Blaine A. Christiansen

Osteocytes regulate skeletal homeostasis and respond to oxygen deprivation via hypoxia-inducible factors (HIFs), the principal transcriptional mediators of the cellular hypoxia response. There are 2 transcriptionally active HIF-α paralogs, HIF-1α and HIF-2α, that have overlapping and distinct transcriptional targets. Under normoxic conditions, the E3 ubiquitin ligase Von Hippel–Lindau (VHL) acts as the master regulator of HIFs by targeting them for proteasomal degradation, whereas hypoxia prevents this, allowing for HIF stabilization and accumulation. Osteocyte-enriched deletion of Vhl (Vhl cKO) produces a high bone mass (HBM) phenotype. Vhl loss stabilizes both HIFs, leaving their distinct contributions to this HBM phenotype undefined. To dissect their role in osteocytes, we developed compound Vhl; Hif1a cKO, Vhl; Hif2a cKO mice, mice expressing individual and combined degradation-resistant (cDR) paralogs HIF-1α (HIF-α cDR), HIF-2α (HIF-2α cDR), and HIF-1α; HIF-2α cDR and evaluated their impact on bone structure and function. Micro-CT revealed that Vhl cKO mice exhibited the highest trabecular bone volume fraction, a phenotype not fully reproduced in any HIF-modified genotype. Among partial phenocopies, Vhl; Hif2a cKO produced the largest partial increase, followed by Vhl; Hif1a cKO, and HIF-2α cDR. HIF-1α; HIF-2α cDR displayed modest elevation, while cre-negative and HIF-1α cDR were indistinguishable. Cortical area expansion required Vhl deletion and was not reproduced by single-paralog HIF stabilization. While Vhl cKO femora exhibited significantly increased whole-bone mechanical properties relative to sex- and age-matched controls, all HIF-modified genotypes had similar material properties and HIF-2α activation appeared to compromise material integrity. HIF-1α and HIF-2α therefore exert non-redundant, compartment-specific roles in the Vhl cKO HBM phenotype and are each required for maximal bone integrity. Yet, no HIF-modified genotype, including dual HIF-1α/HIF-2α stabilization, reproduced the Vhl cKO phenotype, implicating the possibility of VHL functions beyond HIF stabilization in the full skeletal response.

Alzheimer’s disease (AD) and osteoarthritis (OA) are 2 of the most common health conditions affecting the elderly. Recent studies in mice and humans have described an association between OA and AD, with prevalent OA increasing the risk of developing AD-like cognitive declines. In this study, we investigated brain and knee joint degeneration and neurocognitive dysfunction in a mouse model of AD (APP/PS1 double transgenic mice) in the presence or absence of OA induced by non-invasive ACL rupture. We hypothesized that OA would be associated with increased beta amyloid (Aβ) and phosphorylated tau (pTau) in the injured joint and brain and would be associated with accelerated cognitive decline in APP/PS1-Tg mice. Non-invasive knee injury was performed at 7 mo of age in male and female APP/PS1-Tg mice and non-transgenic (WT) littermates. Neurocognitive tests, including open field, novel object recognition, and Morris water maze, were performed 6 wk post-injury. Knees were also assessed at 6 wk post-injury to quantify OA, and Aβ, Tau, and pTau levels in the brain and knee joints. We found that female APP/PS1-OA mice had significantly more osteophyte formation than WT-OA mice, and significantly greater Aβ and pTau levels in the brain and knee joints than in WT or APP/PS1-Sham mice. These results provide insights about the pathoetiology of AD and OA and suggest possible common mechanisms connecting OA-associated inflammation to cognitive function. Identifying novel mechanisms of crosstalk between these 2 conditions could improve clinical care and quality of life for at-risk patient populations.

Objective: Severe burn injuries can lead to systemic bone loss, increasing the risk of secondary fragility fractures. Evaluating treatments aimed at mitigating post-traumatic systemic bone loss is critically important for preventing secondary fractures in burn patients. This study evaluated the efficacy of bisphosphonate (alendronate), oxandrolone, and vitamin D/calcium supplementation in mitigating post-traumatic bone loss and promoting recovery of bone in a murine model of burn-induced systemic bone loss, in which a full-thickness thermal skin burn involving approximately 15% total body surface area was used to model severe burn injury. Methods: Male C57BL/6J mice were divided into five groups: control, burn with no treatment, burn + alendronate, burn + vitamin D/calcium, and burn + oxandrolone. Full thickness burn injuries were induced on the back of mice by submerging in 65° C water for 20 s; debridement and matrix application were performed 48 hours after injury, and treatments were administered starting on the day of injury. Whole-body and site-specific bone mineral density (BMD) and bone mineral composition (BMC) were assessed weekly using dual energy x-ray absorptiometry. Micro-computed tomography and 3-point bending were performed at weeks 2 and 6 post-injury to assess bone microstructure and mechanical properties. Results: Untreated burned mice exhibited significant losses in BMD (-12.0 ± 7.6% from baseline, p=0.025) and BMC (-14.7 ± 13.4% from baseline, p=0.021), peaking at 3 weeks post-injury. Alendronate treatment ameliorated loss of BMD (-5.7 ± 10.6% from baseline, p=0.006 compared to untreated Burn mice) and BMC (-4.2 ± 11.6% from baseline, p=0.006 compared to untreated Burn mice) and accelerated the recovery process, reaching BMD and BMC values greater than baseline by week 6. Vitamin D/calcium supplementation (peak BMD loss -8.6 ± 8.1%, p=0.974; BMC -17.6 ± 14.2%, p=0.973) and oxandrolone treatments (peak BMD loss -17.1 ± 12.5%, p=0.408; BMC -19.5 ± 16.4%, p=0.404) demonstrated no protective or improved recovery effects for bone loss following burns. Conclusion: Alendronate was the superior protective treatment for preventing burn-induced bone loss and enhancing recovery in a murine model of post-burn systemic bone loss compared with oxandrolone, Vitamin D/ Calcium supplementation, and no treatment. These findings suggest that early administration of bisphosphonates in burn patients may be a promising method for mitigating post-traumatic bone loss.

Background: Treatments for Achilles tendon rupture (ATR) are insufficient because they often do not result in full recovery and return to activity. There is no universal standard protocol for ATR, with surgeons using different repair techniques and materials. Some surgeons prefer nonresorbable (NR) suture due to the superior tensile strength, and others use resorbable (RES) because they decrease foreign body reactions. Methods: To establish a model for future interventional studies, we performed ATRs on 3-month-old female Sprague-Dawley rats using either NR or RES suture to repair the tendon and collected tissues at 1 and 2 months. Results: No significant differences in mechanical or material properties as a function of suture type were noted. Independent of suture type, tendons were wider at 1 month and both wider and thicker at 2 months, resulting in large increases in cross-sectional area (CSA) at each time point. The repaired tendons were stronger than the uninjured contralateral tendons at 1 month, but their strength decreased from the first month to the second despite increasing in CSA during this time, indicating progressive deterioration of tissue quality. Conclusions: Our data suggest that without loads in excess of body weight, the healing process becomes maladaptive after repair regardless of suture material. Level of Evidence/Clinical Relevance: These experiments provide Level I pre-clinical evidence showing that RES suture performs at least as well as NR in all physiological measures following repair, indicating it is the preferable repair material for ATR. See Instructions for Authors for a complete description of levels of evidence.

Purpose: People who sustain joint injuries such as anterior cruciate ligament (ACL) rupture often go on to develop post-traumatic osteoarthritis (PTOA). ACL injuries are often treated with ACL reconstruction, but there is typically a gap of several weeks between injury and surgery. However, it is unclear how loading or unloading of the injured joint during the early postinjury period affects the progression of PTOA. The goal of this study was to determine how unloading between noninvasive ACL injury and surgical restabilization of the injured joint affects PTOA progression in mice. Findings: Mice were subjected to noninvasive ACL injury or no injury followed by 1 week of hindlimb unloading (HLU) or normal cage activity. After 1 week of HLU or cage activity, mice underwent restabilization surgery or no surgery. ACL injury resulted in considerable epiphyseal trabecular bone loss regardless of HLU or cage activity. HLU groups exhibited significantly reduced chondrophyte/osteophyte formation, OA scoring, and synovitis at day 42. Single-cell RNA sequencing revealed that 1 week of HLU resulted in more neutrophils and less monocytes-macrophages in the injured joint. Conclusions: This study establishes that 1 week of HLU after ACL injury effectively slowed PTOA progression, suggesting that the early inflammatory response and joint instability play a key role in PTOA initiation and progression, and neutrophils and monocytes-macrophages play roles in the modulation. However, subsequent joint restabilization surgery caused greater inflammatory protease activity in the joint and exacerbated the loss of epiphyseal trabecular bone but did not significantly diminish OA score or synovitis.

Injury of the anterior cruciate ligament (ACL) is a common sports injury that can lead to post-traumatic osteoarthritis (PTOA) within 10-20 years. Surgical ACL reconstruction is often performed several weeks or months after injury, and this period between injury and ACL reconstruction may be a critical time for determining the risk of long-term PTOA progression. However, few (if any) studies in human patients have investigated the long-term effects of exercise or unloading between ACL injury and surgery. Early mobilization is often recommended to maintain range of motion and muscle strength, which are beneficial for positive outcomes of ACL reconstruction, but it is unknown what effects early mobilization or unloading have on long-term PTOA progression. In preclinical animal studies, a brief period of joint unloading immediately after ACL injury significantly decreased osteophyte formation and articular cartilage degeneration, while longer-term non-weightbearing caused muscle atrophy and articular cartilage degradation. Similarly, preclinical studies have shown that different intensities of exercise after knee injury can have divergent effects on PTOA development. Low intensity exercise was protective against joint degeneration, while higher intensity exercise accelerated PTOA progression. The beneficial or detrimental effects of exercise and unloading following ACL injury are likely dependent on the timing, duration, and intensity of these biomechanical interventions. This review summarizes the effects of these biomechanical interventions after ACL injury in both humans and animal models, with the goal of informing therapeutic and rehabilitation strategies for slowing or preventing PTOA progression after injury.