Inherent engraftment differences between CD45.1 and CD45.2 HSCs are caused by differential expression of Cxcr4

The CD45.1/2 system is commonly used in mouse to discriminate the progeny of transplanted cells from those of the recipient. To distinguish the two genetic alleles, discriminatory antibodies specific for a single lysine to glutamic acid amino acid difference are available (Mercier et al; Stem Cell Reports, 2016). Since CD45 is expressed on most nucleated cell of the hematopoietic lineage, it is possible to analyse the origin of various stem, progenitor and differentiated cell populations in bone marrow reconstitution experiments by FACS and compare their respective contributions under competitive conditions. However, an important concern in the field is the observation, that CD45.2 cells are inherently advantageous to CD45.1 ones, even after extensive backcrossing probably due to genetic linkage (Waterstrat et al; Blood, 2010). This complicates the interpretation of published transgenic animal studies and requires carefully chosen controls. To investigate the functional reason for this competitive advantage, we performed RNA-seq of CD45.2 and CD45.1/2 hematopoietic stem cells (HSCs) isolated from competitive transplanted chimeras. Interestingly, one of the top differentially expressed genes was Cxcr4, a well known cell surface receptor that has been shown to be important for HSC maintenance and mobilization. In agreement, we found a gradual decrease of Cxcr4 mRNA and protein expression in CD45.1 compared to CD45.2 mice obtained from different suppliers. Next, we performed competitive transplantations, while treating recipient mice with a Cxcr4-inhibitor. Strikingly, this treatment led to the alleviation of the advantage seen upon transplantation of CD45.2 HSCs. Collectively, this data demonstrates the importance of using correct control mice for transplantation and expression analysis to discriminate mouse background-induced artefacts from genuine phenotypes.