Fig. 6
The decrease of LaminA/C expression in HSCs determines changes in nuclear volume and shape, H4K16ac polarity, and chromosome 11 homolog proximity. a Representative confocal 3D reconstruction of the nucleus (stained with DAPI, blue) of Lmnafl/fl and LmnaΔ/Δ/Vav-Cre HSCs. Bar = 2 μm. b Nuclear volume measurements by Volocity 3D Image Analysis (based on DAPI staining). **p < 0.01; n = 8 biological repeats; 100 single HSCs for Lmnafl/fl, 115 single HSCs for LmnaΔ/Δ/Vav-Cre in total. c Nuclear shape factor (NSF) measurements. *p < 0.05; n = 4 biological repeats; 99 HSCs for Lmnafl/fl, 100 HSCs for LmnaΔ/Δ/Vav-Cre in total. d Representative distribution of H4K16ac in Lmnafl/fl, LmnaΔ/Δ/Vav-Cre, LmnaΔ/Δ/Vav-Cre + CASIN 5 μM, aged, and aged+CASIN 5 μM HSCs. Bar = 2 μm. Panels show DAPI (nucleus, blue), H4K16ac (red). e Percentage of Lmnafl/fl, LmnaΔ/Δ/Vav-Cre, LmnaΔ/Δ/Vav-Cre + CASIN 5 μM, aged, and aged+CASIN 5 μM HSCs with a polar distribution of H4K16ac. Shown are mean values + 1 SE; n = 3–5 biological repeats, ~ 30–100 single HSCs scored per sample in total; *p < 0.05, ***p < 0.001. f Representative FACS histograms of an isotype control stained sample and of H4K16ac in Lmnafl/fl and LmnaΔ/Δ7Vav-Cre samples. HSCs gates are shown. g Median fluorescence intensity of H4K16ac in HSCs of control Lmnafl/fl and LmnaΔ/Δ/Vav-Cre mice; n = 3 biological repeats. h Measurements of the distances between chromosome 11 homologs in Lmnafl/fl and LmnaΔ/ΔVav-Cre HSCs by Volocity 3D Image Analysis; **p < 0.01; n = 3 biological repeats, 27 cells for Lmnafl/fl, 30 cells for LmnaΔ/Δ/Vav-Cre in total. i Percentage of Lmnafl/fl and LmnaΔ/Δ/Vav-Cre HSCs with chromosome homologs proximity for chromosome 11; n = 3 biological repeats; *p < 0.05. j Representative confocal 3D reconstruction of the nucleus (stained with DAPI, blue) and of chromosome 11 (red, FISH stained with a full-paint probe) in Lmnafl/fl and LmnaΔ/Δ/Vav-Cre HSCs. Bar = 2 μm. k 3D reconstruction of the chromosome 11 homologs (red and green) and the distance measurement between them by Volocity 3D Image Analysis. In Lmnafl/fl HSCs green, color for two homologs shows that they are distributed very close to each other and are not recognized as two objects by the software. While in LmnaΔ/Δ/Vav-Cre HSCs, they are marked as two objects in green and red. Bar = 2 μm. l Summary of premature aging-like phenotypes in the hematopoietic system of LaminA/C conditional knockout mice. m Aging of HSCs compartment. Cartoon scheme integrating the novel main findings. The newly identified HSC aging phenotypes are highlighted in red