The aging mouse partially models the aging human spine: lumbar and coccygeal disc height, composition, mechanical properties, and Wnt signaling in young and old mice

Abstract

Murine lumbar and coccygeal (tail) regions of spines are commonly used to study cellular signaling of age-related disc diseases, but the tissue-level changes of aging intervertebral discs and vertebrae of each spinal region remain unclear. Furthermore, the impact of aging lumbar and coccygeal discs on Wnt/β-catenin signaling, which is putatively involved in the catabolism of intervertebral discs, is also unclear. We compared disc/vertebrae morphology and mechanics and biochemical composition of intervertebral discs from lumbar and coccygeal regions between young (4-5 mo) and old (20-22 mo) female C57BL/6 mice. Center intervertebral disc height from both regions was greater in old discs than young discs. Compared with young, old lumbar discs had a lower early viscous coefficient (a measure of stiffness) by 40%, while conversely old coccygeal discs were stiffer by 53%. Biochemically, old mice had double the collagen content in lumbar and coccygeal discs of young discs, greater glycosaminoglycan in lumbar discs by 37%, but less glycosaminoglycan in coccygeal discs by 32%. Next, we compared Wnt activity of lumbar and coccygeal discs of 4- to 5-mo and 12- to 14-mo TOPGAL mice. Despite the disc-specific changes, aging decreased Wnt signaling in the nucleus pulposus from both spinal regions by ≥64%. Compared with young, trabecular bone volume/tissue volume and ultimate force were less in old lumbar vertebrae, but greater in old coccygeal vertebrae. Thus intervertebral discs and vertebrae age in a spinal region-dependent manner, but these differential age-related changes may be uncoupled from Wnt signaling. Overall, lumbar and coccygeal regions are not interchangeable in modeling human aging.

Keywords: WNT/β-catenin; aging; caudal; mouse; tail.

Fig. 1.

A: the 20th loading and unloading tension-compression cycle (shaded diamonds) served as the data for the polynomial fit, which was used to determine the compressive, tensile, and neutral zone stiffness of bone-disc-bone motion segments. B: following tension-compression testing, motion segments were compressed under force control for 1 h to determine creep properties.

Fig. 2.

Vertebrae were compressed to failure to determine stiffness, yield force, ultimate force, and energy-to-ultimate force.

Fig. 3.

Regional height of lumbar and coccygeal intervertebral discs from young and old mice. A: both lumbar and coccygeal discs were greater with aging. B: disc shape (convexity) was greater in old coccygeal discs. Disc shape represents the ratio of center height to the mean of anterior and posterior height. Values are means ± SD. *Significant difference vs. young, P < 0.05.

Fig. 4.

Tensile stiffness (A) and early viscous damping (B) coefficient of lumbar and coccygeal intervertebral discs from young and old mice. These mechanical properties were less in old lumbar discs and greater in old coccygeal discs than young discs. Values are means ± SD. *Significant difference vs. young, P < 0.05.

Fig. 5.

Collagen (A) and glycosaminoglycan (B) of lumbar and coccygeal intervertebral discs from young and old mice. Old lumbar and coccygeal discs had greater collagen. Old lumbar discs had greater glycosaminoglycan, but old coccygeal discs had less glycosaminoglycan. sGAG, sulfated-glycosaminoglycans. Values are means ± SD. *Significant difference vs. young, P < 0.05. Toluidine blue/Fast green stained for glycosaminoglycan of young (C and D) and old (E and F) lumbar spines and of young (G and H) and old (I and J) coccygeal spines is shown. Old lumbar discs had greater staining in the nucleus pulposus (C and E), and old coccygeal discs had less staining than young discs of the respective region (G and I). D, F, H, and J: regions of interest within the nucleus pulposus are highlighted in white boxes. D and F: in lumbar discs, aging disorganized the notochordal band and blurred its distinction from the matrix. H and J: in coccygeal discs, aged discs lost notochordal cells. Scale bars: C, E, G, I = 500 μm; D, F, H, J = 20 μm.

Fig. 6.

Histological staining of young (A and C) and middle-aged (B and D) lumbar (A and B) and coccygeal (C and D) discs from TOPGAL mice. Positive X-gal staining (blue cells) is indicative of activated Wnt signaling. Quantification of the staining for the nucleus pulposus (E) and annulus fibrosus (F) shows that aging decreased the percentage of the Wnt active cells in the nucleus pulposus in lumbar and coccygeal discs, whereas aging did not alter the number of Wnt active cells in the annulus fibrosus. Scale bar is 500 μm. Values are means ± SD. *Significant difference vs. young, P < 0.05.

Fig. 7.

Gene expression of β-catenin of young and middle-aged lumbar and coccygeal discs. Values are means ± SD. *Significant difference vs. young, P < 0.05.

Fig. 8.

A: trabecular bone volume (BV) fraction of lumbar and coccygeal intervertebral discs from young and old mice. Trabecular BV fraction was less in old lumbar discs and greater in old coccygeal discs than the respective young disc. TV, tissue volume. B: ultimate force. Values are means ± SD. *Significant difference vs. young, P < 0.05.