The clinical features, muscle pathology, and role of autophagy in anti-Ku-positive patients

Abstract

Aims: This study aimed to examine the clinical and muscle histological characteristics of anti-Ku-positive patients. A preliminary investigation into the involvement of autophagy was conducted as well.

Methods: Clinical characteristics, laboratory findings, and muscle histological features were collected from patients with isolated anti-Ku antibodies at the Department of Neurology, First Medical Center of the PLA General Hospital, between February 2011 to June 2024. Autophagy-related protein levels were semi-quantitatively assessed on muscle tissue samples using western blot (WB), with sporadic inclusion body myositis (sIBM) and immune-mediated necrotizing myopathy (IMNM) patients as comparison groups.

Results: A total of 6 patients were recruited in the study (50% female, mean age at onset 47.6 ± 15.56 years, mean disease duration 7 ± 5.58 months). Extramuscular involvement was observed in most cases, including subcutaneous edema (33.3%), skin rash (33.3%), hyperpigmentation (33.3%), hair loss (33.3%), arthralgia (50%), and interstitial lung disease (ILD) (33.3%), etc. Coexisting connective tissue diseases included systemic sclerosis (SSc) (83.3%), systemic lupus erythematosus (SLE) (16.7%), and arthritis (16.7%). The distribution of muscle weakness was generally symmetrical and proximal (83.3%). Distal (50%) and axial (50%) muscle weakness could also be found. 2 patients exhibited peripheral nerve damage and myogenic damage in EMG, while 4 showed myogenic damage. Creatine kinase (CK) was mildly or moderately elevated. Muscle biopsy demonstrated two patterns: a neurogenic atrophy pattern and a myositis pattern characterized by a varying degree of necrotizing fibers (100%) with rimmed vacuoles (50%) or non-rimmed vacuoles (50%). Immunohistochemical (IHC) analysis revealed sarcolemma deposition of major histocompatibility complex class I (MHC-I) (83.3%) and MHC-II (83.3%), as well as predominant CD68-positive inflammatory infiltrates (66.7%). IHC for p62 revealed a sarcoplasmic punctate pattern (50%), along with a focal coarse staining pattern (50%) and occasional fine granular staining (33.3%). Electron microscopy (EM) demonstrated filamentous and lipid accumulation within vacuoles. WB analysis showed that p62 levels significantly differed between the anti-Ku and IMNM groups. Additionally, Parkin levels were highest in sIBM, while lysosome-associated membrane protein 2 (LAMP2) and microtubule-associated protein 1A/1B-light chain 3 (LC3) expression was highest in the anti-Ku-positive group in tendency.

Conclusion: The muscular features were heterogeneous in anti-Ku-positive patients. A predominant myositis pattern was characterized by necrotizing fibers and vacuolar changes in muscle histology, which differ from sIBM and IMNM. Autophagy appeared to be a key mechanism implicated in the pathogenesis.

Keywords: anti-Ku antibody; autoimmune disease; autophagy; myositis; skeletal muscle pathology.

Figure 1

Myopathological Patterns in Anti-Ku Antibody-Positive Patients: (1) Neurogenic atrophy pattern: HE staining (A) reveals angular and irregular atrophic muscle fibers (A). ATPase staining at pH 4.3 (B) and pH 11.4 (C) shows fiber-type grouping. Immunohistochemistry for MHC-I demonstrates sarcolemma deposition (D). (2) Myositis with vacuoles/rimmed vacuoles pattern: HE staining shows necrosis and degeneration of myofibers, with some fibers showing phagocytosis (thick arrow, E), accompanied by vacuoles and rimmed vacuoles (E–H). Markedly thickened endomysial capillaries is seen in some cases (black circles in G). Lipid deposition was visualized through ORO staining (H). MAC sarcolemma deposition and CD68-positive macrophage endomysial infiltration are identified (I, J). MHC-II expression is upregulated in sarcolemma and cytoplasm of some fibers (K, L) (3) Vacuole features: Several cases exhibit vacuolar pathology with rimmed vacuoles (thin arrow, M) and non-rimmed vacuoles (thick arrow, N). Most non-rimmed vacuoles display a more pronounced peripheral distribution. Immunohistochemical of p62 revealed a focal coarse pattern (thin arrows) and a sarcoplasmic punctate staining pattern (thick arrows) (O, P). Immunostaining for LAMP2 and LC3 exhibited autophagic membrane-bound vacuoles (Q, R). Analysis by EM demonstrated tubulofilamentous inclusion bodies and myelin figures in vacuolar localization (thick arrow, S) and lipid vacuoles enclosed by single-layer membranes (thin arrow, T). Magnification: x200 (A, D, E, F, I, G, H–J, N–R); x100 (B, C, K, L); x400 (M); x6000 (S); x5000 (T). HE, hematoxylin-eosin staining; mGT, modified Gomori trichrome staining; ATPase, adenosine triphosphatase staining; ORO, oil red O staining; MHC-I, major histocompatibility complex class I; MHC-II, major histocompatibility complex class II; MAC, membrane-attack complex; LAMP2, lysosome-associated membrane protein 2; LC3, microtubule-associated protein 1A/1B-light chain 3; EM, Electron microscopy.

Figure 2

(A) Uncropped WB original images. (B) WB was semi-quantified using densitometry analysis by ImageJ, and quantified data were presented as mean ± SD. P62 levels significantly differed between the anti-Ku and IMNM groups. Parkin levels significantly differed between the anti-Ku and sIBM groups. LAMP2 and LC3 expression were highest in the anti-Ku-positive group, in tendency. (* p < 0.05). WB, western blots; LAMP2, Lysosome-associated membrane protein 2; LC3, microtubule-associated protein 1A/1B-light chain 3.