Skeletal Mineralization Deficits and Impaired Biogenesis and Function of Chondrocyte-Derived Matrix Vesicles in Phospho1-/- and Phospho1/Pit1 Double Knockout Mice

Authors

Manisha C. Yadav, Massimo Bottini., Esther Cory, Kunal Bhattacharya., Pia Kuss, Sonoko Narisawa, Robert L. Sah, Laurent Beck, Bengt Fadeel, Colin Farquharson and José Luis Millán

Abstract

We have previously shown that ablation of either the Phospho1 or Alpl gene, encoding PHOSPHO1 and tissue-nonspecific alkaline phosphatase (TNAP) respectively, lead to hyperosteoidosis but that their chondrocyte- and osteoblast-derived matrix vesicles (MVs) are able to initiate mineralization. In contrast, the double ablation of Phospho1 and Alpl completely abolish initiation and progression of skeletal mineralization. We argued that MVs initiate mineralization by a dual mechanism: PHOSPHO1-mediated intravesicular generation of Pi and phosphate transporter-mediated influx of Pi. To test this hypothesis, we generated mice with col2a1-driven cre-mediated ablation of Slc20a1, hereafter referred to as Pit1, alone or in combination with a Phospho1 gene deletion. Pit1col2/col2 mice did not show any major phenotypic abnormalities, while severe skeletal deformities were observed in the [Phospho1-/-; Pit1col2/col2] double knockout mice that were more pronounced than those observed in the Phospho1-/- mice. Histological analysis of [Phospho1-/-; Pit1col2/col2] bones showed growth plate abnormalities with a shorter hypertrophic chondrocyte zone and extensive hyperosteoidosis. The [Phospho1-/-; Pit1col2/col2] skeleton displayed significantly decreases in BV/TV%, trabecular number and bone mineral density, as well as decreased stiffness, decreased strength, and increased post-yield deflection compared to Phospho1-/- mice. Using atomic force microscopy we found that ∼80% of [Phospho1-/-; Pit1col2/col2] MVs were devoid of mineral in comparison to ∼50% for the Phospho1-/- MVs and ∼25% for the WT and Pit1col2/col2 MVs. We also found a significant decrease in the number of MVs produced by both Phospho1-/- and [Phospho1-/-; Pit1col2/col2] chondrocytes. These data support the involvement of PiT-1 in the initiation of skeletal mineralization and provide compelling evidence that PHOSPHO1 function is involved in MV biogenesis.

Link to Article

http://dx.doi.org/10.1002/jbmr.2790

Injury response of geriatric mouse patellar tendons

Authors

Michael J. Mienaltowski, Andrew A. Dunkman, Mark R. Buckley, David P. Beason, Sheila M. Adams, David E. Birk and Louis J. Soslowsky

Abstract

Injury adversely impacts the structure and mechanical properties of a tendon, thus causing pain and disability. Previously, we demonstrated that patellar tendons in mature (P150) and aged (P300) mice do not recover original functionality, even 6 weeks after injury, and that uninjured geriatric tendons (P570) are functionally inferior to uninjured mature tendons. In this study, we hypothesized that the repair response in injured geriatric mice would be further compromised, thus undermining patellar tendon function post-injury. Patellar tendons from wild-type mice were injured at 540 days. At 3 and 6 weeks post-surgery, structural, mechanical, and biochemical analyses were performed and compared to uninjured controls. Mechanical properties of geriatric tendons failed to improve after injury. When compared to mature and aged tendons post-injury, it was determined that at no age was there a suitable repair response. In previous studies, we were able to associate the absence of SLRPs with phenotypic changes both early and late in repair. Here we found that SLRPs were significantly decreased after injury, thus offering a possible explanation for why geriatric tendons were unable to mount an adequate repair response. Thus, we conclude that regardless of age after maturity, tendon healing ultimately results in a substandard outcome.

Link to Article

http://dx.doi.org/10.1002/jor.23144

Stromal-Initiated Changes in the Bone Promote Metastatic Niche Development

Authors

Xianmin Luo, Yujie Fu, Andrew J. Loza, Bhavna Murali, Kathleen M. Leahy, Megan K. Ruhland, Margery Gang, Xinming Su, Ali Zamani, Yu Shi, Kory J. Lavine, David M. Ornitz, Katherine N. Weilbaecher, Fanxin Long, Deborah V. Novack, Roberta Faccio, Gregory D. Longmore, Sheila A. Stewart

Abstract

More than 85% of advanced breast cancer patients suffer from metastatic bone lesions, yet the mechanisms that facilitate these metastases remain poorly understood. Recent studies suggest that tumor-derived factors initiate changes within the tumor microenvironment to facilitate metastasis. However, whether stromal-initiated changes are sufficient to drive increased metastasis in the bone remains an open question. Thus, we developed a model to induce reactive senescent osteoblasts and found that they increased breast cancer colonization of the bone. Analysis of senescent osteoblasts revealed that they failed to mineralize bone matrix and increased local osteoclastogenesis, the latter process being driven by the senescence-associated secretory phenotype factor, IL-6. Neutralization of IL-6 was sufficient to limit senescence-induced osteoclastogenesis and tumor cell localization to bone, thereby reducing tumor burden. Together, these data suggest that a reactive stromal compartment can condition the niche, in the absence of tumor-derived signals, to facilitate metastatic tumor growth in the bone.

Link to Article

http://dx.doi.org/10.1016/j.celrep.2015.12.016

Manual therapy as an effective treatment for fibrosis in a rat model of upper extremity overuse injury

Authors

Geoffrey M. Bove, Michele Y. Harris, Huaqing Zhao, Mary F. Barbe

Abstract

Key clinical features of carpal tunnel syndrome and other types of cumulative trauma disorders of the hand and wrist include pain and functional disabilities. Mechanistic details remain under investigation but may involve tissue inflammation and/or fibrosis. We examined the effectiveness of modeled manual therapy (MMT) as a treatment for sensorimotor behavior declines and increased fibrogenic processes occurring in forearm tissues of rats performing a high repetition high force (HRHF) reaching and grasping task for 12 weeks. Young adult, female rats were examined: food restricted control rats (FRC, n = 12); rats that were trained for 6 weeks before performing the HRHF task for 12 weeks with no treatment (HRHF-CON, n = 11); and HRHF task rats received modeled manual therapy (HRHF-MMT, n = 5) for 5 days/week for the duration of the 12-week of task. Rats receiving the MMT expressed fewer discomfort-related behaviors, and performed progressively better in the HRHF task. Grip strength, while decreased after training, improved following MMT. Fibrotic nerve and connective tissue changes (increased collagen and TGF-β1 deposition) present in 12-week HRHF-CON rats were significantly decreased in 12-week HRHF-MMT rats. These observations support the investigation of manual therapy as a preventative for repetitive motion disorders.

Link to Article

http://dx.doi.org/10.1016/j.jns.2015.12.029

Arctic Ground Squirrels Limit Bone Loss during the Prolonged Physical Inactivity Associated with Hibernation

Authors

Samantha J. Wojda, Richard A. Gridley, Meghan E. McGee-Lawrence, Thomas D. Drummer, Ann Hess, Franziska Kohl, Brian M. Barnes, and Seth W. Donahue

Abstract

Prolonged disuse (e.g., physical inactivity) typically results in increased bone porosity, decreased mineral density, and decreased bone strength, leading to increased fracture risk in many mammals. However, bears, marmots, and two species of ground squirrels have been shown to preserve macrostructural bone properties and bone strength during long seasons of hibernation while they remain mostly inactive. Some small hibernators (e.g., 13-lined ground squirrels) show microstructural bone loss (i.e., osteocytic osteolysis) during hibernation, which is not seen in larger hibernators (e.g., bears and marmots). Arctic ground squirrels (Urocitellus parryii) are intermediate in size between 13-lined ground squirrels and marmots and are perhaps the most extreme rodent hibernator, hibernating for up to 8 mo annually with body temperatures below freezing. The goal of this study was to quantify the effects of hibernation and inactivity on cortical and trabecular bone properties in arctic ground squirrels. Cortical bone geometrical properties (i.e., thickness, cross-sectional area, and moment of inertia) at the midshaft of the femur were not different in animals sampled over the hibernation and active seasons. Femoral ultimate stress tended to be lower in hibernators than in summer animals, but toughness was not affected by hibernation. The area of osteocyte lacunae was not different between active and hibernating animals. There was an increase in osteocytic lacunar porosity in the hibernation group due to increased lacunar density. Trabecular bone volume fraction in the proximal tibia was unexpectedly greater in the hibernation group than in the active group. This study shows that, similar to other hibernators, arctic ground squirrels are able to preserve many bone properties during hibernation despite being physically inactive for up to 8 mo.

Link to Article

http://dx.doi.org/10.1086/684619

Dual function of Bmpr1a signaling in restricting preosteoblast proliferation and stimulating osteoblast activity in the mouse

Authors

Joohyun Lim, Yu Shi, Courtney M. Karner, Seung-Yon Lee, Wen-Chih Lee, Guangxu He, Fanxin Long

Abstract

Exogenous bone morphogenetic proteins (Bmp) are well known to induce ectopic bone formation, but the physiological effect of Bmp signaling on normal bone is not completely understood. By deleting the receptor Bmpr1a in osteoblast-lineage cells with Dmp1-Cre, we observed a dramatic increase in trabecular bone mass in postnatal mice, due to a marked increase in osteoblast number likely driven by hyperproliferation of Sp7+ preosteoblasts. Similarly, inducible deletion of Bmpr1a in Sp7-positive cells specifically in postnatal mice increased trabecular bone mass. However, deletion of Smad4 by the same approaches had only a minor effect, indicating that Bmpr1a signaling suppresses trabecular bone formation through effectors beyond Smad4. Besides increasing osteoblast number in the trabecular bone, deletion of Bmpr1a by Dmp1-Cre also notably reduced osteoblast activity, resulting in attenuation of periosteal growth. The impairment in osteoblast activity correlated with reduced mTORC1 signaling in vivo, whereas inhibition of mTORC1 activity abolished the induction of protein anabolism genes by Bmp2 in vitro. Thus, physiological Bmpr1a signaling in bone exerts dual function in both restricting preosteoblast proliferation and promoting osteoblast activity.

Link to Article

http://dx.doi.org/10.1242/dev.126227