Fig. 3: Transcriptional profiling of competitively transplanted HSPCs and identification of relevant signal transduction pathways. | Nature Communications

Fig. 3: Transcriptional profiling of competitively transplanted HSPCs and identification of relevant signal transduction pathways.

From: Haploinsufficiency at the CX3CR1 locus of hematopoietic stem cells favors the appearance of microglia-like cells in the central nervous system of transplant recipients

Fig. 3

A Frequency of donor CD45.2+ cells in the BM and brain of competitive-transplant recipients sacrificed at 15 days post-transplant. B Frequency of Cx3cr1+/+ and Cx3cr1−/+ cells within total donor cells in the BM and brain of competitive-transplant recipients, compared to the infused HSPCs (input). For (A) and (B), n = 8, 3 independent experiments. Mean and individual values ± SEM are shown. Statistical analysis: Mann–Whitney test; *p < 0.05, **p < 0.01, ***p < 0.001. C Two-dimensional UMAP representation of the microglia single-cell transcriptome dataset. Each dot represents a cell, and colors indicate the genotypes, as indicated. D UMAP plot showing the clusters identified using the Louvain algorithm. E Bar graph showing the genotype (Cx3cr1+/+ and Cx3cr1−/+) frequency in each cluster. F Gene ontology analysis on cluster 4, enriched in Cx3cr1−/+ cells, showing a significant upregulation of Cdc42 pathway-related genes, microglia migration, and prototypical microglia differentiation pathways. G UMAP plots showing gene expression of 4 (out of 16) genes of the Cdc42 protein signal transduction pathway. H Mean fluorescent intensity (MFI) of Cdc42 expression in wild type and Cx3cr1−/+ microglia cells isolated from naïve mice. n = 10/group. Mean and individual values ± SEM are shown. Statistical analysis: Mann–Whitney test; *p < 0.05, **p < 0.01, ***p < 0.001. I Migration of Cx3cr1+/+ and Cx3cr1−/+ HSPCs isolated from naïve Cx3cr1+/+and Cx3cr1−/+ mice towards fractalkine (FLK) added at different concentrations (100 and 200 ng/mL) in a trans-well system. SDF-1 (50 nM) was employed as a positive control for HSPC migration. RAW 264.7, a murine macrophage cell line expressing high levels of Cx3cr1, was used as a positive control for migration toward FKN. The percentage of migrating cells was analyzed after 24 h. n ≥ 5 per group, 3 independent experiments. Mean v and individual values ± SD are shown. Statistical analysis: Mann–Whitney test; *p < 0.05, **p < 0.01, ***p < 0.001. Source data are provided as a Source Data file.

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