Microinjection of human mesenchymal stem cells to mouse blastocysts

Purpose: 1. Compare transcriptomes of hESC-derived MSCs (EMSCs) and somatic MSCs (SMCS); 2. Reveal transriptomic profiles of EMSCs post injection into mouse blastocyst. Methods: For bulk seq on cultured human MSCs, mRNA was isolated using RNeasy Mini Kit (QIAGEN) and libraries was constructed using an Illumina kit, and then sequenced on NovaSeq platform. For single cell (sc) RNAseq on cultured human MSCs, cells were dissociated and lysed in the Oil-droplet Single Cell Capture System (Nadia Dolomite) and libraries were constructed using Nextera kits (Illumina), and then sequenced on Hiseq 150PE (Illumina). For EMSCs injected into mouse blastocyst, 30~70 embryos were pulled and lyzed in 10uL of Lysis Buffer for SmartSeq. Results: 1. Both in bulk and in single cell level, all types of MSCs share typical common markers; 2. Unignorable differences exhist among hES-derived and somatic MSCs; 3. Rapid mRNA profile shifts in metabolism, cellular skeleton, and immunoregulatory response happen in hES-derived MSCs post injection to mouse embryos. Conclusions: These experiments reveal transcriptomic alterations of EMSCs after injection into mouse blastocyst and transcriptomic profiles of human MSCs from different sources. Examination of transcriptomic profiles of human MSCs maintained in culture and injected into mouse blastocyst