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Ttc7a controls HSC functions
Furthermore, histological assessment of splenic sections revealed extramedullary hematopoiesis as evidenced by elevated counts of megakaryocytes (Figure 1C) and of hematopoietic stem and progenitor cells (HSPC) (Online Supplementary Figure S1). Relative to ctrl mice, the absolute splenic T-cell count in fsn mice was slightly lower at 3 weeks of age but higher at 6 and 12 weeks of age (Figure 1D). A large proportion of Ttc7a-deficient T lymphocytes had an effector memory phenotype (CD44+ CD62L-) (Figure 1E). Splenic B-cell counts were slightly elevated, and B cells presented the impaired maturation phenotype previously described in fsn mice6 (Figure 1F). The lym- phoid alterations were accompanied by massive myelo- proliferation, with an increase over time in the numbers of
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splenic granulocytes (both neutrophils and eosinophils) and resident and inflammatory monocytes (Figure 1G, H). Thus, Ttc7a-deficient mice displayed a number of persist- ent hematopoietic alterations (i.e., leukocytosis, T-lym- phocyte activation and anemia) at a very early age, where- as other manifestations appeared later in life and/or were exacerbated with age (i.e., myeloproliferation and elevat- ed T-cell counts).
Since all the peripheral hematopoietic lineages were affected in fsn mice, we next looked at whether the HSPC compartment was also altered. BM cellularity in fsn mice, in contrast to ctrl mice, increased between 3 and 12 weeks of age (Figure 2A). The LSK stem cell population was slightly higher in fsn mice than in ctrl mice at all the time
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Figure 1. Ttc7a-deficiency perturbs homeostasis of all immune populations. Control littermates (ctrl – black bars) and Ttc7a-deficient (fsn – red bars) mice were analyzed at 3, 6 and 12 weeks of age (mean ± standard error of mean) *P<0.05; **P<0.01; ***P<0.001; ****P<0.0001 (two-tailed t-test). (A) White blood cell count (n≥6). (B) Spleen size determined as percent of body weight (n≥6). (C) Histological sections of spleen stained with hematoxylin and eosin showing megakaryo- cytes (black arrow) (left panel) and quantification of spleen megakaryocytes (right panel). (D) Total number of T cells in the spleen (n≥7). (E) Flow cytometry repre- sentative of T-cell activation (according to CD44 and CD62L expression) at 12 weeks (left panel), and activation index of T cells [ratio of effector memory T cells (CD44+ CD62L-) to naïve T cells (CD44- CD62L+)] (right panel) of fsn and ctrl mice (n≥7). Numbers adjacent to the outlined areas indicate percent cells in the parent gate (mean). (F) Total number of B cells in the spleen (n=4) (left panel) and flow cytometry representative of B-cell maturation (according to IgM and IgD expression) (right panel). (G) Representative flow cytometry at 12 weeks (left panel) and total number of neutrophils (CD11b+ Ly6Ghi) and eosinophils (CD11b+ Ly6Gint SSChi). (H) Total number of inflammatory (CD11b+ Ly6G- Ly6C+) and resident monocytes (CD11b+ Ly6G- Ly6C-) (n≥6). Numbers adjacent to the outlined areas indicate percent cells among leukocytes in the spleen (mean).
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