Figure 5. Solid tumors: Low–lamin A cells show higher curvature nuclei, with increased KU80 mislocalization and DNA damage. (A) A549 Ctl and shLMNA subcutaneous xenografts in NSG mice are monitored by tdTomato fluorescence. Bar, 1 cm. (B) Confocal images of A549 Ctl and shLMNA tumors. Red arrows indicate cytoplasmic KU80. The white arrow indicates mouse nucleus with chromocenters. (C) Intensity profile of a region of interest in B shows cytoplasmic mislocalization of KU80. (D) Top: Cytoplasmic/nuclear KU80 is higher in A549 shLMNA tumors versus Ctl tumors. DNA stain sets baseline for cytoplasm. Middle: Cytoplasmic/nuclear mislocalization in tumors compare well with 2D cultures. Bottom: Nuclear circularity in shLMNA cells is lower than Ctl, while nuclear areas are similar. n = 10 images per tumor from three tumors per group; n > 100 cells for circularity and area. Dark and light gray asterisks indicate significance relative to Ctl. (E) Electrophoretic comet assay shows higher DNA damage level in shLMNA tumors compared with Ctl tumors (i) and in vitro shLMNA cultures (ii) treated with blebbistatin but lower than ones treated with DMSO (data from Fig. 4 C). In vitro data are normalized to A549 Ctl cells treated with DMSO. n = 5 tumors. *, P < 0.05. Bars, 10 µm. All data are presented as mean ± SEM. (F) Schematic of processes and factors that affect nuclear envelope rupture and excess DNA damage.
Figure 5.

Solid tumors: Low–lamin A cells show higher curvature nuclei, with increased KU80 mislocalization and DNA damage. (A) A549 Ctl and shLMNA subcutaneous xenografts in NSG mice are monitored by tdTomato fluorescence. Bar, 1 cm. (B) Confocal images of A549 Ctl and shLMNA tumors. Red arrows indicate cytoplasmic KU80. The white arrow indicates mouse nucleus with chromocenters. (C) Intensity profile of a region of interest in B shows cytoplasmic mislocalization of KU80. (D) Top: Cytoplasmic/nuclear KU80 is higher in A549 shLMNA tumors versus Ctl tumors. DNA stain sets baseline for cytoplasm. Middle: Cytoplasmic/nuclear mislocalization in tumors compare well with 2D cultures. Bottom: Nuclear circularity in shLMNA cells is lower than Ctl, while nuclear areas are similar. n = 10 images per tumor from three tumors per group; n > 100 cells for circularity and area. Dark and light gray asterisks indicate significance relative to Ctl. (E) Electrophoretic comet assay shows higher DNA damage level in shLMNA tumors compared with Ctl tumors (i) and in vitro shLMNA cultures (ii) treated with blebbistatin but lower than ones treated with DMSO (data from Fig. 4 C). In vitro data are normalized to A549 Ctl cells treated with DMSO. n = 5 tumors. *, P < 0.05. Bars, 10 µm. All data are presented as mean ± SEM. (F) Schematic of processes and factors that affect nuclear envelope rupture and excess DNA damage.

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