title: Senescence mechanisms of nucleus pulposus chondrocytes in human intervertebral discs.
abstract: BACKGROUND
  The population of senescent disc cells has been shown to increase in degenerated or herniated discs. However, the mechanism and signaling pathway involved in the senescence of nucleus pulposus (NP) chondrocytes are unknown.
  
  
  OBJECTIVE
  To demonstrate the mechanisms involved in the senescence of NP chondrocytes.
  
  
  METHODS
  Senescence-related markers were assessed in the surgically obtained human NP specimens.
  
  
  METHODS
  NP specimens remaining in the central region of the intervertebral disc were obtained from 25 patients (mean: 49 years, range: 20-75 years) undergoing discectomy. Based on the preoperative magnetic resonance images, there were 3 patients with Grade II degeneration, 17 patients with Grade III degeneration, and 5 patients with Grade IV degeneration.
  
  
  METHODS
  We examined cell senescence markers (senescence-associated beta-galactosidase [SA-beta-gal], telomere length, telomerase activity, p53, p21, pRB, and p16) and the hydrogen peroxide (H(2)O(2)) content as a marker for an oxidative stress in the human NP specimens.
  
  
  METHODS
  SA-beta-gal expression, telomere length, telomerase activity, and H(2)O(2) content as well as their relationships with age and degeneration grades were analyzed. For the mechanism involved in the senescence of NP chondrocytes, expressions of p53, p21, pRB, and p16 in these cells were assessed with immunohistochemistry and Western blotting.
  
  
  RESULTS
  The percentages of SA-beta-gal-positive NP chondrocytes increased with age (r=.82, p<.001), whereas the telomere length and telomerase activity declined (r=-.41, p=.045; r=-.52, p=.008, respectively) However, there was no significant correlation between age and H(2)O(2) contents (p=.18). The NP specimens with Grade III or Grade IV degeneration showed significantly higher percentages of SA-beta-gal-positive NP chondrocytes than those with Grade II degeneration (p=.01 and p=.025, respectively). Immunohistochemistry showed that the senescent NP chondrocytes in all the specimens expressed p53, p21, and pRB, but a few NP chondrocytes in only two specimens expressed p16. Western blotting showed that the expressions of p53, p21, and pRB displayed a corresponding pattern, that is, a strong p53 expression led to strong p21 and pRB expressions and vice versa.
  
  
  CONCLUSIONS
  Our in vivo study demonstrated that senescent NP chondrocytes increased or accumulated in the NP with increasing age and advancing disc degeneration. The NP chondrocytes in the aging discs exhibited characteristic senescent features such as an increased SA-beta-gal expression, shortened telomeres, and decreased telomerase activity. We further demonstrated that the telomere-based p53-p21-pRB pathway, rather than the stress-based p16-pRB pathway, plays a more important role in the senescence of NP chondrocytes in an in vivo condition. Our results suggest that prevention or reversal of the senescence of NP chondrocytes can be a novel therapeutic target for human disc degeneration.
authors.affiliation: Department of Orthopedic Surgery, St. Mary's Hospital, The Catholic University of Korea, 62 Yoido-dong, Youngdeungpo-ku, Seoul, Republic of Korea. kiwonk62@yahoo.co.kr
authors.forename: Ki-Won
authors.initials: KW
authors.lastname: Kim
authors.forename: Ha-Na
authors.initials: HN
authors.lastname: Chung
authors.forename: Kee-Yong
authors.initials: KY
authors.lastname: Ha
authors.forename: Jun-Seok
authors.initials: JS
authors.lastname: Lee
authors.forename: Young-Yul
authors.initials: YY
authors.lastname: Kim
mesh_terms: D000328:Adult; D000368:Aged; D015153:Blotting, Western; D002478:Cells, Cultured; D016922:Cellular Senescence / Q000502:physiology*; D019902:Chondrocytes / Q000378:metabolism* / Q000473:pathology; D005260:Female; D006801:Humans; D006861:Hydrogen Peroxide / Q000378:metabolism; D007150:Immunohistochemistry; D007403:Intervertebral Disc / Q000378:metabolism* / Q000473:pathology; D007405:Intervertebral Disc Displacement / Q000378:metabolism / Q000473:pathology; D008297:Male; D008875:Middle Aged; D019098:Telomerase / Q000378:metabolism; D016615:Telomere / Q000378:metabolism / Q000473:pathology; D001616:beta-Galactosidase / Q000378:metabolism
chemical_list: D006861:Hydrogen Peroxide; D019098:Telomerase; D001616:beta-Galactosidase