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. 2020 Mar 3;9:e52570. doi: 10.7554/eLife.52570

Figure 4. p16 deletion delayed mouse intervertebral disc degeneration (IVDD).

WT and p16 KO mice were fed on the ground or with tail suspension (TS). (A) Radiographs of overall mouse length. (B) After H and E staining and Safranin O staining, collagen and NP cells are orange, and fibers are blue. (C) The intervertebral disc height index as calculated on the basis of lumbar vertebrae. (D) Associated protein levels as assessed by western blotting and (E) as measured by densitometric analysis, with results expressed relative to those in WT mice. (F) Target mRNA expression assessed by RT-PCR relative to GAPDH expression. (G) IL-1β, IL-6 and TNF-α levels in disc tissues as determined by ELISA. (H) The modified Thompson classification as assessed on the basis of lumbar disc signals. (I) Safranin O-positive area (%). Data are presented as mean ± SD (n = 3); *p<0.05; **p<0.01; ***p<0.001.

Figure 4—source data 1. Source data and related summary statistics for Figure 4B, C, D, G and H.

Figure 4.

Figure 4—figure supplement 1. Establishment of TS-induced mouse IVDD model.

Figure 4—figure supplement 1.

(A) A specialized cage used to suspend the tails of mice. The mice could obtain food and water freely in the cage. (B) Mice were sacrificed after 4 weeks of suspension. The muscles around the spine were bloodshot with varying degrees of injury. Tail suspension (TS) caused visibly bloodshot muscles, which were alleviated by p16 KO.
Figure 4—figure supplement 2. Representative micro-MRI of mouse intervertebral discs.

Figure 4—figure supplement 2.

The larger white area that the disc has, the more water it contains, and this is positively related to the degree of disc degeneration. The yellow arrow indicates the disc site.
Figure 4—figure supplement 3. Pffirmann grade of degenerated disc from the mice specimens.

Figure 4—figure supplement 3.

*p<0.05; ***p<0.001.
Figure 4—figure supplement 3—source data 1. Related summary statistics.
Figure 4—figure supplement 4. The intervertebral disc height index (DHI) was calculated by averaging the measurements obtained from the (A) posterior, (B) middle, and (C) anterior portions of the intervertebral disc and dividing these values by the average height of the adjacent (D–I) posterior, middle, and anterior portions of the vertebral body.

Figure 4—figure supplement 4.