1. Distinct phases in recovery of reconstituted innate cellular-mediated immunity after murine syngeneic bone marrow transplantation 
Jeffery J. Auletta, Jennifer L. Devecchio, James L.M. Ferrara, Frederick P. Heinzel 
Biology of Blood and Marrow Transplantation 
Volume 10, Issue 12, Pages (December 2004)
DOI: /j.bbmt
Copyright © 2004 American Society for Blood and Marrow Transplantation Terms and Conditions

2. Figure 1
Macrophage and polymorphonuclear cell reconstitution. Representative FACS histograms for nontransplant control and syngeneic transplant recipients (7 and 21 days after transplantation) are illustrated to show enumeration strategies for macrophages (MAC) and polymorphonuclear cells (PMN). A hematopoietic-derived (CD45+) population was gated to remove FITC-CD3, CD19, and IgM-positive T and B cells, as indicated by the window (R1), followed by analysis of CD11b+ and anti-NIMP-positive cells. PMN cells (defined as CD11b+NIMP+) and macrophages (CD11b+NIMP−) were determined from events. In addition to more donor-derived (CD45.1+) MAC and PMN cells, early reconstitution at 7 days contained a significant population of CD45− cells and nonviable cells versus late reconstitution (see Figure 3 and Table 1 for quantifications and comparisons).
Biology of Blood and Marrow Transplantation  , DOI: ( /j.bbmt )
Copyright © 2004 American Society for Blood and Marrow Transplantation Terms and Conditions

3. Figure 2
Dendritic cell reconstitution. FACS histograms used to enumerate total and myeloid and lymphoid subsets of dendritic cells (DCs) in control and transplant recipients are illustrated. In control animals, non-B and non-T cells were selected in gate R1 (top row, left panel), and CD11c and CD11b were used to define myeloid (M-DC; CD11c+CD11bhigh) and lymphoid (L-DC; CD11c+CD11bintermediate/neg) subpopulations of dendritic cells. Alternatively, these subtypes were additionally separated by their expression of CD8α (lymphoid DCs defined as CD11c+CD8α+; myeloid DCs defined as CD11c+CD8α−). For transplanted mice, the initial gate R1 was modified to include only donor-derived CD45.1+ cells (middle and bottom rows; left panels.) Myeloid DCs are absent early (7 days) after transplantation but are present by 21 days after transplantation.
Biology of Blood and Marrow Transplantation  , DOI: ( /j.bbmt )
Copyright © 2004 American Society for Blood and Marrow Transplantation Terms and Conditions

4. Figure 3
Splenocyte reconstitution after syngeneic bone marrow transplantation: early innate versus late adaptive recovery. Composite histograms of CD45+ splenocyte phenotypes are illustrated for controls and for syngeneic BMT recipients. Percentages of donor versus recipient CD45+ splenocytes at different recovery times are indicated below the figure. The predominant hematopoietic-derived cellular populations differ at early and late recovery times. Notably, a significant CD45− population of splenocytes is present early after transplantation (∼60% of total splenocytes) but is absent by 14 days. MAC indicates macrophage; PMN, polymorphonuclear cells; DC, dendritic cells; NK, natural killer cells.
Biology of Blood and Marrow Transplantation  , DOI: ( /j.bbmt )
Copyright © 2004 American Society for Blood and Marrow Transplantation Terms and Conditions

5. Figure 4
Induction of IL-12p70 (A), IFN-γ (B), and IFN-α (C) from splenocyte cell culture. Mean ± SD cytokine levels are shown for control (CL), irradiated-only animals, and transplant-recipient animals (days after transplantation) after 24 hours of splenocyte stimulation with PAMP (left) and cell-specific agonists (right). Splenocytes from irradiated animals were collected and stimulated 7 days after irradiation. Data from 1 of 4 transplants with similar results are shown to illustrate the phasic recovery of control-level, inducible cytokines from reconstituted splenocytes. Statistical comparisons at defined time points with cytokine data pooled from multiple time courses are provided in Table 2. Inducible IL-12p70 and IFN-α production occurs in the early posttransplantation period (7 days), followed by later IFN-γ production at 21 days after transplantation.
Biology of Blood and Marrow Transplantation  , DOI: ( /j.bbmt )
Copyright © 2004 American Society for Blood and Marrow Transplantation Terms and Conditions

6. Figure 5
IL-6 and TNF-α induction from reconstituted splenocyte cell culture. PAMP-inducible cytokine levels (mean ± SD) are shown for control (CL), irradiated-only (XRT), and transplant splenocytes. Splenocytes from irradiated animals were collected and stimulated 7 days after irradiation. Data from 1 of 4 transplants with similar results are shown. Early inducible IL-6 and TNF-α synthetic production by reconstituted splenocytes was measured.
Biology of Blood and Marrow Transplantation  , DOI: ( /j.bbmt )
Copyright © 2004 American Society for Blood and Marrow Transplantation Terms and Conditions

7. Figure 6
Systemic IL-12p70 (A), IFN-γ (B), and IL-6 (C) after in vivo challenge. Mean ± SD systemic cytokine levels from control (CL), irradiated-only (XRT), and transplant animals 5 hours after intraperitoneal challenge with phosphate-buffered saline (Sham), lipopolysaccharide (LPS), and CpG oligonucleotide are illustrated. In contrast to phasic recovery of inducible cytokine production from cell culture, systemic cytokine production is similar across posttransplantation times (7 versus 21 days) and animal groups. Systemic IFN-α induction was similarly increased across all experimental groups after TLR challenge (data not shown).
Biology of Blood and Marrow Transplantation  , DOI: ( /j.bbmt )
Copyright © 2004 American Society for Blood and Marrow Transplantation Terms and Conditions

8. Figure 7
IL-6 induction in cell culture (A) and after LPS challenge (B): hematopoietic versus systemic disassociation. Mean ± SD IL-6 levels were compared among nontransplant control (CL) mice and 3 groups of syngeneic transplant mice (A–C): group A wild-type (WT) mice received WT marrow; group B WT mice received IL-6 KO marrow; and group C IL-6 KO mice received normal marrow. Twenty-one days after syngeneic chimeric transplantation, there were similar differences in ex vivo and systemic IL-6 cytokine induction among the 3 transplant groups (data not shown). In addition, FACS data revealed similar spleen composites for all transplant groups at each harvest time. However, group B mice had 2-fold decreases in total numbers of reconstituted splenocytes relative to other transplant groups (data not shown).
Biology of Blood and Marrow Transplantation  , DOI: ( /j.bbmt )
Copyright © 2004 American Society for Blood and Marrow Transplantation Terms and Conditions