Discovery of a bone-like blood particle in the peripheral circulation of humans and rodents.

OBJECTIVE: To characterize ossified bone marrow blood vessels and confirm the presence of ossified particles (OSP) in humans and rodents. METHODS: Human bone marrow blood vessels were processed for scanning and transmission electron microscopy. Whole blood samples were collected from younger (26-39 years; n = 6) and older (55-63 years; n = 6) volunteers and male Fischer-344 rats (1 month, n = 7; 6 months, n = 7; 12 months, n = 7; 18-months, n = 6; 24 months, n = 8). OSP in the whole blood samples were sorted and imaged with microscopy to determine diameter, circularity, and solidity. Additionally, the chemical composition of OSP was determined via elemental analysis. RESULTS: SEM revealed two types of ossified bone marrow blood vessels: that is, "transitioning" and "ossified." OSP were adhered to the surface of transitioning vessels and theoretically gain access to and circulate within the blood. The majority of OSP were ≤15 µm in diameter, but many were of sufficient size to serve as emboli (ie, >15 µm).OSP were predominately oblong in shape and several had jagged tips and edges. CONCLUSIONS: We introduce a novel, bone-like blood particle that may be diagnostic of bone marrow blood vessel ossification. Further, OSP may associate with several disease states (eg, atherosclerosis).

Progressive ossification of the bone marrow vasculature with advancing age corresponds with reduced red blood cell count and percentage of circulating lymphocytes in male Fischer-344 rats.

OBJECTIVE: Assess the link between bone marrow blood vessel ossification, Tb. and cortical bone, and hematological parameters across the lifespan in rats. METHODS: Right femora and whole blood samples were taken from male Fischer-344 rats at 1, 6, 12, 18 and 24 months. Femora were scanned by micro-computed tomography (MicroCT) to determine bone marrow blood vessel ossification (ie, ossified vessel volume [OsVV], ossified vessel thickness (OsV.Th), ossified vessel density (OsV density), and structural model index [SMI]). Bone microarchitecture (ie, bone volume [BV/TV], trabecular thickness, trabecular number, and trabecular separation), density and SMI, and cortical bone parameters (ie, cortical shell thickness, porosity, and density) were also determined by MicroCT. Complete blood counts with differentials were conducted. RESULTS: Ossified vessel volume increased throughout the lifespan, coinciding with reduced trabecular BV/TV and cortical shell thickness at 24 months. Many of the hematological parameters were unchanged (ie, white blood cells, lymphocyte number) or increased (monocyte number, percent monocyte, granulocyte number, percent granulocyte, hemoglobin, hematocrit, mean corpuscular hemoglobin concentration, red blood cell distribution width, platelet, mean platelet volume) with advancing age; however, red blood cells (RBC) and percent lymphocytes (LY%) were reduced at 24 months. In addition, OsV density was similar to trabecular bone density. CONCLUSIONS: Declines in trabecular BV/TV, cortical shell thickness, RBC, and LY% with advanced age coincided with augmented ossification of bone marrow blood vessels.

Lateral Ankle Sprain in a Mouse Model: Lifelong Sensorimotor Dysfunction.

CONTEXT: Ankle sprains are the most common orthopaedic injury that occurs during sport and physical activity. Many individuals who sprain their ankles develop chronic ankle instability (CAI), a condition characterized by recurrent injury, decreased physical activity, and decreased quality of life. These residual impairments are believed to persist for the remainder of the patient's life, in part due to the link between CAI and posttraumatic ankle osteoarthritis. However, this belief remains speculative due to the lack of long-term prospective investigations. OBJECTIVE: To use a mouse model of mild (MILD) and severe (SEVERE) ankle sprains to quantify balance and locomotor adaptations across the lifespan. DESIGN: Cohort study. SETTING: University research laboratory. PATIENTS OR OTHER PARTICIPANTS: Fifty male mice (CBA/J) were randomly placed into a control (SHAM), MILD, or SEVERE group and housed individually. INTERVENTION(S): The MILD group underwent surgical transection of a single right hind-limb lateral ankle ligament, and the SEVERE group had 2 of the lateral ligaments transected. The SHAM group underwent a sham surgery during which no lateral ligaments were transected. MAIN OUTCOME MEASURE(S): After surgically inducing the ankle sprain, we measured balance and gait using a balance beam and footprint test before and every 6 weeks for 78 weeks. RESULTS: Age-related declines in balance but not stride length were exacerbated by an ankle sprain ( P < .001). Balance and stride lengths changed with age ( P < .001). Foot slips were worse in the SEVERE (4.32 ± 0.98) and MILD (3.53 ± 0.98) groups than in the SHAM group (2.16 ± 0.99; P < .001). Right-limb stride length was shorter in the SEVERE group (6.45 cm ± 0.41 cm) than in the SHAM group (6.87 cm ± 0.40 cm; P = .04). CONCLUSIONS: Transecting the lateral ligaments of a mouse hind foot resulted in lifelong sensorimotor dysfunction. Declines starting at 42 weeks postinjury may have represented the onset of posttraumatic osteoarthritis.

Acute Ankle Sprain in a Mouse Model: Changes in Knee-Joint Space.

CONTEXT: Ankle sprains remain the most common orthopaedic injury. Conducting long-term studies in humans is difficult and costly, so the long-term consequences of an ankle sprain are not entirely known. OBJECTIVE: To measure knee-joint space after a single surgically induced ankle sprain in mice. DESIGN: Randomized controlled trial. SETTING: University research laboratory. PATIENTS OR OTHER PARTICIPANTS: Thirty male mice (CBA/2J) were randomly placed into 1 of 3 surgical groups: the transected calcaneofibular ligament (CFL) group, the transected anterior talofibular ligament/CFL group, or a sham treatment group. The right ankle was operated on in all mice. MAIN OUTCOME MEASURE(S): Three days after surgery, all of the mice were individually housed in cages containing a solid-surface running wheel, and daily running-wheel measurements were recorded. Before surgery and every 6 weeks after surgery, a diagnostic ultrasound was used to measure medial and lateral knee-joint space in both hind limbs. RESULTS: Right medial (P = .003), right lateral (P = .002), left medial (P = .03), and left lateral (P = .002) knee-joint spaces decreased across the life span. The mice in the anterior talofibular ligament/CFL group had decreased right medial (P = .004) joint space compared with the sham and CFL groups starting at 24 weeks of age and continuing throughout the life span. No differences occurred in contralateral knee-joint degeneration among any of the groups. CONCLUSIONS: Based on current data, mice that sustained a surgically induced severe ankle sprain developed greater joint degeneration in the ipsilateral knee. Knee degeneration could result from accommodation to the laxity of the ankle or biomechanical alterations secondary to ankle instability. A single surgically induced ankle sprain could significantly affect knee-joint function.

Altered left ventricular performance in aging physically active mice with an ankle sprain injury.

We assessed the impact of differing physical activity levels throughout the lifespan, using a musculoskeletal injury model, on the age-related changes in left ventricular (LV) parameters in active mice. Forty male mice (CBA/J) were randomly placed into one of three running wheel groups (transected CFL group, transected ATFL/CFL group, SHAM group) or a SHAM Sedentary group (SHAMSED). Before surgery and every 6 weeks after surgery, LV parameters were measured under 2.5 % isoflurane inhalation. Group effects for daily distance run was significantly greater for the SHAM and lesser for the ATLF/CFL mice (p = 0.013) with distance run decreasing with age for all mice (p < 0.0001). Beginning at 6 months of age, interaction (group × age) was noted with LV posterior wall thickness-to-radius ratios (h/r) where h/r increased with age in the ATFL/CFL and SHAMSED mice while the SHAM and CFL mice exhibited decreased h/r with age (p = 0.0002). Passive filling velocity (E wave) was significantly greater in the SHAM mice and lowest for the ATFL/CFL and SHAMSED mice (p < 0.0001) beginning at 9 months of age. Active filling velocity (A wave) was not different between groups (p = 0.10). Passive-to-active filling velocity ratio (E/A ratio) was different between groups (p < 0.0001), with higher ratios for the SHAM mice and lower ratios for the ATFL/CFL and SHAMSED mice in response to physical activity beginning at 9 months of age. Passive-to-active filling velocity ratio decreased with age (p < 0.0001). Regular physical activity throughout the lifespan improved LV structure, passive filling velocity, and E/A ratio by 6 to 9 months of age and attenuated any negative alterations throughout the second half of life. The diastolic filling differences were found to be significantly related to the amount of activity performed by 9 months and at the end of the lifespan.

An Acute Lateral Ankle Sprain Significantly Decreases Physical Activity across the Lifespan.

We do not know the impact an ankle sprain has on physical activity levels across the lifespan. With the negative consequences of physical inactivity well established, understanding the effect of an ankle sprain on this outcome is critical. The objective of this study was to measure physical activity across the lifespan after a single ankle sprain in an animal model. Thirty male mice (CBA/J) were randomly placed into one of three groups: the transected calcaneofibular ligament (CFL) group, the transected anterior talofibular ligament (ATFL)/CFL group, and a SHAM group. Three days after surgery, all of the mice were individually housed in a cage containing a solid surface running wheel. Physical activity levels were recorded and averaged every week across the mouse's lifespan. The SHAM mice ran significantly more distance each day compared to the remaining two running groups (post hoc p = 0.011). Daily duration was different between the three running groups (p = 0.048). The SHAM mice ran significantly more minutes each day compared to the remaining two running groups (post hoc p=0.046) while the ATFL/CFL mice ran significantly less minutes each day (post hoc p = 0.028) compared to both the SHAM and CFL only group. The SHAM mice ran at a faster daily speed versus the remaining two groups of mice (post hoc p = 0.019) and the ATFL/CFL mice ran significantly slower each day compared to the SHAM and CFL group (post hoc p = 0.005). The results of this study indicate that a single ankle sprain significantly decreases physical activity across the lifespan in mice. This decrease in physical activity can potentially lead to the development of numerous chronic diseases. An ankle sprain thus has the potential to lead to significant long term health risks if not treated appropriately. Key pointsA single ankle significantly decreased physical activity levels in mice across the lifespan.Decreased physical activity could significantly negatively impact overall health if not modified.Initial treatment and rehabilitation of ankle sprains needs to be studied to determine ways to keep physical activity levels up after injury.

Developing a mouse model of chronic ankle instability.

INTRODUCTION: Ankle sprains are the most common orthopedic pathology experienced during sport and physical activity and often result in chronic ankle instability (CAI). Understanding how to prevent CAI is difficult because of the costs and logistics associated with clinical trials aimed at preventing the heterogeneous symptoms associated with CAI. Thus, a need exists to develop an animal model that presents similar long-term consequences as CAI to assess preclinical data. Thus, the purpose was to determine whether surgically transecting the lateral ligaments of a mouse hind limb results in the development of CAI-like symptoms 12 months after injury. METHODS: Thirty male mice (CBA/J) were randomly placed into a SHAM (control), CFL (calcaneofibular ligament; mild ankle sprain), or ATFL/CFL (anterior talofibular ligament/CFL; severe) ankle sprain group and housed individually. Three days after surgically transecting the respective lateral ligaments, mice were given a solid surface running wheel and daily running wheel measurements were recorded. Outcome measures of balance and gait were obtained before and at 4, 48, 54, and 60 wk after injury. RESULTS: The ATFL/CFL group had significantly more hind foot slips than the CFL and SHAM groups (P < 0.05). The CFL also had more hind foot slips relative to the SHAM group (P < 0.05). The ATFL/CFL group was significantly less physically active relative to the SHAM and CFL groups (P < 0.05). A cut score of 4.75 foot slips had a sensitivity of 0.68 and specificity of 1.00 and indicates that 70% (14/20) of mice with an ankle sprain had developed CAI. CONCLUSIONS: The results of this study indicate that an acute ankle sprain in mice can result in the development of CAI-like symptoms 12 months after injury.

Lifelong physical activity and knee osteoarthritis development in mice.

AIM: To examine the effect of lifelong physical activity on the development of knee osteoarthritis (OA). METHODS: Fifteen C57Bl/6J mice (8 male and 7 female) were individually caged with a running wheel, magnetic sensor and digital odometer, and 15 mice (7 male and 8 female) were individually housed without a running wheel. Each mouse was provided with standard chow and water ad libitum. Running wheel distance, duration and speed were recorded daily. A diagnostic ultrasound (SONOS 5500 Ultrasound and 15-6L ultrasound probe) was used to measure medial and lateral knee joint space in both hind limbs every month until the end of the mice's lifespans. RESULTS: Right medial (P = 0.002), right lateral (P < 0.0001), left medial (P = 0.01), and left lateral (P = 0.007) knee joint spaces were significantly larger from months 3 to 12 in the physically active C57Bl/6J mice. However, there was no significant difference between all knee joint spaces of the running and sedentary C57Bl/6J mice throughout the remainder of the lifespan. All hindlimb knee joint spaces significantly declined with aging (P < 0.0001). CONCLUSIONS: During the first 12 months of the lifespan, physical activity seemed to protect the joint from degeneration; however; after that there was no protective benefit. This may be due to the fact that C57Bl/6J mice are genetically known to develop OA at 9 months of age. Physical activity may have no impact on this genetic predisposition. Further study in mice not genetically predisposed to develop OA is needed.

Acute ankle sprain in a mouse model.

INTRODUCTION: Ankle sprains remain the most common orthopedic pathology. Conducting long-term studies in humans is difficult and costly, so the long-term consequences of an ankle sprain are not entirely known. PURPOSE: The objective of this study is to develop and test a mechanical ankle instability model in mice. METHODS: Thirty male mice (CBA/2J) were randomly placed into one of three groups: the transected calcaneal fibular ligament (CFL) group, the transected anterior talofibular ligament (ATFL)/CFL group, and a SHAM group. Three days after surgery, all of the mice were individually housed in a cage containing a solid surface running wheel, and daily running wheel measurements were recorded. Before and after surgery, measures of balance and gait were measured on all mice for 4 wk. RESULTS: The mice in the ATFL/CFL group had significantly decreased duration (P = 0.0239), distance (P = 0.013), and speed (P = 0.003) compared with the SHAM group during week 1. During weeks 2 and 3, the ATFL/CFL group had significantly less distance (P = 0.0001) and duration (P = 0.002) compared with the SHAM and CFL-only group. The transection of the lateral ankle ligaments did affect the number of slips experienced during the balance test. The ATFL/CFL group had greater slips at 1 and 4 wk postsurgery (P = 0.05), whereas the CFL-only group had greater slips at 3 d and 1 wk postsurgery (P = 0.05). Relative to the SHAM group, the ATFL/CFL group and CFL-only group had smaller right-stride lengths (involved limb) at 3 d postsurgery (P = 0.05). The ATFL/CFL group also had smaller right-side stride lengths at 1 and 4 wk postsurgery (P = 0.05). CONCLUSIONS: The results of this study indicate that a mouse model can be used to induce mechanical instability in the ankle.