Bulk RNA seq data for sorted human CD34+CD38low cells after DMSO or 78c treatment
Hematopoietic stem cells (HSCs) balance self-renewal and differentiation within the bone marrow, a process that is disrupted under in vitro conditions, posing challenges for ex vivo studies and clinical applications such as gene therapy. This study examines the role of CD38, a glycoprotein involved in NAD+ metabolism, in regulating HSC function. Inhibition of CD38 activity with the small molecule 78c significantly increased the percentage of phenotypic HSCs and maintained their engraftment potential in mice, akin to uncultured cells.Here we performed bulk RNA seq on DMSO and 78c treated human CD34+CD38low cells to look for the transcriptomic differences between the two groups. Overall design: Human CD34+CD38low hematopoietic stem and progenitor cells isolated from mobilized peripheral blood CD34+ cells and cultured for 24 hours in the presence of cytokines (SCF, FLT3, and TPO) with either 5 µM of the CD38 inhibitor 78c or a DMSO vehicle control. Following culture, cells were harvested, and RNA was extracted using the RNeasy Plus Mini Kit (Qiagen) as per the manufacturer's protocol. The sequencing libraries were constructed from 800 ng of total RNA using the Illumina Stranded mRNA library prep kit (#20040534, Illumina) following the manufacturer instruction. The fragment size of RNA seq libraries was verified using the Agilent 2100 Bioanalyzer (Agilent) and the concentrations were determined using Qubit instrument (LifeTech). The libraries were loaded onto the Illumina Novaseq 6000 SP flow cell for 2x100 bp paired end read sequencing, generating approximately 50 million reads per sample. The fastq files were generated using the bcl2fastq software for further analysis. Subsequent analysis involved aligning the reads to a reference genome, followed by differential gene expression analysis to identify key pathways and molecular mechanisms modulated by CD38 inhibition.