Fig 1.
BDNF and TrkB expression in the bone marrow niche.
(A) Bdnf mRNA expression in the thymus, spleen, bone marrow CD31+ endothelial cells (ECs), and bone marrow PDGFRa+ mesenchymal stromal cells (MSCs). (B) TrkB mRNA expression in the same populations. (C) p75NTR (NGFR) and TrkB mRNA expression in sorted hematopoietic stem and progenitor cell populations. Data are based on cells sorted from three mice per group. (D) ELISA analysis of BDNF and pro-BDNF protein concentration in the bone marrow supernatant following 5 Gy total body irradiation (TBI). n = 3 mice per timepoint. (E) Representative distribution of BDNF (green) in longitudinal femur sections labeled with VE-cadherin (red) and DAPI (blue) of non-irradiated control (left) and at 24 hours following 5 Gy TBI. Scale bar = 50 μm. (F) Fold change in MSC cell culture supernatant levels of cytokines IL-6, LIF, Eotaxin-2, IP-10, and IL-9 following either 8 Gy (black bar) or 8 Gy + 100 ng/ml BDNF (red bar). Fold change is relative to non-irradiated control MSCs. n = 3 replicates per treatment. Significance was determined using multiple t-tests with Bonferroni correction. Shown are mean values +/- SEM. *P < .05, ***P < .001, ****P < .0001.
Fig 2.
Pharmacologic activation of BDNF-TrkB signaling promotes recovery of the bone marrow niche following radiation injury.
(A) Schematic representation of the experimental design: adult C57BL/6 mice were irradiated with 5 Gy and treated starting at 24 hours with vehicle, 0.5 mg/kg BDNF, or 1 mg/kg 7,8-DHF. (B) Total bone marrow cells per femur at day 10 (n = 5 mice per treatment). (C) Representative longitudinal femur sections labeled with VE-cadherin (white) and DAPI (blue) at day 10 following 5 Gy TBI. Scale bar = 100 μm. (D) Total ECs per femur at day 10. (E) EBD permeability at day 2. (F) Percent ECs positive for active caspase 3/7 at day 2. (G) Percent ECs positive for Mitosox at day 2. (n = 4 mice per treatment). *P < .05, **P < .01, ***P < .001, ****P < .0001.
Fig 3.
Pharmacologic activation of BDNF-TrkB signaling promotes reconstitution of peripheral blood neutrophils and lymphocytes.
(A) Day 10 CBC analysis of total WBCs, neutrophils (NEU), lymphocytes (LYMPH), platelets (PLT), and hemoglobin (Hb). (B) Day 10 peripheral blood analysis of circulating Lin- c-kit+ cells. (n = 5 mice per group). *P < .05, **P < .01, ***P < .001.
Fig 4.
BDNF administration increases bone marrow stem and progenitor cell recovery following TBI.
(A) Left, representative day 10 bone marrow FACS analysis of lin-c-kit+ myeloid progenitors (MP) and lin-sca-1+c-kit+ (LSK) cells. Right, %MP and %LSK of the total viable bone marrow cells. (B) Schematic representation of the experimental design: adult C57BL/6 mice were irradiated with 5 Gy and treated starting at 24 hours with vehicle, 0.5 mg/kg BDNF. At day 7, bone marrow was harvested and transplanted competitively into congenic recipient B6.SJL mice. (C) Total bone marrow engraftment at week 20 post-transplant. (D) Representative FACS analysis of bone marrow myeloid, B-lymph, T-lymph, and erythroid engraftment at 20 weeks post-transplant. E, %Donor-derived myeloid, B-lymph, T-lymph, and erythroid engraftment at 20 weeks post-transplant. (n = 7 recipient mice per group). Shown are mean values +/- SEM. *P < .05, **P < .01.
Fig 5.
BDNF administration promotes thymic regeneration following TBI.
(A) Thymus weights at days 10 and 120 following irradiation and treatment with BDNF. (B) Representative day 120 thymi. (C) Total viable thymocytes at days 10 and 120. (D) Total numbers of double negative (DN), double positive (DP), CD4+ single positive, and CD8+ single positive cells at day 10. E, Total numbers of DN1, DN2, DN3, and DN4 thymocyte precursors at day 10. (Day 10: n = 5 mice per group). Shown are mean values + SEM. *P < .05, **P < .01, ****P < .0001.