Regenerative Capacity of Old Muscle Stem Cells Declines without Significant Accumulation of DNA Damage
Figure 1
No difference in DNA DSBs in muscle stem cells with aging.
(A-F) Satellite cells from uninjured muscle or activated by injury for different times were isolated from hind leg muscle of young and old C57BL/6 mice and young SCID mice and were immunostained for γ-H2AX (green) (A-D) or for 53BP1 (green) (E-F), and counterstained with DAPI (blue). (A) Representative pictures of binned ranges of γ-H2AX foci/nuclei. Scale bar represents 5 µm. (B-C) Quantification of γ-H2AX foci in activated satellite cells 72 hours post muscle injury. At least 140 cells were counted per mouse. Data represent the mean +/- SEM, n = 4 mice per group; two-tailed unpaired Student’s t- test, *: p value < 0.05. (B) Binned ranges of foci/nuclei. (C) Percentage of γ-H2AX-positive cells ( = 5 foci/nuclei). (D) Quantification of γ-H2AX positive nuclei in satellite cells from uninjured muscle (uninj) and injured muscle 12, 24, and 36 hours post injury. Young and old mice are symbolized by black dots and red squares, respectively. Each dot or square represents one animal and the lines correspond to the mean. 2 young and 2 old mice were used per time point and at least 100 cells were counted per mouse, two-tailed unpaired Student’s t-test, *: p value < 0.05, **: p value < 0.01 (black * young uninj. vs 12, 24 or 36 hours, red * old uninj. vs 12, 24 or 36 hours). (E) Representative picture of 53BP1 positive cell. Scale bar represents 5 µm. (F) 53BP1 positive cell numbers ( = 5 foci/nuclei) were compared between young and old mice from uninjured muscle and 36 hours after injury. At least 100 cells were counted per mouse. Data are presented as mean +/- SEM, n = 2 mice per condition, two-tailed unpaired Student's t-test, *: p value < 0.05 (uninjured vs. injured), no age-specific difference was observed. (G-H) Tibialis anterior muscles of young and old wild-type mice as well as young SCID mice were cryosectioned. 10 ?m cryosections were immunostained for γ-H2AX and counterstained with DAPI. Confocal Z-series images were projected into a single plan using maximum pixel intensity. (G) Magnification of a representative picture, showing a γ-H2AX positive satellite cell (white arrowhead). DAPI is represented in blue and γ-H2AX is represented in red. Scale bar represents 10 µm. (H) The percentages of ?-H2AX positive cells in satellite positions relative to young were compared between young, old, and SCID mice. At least 150 nuclei were scored per mouse on Z-stack projected images. Data represent the mean +/- SEM, n = 3 mice per group; two-tailed unpaired Student’s t- test, *: p value < 0.05.