GFP MSCV Retrovirus

Nucleotides that bind to P2 receptors have emerged as a member of the family of intercellular communication mediators. P2X7 is a member of the P2X family of ligand-gated ion channels responsible for responding to extracellular ATP. High levels of P2X7 expression were detected in leukemia samples, especially in relapsed cases. However, the role of P2X7-mediated signaling in hematopoietic stem/progenitor cells (HSPCs) and its potential role in leukemogenesis have not been determined. Here, researchers analyzed the expression of P2X7 in hematopoietic cells of different lineages and stages. P2X7 was overexpressed in HSPCs by retroviral infection to study its effects on HSPCs. The results showed that low levels of P2X7 expression were detected in HSPCs. Overexpression of P2X7 in HSPCs resulted in decreased in vitro clonogenicity and in vivo engraftment potential. These results suggest that high levels of purinergic signaling by P2X7 impair the function of HSPCs.

Since high expression of P2X7 was detected in samples from leukemia patients, it is crucial to explore whether overexpression of P2X7 in hematopoietic stem cells (HSPCs) affects their function. Mouse LKS cells were infected with blank MSCV-GFP retrovirus, retrovirus carrying WP2X7 or MP2X7, respectively. Flow cytometry analysis showed that 32.6%, 28.4% and 29.6% of GFP+ cells were detected in the three groups of cells, respectively (Figure 1a). After sorting, the GFP+ cells were named control group, WP2X7 group and MP2X7 group, respectively. The expression of P2X7 was verified by RT-PCR (Figure 1b) and confocal microscopy (Figure 1c).

Figure 1. Over-expression of P2X7 in LKS cells. (Feng W, et al., 2016)