Tibial Loading Increases Osteogenic Gene Expression and Cortical Bone Volume in Mature and Middle-Aged Mice

Abstract

There are conflicting data on whether age reduces the response of the skeleton to mechanical stimuli. We examined this question in female BALB/c mice of different ages, ranging from young to middle-aged (2, 4, 7, 12 months). We first assessed markers of bone turnover in control (non-loaded) mice. Serum osteocalcin and CTX declined significantly from 2 to 4 months (p<0.001). There were similar age-related declines in tibial mRNA expression of osteoblast- and osteoclast-related genes, most notably in late osteoblast/matrix genes. For example, Col1a1 expression declined 90% from 2 to 7 months (p<0.001). We then assessed tibial responses to mechanical loading using age-specific forces to produce similar peak strains (−1300 µε endocortical; −2350 µε periosteal). Axial tibial compression was applied to the right leg for 60 cycles/day on alternate days for 1 or 6 weeks. qPCR after 1 week revealed no effect of loading in young (2-month) mice, but significant increases in osteoblast/matrix genes in older mice. For example, in 12-month old mice Col1a1 was increased 6-fold in loaded tibias vs. controls (p = 0.001). In vivo microCT after 6 weeks revealed that loaded tibias in each age group had greater cortical bone volume (BV) than contralateral control tibias (p<0.05), due to relative periosteal expansion. The loading-induced increase in cortical BV was greatest in 4-month old mice (+13%; p<0.05 vs. other ages). In summary, non-loaded female BALB/c mice exhibit an age-related decline in measures related to bone formation. Yet when subjected to tibial compression, mice from 2–12 months have an increase in cortical bone volume. Older mice respond with an upregulation of osteoblast/matrix genes, which increase to levels comparable to young mice. We conclude that mechanical loading of the tibia is anabolic for cortical bone in young and middle-aged female BALB/c mice.

Aging diminishes lamellar and woven bone formation induced by tibial compression in adult C57BL/6

“…We have previously applied tibial compression to BALB/c mice across their life span [17, 18]. Our initial study reported no decline in periosteal mechano-responsiveness with aging when comparing 7– to 22-month-old male BALB/c mice [17], a finding that was unexpected and at odds with prior reports in humans, rats and C57Bl/6 mice [5, 8].…”

“…Scan resolution was 21 μm (70 kV, 114 μA, 100 ms integration time). This scanning protocol has proven sensitive enough to detect longitudinal bone changes [17, 18], and represents a compromise that sacrifices optimal image quality in order to reduce radiation exposure. Negative radiation side effects have been noted by others who scanned at a higher resolution (10 μm) and more frequently (every 5 days) [23].…”

“…In contrast to most direct loading studies [12–14], tibial compression of aged (22 month), male BALB/c mice induced greater endocortical bone formation and equivalent periosteal bone formation compared to young-adult (7 month) mice [17]. On the other hand, accrual of cortical bone volume was greater in 4-month old female BALB/c mice than in 7– and 12–month-old mice loaded by tibial compression [18]. The basis underlying these age effects in BALB/c mice is unclear but while BALB/c mice lose a modest amount of bone with advanced aging [3], it may be more appropriate to use C57BL/6 mice, which display greater cortical and trabecular bone loss with aging [19-21].…”

Transcriptional profiling of cortical versus cancellous bone from mechanically-loaded murine tibiae reveals differential gene expression

“…RNA isolation was performed using Trizol (Life Technologies, Carlsbad, CA) and RNeasy Mini kit (Qiagen, Germantown, MD) as described previously [13]. Briefly, individual cancellous and cortical samples were pulverized in liquid nitrogen-cooled flasks (Mikrodismembrator S, Sartorius Stedim Biotech, Bohemia, NY, USA).…”

Monitoring in vivo (re)modeling: A computational approach using 4D microCT data to quantify bone surface movements

“…MicroCT images are widely used to determine 3D micro-morphological parameters of rodent bone [31][32][33]. Being a non-invasive technique microCT can be used to monitor structural changes of bone in longitudinal studies of living mice [34][35][36][37]. Recently longitudinal in vivo microCT was combined with 3D image registration, automated volumetric segmentation and morphological evaluation to volumetrically quantify spatial bone (re)modeling (formation and resorption) events on the bone surfaces of living mice [35,36,38].…”