Aging impairs human bone marrow function and cardiac repair following myocardial infarction in a humanized chimeric mouse

Figures

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  FIGURE 1. Old CD34+ cells exhibit reduced colony formation and lower lymphoid cell repopulation following transplant. (a) Mononuclear cell concentration (n = 40, 56.4 ± 0.97 years old [Y‐CD34+], and n = 48, 72.7 ± 0.59 years old [O‐CD34+]) and (b) CD34+ cell frequency in younger and old patient BM (n = 51, 56.8 ± 1.2 years old [Y‐CD34+] vs. 73 ± 0.7 years old [O‐CD34+]). (c) Colony‐forming ability of CD34+ cells from younger and old patient BM (n = 25, 52.7 ± 2.5 years old [Y‐CD34+] vs. 75.1 ± 1.4 years old [O‐CD34+]). (d) And total number of colonies formed stratified based on age group (n = 13, 52.7 ± 2.5 years old [Y‐CD34+] and n = 12, 75.1 ± 1.4 years old [O‐CD34+]). (e) Schematic representation of experimental design for reconstitution model. (f) hCD45+, and (g) CD34+ frequency in the BM at 3 months postreconstitution (n = 15, 56.2 ± 1.6 years old [Y‐CD34+], n = 17, 71.8 ± 0.7 years old [O‐CD34+], n = 14, 68 ± 2.6 years old [CD34−]). (h) Engrafted human hematopoietic progenitor cell frequencies in the BM of NSG mice transplanted with younger and old CD34+ bone marrow cells and (i) engrafted BM human HSCs 3 months after reconstitution (n = 7, 56.9 ± 1.4 years old [Y‐CD34+], n = 8, 73.1…

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  FIGURE 2. Age and cell type affect the long‐term functional outcome post‐MI. (a) Schematic representation of cardiac function analysis after reconstitution and coronary ligation. (b) Survival curve of Y/O‐CD34+ mice versus CD34− mice by 4 weeks post‐MI, *p < 0.05 Y‐CD34+ vs. all groups. (c) Functional analysis post‐MI measured by echocardiography demonstrating dimensions at (c) diastole and (d) systole and cardiac function demonstrated by (e) ejection fraction and (f) fractional shortening at 4 weeks post‐MI (55.3 ± 2.1 years old [Y‐CD34+] vs. 74.1 ± 1.7 years old [O‐CD34+], n = 7‐10/group). All timepoint values are provided in Table S1. (g) Gross tissue and (h) histological analyses with trichrome staining at 4 weeks post‐MI for analysis of scar size. Two different stained hearts are shown for each group. (i) Quantification of infarct expansion comparison (53.6 ± 2.5 years old [Y‐CD34+] vs.73 ± 0.6 years old [O‐CD34+], n = 4‐5/group). *p < 0.05 vs. Y‐CD34+ all groups at same time point, † p < 0.05 O‐CD34+ vs. CD34− at same time point, and **p < 0.05 Y‐CD34+ vs. CD34− at same time point. Values are mean ± SEM

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  FIGURE 3. Younger CD34+ cells reduce remodeling and improve scar angiogenesis by 4 weeks post‐MI. (a) WGA staining of younger CD34+, old CD34+, and CD34− myocytes for cross‐sectional area analysis at 4w post‐MI, and quantification (52.2 ± 3.0 years old [YCD34+] vs. 71.2 ± 1.6 years old [O‐CD34+], vs. 59.2 ± 2.9 years old [CD34‐], n = 4/group). (b) Isolectin‐IB4 staining of the infarct region. (c) Mouse CD31+ cell staining in the infarct regions. (d) Quantification of isolectin staining for comparison between the three groups (50.6 ± 2.8 years old [Y‐CD34+] vs. 70.8 ± 1.3 years old [O‐CD34+], vs. 57.4 ± 2.1 years old [CD34‐], n = 5/group). (e) Quantification of mCD31 cell staining for angiogenesis comparison between the groups (50.6 ± 2.8 years old [Y‐CD34+] vs. 70.8 ± 1.3 years old [O‐CD34+], n = 5/group). (f) Human CD45+ staining demonstrating inflammatory cell infiltration into the myocardial infarct region at 3d post‐MI. (g) Quantification of the hCD45+ staining at 3d post‐MI among the younger CD34+ (50.6 ± 2.8 years old), old CD34+ (70.8 ± 1.3 years old), and CD34− hearts (n = 5/group). *p < 0.05 vs. all groups, † p < 0.05 vs. CD34−. Values are mean ± SEM

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  FIGURE 4. Aged CD34+ cells exhibit impaired human cell recruitment to the heart post‐MI. (a) Human CD45 cells (hCD45), (b) human T cells (hCD3), and (c) human myeloid (hCD33) cells in the mouse myocardium at baseline, 3, and 7 d post‐MI as measured by flow cytometry (56.5 ± 3.8 years old [Y‐CD34+] vs. 72.2 ± 0.8 years old [O‐CD34+] at baseline, 57 ± 1.2 years old [Y‐CD34+] vs. 71.9 ± 1.0 years old [O‐CD34+] at 3 d and 55.4 ± 1.7 years old [Y‐CD34+] vs. 72.7 ± 1.2 years old [O‐CD34+] at 7 d post‐MI, n = 4–10/group). (d) Representative flow cytometry plots showing differences in T‐cell cell number. (e) Circulatory hCD45+, (f) hCD3+, (g) CD33+ cell frequency in the blood of Y/O‐CD34+ reconstituted mice at baseline, 3 and 7 d post‐MI (56.5 ± 1.6 years old [Y‐CD34+] vs. 73.1 ± 1.0 years old [O‐CD34+] at baseline, 56.8 ± 0.7 years old [Y‐CD34+] vs. 72.2 ± 1.3 years old [O‐CD34+] at 3 d, and 58 ± 1.4 years old [Y‐CD34+] vs. 72.4 ± 1.1 years old [O‐CD34+] at 7 d post‐MI, n = 7–10/group). (h) Spleen hCD3+ cell frequency in chimeric mice at baseline, 3 and 7 d post‐MI (56.5 ± 1.6 years old [Y‐CD34+] vs. 73.1 ± 1.0 years old [OCD34+] at baseline, 56.5 ± 0.7 years old [Y‐CD34+] vs. 72.2 ± 1.3 years…

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  FIGURE 5. CD34+ cell age affects early mediators of cardiac repair. (a) T‐cell subtype analysis of CD4+ frequency 3 and 7 d post‐MI (58.2 ± 1.9 years old [Y‐CD34+] vs. 71.9 ± 1.0 years old [O‐CD34+] at 3 d and 57 ± 1.6 years old [Y‐CD34+] vs. 73 ± 0.9 years old [OCD34+] at 7 d post‐MI, n = 5–8/group). (b) Human cytokine analysis of pro‐ and anti‐inflammatory cytokines in younger and older CD34+ hearts at 3 d post‐MI (56 ± 2.1 years old [Y‐CD34+] vs. 69.5 ± 1.7 years old [O‐CD34+], n = 6/group). Myocardial mouse cytokines (c) IL‐6, (d) IL‐10, and (e) IL‐1β in Y/O CD34+ and CD34− myocardium (56 ± 2.1 years old [Y‐CD34+] vs. 69.5 ± 1.7 years old [O‐CD34+], n = 7/group). (f) MMP2 and MMP9 activity at 3 d post‐MI in in Y/O CD34+ and CD34− infarcted myocardium measured using zymography (55.6 ± 2.2 years old [Y‐CD34+] vs. 69.8 ± 1.9 years old [O‐CD34+], n = 5/group). *p < 0.05 vs all other groups, † p < 0.05 groups as indicated−. Values are mean ± SEM

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  FIGURE 6. CD34+ cell age alters the infiltration of mouse (host) myeloid cells post‐MI. (a) Quantification of mouse CD45+ (mCD45) cells in the heart 3‐ and 7 d post‐MI (56.3 ± 1.1 years old [Y‐CD34+] vs. 70.1 ± 1.6 years old [O‐CD34+], vs. 68.5 ± 3.5 years old [CD34‐] at 3 d and 61.5 ± 1.4 years old [Y‐CD34+] vs. 70.3 ± 1.4 years old [O‐CD34+], vs. 65.5 ± 3.1 years old [CD34‐] at 7 d, n = 3–7/group). Quantification of mouse (b) neutrophil, (c) Ly6Chi monocytes, and (d) Ly6Cneg monocytes 3 and 7 d post‐MI in Y‐CD34+, O‐CD34+, CD34−, and WT NSG mice (n = 3–7/group). (e) Quantification of total macrophages in heart 3‐ and 7‐d post‐MI and (f) frequency of CD206+ macrophages in the heart at 3 d post‐MI. † p < 0.05 vs. CD34− at the same timepoint, †† p < 0.05 vs. WT NSG at the same timepoint, *p < 0.05 vs indicated groups at same timepoint, €p < 0.05 vs. O‐CD34+ at the same timepoint. Values are mean ± SEM