RNA-seq in human hematopoietic stem cells (HSCs) cultures on 3 different materials

Hematopoietic stem cell transplantation (HSCT) is successfully applied since the late 1950s, however, its efficacy still needs to be improved. A promising strategy is to transplant high numbers of pluripotent hematopoietic stem cells (HSCs). Therefore, an advanced ex vivo culture system is needed that supports the proliferation and maintains the pluripotency of HSC to override possible limitations in cell numbers gained from donors. To model the natural HSC niche in vitro and thus, to amplify high numbers of undifferentiated HSCs, we used an optimized HSC cell culture medium in combination with artificial 3D bone marrow-like scaffolds made of polydimethylsiloxane (PDMS). After 14 days in vitro (DIV) cell culture, we performed transcriptome and proteome analysis of the whole cell populations. Ingenuity pathway analysis (IPA) indicated that our 3D PDMS cell culture scaffolds altered PI3K/AKT/mTOR pathways and activated SREBP, HIF1α and FOXO signaling leading to metabolic adaptations, which we confirmed by ELISA, Western blot and metabolic flux analysis. These molecular signaling pathways can promote the expansion HSCs and are involved in their pluripotency maintenance. Thus, we could show that our 3D bone marrow-like PDMS scaffolds activate key molecular signaling pathways to amplify the numbers of undifferentiated HSCs efficiently ex vivo. 16 samples: HSCs of 4 anonymous human indivduals were used. Of each donor, a control sample was take before in vitro amplification ("Day0") while 3 fractions of HSCs were cultured on different materials ("2D PS", "3D PDMS" and "3D SiOn") for 14 days. The study has been approved by the Jena University Hospital Ethics Committee (#3595-10/12, #5320-10/17). ***Please note that raw data is not provided as consent forms do not allow for public access to raw data****