Longitudinal study revealing motor, cognitive and behavioral decline in a transgenic minipig model of Huntington's disease

Abstract

Huntington's disease (HD) is an inherited devastating neurodegenerative disease with no known cure to date. Several therapeutic treatments for HD are in development, but their safety, tolerability and efficacy need to be tested before translation to bedside. The monogenetic nature of this disorder has enabled the generation of transgenic animal models carrying a mutant huntingtin (mHTT) gene causing HD. A large animal model reflecting disease progression in humans would be beneficial for testing the potential therapeutic approaches. Progression of the motor, cognitive and behavioral phenotype was monitored in transgenic Huntington's disease minipigs (TgHD) expressing the N-terminal part of human mHTT. New tests were established to investigate physical activity by telemetry, and to explore the stress-induced behavioral and cognitive changes in minipigs. The longitudinal study revealed significant differences between 6- to 8-year-old TgHD animals and their wild-type (WT) controls in a majority of the tests. The telemetric study showed increased physical activity of 4.6- to 6.5-year-old TgHD boars compared to their WT counterparts during the lunch period as well as in the afternoon. Our phenotypic study indicates progression in adult TgHD minipigs and therefore this model could be suitable for longstanding preclinical studies of HD.This article has an associated First Person interview with the first author of the paper.

Keywords: Cognitive and behavioral studies; Huntington's disease; Large animal model; Motor; Phenotyping.

Fig. 1.

Motor impairment. (A) Walking test. Impairment in gait was observed in TgHD animals in comparison to their WT controls. The difference was significant in 6- to 7.9-year-old TgHD boars (P=0.013) and 4- to 5.9-year-old TgHD sows (P=0.015). The decline was more obvious in TgHD boars than in TgHD sows. Clear sex-related differences were noticed between TgHD boars and TgHD sows of matching ages (P=0.003 and P=0.024, respectively). WT boars 4-5.9 years n=4, WT boars 6-7.9 years n=6, TgHD boars 4-5.9 years n=6, TgHD boars 6-7.9 years n=5, WT sows 4-5.9 years n=8, WT sows 6-7.9 years n=9, TgHD sows 4-5.9 years n=8, TgHD sows 6-7.9 years n=8. (B) Hurdle test. The ability to cross a barrier declined in TgHD boars with age. TgHD sows obtained a worse score at the age of 4-5.9 years, but a better score at the age of 6-7.9 years, in comparison to their age-matched WT controls. Scores were lower in TgHD boars than in TgHD sows at the age of 6-7.9 years, but this difference was not significant. WT boars 4-5.9 years n=3, WT boars 6-7.9 years n=6, TgHD boars 4-5.9 years n=6, TgHD boars 6-7.9 years n=5, WT sows 4-5.9 years n=8, WT sows 6-7.9 years n=5, TgHD sows 4-5.9 years n=8, TgHD sows 6-7.9 years n=5. *P≤0.05, **P≤0.01.

Fig. 2.

Tongue protrusion persistence. (A) In the number-of-reached-treats test, a significantly (P=0.004) lower score was observed in TgHD boars than in WT boars. (B) The deepest-hole test revealed a lower score in TgHD boars, and a similar score in TgHD sows, in comparison to their age-matched WT controls. WT boars 6-7.9 years n=5, TgHD boars 6-7.9 years n=6, WT sows 6-7.9 years n=10, TgHD sows 6-7.9 years n=7. *P≤0.05, **P≤0.01.

Fig. 3.

Cognitive changes. (A) In the skittles test, a significantly (P=0.03) decreased score was detected in TgHD boars compared to WT boars at the age of 6-7.9 years, but scores were similar among all sows. WT boars 4-5.9 years n=2, WT boars 6-7.9 years n=6, TgHD boars 4-5.9 years n=5, TgHD boars 6-7.9 years n=5, WT sows 4-5.9 years n=6, WT sows 6-7.9 years n=9, TgHD sows 4-5.9 years n=6, TgHD sows 6-7.9 years n=7. (B) In the cover pan test, scores were lower in 6- to 7.9-year-old TgHD boars than in their WT controls, but this difference was not statistically significant. Impairment in the ability to perform the test with age was observed. WT boars 4-5.9 years n=2, WT boars 6-7.9 years n=5, TgHD boars 4-5.9 years n=5, TgHD boars 6-7.9 years n=3, WT sows 4-5.9 years n=6, WT sows 6-7.9 years n=9, TgHD sows 4-5.9 years n=6, TgHD sows 6-7.9 years n=7. *P≤0.05, **P≤0.01.

Fig. 4.

Decline in stress-induced performance. (A) In the balance beam test, significant (P=0.033) deterioration was observed in TgHD boars compared to their WT controls at the age of 6-7.9 years. Both TgHD and WT sows had lower scores than boars at older age (P=0.012 for TgHD at 4-5.9 years; P=0.021 for WT at 6-7.9 years). The ability to pass the assessment decreased with age in all TgHD animals. WT boars 4-5.9 years n=4, WT boars 6-7.9 years n=7, TgHD boars 4-5.9 years n=6, TgHD boars 6-7.9 years n=7, WT sows 4-5.9 years n=9, WT sows 6-7.9 years n=9, TgHD sows 4-5.9 years n=8, TgHD sows 6-7.9 years n=8. (B) In the seesaw test, TgHD boars obtained a significantly (P=0.004) lower score than their WT controls at 6-7.9 years. The attained score decreased with age in all TgHD animals, significantly (P=0.024) in TgHD sows. WT boars 4-5.9 years n=2, WT boars 6-7.9 years n=6, TgHD boars 4-5.9 years n=4, TgHD boars 6-7.9 years n=5, WT sows 4-5.9 years n=5, WT sows 6-7.9 years n=9, TgHD sows 4-5.9 years n=4, TgHD sows 6-7.9 years n=7. *P≤0.05, **P≤0.01.

Fig. 5.

Altered physical activity. (A) In TgHD animals, physical activity was lower in the morning than in other periods. (B) During the lunch period, 2.6- to 4.5-year-old TgHD animals had comparable physical activity to that of their WT controls. The activity of 4.6- to 6.5-year-old TgHD animals was significantly (P=0.026) higher than that of the WT controls. Activity significantly (P=0.003) increased with age in TgHD animals. (C) In the afternoon period, 2.6- to 4.5-year-old TgHD animals had decreased physical activity, whereas 4.6- to 6.5-year-old TgHD animals had increased physical activity, in comparison to that of their WT controls, although these differences were not statistically significant. Significantly increased (P=0.002) physical activity was observed with aging in TgHD animals. WT boars 2.6-4.5 years n=7, WT boars 4.6-6.5 years n=17, TgHD boars 2.6-4.5 years n=10, TgHD boars 4.6-6.5 years n=14. *P≤0.05, **P≤0.01.

Fig. 6.

Equipment for motor, cognitive and behavioral studies. (A) Hurdle. (B) Tongue board. (C) Skittles pan. (D) Cover pan. (E) Balance beam. (F) Seesaw.