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Specific bone cells produce DLL4 to generate thymus-seeding progenitors from bone marrow.

Yu VW, Saez B, Cook C, Lotinun S, Pardo-Saganta A, Wang YH, Lymperi S, Ferraro F, Raaijmakers MH, Wu JY, Zhou L, Rajagopal J, Kronenberg HM, Baron R, Scadden DT - J. Exp. Med. (2015)

Bottom Line: Here, we report that specific deletion of bone-producing osteocalcin (Ocn)-expressing cells in vivo markedly reduces T-competent progenitors and thymus-homing receptor expression among bone marrow hematopoietic cells.Decreased intrathymic T cell precursors and decreased generation of mature T cells occurred despite normal thymic function.These data indicate that specific mesenchymal cells in bone marrow provide key molecular drivers enforcing thymus-seeding progenitor generation and thereby directly link skeletal biology to the production of T cell-based adaptive immunity.

View Article: PubMed Central - HTML - PubMed

Affiliation: Center for Regenerative Medicine, Massachusetts General Hospital, Boston, MA 02215 Harvard Stem Cell Institute, Cambridge, MA 02215 Department of Stem Cell and Regenerative Biology, Harvard University, Cambridge, MA 02215.

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Ocn+ cell–specific deletion in vivo without altering osteoclastogenesis and mesenchymal progenitors. (A) WT mice (Ctrl) Ocn+ osteolineage cell deletion mice (Mut) were monitored for body size and weight; n = 8–10 mice/group. Data show mean ± SEM. (B) Femurs and tibiae in the OcnCre+/−;iDTR mutants or WT (Ctrl) mice were assessed histologically. Bottom images are at a higher magnification with arrows pointing to empty lacunae within the cortex and altered endosteal surface; images reflect comparable findings in all animals; n = 8/experiment. (C) Osteoblasts in the OcnCre+/−;iDTR and WT mice were quantitated by histomorphometry; n = 7–8 mice/group. Data show mean ± SEM. (D and E) Ocn and DTR expression was examined in bone sections from untreated OcnCre+/−;iDTR by immunohistochemistry using Ocn- and DTR-specific antibodies (D), or by immunofluorescence using Ocn-specific antibodies and TUNEL staining after DT treatment (E); n = 6 mice/group. (F) Osteoclast numbers were assessed by TRAP staining (n = 6 mice/group) and (G) osteoclast activity by collagen breakdown in sera using ELISA assay; n = 7–8 mice/group. Data show mean ± SEM. (H) Mesenchymal progenitor activity in the bone marrow of OcnCre+/−;iDTR mutants or WT controls was assessed by CFU-Ob assay; n = 9 mice/group. Data show mean ± SEM. (I) CD31−CD45−Ter119−LepR+ cells in the bone marrow stroma of OcnCre+/−;iDTR mutants and controls were quantified by flow cytometry; n = 6–7 mice/group. Data show mean ± SEM. (A–I) For each experiment, 3–6 independent repeats were performed.
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fig1: Ocn+ cell–specific deletion in vivo without altering osteoclastogenesis and mesenchymal progenitors. (A) WT mice (Ctrl) Ocn+ osteolineage cell deletion mice (Mut) were monitored for body size and weight; n = 8–10 mice/group. Data show mean ± SEM. (B) Femurs and tibiae in the OcnCre+/−;iDTR mutants or WT (Ctrl) mice were assessed histologically. Bottom images are at a higher magnification with arrows pointing to empty lacunae within the cortex and altered endosteal surface; images reflect comparable findings in all animals; n = 8/experiment. (C) Osteoblasts in the OcnCre+/−;iDTR and WT mice were quantitated by histomorphometry; n = 7–8 mice/group. Data show mean ± SEM. (D and E) Ocn and DTR expression was examined in bone sections from untreated OcnCre+/−;iDTR by immunohistochemistry using Ocn- and DTR-specific antibodies (D), or by immunofluorescence using Ocn-specific antibodies and TUNEL staining after DT treatment (E); n = 6 mice/group. (F) Osteoclast numbers were assessed by TRAP staining (n = 6 mice/group) and (G) osteoclast activity by collagen breakdown in sera using ELISA assay; n = 7–8 mice/group. Data show mean ± SEM. (H) Mesenchymal progenitor activity in the bone marrow of OcnCre+/−;iDTR mutants or WT controls was assessed by CFU-Ob assay; n = 9 mice/group. Data show mean ± SEM. (I) CD31−CD45−Ter119−LepR+ cells in the bone marrow stroma of OcnCre+/−;iDTR mutants and controls were quantified by flow cytometry; n = 6–7 mice/group. Data show mean ± SEM. (A–I) For each experiment, 3–6 independent repeats were performed.

Mentions: We generated mouse strains in which Cre recombinase produced by either the Osteocalcin promoter expressed in mature osteoblasts and osteocytes, or the Osterix promoter expressed in distinct, more immature subsets of bone cells, drives expression of the diphtheria toxin (DT) receptor (DTR) on cell surface (OcnCre+/−;iDTR and OsxCre+/−;iDTR, respectively; OcnCre+/− and OsxCre+/− served as controls). Specific in vivo cell ablation was achieved by intraperitoneal injection of DT. Daily injections into both control and mutant animals began at age 4 wk, and by 6 wk a difference in body size was noted in both the OsxCre+/−;iDTR and OcnCre+/−;iDTR mutant mice compared with littermate controls, which is consistent with inhibition of bone formation (Fig. 1 A). In early experiments, OsxCre+/−;iDTR and OcnCre+/−;iDTR animals without DT treatment were assessed and no phenotypic difference with the OsxCre+/− and OcnCre+/− controls were noted and therefore are not presented further. The T lymphopenic effect was observed only in the OcnCre+/−;iDTR strain and not the OsxCre+/−;iDTR strain, and thus it is the focus of this work.


Specific bone cells produce DLL4 to generate thymus-seeding progenitors from bone marrow.

Yu VW, Saez B, Cook C, Lotinun S, Pardo-Saganta A, Wang YH, Lymperi S, Ferraro F, Raaijmakers MH, Wu JY, Zhou L, Rajagopal J, Kronenberg HM, Baron R, Scadden DT - J. Exp. Med. (2015)

Ocn+ cell–specific deletion in vivo without altering osteoclastogenesis and mesenchymal progenitors. (A) WT mice (Ctrl) Ocn+ osteolineage cell deletion mice (Mut) were monitored for body size and weight; n = 8–10 mice/group. Data show mean ± SEM. (B) Femurs and tibiae in the OcnCre+/−;iDTR mutants or WT (Ctrl) mice were assessed histologically. Bottom images are at a higher magnification with arrows pointing to empty lacunae within the cortex and altered endosteal surface; images reflect comparable findings in all animals; n = 8/experiment. (C) Osteoblasts in the OcnCre+/−;iDTR and WT mice were quantitated by histomorphometry; n = 7–8 mice/group. Data show mean ± SEM. (D and E) Ocn and DTR expression was examined in bone sections from untreated OcnCre+/−;iDTR by immunohistochemistry using Ocn- and DTR-specific antibodies (D), or by immunofluorescence using Ocn-specific antibodies and TUNEL staining after DT treatment (E); n = 6 mice/group. (F) Osteoclast numbers were assessed by TRAP staining (n = 6 mice/group) and (G) osteoclast activity by collagen breakdown in sera using ELISA assay; n = 7–8 mice/group. Data show mean ± SEM. (H) Mesenchymal progenitor activity in the bone marrow of OcnCre+/−;iDTR mutants or WT controls was assessed by CFU-Ob assay; n = 9 mice/group. Data show mean ± SEM. (I) CD31−CD45−Ter119−LepR+ cells in the bone marrow stroma of OcnCre+/−;iDTR mutants and controls were quantified by flow cytometry; n = 6–7 mice/group. Data show mean ± SEM. (A–I) For each experiment, 3–6 independent repeats were performed.
© Copyright Policy - openaccess
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4419348&req=5

fig1: Ocn+ cell–specific deletion in vivo without altering osteoclastogenesis and mesenchymal progenitors. (A) WT mice (Ctrl) Ocn+ osteolineage cell deletion mice (Mut) were monitored for body size and weight; n = 8–10 mice/group. Data show mean ± SEM. (B) Femurs and tibiae in the OcnCre+/−;iDTR mutants or WT (Ctrl) mice were assessed histologically. Bottom images are at a higher magnification with arrows pointing to empty lacunae within the cortex and altered endosteal surface; images reflect comparable findings in all animals; n = 8/experiment. (C) Osteoblasts in the OcnCre+/−;iDTR and WT mice were quantitated by histomorphometry; n = 7–8 mice/group. Data show mean ± SEM. (D and E) Ocn and DTR expression was examined in bone sections from untreated OcnCre+/−;iDTR by immunohistochemistry using Ocn- and DTR-specific antibodies (D), or by immunofluorescence using Ocn-specific antibodies and TUNEL staining after DT treatment (E); n = 6 mice/group. (F) Osteoclast numbers were assessed by TRAP staining (n = 6 mice/group) and (G) osteoclast activity by collagen breakdown in sera using ELISA assay; n = 7–8 mice/group. Data show mean ± SEM. (H) Mesenchymal progenitor activity in the bone marrow of OcnCre+/−;iDTR mutants or WT controls was assessed by CFU-Ob assay; n = 9 mice/group. Data show mean ± SEM. (I) CD31−CD45−Ter119−LepR+ cells in the bone marrow stroma of OcnCre+/−;iDTR mutants and controls were quantified by flow cytometry; n = 6–7 mice/group. Data show mean ± SEM. (A–I) For each experiment, 3–6 independent repeats were performed.
Mentions: We generated mouse strains in which Cre recombinase produced by either the Osteocalcin promoter expressed in mature osteoblasts and osteocytes, or the Osterix promoter expressed in distinct, more immature subsets of bone cells, drives expression of the diphtheria toxin (DT) receptor (DTR) on cell surface (OcnCre+/−;iDTR and OsxCre+/−;iDTR, respectively; OcnCre+/− and OsxCre+/− served as controls). Specific in vivo cell ablation was achieved by intraperitoneal injection of DT. Daily injections into both control and mutant animals began at age 4 wk, and by 6 wk a difference in body size was noted in both the OsxCre+/−;iDTR and OcnCre+/−;iDTR mutant mice compared with littermate controls, which is consistent with inhibition of bone formation (Fig. 1 A). In early experiments, OsxCre+/−;iDTR and OcnCre+/−;iDTR animals without DT treatment were assessed and no phenotypic difference with the OsxCre+/− and OcnCre+/− controls were noted and therefore are not presented further. The T lymphopenic effect was observed only in the OcnCre+/−;iDTR strain and not the OsxCre+/−;iDTR strain, and thus it is the focus of this work.

Bottom Line: Here, we report that specific deletion of bone-producing osteocalcin (Ocn)-expressing cells in vivo markedly reduces T-competent progenitors and thymus-homing receptor expression among bone marrow hematopoietic cells.Decreased intrathymic T cell precursors and decreased generation of mature T cells occurred despite normal thymic function.These data indicate that specific mesenchymal cells in bone marrow provide key molecular drivers enforcing thymus-seeding progenitor generation and thereby directly link skeletal biology to the production of T cell-based adaptive immunity.

View Article: PubMed Central - HTML - PubMed

Affiliation: Center for Regenerative Medicine, Massachusetts General Hospital, Boston, MA 02215 Harvard Stem Cell Institute, Cambridge, MA 02215 Department of Stem Cell and Regenerative Biology, Harvard University, Cambridge, MA 02215.

Show MeSH
Related in: MedlinePlus