Laser printing based on laser-induced forward transfer (LIFT) is a new biofabrication technique for the arrangement of biological materials or living cells in well-defined patterns. In the current study, skin cell lines (fibroblasts/keratinocytes) and human mesenchymal stem cells (hMSC) were chosen for laser printing experiments due to their high potential in regeneration of human skin and new application possibilities of stem cell therapy. To evaluate the influence of LIFT on the cells, their survival rate, their proliferation and apoptotic activity, and the DNA damages and modifications of their cell surface markers were assessed and statistically evaluated over several days. The cells survived the transfer procedure with a rate of 98%  +/- 1% standard error of the mean (skin cells) and 90%  +/- 10% (hMSC), respectively. All used cell types maintain their ability to proliferate after LIFT. Further, skin cells and hMSC did not show an increase of apoptosis or DNA fragmentation. In addition, the hMSC keep their phenotype as proven by fluorescence activated cell sorting (FACS) analysis. This study demonstrates LIFT as a suitable technique for unharmed computer-controlled positioning of different cell types and a promising tool for future applications in the ex vivo generation of tissue replacements.

No abstract

No abstract

The possible different therapeutic efficacy of human mesenchymal stem cells (hMSC) derived from umbilical cord blood (CB), adipose tissue (AT) or bone marrow (BM) for the treatment of myocardial infarction (MI) remains unexplored. This study was to assess the regenerative potential of hMSC from different origins and to evaluate the role of CD105 in cardiac regeneration. Male SCID mice underwent LAD-ligation and received the respective cell type (400.000/per animal) intramyocardially. Six weeks post infarction, cardiac catheterization showed significant preservation of left ventricular functions in BM and CD105+-CB treated groups compared to CB and nontreated MI group (MI-C). Cell survival analyzed by quantitative real time PCR for human GAPDH and capillary density measured by immunostaining showed consistent results. Furthermore, cardiac remodeling can be significantly attenuated by BM-hMSC compared to MI-C. Under hypoxic conditions in vitro, remarkably increased extracellular acidification and apoptosis has been detected from CB-hMSC compared to BM and CD105 purified CB-derived hMSC. Our findings suggests that hMSC originating from different sources showed a different healing performance in cardiac regeneration and CD105+ hMSC exhibited a favorable survival pattern in infarcted hearts, which translates into a more robust preservation of cardiac function.

Human Mesenchymal Stem Cells (hMSCs) present a promising tool for regenerative medicine. However, ex vivo expansion is necessary to obtain sufficient cells for clinical therapy. Conventional growth media usually contain the critical component fetal bovine serum. For clinical use, chemically defined media will be required. In this study, the capability of two commercial, chemically defined, serum-free hMSC growth media (MSCGM-CD and PowerStem) for hMSC proliferation was examined and compared to serum-containing medium (MSCGM). Immunophenotyping of hMSCs was performed using flow cytometry, and they were tested for their ability to differentiate into a variety of cell types. Although the morphology of hMSCs cultured in the different media differed, immunophenotyping displayed similar marker patterns (high expression of CD29, CD44, CD73, and CD90 cell surface markers and absence of CD45). Interestingly, the expression of CD105 was significantly lower for hMSCs cultured in MSCGM-CD compared to MSCGM. Both groups maintained mesenchymal multilineage differentiation potential. In conclusion, the serum-free growth medium is suitable for hMSC culture and comparable to its serum-containing counterpart. As the expression of CD105 has been shown to positively influence hMSC cardiac regenerative potential, the impact of CD105 expression onto clinical use after expansion in MSCGM-CD will have to be tested.

Background Bone marrow stem cell clonal dysfunction by somatic mutation is suspected to affect post-infarction myocardial regeneration after coronary bypass surgery (CABG). Methods Transcriptome and variant expression analysis was studied in the phase 3 PERFECT trial post myocardial infarction CABG and CD133 + bone marrow derived hematopoetic stem cells showing difference in left ventricular ejection fraction (∆LVEF) myocardial regeneration Responders ( n= 14; ∆LVEF +16% day 180/0) and Non-responders ( n= 9; ∆LVEF -1.1% day 180/0). Subsequently, the findings have been validated in an independent patient cohort ( n= 14) as well as in two preclinical mouse models investigating SH2B3 /LNK antisense or knockout deficient conditions. Findings 1. Clinical: R differed from NR in a total of 161 genes in differential expression ( n= 23, q <0•05) and 872 genes in coexpression analysis ( n= 23, q<0•05). Machine Learning clustering analysis revealed distinct R vs NR preoperative gene-expression signatures in peripheral blood acorrelated to SH2B3 ( p< 0.05). Mutation analysis revealed increased specific variants in R vsN R. (R: 48 genes; NR: 224 genes). 2. Preclinical: SH2B3 /LNK-silenced hematopoietic stem cell (HSC) clones displayed significant overgrowth of myeloid and immune cells in bone marrow, peripheral blood, and tissue at day 160 after competitive bone-marrow transplantation into mice. SH2B3 /LNK −/− mice demonstrated enhanced cardiac repair through augmenting the kinetics of bone marrow-derived endothelial progenitor cells, increased capillary density in ischemic myocardium, and reduced left ventricular fibrosis with preserved cardiac function. 3. Validation: Evaluation analysis in 14 additional patients revealed 85% R vs NR (12/14 patients) prediction accuracy for the identified biomarker signature. Interpretation Myocardial repair is affected by HSC gene response and somatic mutation. Machine Learning can be utilized to identify and predict pathological HSC response. Funding German Ministry of Research and Education (BMBF): Reference and Translation Center for Cardiac Stem Cell Therapy - FKZ0312138A and FKZ031L0106C, German Ministry of Research and Education (BMBF): Collaborative research center - DFG:SFB738 and Center of Excellence - DFG:EC-REBIRTH), European Social Fonds: ESF/IV-WM-B34-0011/08, ESF/IV-WM-B34-0030/10, and Miltenyi Biotec GmbH, Bergisch-Gladbach, Germany. Japanese Ministry of Health : Health and Labou...

Different subtypes of bone marrow-derived stem cells are characterized by varying functionality and activity after transplantation into the infarcted heart. Improvement of stem cell therapeutics requires deep knowledge about the mechanisms that mediate the benefits of stem cell treatment. Here, we demonstrated that co-transplantation of mesenchymal stem cells (MSCs) and hematopoietic stem cells (HSCs) led to enhanced synergistic effects on cardiac remodeling. While HSCs were associated with blood vessel formation, MSCs were found to possess transdifferentiation capacity. This cardiomyogenic plasticity of MSCs was strongly promoted by a gap junction-dependent crosstalk between myocytes and stem cells. The inhibition of cell-cell coupling significantly reduced the expression of the cardiac specific transcription factors NKX2.5 and GATA4. Interestingly, we observed that small non-coding RNAs are exchanged between MSCs and cardiomyocytes in a GJ-dependent manner that might contribute to the transdifferentiation process of MSCs within a cardiac environment. Our results suggest that the predominant mechanism of HSCs contribution to cardiac regeneration is based on their ability to regulate angiogenesis. In contrast, transplanted MSCs have the capability for intercellular communication with surrounding cardiomyocytes, which triggers the intrinsic program of cardiogenic lineage specification of MSCs by providing cardiomyocyte-derived cues.

Myocardial infarction (MI) is a major condition causing heart failure (HF). After MI, the renin angiotensin system (RAS) and its signalling octapeptide angiotensin II (Ang II) interferes with cardiac injury/repair via the AT1 and AT2 receptors (AT1R, AT2R). Our study aimed at deciphering the mechanisms underlying the link between RAS and cellular components of the immune response relying on a rodent model of HF as well as HF patients. Flow cytometric analyses showed an increase in the expression of CD4+ AT2R+ cells in the rat heart and spleen post-infarction, but a reduction in the peripheral blood. The latter was also observed in HF patients. The frequency of rat CD4+ AT2R+ T cells in circulating blood, post-infarcted heart and spleen represented 3.8 ± 0.4%, 23.2 ± 2.7% and 22.6 ± 2.6% of the CD4+ cells. CD4+ AT2R+ T cells within blood CD4+ T cells were reduced from 2.6 ± 0.2% in healthy controls to 1.7 ± 0.4% in patients. Moreover, we characterized CD4+ AT2R+ T cells which expressed regulatory FoxP3, secreted interleukin-10 and other inflammatory-related cytokines. Furthermore, intramyocardial injection of MI-induced splenic CD4+ AT2R+ T cells into recipient rats with MI led to reduced infarct size and improved cardiac performance. We defined CD4+ AT2R+ cells as a T cell subset improving heart function post-MI corresponding with reduced infarction size in a rat MI-model. Our results indicate CD4+ AT2R+ cells as a promising population for regenerative therapy, via myocardial transplantation, pharmacological AT2R activation or a combination thereof.