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Abstract “Medical scientists” are postgraduate investigators who are engaged in biomedical research, and either hold a biomedical PhD or are qualified in medicine but do not participate in patient care. Medical scientists constitute ~40% of staff at medical faculties and >90% at nonuniversity medical research institutions in Germany. However, medical scientists in Germany face limited long‐term career prospects and a lack of dedicated training and support programmes. They also face time limits on their career progression arising from national academic employment legislation, and imminent reforms by the German government are likely to make this worse. Nevertheless, recent developments in the educational landscape including the introduction of increasingly focused MSc, pre‐PhD, and doctoral programmes to train medically aware basic scientists, as well as improved general recognition of the roles and relevance of medical scientists in health research, are encouraging. Physiologists have taken essential steps to improve the recognition of medical scientists in Germany by introducing a “specialist physiologist” qualification; this initiative could be applied to support medical scientists in other fields and countries. In this review, we describe the particular challenges facing medical scientists in Germany and make recommendations that may apply to other academic systems.

The fluorescence ubiquitination cell cycle inhibitor (FUCCI) has been introduced to monitor cell cycle activity in living cells, including human induced pluripotent stem cells (hiPSC) and derived cell types. We have recently developed hiPSC with stable expression of dCas9VPR for endogenous gene activation and a Citrine-tagged ACTN2 cell line to monitor sarcomere development and function in muscle cells. Here, we present dual and triple transgenic hiPSC lines developed by genomic integration of FUCCI with and without dCas9VPR into the ROSA26 and AAVS1 loci, respectively, in the previously introduced ACTN2-Citrine line. Functionality of the transgenes was demonstrated in the novel hiPSC line, which we introduce as Myo-CCER and CraCCER.

Gene variants in LZTR1 are implicated to cause Noonan syndrome associated with a severe and early-onset hypertrophic cardiomyopathy. Mechanistically, LZTR1 deficiency results in accumulation of RAS GTPases and, as a consequence, in RAS-MAPK signaling hyperactivity, thereby causing the Noonan syndrome-associated phenotype. Despite its epidemiological relevance, pharmacological as well as invasive therapies remain limited. Here, personalized CRISPR-Cas9 gene therapies might offer a novel alternative for a curative treatment in this patient cohort. In this study, by utilizing a patient-specific screening platform based on iPSC-derived cardiomyocytes from two Noonan syndrome patients, we evaluated different clinically translatable therapeutic approaches using small Cas9 orthologs targeting a deep-intronic LZTR1 variant to cure the disease-associated molecular pathology. Despite high editing efficiencies in cardiomyocyte cultures transduced with lentivirus or all-in-one adeno-associated viruses, we observed crucial differences in editing outcomes in proliferative iPSCs vs. non-proliferative cardiomyocytes. While editing in iPSCs rescued the phenotype, the same editing approaches did not robustly restore LZTR1 function in cardiomyocytes, indicating critical differences in the activity of DNA double-strand break repair mechanisms between proliferative and non-proliferative cell types and highlighting the importance of cell type-specific screens for testing CRISPR-Cas9 gene therapies.

Abstract Aberrant Wnt activation has been reported in failing cardiomyocytes. Here we present single cell transcriptome profiling of hearts with inducible cardiomyocyte-specific Wnt activation (β-catΔex3) as well as with compensatory and failing hypertrophic remodeling. We show that functional enrichment analysis points to an involvement of extracellular vesicles (EVs) related processes in hearts of β-catΔex3 mice. A proteomic analysis of in vivo cardiac derived EVs from β-catΔex3 hearts has identified differentially enriched proteins involving 20 S proteasome constitutes, protein quality control (PQC), chaperones and associated cardiac proteins including α-Crystallin B (CRYAB) and sarcomeric components. The hypertrophic model confirms that cardiomyocytes reacted with an acute early transcriptional upregulation of exosome biogenesis processes and chaperones transcripts including CRYAB, which is ameliorated in advanced remodeling. Finally, human induced pluripotent stem cells (iPSC)-derived cardiomyocytes subjected to pharmacological Wnt activation recapitulated the increased expression of exosomal markers, CRYAB accumulation and increased PQC signaling. These findings reveal that secretion of EVs with a proteostasis signature contributes to early patho-physiological adaptation of cardiomyocytes, which may serve as a read-out of disease progression and can be used for monitoring cellular remodeling in vivo with a possible diagnostic and prognostic role in the future.

Abstract Wnt/β‐catenin signalling controls adult heart remodelling in part via regulation of cardiac progenitor cell (CPC) differentiation. An enhanced understanding of mechanisms controlling CPC biology might facilitate the development of new therapeutic strategies in heart failure. We identified and characterized a novel cardiac interaction between Krueppel‐like factor 15 and components of the Wnt/β‐catenin pathway leading to inhibition of transcription. In vitro mutation, reporter assays and co‐localization analyses revealed that KLF15 requires both the C‐terminus, necessary for nuclear localization, and a minimal N‐terminal regulatory region to inhibit transcription. In line with this, functional Klf15 knock‐out mice exhibited cardiac β‐catenin transcriptional activation along with functional cardiac deterioration in normal homeostasis and upon hypertrophy. We further provide in vivo and in vitro evidences for preferential endothelial lineage differentiation of CPCs upon KLF15 deletion. Via inhibition of β‐catenin transcription, KLF15 controls CPC homeostasis in the adult heart similar to embryonic cardiogenesis. This knowledge may provide a tool for reactivation of this apparently dormant CPC population in the adult heart and thus be an attractive approach to enhance endogenous cardiac repair.

The Rho guanine nucleotide exchange factor RhoGEF17 was described to reside in adherens junctions (AJ) in endothelial cells (EC) and to play a critical role in the regulation of cell adhesion and barrier function. The purpose of this study was to analyze signal cascades and processes occurring subsequent to AJ disruption induced by RhoGEF17 knockdown. Primary human and immortalized rat EC were used to demonstrate that an adenoviral-mediated knockdown of RhoGEF17 resulted in cell rounding and an impairment in spheroid formation due to an enhanced proteasomal degradation of AJ components. In contrast, β-catenin degradation was impaired, which resulted in an induction of the β-catenin-target genes cyclin D1 and survivin. RhoGEF17 depletion additionally inhibited cell adhesion and sheet migration. The RhoGEF17 knockdown prevented the cells with impeded cell–cell and cell–matrix contacts from apoptosis, which was in line with a reduction in pro-caspase 3 expression and an increase in Akt phosphorylation. Nevertheless, the cells were not able to proliferate as a cell cycle block occurred. In summary, we demonstrate that a loss of RhoGEF17 disturbs cell–cell and cell–substrate interaction in EC. Moreover, it prevents the EC from cell death and blocks cell proliferation. Non-canonical β-catenin signaling and Akt activation could be identified as a potential mechanism.