell. eYFP activity is regulated by the iodide concentration, which is travels in and out the cell through the CFTR channel (figure 2). So when a colony of iPSCs is both fluorescent for the red tdtomato and yellow YFP, Martin can use it for further research. Now, a protocol is needed in which iPSCs can be differentiated in functional lung cells. There is a protocol proposed by Kadzik and Morrisey (2012) for this differentiation. However, Martin wants to use an upscaled version. At the moment, the upscaled procedure is not as efficient as the original variant, but it is still work in progress.

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Figure 2: YFP is sensitive for anions like iodide. It will get activated when iodide enters the cell. Which happens when the CFTR channel is functional.  Adapted from Vijftigschild, L.A.W., Ent, C.K. van der, Beekman, J.M. (2013) A novel fluorescent sensor for measurement of CFTR function by flow cytometry. Cytometry Part A 83A: 578 fig. 1A

Besides CF, Martins research also focusses on iPSCs derived cardiomyocytes as cellular therapeutic for heart repair. In order for therapies like these to exist, cardiomyocytes need to be produced in a safe, efficient and large scale production. In this case large scale production is a real must, since humans lose 1-2 billion cardiomyocytes after a myocardial infarction. Martin and his team are testing these large-scale productions. One important finding is the relationship between the state of differentiation and the density in which the cells where located. This stresses the importance of an equally divide density of cells over the entire tank.

It is now possible to form small amounts of differentiated tissue (few centimeter in diameter), which can beat like heart-tissue. However, the tissue is far from the strength of adult heart tissue. On the bright side, the tissue can deal with higher forces than the heart-tissue of new-borns. At the moment, stem cell derived cardiomyocyte tissue is implanted in monkeys who have suffered a myocardial infarction. This means it is time to prepare for clinical application. The start of ‘iPSCs for Clinically Applicable heart Repair’ or iCARE will help in the realization of clinical application.

I thought this seminar was one of the most interesting ones I have attended. It made me realise how incredible fast the progress in stem cell studies goes. I liked to see the connection between research and clinic. Martin used many clear examples which made his talk easy to follow. I also liked to see the enthusiasm with which he talks about his work. This mainly came back at the end when he got some interesting questions. All in all a good seminar.



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