G-CSF Induces Alterations in the Bone Marrow Microenvironment That Suppress B Lymphopoiesis,

Abstract 1246 Infectious stress is associated with a shift in the bone marrow from lymphopoiesis to granulopoiesis. Expression of granulocyte colony-stimulating factor (G-CSF), the principal cytokine regulating granulopoiesis, is often induced during infection. We previously reported that G-CSF treatment is associated with marked suppression of B lymphopoiesis in murine bone marrow. After 5 days of G-CSF treatment (250 μg/kg), total B cells in the bone marrow were reduced 8.1 ± 0.9-fold. Pre-pro-B cells were reduced 1.6 ± 0.3-fold, pro-B cells 12.4 ± 1.9-fold, pre-B cells 5.6 ± 0.8-fold, immature B cells 7.5 ± 1.2-fold, and mature naive B cells 83 ± 7.6-fold. B-committed lymphoid progenitors (BLP) were modestly but significantly decreased (1.4 ± 0.2-fold), while common lymphoid progenitors (CLP) were not affected by G-CSF treatment. Increased apoptosis of mature naive B cells in the bone marrow was observed. Studies of G-CSF receptor deficient ( Csf3r −/− ) bone marrow chimeras show that G-CSF acts in a non-cell intrinsic fashion to suppress B lymphopoiesis. Consistent with this observation, we show that G-CSF treatment results in decreased expression in the bone marrow microenvironment of multiple B-supportive factors including CXCL12, interleukin-6, interleukin-7, and B cell activating factor (BAFF). Prior studies have established that CXCL12-abundant reticular (CAR) cells in the bone marrow play a key role in B cell development. CAR cells are perivascular stromal cells that express very high levels of CXCL12 and are in direct contact with pre-pro-B cells. G-CSF treatment did not affect CAR cell number. However, RNA expression profiling of sorted CAR cells showed that expression of several genes associated with B cell development are significantly decreased by G-CSF, including CXCL12 (4.2 ± 1.5-fold). In addition to CAR cells, other stromal cells in the bone marrow express CXCL12, including osteoblasts and endothelial cells. To assess the role of CXCL12 production by each of these cell types to B lymphopoiesis, we generated Cxcl12 flox mice and crossed them with mice expressing the following tissue-specific Cre-recombinase transgenes: Osteocalcin-Cre ( Oc-Cre ) targeting mature mineralizing osteoblasts; Osterix-Cre ( Osx-Cre ) targeting CAR cells and all osteolineage cells; or Prx1-Cre targeting mesenchymal progenitors and their progeny. Deletion of Cxcl12 using Oc-Cre or Osx-Cre had a similar effect on B cell development, with an isolated loss of mature naive B cells in the bone marrow (2.7 ± 0.5 and 4.1 ± 1.7-fold, respectively). In contrast, deletion of Cxcl12 using Prx1-Cre resulted in severe suppression of B lymphopoiesis that included a loss of CLP (3.3 ± 2.0-fold), BLP (5.6 ± 4.3-fold), and pre-pro-B cells (12.4 ± 5.1-fold). Interestingly, treatment of Prx1-Cre Cxcl12 flox/- mice with G-CSF resulted in additional B cell loss, indicating that deletion of Cxcl12 in mesenchymal stromal cells is not sufficient to fully recapitulate G-CSF-induced B cell suppression. In summary, G-CSF treatment results in marked changes in the bone marrow microenvironment that lead to a suppression of B lymphopoiesis. While G-CSF-induced inhibition of CXCL12 expression from stromal cells contributes to B cell suppression, additional alterations in the microenvironment also contribute to this phenotype. Disclosures: No relevant conflicts of interest to declare.