Genomic / Proteomic Research

The research group of Prof Steinhoff has established an animal model for acute myocardial infarction using the NOD SCID mice strain (mice with immunodeficiency, suitable for transplantation of human cells). The scientists have shown that injection of different types of stem cells into the myocardium causes an increased capillary density. It is known from other studies that bone-marrow-derived CD133+ cellsreduce for example the apoptosis of cardiomyocytes and can consequently increase myocardial contractility. Until now, however, there is a lack of differential genomic and proteomic analyses of stem cells in order to understand previously described and further effects in details.

One important current research focus at the RTC is the proteomic analysis of stem cell fractions. This work is carried out in close collaboration with Prof. Takayuki Asahara (RIKEN Center for Developmental Biology, Japan) who is one of the worldwide leading scientist in the field of endothelial progenitor cells for angiogenesis. This work includes the characterisation of endothelial progenitor cells by 2D SDS-PAGE, MALDI-TOF Mass-Spectrometry (Matrix-assisted-Laser-Desorption/Ionization –Time-Of-Flight-Mass-Spectrometry, automatic determination of mass of proteins, peptides and sequences of amino acids) and MudPIT (Multidimension protein identification technology, separation and identification of complex protein- and peptide compounds)

Animal models

Until now, it has never been observed that application of CD133+ stem cells has a toxic effect or causes tumours of the heart. Nevertheless, extended and systematic research regarding the developement of tumors and toxicity of intracardially applied CD133+ stem cells will be performed at the RTC for a further validation of the therapy. The efficacy and safety of human CD133+stem cells are to be proven with new methods.

Series of studies using NOD SCID mice to investigate the toxicity of tissues (LD50) and their potential to develope tumors are planned. Different concentrations of stem cells will be injected into normal and infarcted myocardium. Then, the development of tissue and tumors will be analysed in a long-term study. This model should provide a basis for a standard test of stem cell preparations in pre-clinical studies in the future. Corresponding test procedures and parameters will be developed in the research laboratory of the RTC.

The quality of cell products and the standardisation of their application must be ensured before the therapy can be applied in a large scale context. Furthermore, the function of bone-marrow-derived stem cells should be analysed regarding their capacity of migration using the following two methods:

Boyden chamber assay (analysis of chemotaxis by SDF-1α gradient, interindividual variabilities)
Intravital fluorescence microscopy in mouse cremaster model (regulation of intravascular adhesion of stem cells by SDF-1α und TNF α)

Interindividual differences can be investigated during the tests.

Differentiation of stem cells

Here, we analyse the role of microRNA (miRNA, small single-stranded RNA molecules) in the differentiation of human stem cells in endothelial cells. Such fundamental knowledge is essential for a further development of stem cell therapies in order to treat a variety of age-independent cardiovascular diseases.

Among other things, the miRNA molecules regulate gene expression during heart and skeletal muscle developing. These regulatory molecules have an extensive function in controlling different aspects of cardiac function and dysfunction. These include the growth of myocytes, the integrity of heart chamber walls, contractility, gene expression, and maintenance of heart rhythm. Studies of specific miRNAs in diseased hearts revealed deficient gene expression. Experiments with mice regarding improvement and degradation of heart function demonstrate that many types of heart diseases are dependent of miRNA function.

However, the role of miRNA function especially in heart diseases is completely unknown until now. Therefore, a number of research approaches are planned to address the issues such as:

Clarify how and to which extent miRNAs mediate the differentiation of CD133+ stem cells to endothelial cells and if miRNAs play any role in the decision process of proliferation vs. differentiation of endothelial cells.

Clarify whether miRNAs suppress the differentiation of CD133+ cells.

Identification of mechanisms by which miRNA control mesodermal tissue and the differentiation or proliferation of endothelial cells and their potential intercation via specific signal pathways.
Evaluation of miRNA-modified CD133+ cells using the model of acute myocard infarcts and/or ischemia of the lower extremeties.

These research approaches aim to optimise the technology of microRNA Microarrays in order to analyse the global gene expression of miRNAs in different stem cell populations.