A novel autoantibody recognizing 200-kd and 100-kd proteins is associated with an immune-mediated necrotizing myopathy

Abstract

Objective: Myofiber necrosis without prominent inflammation is a nonspecific finding in patients with dystrophies and toxic or immune-mediated myopathies. However, the etiology of a necrotizing myopathy is often obscure, and the question of which patients would benefit from immunosuppression remains unanswered. The aim of this study was to identify novel autoantibodies in patients with necrotizing myopathy.

Methods: Muscle biopsy specimens and serum samples were available for 225 patients with myopathy. Antibody specificities were determined by performing immunoprecipitations from (35)S-methionine-labeled HeLa cell lysates. Selected biopsy specimens were stained for membrane attack complex, class I major histocompatibility complex (MHC), and endothelial cell marker CD31.

Results: Muscle biopsy specimens from 38 of 225 patients showed predominantly myofiber necrosis. Twelve of these patients had a known autoantibody association with or other etiology for their myopathy. Sixteen of the remaining 26 sera immunoprecipitated 200-kd and 100-kd proteins; this specificity was observed in only 1 of 187 patients without necrotizing myopathy. Patients with the anti-200/100 autoantibody specificity had proximal weakness (100%), high creatine kinase levels (mean maximum 10,333 IU/liter), and an irritable myopathy on electromyography (88%). Sixty-three percent of these patients had been exposed to statins prior to the onset of weakness. All patients responded to immunosuppressive therapy, and many experienced a relapse of weakness when the medication was tapered. Immunohistochemical studies showed membrane attack complex on small blood vessels in 6 of 8 patients and on the surface of non-necrotic myofibers in 4 of 8 patients. Five of 8 patients had abnormal capillary morphology, and 4 of 8 patients expressed class I MHC on the surface of non-necrotic myofibers.

Conclusion: An anti-200/100-kd specificity defines a subgroup of patients with necrotizing myopathy who previously were considered to be autoantibody negative. We propose that these patients have an immune-mediated myopathy that is frequently associated with prior statin use and should be treated with immunosuppressive therapy.

Figure 1. Sera from patients with a necrotizing myopathy immunoprecipitate ~200 and ~100 kDa proteins

Patient sera were used to immunoprecipitate radioactively labeled proteins from HeLa cell extracts which had been incubated with ³⁵S-methionine. Immunoprecipitated proteins were separated by electrophoresis on 10% SDS-polyacrylamide gels. The left and right panels are autoradiographs from 2 separate experiments; data shown on the right panel is from a single autoradiograph which has been cropped between lanes 7 and 8 to exclude immunoprecipitations irrelevant to the current study. The numbers at the top of lanes 1–4 and 6–9 correspond to the patient numbers in Supplemental Table 1. Two different normal control sera (cont 33 and cont 35) were used in the immunoprecipitations shown in lanes 5 and 10. The migration of molecular weight marker standards is shown on the right side.

Figure 2. Abnormal capillary morphology in anti-200/100 muscle biopsies

A normal (A) and an anti-200/100 (B) muscle biopsy specimen were stained with anti-CD31, an endothelial cell marker. Arrows indicate endomysial capillaries with normal morphology in the control specimen (A) and those with thickened walls and dilated lumens in a patient with anti-200/100 autoantibodies (B). These biopsy specimens were processed simultaneously under identical conditions.

Figure 3. Membrane attack complex deposition on small blood vessels and non-necrotic myofibers

Serial sections from an anti-200/100 necrotizing myopathy muscle biopsy (from patient 8076) stained with anti-MAC (A) or H&E (B) demonstrate a perimysial blood vessel with marked complement deposition. Another anti-200/100 muscle specimen (from patient 8024) shows MAC deposition on scattered non-necrotic fibers (C); a higher power detail from the same field is also shown (D; asterisks mark matching myofibers). Note the absence of MAC staining on endomysial capillaries (D; white arrows.)

Figure 4. MHC I deposition on non-necrotic fibers in anti-200/100 biopsies

The endomysial capillaries of normal human muscle stain with anti-MHC I antibodies (arrow), but the sarcolemma do not (A). In contrast, the sarcolemma of scattered muscle fibers in two anti-200/100 patients (B and C; single asterisks) are stained by anti-MHC I. The cytoplasm of an anti-200/100 fiber also stains with anti-MHC I (B; double asterisk); this likely represents a regenerating fiber. These biopsy specimens were processed simultaneously under identical conditions.