Fig. 2
From: TGF-β1-triggered maladaptive bone marrow endothelium impedes hematopoietic recovery

TGF-βRI overexpression in BM ECs exacerbates endothelial damage and delays hematopoietic recovery in mice. a Illustrative diagram of the study design involving mice with BM EC-specific overexpression of TGF-βRI. Adult female C57BL/6 J mice (8–10-weeks-old) were administered the recombinant AAV-VEC via intraosseous injection. AAV-TGF-βRI mice (TGF-βRI) received AAV-VEC encoding the TGF-βRI, Flag tag and ZsGreen genes under the regulation of a Tie promoter that is specific to ECs. TGF-βRI (T204D) indicates that the threonine at position 204 is replaced by aspartic acid, mimicking the persistent activation of TGF-βRI receptor. AAV-CTL mice (CTL), which were sex- and age-matched, received AAV-VEC encoding only the Flag tag and ZsGreen genes under the regulation of the same Tie promoter. Cohorts of mice (TGF-βRI + LY) were subjected to treatment with 5-FU at a dose of 250 mg/kg via tail vein injection on day 0. LY2157299 (LY) or dimethyl sulfoxide (DMSO) was subsequently administered daily via intragastric gavage to the mice. On day 14, the mice were sacrificed. b Recovery kinetics of WBCs, Hb, and PLTs after the given treatments were analyzed (N = 6 per group, n = 3 per sample). c Representative fluorescent multiplex immunohistochemistry analysis of Endomucin+ (white) and SCA1+ (red) BM vessels and nuclei (blue) in femurs from the specified groups. Scale bars represent 100 μm. d The frequencies of CD31+VE-Cadherin+ECs within CD45-Ter119- BM cells from mice subjected to the specified treatments were assessed through flow cytometry (N = 6 per group, n = 1 per sample). The frequencies of lineage-cKIT+SCA1+ HSPCs (e) and lineage-cKIT+SCA1- MPs (f) in WBM cells were assessed via flow cytometry (N = 6 per group, n = 1 per sample). N represents biological replicates; n represents technical replicates. Data are expressed as mean ± SEM and analyzed by the Mann–Whitney U test. *p ≤ 0.05. EC endothelial cell, Hb hemoglobin, HSPC hematopoietic stem and progenitor cell, MP myeloid progenitor, PLT platelet, SEM standard error of the mean, WBM whole bone marrow, WBC white blood cell, 5-FU 5-fluorouracil