Deletion of Ku86 causes early onset of senescence in mice

Abstract

DNA double-strand breaks formed during the assembly of antigen receptors or after exposure to ionizing radiation are repaired by proteins important for nonhomologous end joining that include Ku86, Ku70, DNA-PK(CS), Xrcc4, and DNA ligase IV. Here we show that ku86-mutant mice, compared with control littermates, prematurely exhibited age-specific changes characteristic of senescence that include osteopenia, atrophic skin, hepatocellular degeneration, hepatocellular inclusions, hepatic hyperplastic foci, and age-specific mortality. Cancer and likely sepsis (indicated by reactive immune responses) partly contributed to age-specific mortality for both cohorts, and both conditions occurred earlier in ku86(-/-) mice. These data indicate that Ku86-dependent chromosomal metabolism is important for determining the onset of age-specific changes characteristic of senescence in mice.

Figure 1

Lifespan and potential causes of age-specific death. (A) Survival curve (100%× number of mice alive after each week/total number of mice at beginning of study). The survival curve begins after weaning (3 wk) because ku86^−/− pups are less fit to compete for resources than control littermates, and about 50% die before weaning unless control pups are removed soon after birth (16). Control mice, blue squares; ku86^−/− mice, green triangles. A dashed blue line representing the control survival curve is superimposed onto the ku86^−/− curve, and a dashed green line representing the ku86^−/− survival curve is superimposed onto the control curve, to illustrate their differences. The percent of population that dies each week is displayed to the left of the ku86^−/− survival curve and the right of the control survival curve for each interval of 20% starting at the onset of age-specific mortality. Number of mice observed: control, 47; ku86^−/−, 89. (B and C) Section of liver with reactive immune response from (B) 89-wk-old control mouse and (C) 61-wk-old ku86^−/− mouse. Note infiltration of mononuclear cells for B and neutrophils for C. (D and E) Section of malignant lymphoma from (D) 71-wk-old control mouse (infiltrating intestine) and (E) 37-wk-old ku86^−/− mouse (subjacent to bronchial epithelium). (B–E, ×205.)

Figure 2

Age-specific changes in control and ku86^−/− mice. (A–D) Outward signs of senescence. The dorsal region was shaved to enhance visualization of kyphosis. (A) Control (+/+ and +/−) and ku86^−/− (−/−) mice at 2.5 wk. No kyphosis observed. (B) Control and ku86^−/− mice at 31 wk. Kyphosis observed in only ku86^−/− mouse. (C) Control and ku86^−/− mice at 75 and 79 wk, respectively. Kyphosis observed in only ku86^−/− mouse. (D) Control mouse at 120 wk. Kyphosis observed. (E and F) Section of vertebral articular processes from (E) 45-wk-old control and (F) 49-wk-old ku86^−/− mouse. For ku86^−/− bone, compared with control, the cortical wall (cw) and trabeculae (t) is thinner, the number of trabeculae are reduced and the medullary cavity (mc) is expanded. (G and H) Section of epiphysis from 22-wk-old (G) control and (H) ku86^−/− mouse. For ku86^−/− epiphysis, compared with control, the number of chondrocytes is reduced and the columnar organization of chondrocytes is lost. (I–L) Section of skin (dorsal region over cranial to mid-thorax) from (I and J) 45-wk-old control and (K and L) 49-wk-old ku86^−/− mouse. For ku86^−/− skin, compared with control, all subcutaneous elements, including superficial collagen (sc), subcutaneous adipose (sa), and skeletal muscle (sm) are reduced and hair follicles and sebaceous glands (arrow in J and L) atrophied. (Bar = A–D 1 cm.) (E and F, ×65; G and H, ×200; I and K, ×35; J and L, ×175.)

Figure 3

Age-specific changes in liver. (A and B) Liver nodules from (A) 110-wk-old control mouse and (B) 61-wk-old ku86^−/− mouse. Normal part of liver (Upper Left) is compressed by nodule (Lower Right), separated by dashed line. (C) Expression of GS in normal part of liver. GS present in cells only two deep from portal vein (pv). (D–G) Dysregulation, proliferation, and dedifferentiation of cells in a liver nodule from a 61-wk-old ku86^−/− mouse. (D) GS stain. Diffuse hepatocellular staining in liver nodule, but not normal liver. (E) FAH stain. Loss of FAH diffuse positivity resulting in mosaic expression pattern in liver nodule but not normal liver. (F) Anti-Ki-67 stain. Cellular proliferation in nodular hepatocytes, but not normal liver. (G) MAFP stain. MAFP is not normally expressed in adult hepatocytes. (H and I) Hepatocellular cytoplasmic inclusions (arrows) from: (H) 111-wk-old control mouse and (I) 65-wk-old ku86^−/− mouse. (A–F ×115; G, ×210; H and I, ×420.)