Establishment of a Neurodegenerative Charcot Mouse Model

Abstract

Background: This study aimed to mimic the changes from Charcot neuropathic arthropathy in humans by examining the effects of exposing diet-induced obese (DIO) mice to neurotrauma through a regimented running protocol.

Methods: Forty-eight male wild-type C57BL/6J mice were obtained at age 6 weeks and separated into 2 groups for diet assignment. After a 1-week acclimation period, half of the mice consumed a high-fat diet (60% fat by kcal) ad libitum to facilitate neuropathic diet-induced obesity whereas the other half were control mice and consumed an age-matched standard low-fat control diet (10% fat by kcal). At age 12 weeks, half of the animals from each group were subjected to a high-intensity inclined treadmill running protocol, which has been previously demonstrated to induce neurotrauma. Sensory testing and radiographic analyses were periodically performed. Histopathologic analyses were performed post killing.

Results: DIO mice had significantly higher bodyweights, higher body fat percentages, and lower bone mineral density than wildtype control mice that were fed a normal diet throughout the experiment (P < .001 for each). DIO mice displayed significantly reduced sensory function in week 1 (P = .005) and this worsened over time, requiring 20.6% more force for paw withdrawal by week 10 (P < .001). DIO mice that ran demonstrated greater midfoot subluxation and tarsal instability over all time points compared with normal-diet mice that ran (P < .001). Histopathologic analyses revealed that DIO mice that ran demonstrated significant changes compared with controls that ran (P < .001 for each parameter).

Conclusion: Changes akin to the earliest changes observed in or before joint destruction identified in diabetic Charcot neuropathic arthropathy in humans were observed.

Clinical relevance: There is currently no standard of treatment for patients with Charcot neuropathic arthropathy. This study establishes a protocol for an animal model that can be used to study and compare interventions to treat this disease.

Keywords: Charcot neuroarthropathy; microtrauma; neuropathic mouse; obese animal model; treadmill running.

Figure 1.

Comparison of (A) average weight, (B) bodyfat percentage, and (C) bone mineral density throughout the experiment (n = 12 per group). Error bars indicate standard error of the mean. *, **, and *** indicate significant differences at P < .05, P < .01, and P < .001, respectively.

Figure 2.

Comparison of the force required to elicit a positive response in mouse hind paw using von Frey filament over the 10-week time period (n = 12 per group). Error bars indicate standard error of the mean. ** and *** indicate significant differences at P < .01 and P < .001, respectively.

Figure 3.

Comparisons of (A) subluxation percentage of the tarsal over the metatarsal bones (B) Meary angle, and (C) calcaneus–fifth metatarsal angle between each group over the duration of the study (n = 12 per group). Error bars indicate standard error of the mean. *, **, and *** indicate significant differences at P < .05, P < .01, and P < .001, respectively.

Figure 4.

Graphic representation of histopathologic changes of Charcot neuropathic arthropathy model (n = 12 per group). This includes assessments of nerve intraneural myxoid (edema) change and fibrosis (column 1), small vessel arteriolosclerosis (column 2), articular peculiar perichondral ossification (column 3), subchondral osteosclerosis (microfracture endochondral ossification) (column 4), increased distance from the articular surface to closest marrow space due to osteosclerosis (column 6), and cortex endochondral ossification remodeling (column 5). The level of degenerative changes on this plot correlates to the scores for the respective grading scale for each measurement.

Figure 5.

Representative radiographic images in the sagittal plane of (A) diet-induced obese runner (DIO+R) with Meary angle and calcaneal–fifth metatarsal angle and (B) midfoot subluxation. As a comparison, wildtype control nonrunner (WTC) mouse (C) Meary angle and calcaneal–fifth metatarsal angles, and (D) measured midfoot subluxation.