Developmental Vitamin D Availability Impacts Hematopoietic Stem Cell Production

SUMMARY Vitamin D insufficiency is a worldwide epidemic affecting billions of individuals, including pregnant women and children. Despite its high incidence, the impact of active vitamin D3 (1,25(OH)D3) on embryonic development beyond osteo-regulation remains largely undefined. Here, we demonstrate that 1,25(OH)D3 availability modulates zebrafish hematopoietic stem and progenitor cell (HSPC) production. Loss of Cyp27b1-mediated biosynthesis or vitamin D receptor (VDR) function by gene knockdown resulted in significantly reduced runx1 expression and Flk1+cMyb+ HSPC numbers. Selective modulation in vivo and in vitro in zebrafish indicated that vitamin D3 acts directly on HSPCs, independent of calcium regulation, to increase proliferation. Notably, ex vivo treatment of human HSPCs with 1,25(OH)D3 also enhanced hematopoietic colony numbers, illustrating conservation across species. Finally, gene expression and epistasis analysis indicated that CXCL8 (IL-8) was a functional target of vitamin D3-mediated HSPC regulation. Together, these findings highlight the relevance of developmental 1,25(OH)D3 availability for definitive hematopoiesis and suggest potential therapeutic utility in HSPC expansion

Inflammatory signaling regulates embryonic hematopoietic stem and progenitor cell production

Identifying signaling pathways that regulate hematopoietic stem and progenitor cell (HSPC) formation in the embryo will guide efforts to produce and expand HSPCs ex vivo. Here we show that sterile tonic inflammatory signaling regulates embryonic HSPC formation. Expression profiling of progenitors with lymphoid potential and hematopoietic stem cells (HSCs) from aorta/gonad/mesonephros (AGM) regions of midgestation mouse embryos revealed a robust innate immune/inflammatory signature. Mouse embryos lacking interferon γ (IFN-γ) or IFN-α signaling and zebrafish morphants lacking IFN-γ and IFN-ϕ activity had significantly fewer AGM HSPCs. Conversely, knockdown of IFN regulatory factor 2 (IRF2), a negative regulator of IFN signaling, increased expression of IFN target genes and HSPC production in zebrafish. Chromatin immunoprecipitation (ChIP) combined with sequencing (ChIP-seq) and expression analyses demonstrated that IRF2-occupied genes identified in human fetal liver CD34(+) HSPCs are actively transcribed in human and mouse HSPCs. Furthermore, we demonstrate that the primitive myeloid population contributes to the local inflammatory response to impact the scale of HSPC production in the AGM region. Thus, sterile inflammatory signaling is an evolutionarily conserved pathway regulating the production of HSPCs during embryonic development

Lipid-Bicelle-Coated Microfluidics for Intracellular Delivery with Reduced Fouling

Innovative technologies for intracellular delivery are ushering in a new era for gene editing, enabling the utilization of a patient's own cells for stem cell and immunotherapies. In particular, cell-squeezing platforms provide unconventional forms of intracellular delivery, deforming cells through microfluidic constrictions to generate transient pores and to enable effective diffusion of biomolecular cargo. While these devices are promising gene-editing platforms, they require frequent maintenance due to the accumulation of cellular debris, limiting their potential for reaching the throughputs necessary for scalable cellular therapies. As these cell-squeezing technologies are improved, there is a need to develop next-generation platforms with higher throughput and longer lifespan, importantly, avoiding the buildup of cell debris and thus channel clogging. Here, we report a versatile strategy to coat the channels of microfluidic devices with lipid bilayers based on noncovalent lipid bicelle technology, which led to substantial improvements in reducing cell adhesion and protein adsorption. The antifouling properties of the lipid bilayer coating were evaluated, including membrane uniformity, passivation against nonspecific protein adsorption, and inhibition of cell attachment against multiple cell types. This surface functionalization approach was applied to coat constricted microfluidic channels for the intracellular delivery of fluorescently labeled dextran and plasmid DNA, demonstrating significant reductions in the accumulation of cell debris. Taken together, our work demonstrates that lipid bicelles are a useful tool to fabricate antifouling lipid bilayer coatings in cell-squeezing devices, resulting in reduced nonspecific fouling and cell clogging to improve performance

Precision Medicine in Pediatric Neurooncology: A Review

Central nervous system tumors are the leading cause of cancer related death in children. Despite much progress in the field of pediatric neurooncology, modern combination treatment regimens often result in significant late effects, such as neurocognitive deficits, endocrine dysfunction, secondary malignancies, and a host of other chronic health problems. Precision medicine strategies applied to pediatric neurooncology target specific characteristics of individual patients' tumors to achieve maximal killing of neoplastic cells while minimizing unwanted adverse effects. Here, we review emerging trends and the current literature that have guided the development of new molecularly based classification schemas, promising diagnostic techniques, targeted therapies, and delivery platforms for the treatment of pediatric central nervous system tumors