Up-regulation of MHC class I expression accompanies but is not required for spontaneous myopathy in dysferlin-deficient SJL/J mice

Abstract

We found that up-regulation of major histocompatibility complex (MHC) class I expression accompanies, but is not required for, appearance of spontaneous myopathy in SJL/J mice. In some neuromuscular diseases, MHC class I expression is markedly up-regulated in muscles, though the consequences of this up-regulation for pathology are not clear. To study MHC class I in myopathy, we compared muscles of SJL/J mice to muscles of SJL/J mice that were also MHC class I-deficient due to targeted mutation in the beta-2-microglobulin gene (SJL/J B2m (-/-) mice). SJL/J mice show spontaneous myopathy and have a mutation in the dysferlin gene, a gene which is also mutated in human limb-girdle muscular dystrophy type 2B (LGMD2B). Muscles of eight-month-old SJL/J mice had higher levels of MHC class I expression than muscles of either C57BL/6J (wild-type) or SJL/J B2m (-/-) mice. In contrast, the percentage of abnormal muscle fibers was similar in SJL/J and SJL/J B2m (-/-) muscles. Invading Mac-1(+) cells were most abundant in SJL/J B2m (-/-) muscles, moderately abundant in SJL/J muscles, and rare in C57BL/6J muscles. Thus, MHC class I was markedly up-regulated in SJL/J muscles, but this high level of MHC class I was not necessary for the appearance of myopathy.

Figure 1.

A–C: MHC class I expression is up-regulated in muscles of SJL/J mice compared to muscles of SJL/J B2m (−/−) and C57BL/6J mice. A: Immunohistology of C57BL/6J gastrocnemius muscle showed little staining, a result consistent with the very low expression of MHC class I proteins on normal muscle fibers. B: In contrast, SJL/J muscle showed dark staining apparently on the surface of muscle fibers (eg, arrow), as well as staining on cells in between, and sometimes within, muscle fibers. C: In contrast, SJL/J B2m (−/−) muscle showed very little staining, a result expected for tissues lacking β-2-microglobulin. Scale bar, 40 μm. D–F: Muscles from SJL/J and SJL/J B2m (−/−) mice showed similar extents of myopathy. D: H&E staining showed normal fibers in wild-type C57BL/6J gastrocnemius muscle. E and F: In contrast, abnormal fibers were abundant and found at similar percentages in SJL/J (E) and SJL/J B2 m (−/−) (F) muscles as described in Table 1 ▶ . G and H: Mac-1-positive cells were most abundant in myopathic muscles of SJL/J B2m (−/−) mice, moderately abundant in SJL/muscles, and rare in the normal muscles of C57BL/6J mice. G: Immunohistochemistry showed that Mac-1-postive cells (arrow) were rare in the gastrocnemius muscle of C57BL/6J wild-type mice. H and I: In contrast, Mac-1-positive cells were abundant in SJL/J (H) and SJL/J B2m (−/−) (I) muscles, being most abundant in the B2m (−/−) muscles as described in Table 2 ▶ .

Figure 2.

β-2-microglobulin levels were up-regulated in SJL/J muscle. Hindlimb muscles from mice of the indicated genotypes were analyzed by immunoblotting with antibody specific for β-2-microglobulin (Upper panel, B2m). Spleens from C57BL/6J and SJL/J mice had about equal levels of β-2-microglobulin. In contrast, β-2-microglobulin was much more abundant in SJL/J muscle than in C57BL/6J muscle, and was absent from SJL/J B2m (−/−) muscle, a result that was consistent with the MHC class I immunohistochemistry analyses in Figure 1 ▶ . Each lane had 60 μg of cell lysate protein. The loading controls (Lower panel, Load) for the spleen samples show Ponceau S staining of an abundant ∼13-kd protein band, whereas loading controls for the muscle samples show immunoblotting for GAPDH.