The Gne−/−•hGNED176V-Tg mice can be littered in Mendelian rate and no apparent phenotype at birth. The cellular sialic acid levels in various organs other than the brain were remarkably reduced. The weights of the GNE myopathy mice were lighter than control littermates, and this difference in weight becomes more remarkable with age [22]. The motor performance and exercise endurance of the model mice were worse than their littermates, and the impairment of the motor performance became evident from the age of 21 weeks and displayed remarkable disease progression after 31 weeks of age.

In isolated gastrocnemius muscles, peak isometric twitch and maximum tetanic force showed gradual decrements with age: decrease to 90 % by 10 weeks, 80 % by 21 weeks, 70 % by 31 weeks, and 50 % by 41 weeks. The tetanic force was markedly reduced after 41 weeks of age, so the twitch-tetanic ratio is significantly higher in GNE myopathy mice compared with control mice. The gastrocnemius muscles of the GNE myopathy mice were smaller as compared with those of control mice, even in younger age. Furthermore, the muscles demonstrated a steadily decreasing muscle mass of at least 10–20 % from 31 weeks of age and decreasing more remarkably from 41 weeks of age. Cross-sectional area-normalized twitch and tetanic forces showed similar temporal pattern of force reduction in GNE myopathy mice, but interestingly significant differences between GNE myopathy and control mice were only seen from the age of 31 weeks, indicating the only atrophy but not muscle degeneration attributes to muscle weakness in younger age by 30 weeks, and symptomatic muscle degeneration, which would cause the remarkable reduction of tetanic force, begins from 31 weeks of age and results in the severe muscle weakness after 41 weeks.

No remarkable change in morphological appearance on light microscopy was seen in the muscles from the GNE myopathy mice from 10 to 20 weeks of age, except for minimal variation in fiber size in the gastrocnemius, TA, and QF muscles. The number of small size fibers increased from 21 to 30 weeks of age in gastrocnemius and TA muscles, which contribute to the increasing variation in fiber size. From 31 to 40 weeks of age, a few intracellular inclusions, which were Congo red positive and immunoreactive to various antibodies to amyloid, were observed. The appearance of RVs delayed from 42 weeks (Fig. 4.4). In electron microscopy, the protein inclusions were observed between the bundles of myofibrils in 30 weeks of age, and both the protein inclusions and autophagic vacuoles were accumulated in the center of myofibers apart from the myofibrils after 41 weeks as observed in human GNE patients’ muscles. This accumulation would be results of active cellular response to intracellular inclusion proteins, gathering them into central region by intracellular transport system and inducing macroautophagy process for proteolysis of them.

Fiber-type selective atrophy was not observed at least in gastrocnemius, TA, and QF muscle. However, surprisingly, almost all fibers that had either RVs or intracellular depositions were type II fibers. In other murine models of muscular dystrophy and myopathies, the predominant involvement of type II muscle fibers [23] has been estimated to be due to the increased susceptibility of these muscles to eccentric contraction-induced damage [24, 25]. In the GNE myopathy mice, this is interesting since the mechanism underlying the disease is remarkably different from these other types of murine myopathy models. Interestingly, in a transgenic mouse overexpressing β-amyloid precursor protein (β-APP), intracellular amyloid deposition has been reported predominantly in type II fibers [26]. This could suggest that the involvement of type II fibers may be secondary to susceptibility to oxidative stress and/or calcium dysregulation attributed to type II fibers. Moreover, in a mouse which overexpresses β-APP in type II fibers, an increase in resting calcium and relative membrane depolarization in muscle fibers have been observed and are thought to represent a mechanism relating β-APP mismetabolism to altered calcium homeostasis and clinical weakness [27]. Because intracellular amyloid depositions are key events in muscle degeneration of GNE myopathy mice, these findings may be of interest for future investigations.

ManNAc was effective in preventing the myopathic symptoms of the GNE myopathy mice, equally with low dose (20 mg per kg bodyweight per day), medium dose (200 mg per kg bodyweight per day), and high dose (2000 mg per kg bodyweight per day) [28]. The serum creatine kinase concentration was lowered and motor performance, body weight, and muscle mass were increased after treatment. The number of rimmed vacuoles was significantly lower than the mice who received control treatment. Treatment also increased muscle cross-sectional area and force production and substantially diminished congophilic, amyloid-positive, or tau-positive inclusions. There was no apparent dose effect of ManNAc supplementation in preventing the myopathic symptoms of the GNE myopathy mice. NeuAc and sialyllactose were also effective for the prevention of disease with lower dose at 20 mg per kg bodyweight per day.

To aim for the enhancement of cellular uptake of the administered sialic acid metabolites in mice, we used peracetylated ManNAc for prophylactic treatment of the GNE myopathy mice [29]. Peracetylated ManNAc showed similar recovery of cellular sialylation at less than 1/10 concentration of regular ManNAc in culture medium. As expected, cellular sialic acid levels were highly increased to control levels in all organs including serum, and all aspects in muscle phenotypes, such as muscle power, size, and pathology, were well recovered to those in control mice.