1. Aging and age related stresses: a senescence mechanism of intervertebral disc degeneration 
F. Wang, F. Cai, R. Shi, X.-H. Wang, X.-T. Wu 
Osteoarthritis and Cartilage 
Volume 24, Issue 3, Pages (March 2016)
DOI: /j.joca
Copyright © 2015 Osteoarthritis Research Society International Terms and Conditions

2. Fig. 1
Age-related changes of the IVD. During growth and maturation, the IVD undergoes significant histocytological changes that include fibrosis of the gel-like nucleus pulpusos (NP), the transition of vaculated notochondral cells to chondrocyte-like NP cells, thinning of the cartilagenous layers in the endplate, as well as the disorganization and fibrocartilaginous metaplasia of the annulus fibrosis (AF). In the middle transverse section of a rat lumber IVD at 6 weeks old (A1) and 20 weeks old (B1), a distinct demarcation between the NP and the AF (red arrow) can be observed in the young rat IVD. Hematoxylin-Eosin staining shows that the young disc is composed of a gel-like NP containing predominantly vacuolated notochordal cells (A2, red arrows), thick layers of a cartilaginous endplate (A3), and an organized AF with concentric lamella (A4); whereas the adult disc features a fibrotic NP sparsely populated with smaller chondrocyte-like NP cells (B2, black arrow), a narrowed cartilaginous layer in the endplate (B3), and disorganized collagen fibers in the AF (B4). Of note is the hyaluronic acid and proteoglycan rich extracellular matrix in the young NP (A2, red triangle) is negatively stained by hematoxylin, while the increased cartilaginous matrix can be detected in the NP (B2, black triangle) and the AF (B4) within the adult IVD, supporting a fibrocartilaginous change during disc growth and maturation.
Osteoarthritis and Cartilage  , DOI: ( /j.joca )
Copyright © 2015 Osteoarthritis Research Society International Terms and Conditions

3. Fig. 2
Acceleration of IVDD. IVDD is an age-related process that is accelerated by multiple stresses, such as disc injuries, disturbed mechanical loadings, and various oxidative or genotoxic injuries. A. MRI of a 48-year-old male with asymptomatic IVDD at the lumber (L) 4/5 segment. A reduction in the disc height and T2-weighted signal intensity indicates a significant loss of hydrophilic extracellular matrix and water content. B. MRI of a 26-year-old male with symptomatic IVDD at L1/2 and L4/5. The IVDD process was accelerated by a remarkable spinal trauma that may have caused a loss of the IVD integrity. C. MRI of a rabbit with IVDD induced by annulus stabbing. In comparison with normal disc (C1), puncturing at the L2/3, L3/4, and L4/5 annuli resulted in acute loss of the T2-signal intensity and disc height (red arrows) 2 weeks post-stabbing. No significant radiological evidence for self-repair was detected 12 weeks post-stabbing, suggesting accelerated degeneration and compromised self-repair capacity following the disc injuries.
Osteoarthritis and Cartilage  , DOI: ( /j.joca )
Copyright © 2015 Osteoarthritis Research Society International Terms and Conditions

4. Fig. 3
Prolonged culture of a rat NP cell in monolayers. NP cells were isolated from a 6-week-old rat lumber IVD, cultured in monolayers, and expanded until passage (P) 10. At P1, notochord cells were seen in the monolayer cultures (A1), which were larger in size and contained numerous cytoplasmic vacuoles (A2). After they were expanded to P3 (B1), most of the notochord cells had transformed into smaller chondrocyte-like NP cells (B2); in the latter, cellular senescence could be detected by SA-β-gal staining (B1, C2). As the NP cells underwent prolonged replications, cellular senescence was significantly increased and accumulated (C1), forming a similar senescent cluster to that in vivo.
Osteoarthritis and Cartilage  , DOI: ( /j.joca )
Copyright © 2015 Osteoarthritis Research Society International Terms and Conditions

5. Fig. 4
The p53-p21-RB and p16-RB senescent pathways within an IVD. By inhibiting the transcriptional factor E2F, which targets most genes required for S-phase onset, the hypophosphorylated retinoblastoma (RB) protein functions as the core repressor for cell cycle progression. In normal cells the cyclin and cyclin-dependent kinase (CDK) complexes hyperphosphorylate and inactivate RB to allow cell cycle transition. Under various senescence-inducing damages, the sustained activation of the p53-p21 and p16 signals will inhibit CDK2 and CDK4/6 respectively, thus resulting in increased RB and a stable cell cycle arrest. As one of the major risk factors for IVDD, aging indicates a prolonged replication and a shortened telomere, which ultimately initiates the DNA damage response and activate the p53-p21-RB and/or p16-RB pathways to induce chronic RS. By contrast, acute exposure to disturbed mechanical loadings, disc injuries, as well as an altered oxygen tension and nutritional supply might accelerate the kinetics of senescence formation, preferably by activating the p16-RB pathway to induce premature disc cell senescence.
Osteoarthritis and Cartilage  , DOI: ( /j.joca )
Copyright © 2015 Osteoarthritis Research Society International Terms and Conditions

6. Fig. 5
The senescence mechanism of IVD aging and degeneration. Cellular senescence is a critical mechanism that underlies IVD aging and degeneration. After chronic and prolonged replication, cellular senescence might occur as a natural part of disc aging, but can be potentially accelerated by growth factor deficiency, oxidant accumulation, and inflammatory irritation. In the early process of disc degeneration, the proliferation check from hypertonicity might be weakened as the gel-like NP undergoes fibrosis and reduces the aggrecan content. By causing a nutrient deficiency in the avascular IVD, the age-related decrease in endplate permeability might contribute to reducing the ECM synthesis, while increased mechanical overloading generates proinflammatory cytokines and enhances the ECM degradation. Cell clusters are possibly formed, and RS is accelerated, by yielding more disc cells to synthesize the ECM and self-repair tissue damage. Once the self-repair mechanism fails to reverse the pathological changes or rebalance anabolism and catabolism, a degenerating disc is likely to experience increased and accumulated senescence-inducing stresses. These might arise from a vicious cycle that amplifies the tissue destroying and pro-catabolic effects of mechanical stresses or, instead, originate from an acute disc injury that breaches the tissue integrity and disturbs metabolic homeostasis. In addition, the accumulated senescent cells themselves, might also contribute to generating more DNA damage and oxidative stresses, which promote SIPS and accelerate the IVDD. In a diabetic or cigarette-exposed IVD, more of these senescence-inducing stresses are thought to generate and accumulate. Although disc cells may undergo senescence at a distinct pace and demonstrate varied SASP features, the immune-clearance of cellular senescence is hampered by the avascular nature of IVD, which also accounts for senescence accumulation during disc aging and degeneration. In a herniated or revascularized degenerative disc, an invasion of inflammatory cells might accelerate rather than alleviate IVDD, probably by amplifying the inflammatory response and irritating discogenic pain. Additionally, cellular senescence might also be dynamically involved in the formation and development of IVD.
Osteoarthritis and Cartilage  , DOI: ( /j.joca )
Copyright © 2015 Osteoarthritis Research Society International Terms and Conditions