Cell stress molecules in the skeletal muscle of GNE myopathy

Abstract

BackgroundMutations of the UDP-N-acetylglucosamine-2-epimerase/N-acetylmannosamine-kinase (GNE)-gene are causally related to GNE myopathy. Yet, underlying pathomechanisms of muscle fibre damage have remained elusive. In sporadic inclusion body myositis (sIBM), the pro-inflammatory cell-stress mediators αB-crystallin and inducible nitric oxide synthase (iNOS) are crucial markers of the disease pathology.Methods10 muscle biopsies from GNE myopathy patients were analyzed for mRNA-expression of markers of cell-stress, inflammation and β-amyloid and compared to non-myopathic controls. Using double-labeling immunohistochemistry, serial sections of skeletal muscle biopsies were stained for amyloid precursor protein (APP), major histocompatibility complex (MHC)-I, αB-crystallin, neural cell adhesion molecule (NCAM), interleukin (IL)-1β, β-amyloid, iNOS, and phosphorylated neurofilament (P-neurofilament) as well as hematoxylin/eosin histochemistry. Corresponding areas of all biopsies with a total of 2,817 muscle fibres were quantitatively assessed for all markers.ResultsmRNA-expression of APP, NCAM, iNOS, TNF-α and TGF-β was higher in GNE myopathy compared to controls, yet this was not statistically significant. The mRNA-expression of APP and αB-crystallin significantly correlated with the expression of several pro-inflammatory and cell-stress-associated markers as NCAM, IL-1β, TGF-β, CCL-3, and CCL4. By immunohistochemistry, αB-crystallin and iNOS were co-upregulated and the number of fibres positive for αB-crystallin, NCAM, MHC-I and iNOS significantly correlated with each other. A large fraction of fibres positive for αB-crystallin were double positive for iNOS and vice-versa. Moreover, several fibres with structural abnormalities were positive for αB-crystallin and iNOS. Notably, particularly normal appearing fibres displayed an overexpression of these molecules.ConclusionsThe cell-stress molecules αB-crystallin and iNOS are overexpressed in GNE myopathy muscle and may identify early disease mechanisms. The data help to better understand the pathology of GNE myopathy.

Amyloid deposits and inflammatory infiltrates in sporadic inclusion body myositis: the inflammatory egg comes before the degenerative chicken

“…But this vacuolar myopathy is essentially not accompanied by muscle inflammation [ 25 ], except in three cases, from two families [ 5 , 54 ]. Nevertheless and despite the general absence of inflammation, iNOS and αB-crystallin, two markers of cell stress, are present in normal appearing fibres of GNE patients and correlated with pro-inflammatory markers (such as IL-6), which remained expressed at levels comparable to control muscles [ 34 ]. Furthermore, mouse models of GNE do not show features of inflammation [ 105 ].…”

Inclusion body myositis – pathomechanism and lessons from genetics

“…The cellular stress molecules αB-crystallin and iNOS are overexpressed in IBM2 muscle and may identify early disease mechanisms. This result helps us to better understand the pathological processes of IBM2 [12]. Sequence analysis of GNE revealed IBM2 individuals were compound heterozygous for a mutation in the 5’ splice donor site of intron 10 (c.1816+5G>A), and a missense mutation (c.2086G>A, p.V696M), confirming the diagnosis as IBM2.…”

Fighting the Cause of Alzheimer’s and GNE Myopathy

“…However, since GNE is a key sialic acid biosynthetic enzyme, mutation in GNE affects the sialylation of proteins ( Noguchi et al, 2004 ). The immunohistochemistry of GNEM muscle samples revealed upregulation of αβ-crystallin, NCAM, MHC-1, and iNOS levels ( Fischer et al, 2013 ). NCAM was hyposialylated in GNEM and proposed as diagnostic marker for GNEM ( Ricci et al, 2006 ).…”

“…Several proteins such as neural cell adhesion molecule (NCAM), α-dystroglycan, integrin, IGF-1R, and other proteins have been found with altered sialylation in absence of functional GNE ( Huizing et al, 2004 ; Ricci et al, 2006 ; Grover and Arya, 2014 ; Singh et al, 2018 ). However, changes in glycosylation pattern of APP or Aβ are not studied in GNEM despite elevated levels of APP reported in ALS and GNEM ( Koistinen et al, 2006 ; Fischer et al, 2013 ). Thus, there is a need to investigate whether hyposialylation of muscle cells, as effect of mutation in GNE, affects the glycosylation pattern and sialylation of accumulated glycoproteins and proteins like Aβ, presenilin-1 etc.…”

“…In GNEM, upregulation of cell stress molecules, such as Aβ oligomers, αβ-crystallin that signals to elevate APP protein was reported ( Fischer et al, 2013 ). Upregulation of iNOS enzyme suggested that cell stress in GNE myopathy is mainly due to NO-related free radicals ( Fischer et al, 2013 ). In GNEM patients and mouse model, proteins were found to be highly modified with S-nitrosylation ( Cho et al, 2017 ).…”