Neural differentiation potential of human bone marrow-derived mesenchymal stromal cells: misleading marker gene expression

Background: In contrast to pluripotent embryonic stem cells, adult stem cells have been considered to be multipotent, being somewhat more restricted in their differentiation capacity and only giving rise to cell types related to their tissue of origin. Several studies, however, have reported that bone marrow-derived mesenchymal stromal cells (MSCs) are capable of transdifferentiating to neural cell types, effectively crossing normal lineage restriction boundaries. Such reports have been based on the detection of neural-related proteins by the differentiated MSCs. In order to assess the potential of human adult MSCs to undergo true differentiation to a neural lineage and to determine the degree of homogeneity between donor samples, we have used RT-PCR and immunocytochemistry to investigate the basal expression of a range of neural related mRNAs and proteins in populations of non-differentiated MSCs obtained from 4 donors. Results: The expression analysis revealed that several of the commonly used marker genes from other studies like nestin, Enolase2 and microtubule associated protein 1b (MAP1b) are already expressed by undifferentiated human MSCs. Furthermore, mRNA for some of the neural-related transcription factors, e.g. Engrailed-1 and Nurr1 were also strongly expressed. However, several other neural-related mRNAs (e.g. DRD2, enolase2, NFL and MBP) could be identified, but not in all donor samples. Similarly, synaptic vesicle-related mRNA, STX1A could only be detected in 2 of the 4 undifferentiated donor hMSC samples. More significantly, each donor sample revealed a unique expression pattern, demonstrating a significant variation of marker expression. Conclusion: The present study highlights the existence of an inter-donor variability of expression of neuralrelated markers in human MSC samples that has not previously been described. This donor-related heterogeneity might influence the reproducibility of transdifferentiation protocols as well as contributing to the ongoing controversy about differentiation capacities of MSCs. Therefore, further studies need to consider the differences between donor samples prior to any treatment as well as the possibility of harvesting donor cells that may be inappropriate for transplantation strategies

Mapping cardiac remodeling in chronic kidney disease

Patients with advanced chronic kidney disease (CKD) mostly die from sudden cardiac death and recurrent heart failure. The mechanisms of cardiac remodeling are largely unclear. To dissect molecular and cellular mechanisms of cardiac remodeling in CKD in an unbiased fashion, we performed left ventricular single-nuclear RNA sequencing in two mouse models of CKD. Our data showed a hypertrophic response trajectory of cardiomyocytes with stress signaling and metabolic changes driven by soluble uremia-related factors. We mapped fibroblast to myofibroblast differentiation in this process and identified notable changes in the cardiac vasculature, suggesting inflammation and dysfunction. An integrated analysis of cardiac cellular responses to uremic toxins pointed toward endothelin-1 and methylglyoxal being involved in capillary dysfunction and TNFα driving cardiomyocyte hypertrophy in CKD, which was validated in vitro and in vivo. TNFα inhibition in vivo ameliorated the cardiac phenotype in CKD. Thus, interventional approaches directed against uremic toxins, such as TNFα, hold promise to ameliorate cardiac remodeling in CKD.</p

Biofabrication under fluorocarbon: a novel freeform fabrication technique to generate high aspect ratio tissue-engineered constructs

Bioprinting is a recent development in tissue engineering, which applies rapid prototyping techniques to generate complex living tissues. Typically, cell-containing hydrogels are dispensed layer-by-layer according to a computer-generated three-dimensional model. The lack of mechanical stability of printed hydrogels hinders the fabrication of high aspect ratio constructs. Here we present submerged bioprinting, a novel technique for freeform fabrication of hydrogels in liquid fluorocarbon. The high buoyant density of fluorocarbons supports soft hydrogels by floating. Hydrogel constructs of up to 30-mm height were generated. Using 3% (w/v) agarose as the hydrogel and disposable syringe needles as nozzles, the printer produced features down to 570-μm diameter with a lateral dispensing accuracy of 89 μm. We printed thin-walled hydrogel cylinders measuring 4.8 mm in height, with an inner diameter of ∼2.9 mm and a minimal wall thickness of ∼650 μm. The technique was successfully applied in printing a model of an arterial bifurcation. We extruded under fluorocarbon, cellularized alginate tubes with 5-mm outer diameter and 3-cm length. Cells grew vigorously and formed clonal colonies within the 7-day culture period. Submerged bioprinting thus seems particularly suited to fabricate hollow structures with a high aspect ratio like vascular grafts for cardiovascular tissue engineering as well as branching or cantilever-like structures, obviating the need for a solid support beneath the overhanging protrusions