Alzheimer’s Mouse Model Shows Sex-Specific Muscle Fatigue Changes Before Cognitive Decline

In a finding that could reshape how scientists think about the body-brain connection in Alzheimer’s disease, researchers at Brock University have reported that a knock-in mouse model carrying a humanized amyloid beta sequence shows strikingly different skeletal muscle fatigue properties in males and females at 8 months of age. The fast-twitch extensor digitorum longus (EDL) muscle of 8-month-old male hAβ-KI mice fatigued faster during repeated stimulation than that of wild-type littermates, while both the EDL and the slow-twitch soleus of 8-month-old female hAβ-KI mice fatigued slower than controls. Remarkably, none of these fatigue differences were present at 15 months of age, and all of them emerged before the cognitive deficits that have previously been documented in this model at 10 and 14 months. The study, published as a preliminary characterization in a peer-reviewed journal, offers one of the first looks at peripheral muscle function in a model designed to reflect sporadic Alzheimer’s disease.

The hAβ-KI mouse was developed to address a persistent translatability problem in Alzheimer’s research. Familial Alzheimer’s disease accounts for less than 5 percent of all diagnoses and stems from autosomal dominant mutations, yet many widely used preclinical models rely on exactly such mutant genetic manipulations. The new model instead inserts a humanized amyloid beta peptide-coding sequence into the mouse amyloid precursor protein (App) gene without any familial mutation. It exhibits age-dependent memory changes and increased amyloid beta production beginning around 10 to 14 months of age. Because it carries no familial genetic manipulation, researchers regard it as a novel model of sporadic Alzheimer’s disease, the form that makes up the remaining 95 percent of diagnoses and develops after age 65 in association with risk factors including age, sex, diet, and physical inactivity. While the cognitive effects of the hAβ-KI mouse were well characterized upon its generation, its peripheral effects, particularly on skeletal muscle, had never been explored.

The motivation for examining muscle comes from a growing body of evidence that skeletal muscle may act as a modulator of brain health. Age-related muscle loss, or sarcopenia, begins as early as the third decade of life, progressing at roughly 3 to 5 percent per decade and accelerating after age 60. That timeline overlaps substantially with the preclinical phase of Alzheimer’s disease, which can span 10 to 20 years before clinical diagnosis and often begins around midlife. Sarcopenia has been linked to cardiovascular disease, type 2 diabetes, and Alzheimer’s disease itself. The prevalence of sarcopenia is significantly higher in Alzheimer’s patients compared to non-Alzheimer’s controls, and older adults diagnosed with sarcopenia are more likely to develop cognitive impairments, including poorer executive function, calculations, and recall, with performance declining as sarcopenia worsens. Preclinical work has echoed these relationships: hindlimb casting that induced muscle atrophy worsened memory performance in young, presymptomatic 5×FAD mice, while aged triple transgenic Alzheimer’s female mice showed lower muscle mass and poorer neuromuscular junction function than wild-type counterparts.

To probe muscle form and function, the team studied wild-type and hAβ-KI mice at 8 and 15 months of age, with 10 mice per sex per age cohort. From 4 months onward, lean and fat mass percentages were tracked every 2 months using dual X-ray absorptiometry (DXA). At the terminal ages, the soleus and EDL muscles were isolated for contractility testing on an Aurora Scientific apparatus. Muscles underwent an isometric force-frequency protocol consisting of 1, 20, 40, 80, and 160 Hz stimulations to obtain peak isometric force, which was normalized to calculated cross-sectional area. Two minutes later, a fatigue protocol delivered 70 Hz volleys every 2 seconds for 3 minutes, with data expressed as percent of initial force and analyzed via area-under-the-curve. Fiber type composition was assessed by immunofluorescent staining of myosin heavy chain isoforms, and RNA sequencing of the EDL identified differentially expressed genes using thresholds of greater than 50 percent expression change and p less than 0.05.

The body composition data revealed a clear sex difference. Both male and female mice of both genotypes showed the expected age-related pattern of higher body mass, lower lean mass percentage, and higher fat mass percentage between 4 and 15 months. However, male hAβ-KI mice exhibited larger gains in body mass, 11.7 grams compared with 9.08 grams in wild-type males, alongside a significantly greater loss in lean mass percentage, 5.71 percent versus 2.88 percent. No significant genotype differences emerged in the females, who lost roughly 8 percent lean mass regardless of genotype. Food intake, measured weekly and summed across the study, showed no significant differences between genotypes, meaning the altered body composition in males could not be explained by eating more. The authors suggest this pattern points toward a potential sarcopenic phenotype in male hAβ-KI mice, consistent with clinical observations that accelerated lean mass loss occurs during early stages of Alzheimer’s disease in older adults, though they note that in vivo muscle strength should be investigated in future studies.

The isolated muscle contractility results were the study’s centerpiece. In 8-month-old males, EDL muscle mass, cross-sectional area, and both absolute and specific force production were indistinguishable between genotypes. During the fatigue protocol, however, the male hAβ-KI EDL showed a leftward shift in the percent of force produced over time, indicative of greater fatiguability, supported by a significant time-by-genotype interaction, although the area-under-the-curve and time to 60 percent rundown did not reach statistical significance. In 8-month-old females, the opposite occurred: the female hAβ-KI EDL exhibited a rightward shift in the force-time curve, indicative of less fatiguability than wild-type controls, again supported by a significant time-by-genotype interaction. Isolated EDL mass in the females was lower in hAβ-KI mice but did not reach significance at p equal to 0.063. At 15 months, the fatigue differences vanished in both sexes, and force production and fatiguability no longer differed, although in the 15-month-old males the specific EDL mass relative to body mass was significantly smaller in hAβ-KI mice than in wild-type mice, even though isolated cross-sectional area did not differ. Fiber typing showed no changes in the percent of glycolytic or oxidative fibers, and no differences in fiber type-specific cross-sectional area, at either age in either sex, ruling out a fiber type shift as the explanation for the fatigue findings.

The soleus, a primarily slow-twitch muscle offering greater fatigue resistance, told a complementary story. In 8-month-old males, the soleus was entirely unaffected by genotype, with no differences in mass, force production, fatiguability, or fiber composition at either age examined. In 8-month-old females, however, the hAβ-KI soleus produced significantly less absolute force than wild-type controls, with significant differences at 80 and 160 Hz stimulation, although when force was made relative to cross-sectional area, specific force production no longer differed. During the fatigue protocol, the 8-month-old female hAβ-KI soleus again showed a rightward shift in the force-time curve, indicating less fatiguability, supported by a significant time-by-genotype interaction. As with the EDL, fiber type composition and fiber type-specific cross-sectional area were unchanged. By 15 months, all soleus differences in the females had disappeared. Taken together, the results suggest that the EDL of male hAβ-KI mice fatigues faster at 8 months, before cognitive deficit onset, while both muscles of 8-month-old females are protected against fatigue, protection that appears lost by 15 months.

RNA sequencing of the 8-month-old EDL offered potential mechanistic insight into these sex-specific functional differences. In males, 536 differentially expressed genes were identified between hAβ-KI and wild-type comparisons, comprising 200 upregulated and 336 downregulated genes. Gene ontology enrichment analysis showed significant downregulation of pathways related to actin binding, actomyosin, myosin filament, musculoskeletal movement, muscle contraction, and muscle development. Actin binding is an essential component of the sliding filament theory, in which the myosin head binds actin to form the crossbridge and initiate the power stroke, and the actin-myosin interaction has been suggested to be a major contributor to muscle fatigue. In females, 437 differentially expressed genes were identified, with 218 upregulated and 219 downregulated. Female hAβ-KI mice showed no downregulation of muscle development or contraction pathways and instead displayed significant upregulation of pathways relating to aerobic respiration, proton motive force-driven ATP synthesis, oxidative phosphorylation, and cellular respiration. Only 86 genes overlapped between the male and female comparisons, underscoring how divergent the transcriptional responses were.

The transcriptomic patterns align neatly with the functional observations. Slow-twitch muscle fibers have a higher aerobic capacity and rely more heavily on aerobic respiration and oxidative phosphorylation than fast-twitch fibers, making them more fatigue-resistant. The upregulation of aerobic respiration genes in the female hAβ-KI EDL could contribute to enhanced fatigue resistance even though myosin heavy chain-based fiber typing showed no change, which the authors interpret as suggestive of underlying metabolic adaptations potentially paired with a fast-to-slow fiber type shift that dedicated studies would need to confirm. Conversely, the downregulation of contraction-related genes in males, including actin binding and actomyosin pathways, is likely to contribute to the faster fatiguability observed in the male EDL, though directed studies are required to test this hypothesis. The authors also raise the possibility that male hAβ-KI mice express an early-onset aging phenotype, given that healthy male wild-type mice have previously been shown to become more susceptible to EDL fatigue at an earlier age than females.

Important caveats frame the interpretation. Rodents express a much higher percentage of fast-twitch fibers than humans, so translation to human populations should be made with caution. Previous research has documented denervated neuromuscular junctions in old male 3×TgAD mice and impairments to peripheral nerve activity in relatively young 5×FAD mice, so future studies should examine peripheral nerve conductance and neuromuscular junction integrity in the hAβ-KI model. The RNA sequencing used a relatively small sample of three mice per group and requires dedicated follow-up analysis. The authors also note that sporadic Alzheimer’s disease is influenced by interacting risk factors including diet and biological sex, particularly around menopause onset. Female mice do not undergo menopause, which may partially explain the absence of a female-specific body composition phenotype, and high-fat diets have been shown to exacerbate both muscular and cognitive impairments in rodent models. Introducing external metabolic or hormone stressors might unmask more pronounced phenotypes. Why the 8-month-old mice, but not the 15-month-old mice, showed functional alterations also remains open, whether these early changes are transient or whether age-related declines in both genotypes mask genotype-specific effects at the later age. Still, the study highlights a potential temporal relationship between muscle function and cognitive performance, since the fatigue alterations emerged before the previously reported cognitive deficits, and future work should investigate how muscle fatigue and cognition relate in this increasingly translatable model of sporadic Alzheimer’s disease.

Subject of Research: Sex-specific skeletal muscle fatigue resistance in a sporadic Alzheimer's disease mouse model

Article Title: Skeletal muscle fatigue resistance is differentially affected in 8‐month‐old male and female hAβ‐KI mice

Article References: Copeland, E. N., Melekh, E., Fajardo, V. A., & MacPherson, R. E. K. (2026). Skeletal muscle fatigue resistance is differentially affected in 8‐month‐old male and female hAβ ‐ KI mice. Physiological Reports, 14(20), Article e71153. https://doi.org/10.14814/phy2.71153

Image Credits: AI Generated

DOI: 10.14814/phy2.71153

Keywords: Alzheimer's disease, skeletal muscle, hAβ-KI mouse model, sporadic Alzheimer's disease, muscle fatigue, amyloid beta, sarcopenia, EDL muscle, soleus, RNA sequencing, oxidative phosphorylation, sex differences