Background aimsHuman umbilical cord–derived mesenchymal stromal cells (hUC-MSCs) are increasingly used in research and therapy. To obtain hUC-MSCs, a diversity of isolation and expansion methods... Show moreBackground aimsHuman umbilical cord–derived mesenchymal stromal cells (hUC-MSCs) are increasingly used in research and therapy. To obtain hUC-MSCs, a diversity of isolation and expansion methods are applied. Here, we report on a robust and standardized method for hUC-MSC isolation and expansion.MethodsUsing 90 hUC donors, we compared and optimized critical variables during each phase of the multi-step procedure involving UC collection, processing, MSC isolation, expansion and characterization. Furthermore, we assessed the effect of donor-to-donor variability regarding UC morphology and donor attributes on hUC-MSC characteristics.ResultsWe demonstrated robustness of our method across 90 UC donors at each step of the procedure. With our method, UCs can be collected up to 6 h after birth, and UC-processing can be initiated up to 48 h after collection without impacting on hUC-MSC characteristics. The removal of blood vessels before explant cultures improved hUC-MSC purity. Expansion in Minimum essential medium α supplemented with human platelet lysate increased reproducibility of the expansion rate and MSC characteristics as compared with Dulbecco's Modified Eagle's Medium supplemented with fetal bovine serum. The isolated hUC-MSCs showed a purity of ∼98.9%, a viability of >97% and a high proliferative capacity. Trilineage differentiation capacity of hUC-MSCs was reduced as compared with bone marrow-derived MSCs. Functional assays indicated that the hUC-MSCs were able to inhibit T-cell proliferation demonstrating their immune-modulatory capacity.ConclusionsWe present a robust and standardized method to isolate and expand hUC-MSCs, minimizing technical variability and thereby lay a foundation to advance reliability and comparability of results obtained from different donors and different studies. Show less
Lifelong health is thought to be partially set during intrauterine life by persistent epigenetic changes induced by the prenatal environment. To evaluate this hypothesis, we initiated a prospective... Show moreLifelong health is thought to be partially set during intrauterine life by persistent epigenetic changes induced by the prenatal environment. To evaluate this hypothesis, we initiated a prospective longitudinal study in monochorionic (MC) twins: the TwinLIFE study. MC twins are monozygotic, thus in origin genetically identical, and share a single placenta. Although MC twins have many environmental factors in common, in one-third of the MC twin pairs, one fetus has significantly less access to nutrients and resources during pregnancy than its co-twin often resulting in a significant discordance in prenatal growth. Hence, MC twins constitute a unique natural experiment to study the influence of the prenatal environment on health. In TwinLIFE, we will chart intrapair differences in DNA methylation focusing on mesenchymal stromal cells isolated from cord as an advanced proxy of epigenetic dysregulation relevant for long-term health consequences. Next, we will follow up the MC twins for growth, cardiovascular and neurodevelopmental outcomes during childhood and evaluate the impact of an epigenetic signature at birth on future health. The current target is to include 100 MC twin pairs, but we aim to continue enrollment after procuring additional funding. TwinLIFE will not only address an unmet clinical need in the high-risk group of MC twins, but may also advance early-life strategies to prevent adverse growth, cardiovascular and neurodevelopmental outcomes in the general population. Show less
