No abstract

Summary Pluripotency is tightly regulated and is crucial for stem cells and their implementation for regenerative medicine. Non-coding RNAs, especially long non-coding RNAs (lncRNAs) emerged as orchestrators of versatile (patho)-physiological processes on the transcriptional and post-transcriptional level. Cyrano , a well-conserved lncRNA, is highly expressed in stem cells suggesting an important role in pluripotency, which we aimed to investigate in loss-off-function (LOF) experiments. Cyrano was described previously to be essential for the maintenance of mouse embryonic stem cell (ESC) pluripotency. In contrast, using different genetic models, we here found Cyrano to be dispensable in murine and human iPSCs and in human ESCs. RNA sequencing revealed only a moderate influence of Cyrano on the global transcriptome. In line, Cyrano -depleted iPSCs retained the potential to differentiate into the three germ layers. In conclusion, different methods were applied for LOF studies to rule out potential off-target effects. These approaches revealed that Cyrano does not impact pluripotency.

Myocardial infarction causes a massive loss of cardiomyocytes (CMs), which can lead to heart failure accompanied by fibrosis, stiffening of the heart, and loss of function. Heart failure causes high mortality rates and is a huge socioeconomic burden, which, based on diets and lifestyle in the developed world, is expected to increase further in the next years. At present, the only curative treatment for heart failure is heart transplantation associated with a number of limitations such as donor organ availability and transplant rejection among others. Thus, the development of cellular reprogramming and defined differentiation protocols provide exciting new possibilities for cell therapy approaches and which opened up a new era in regenerative medicine. Consequently, tremendous research efforts were undertaken to gain a detailed molecular understanding of the reprogramming processes and the in vitro differentiation of pluripotent stem cells into functional CMs for transplantation into the patient’s injured heart. In the last decade, non-coding RNAs, particularly microRNAs, long non-coding RNAs, and circular RNAs emerged as critical regulators of gene expression that were shown to fine-tune cellular processes both on the transcriptional and the post-transcriptional level. Unsurprisingly, also cellular reprogramming, pluripotency, and cardiac differentiation and maturation are regulated by non-coding RNAs. In here, we review the current knowledge on non-coding RNAs in these processes and highlight how their modulation may enhance the quality and quantity of stem cells and their derivatives for safe and efficient clinical application in patients with heart failure. In addition, we summarize the clinical cell therapy efforts undertaken thus far.

Cardiac fibroblasts constitute the major cell type of the murine and human heart. Once activated, they contribute to an excessive deposition of extracellular matrix (ECM) leading to cardiac fibrosis and subsequently organ dysfunction. With the exception of the pulmonary drugs, nintedanib and pirfenidone, drugs specifically targeting anti-fibrotic pathways are scarce. We recently performed large library screenings of natural occurring compounds and identified first lead structures with anti-fibrotic properties in vitro and in vivo. In line, we now aimed to improve efficacy of these anti-fibrotic lead structures by combining in vitro validation studies and in silico prediction. Next to this combined approach, we performed large OMICs-multi-panel-based mechanistic studies. Applying human cardiac fibroblasts (HCF), we analysed 26 similars of the initially identified anti-fibrotic lead molecules bufalin and lycorine and determined anti-proliferative activity and potential toxicity in an array of in vitro and ex vivo studies. Of note, even at lower concentrations, certain similars were more effective at inhibiting HCF proliferation than nintedanib and pirfenidone. Additionally, selected similars showed low cytotoxicity on human iPS-derived cardiomyocytes and anti-fibrotic gene regulation in human ex vivo living myocardial slices. Further, array and RNA sequencing studies of coding and non-coding RNAs in treated HCFs revealed strong anti-fibrotic properties, especially with the lycorine similar lyco-s (also known as homoharringtonine), that led to a nearly complete shutdown of ECM production at concentrations 100-fold lower than the previously identified anti-fibrotic compound lycorine without inducing cellular toxicity. We thus identified a new natural compound similar with strong anti-fibrotic properties in human cardiac fibroblasts and human living heart tissue potentially opening new anti-fibrotic treatment strategies.

Takotsubo syndrome (TTS), an acute cardiac condition characterized by transient wall motion abnormalities mostly of the left ventricle, results in difficulties in diagnosing patients. We set out to present a detailed blood analysis of TTS patients analyzing novel markers to understand the development of TTS. Significant differences in pro-inflammatory cytokine expression patterns, sex steroid and glucocorticoid receptor (GR) expression levels were observed in the TTS patient collective. Remarkably, the measured catecholamine serum concentrations determined from TTS patient blood could be shown to be two orders of magnitude lower than the levels determined from experimentally induced TTS in laboratory animals. Consequently, exposure of endothelial cells and cardiomyocytes in vitro to such catecholamine concentrations did not damage the cellular integrity or function of neither endothelial cells forming the blood brain barrier, endothelial cells derived from myocardium nor cardiomyocytes in vitro. Computational analysis was able to link the identified blood markers, specifically, the pro-inflammatory cytokines and glucocorticoid receptor GR to microRNA (miR) relevant in the ontogeny of TTS (miR-15), and inflammation (miR-21, miR-146a), respectively. Amongst the well-described risk factors of TTS (older age, female gender), inflammaging-related pathways were identified to add to these relevant risk factors or prediagnostic markers of TTS.