Musk-ilişkili myasthenia gravis’te antikor ve sitokin profili

Myasthenia gravis (MG) asetilkolin reseptörü (AChR) veya “muscle-specific receptor tyrosine kinase” (MuSK) antikoruyla ilişkili, kas zaafıyla karakterize otoimmün bir hastalıktır. MuSK antikorları ağırlıkla IgG4 izotipindendir. MuSK ilişkili MG (MuSK-MG) gelişimini etkileyen immünglobülin (Ig) ve sitokinleri araştırmak amacıyla MuSK ilişkili deneysel otoimmün MG (MuSK-DOMG) modeli C57BL/6 farelerinde oluşturulmuştur.Rekombinan insan MuSK proteini ve “complete Freund’s adjuvant” (CFA) (MuSK+CFA, n=8) ile immünize farelerin immünopatolojik bulguları sadece CFA ile immünize (CFA, n=8) kontrol farelerle karşılaştırılmıştır. Kas zaafı, klinik skorlama, asma ve motor aktivite testleriyle değerlendirilmiştir. Serum anti-MuSK Ig izotip ve kompleman C3 seviyeleri ELISA ile, nöromüsküler bileşke (NMB)’deki Ig ve kompleman depozitleri immünofloresan boyamayla, kas AChR ve MuSK miktarları immünfloresan boyama ve western blot ile değerlendirilmiştir. Serum ve lenf ganglionu hücrelerinin kültür üst sıvılarında sitokin düzeyleri ELISA ve çoklu boncuk yöntemiyle ölçülmüştür.MuSK+CFA grubunda 7 farede (%87.5) kas zaafı gözlenirken, CFA grubunda yoktu. CFA grubuna göre, MuSK+CFA’da serum anti-MuSK Ig izotip ve C3 seviyeleri, NMB Ig izotip, C3, membran atak kompleksi (MAK) depozit sayıları yüksek, NMB’deki AChR düzeyleri düşükken MuSK benzerdi. IgG1 baskın anti-MuSK Ig izotipiydi. MuSK+CFA grubunda serumda IL-4, kültür üst sıvılarında IL-4, IL-10, IFN-γ düzeyleri artmıştı. MuSK-MG’ye benzer şekilde MuSK-DOMG de komplemanı aktive etmeyen Ig izotipi IgG1 baskındır. NMB’de kompleman depozitlerinin görülmesi MuSK-MG patogenezinde kompleman aracılı membran hasarında rol oynar. IgG1, IL-4, IL-10 düzeylerindeki artış MuSK immünitesinde Th2 tipi yardımcı T lenfositlerini düşündürmektedir.

Anahtar Kelimeler:

Myasthenia gravis, sitokin, MuSK, hayvan modeli

The antibody and cytokine profile in Musk-associated myasthenia gravis

Myasthenia gravis (MG) is an autoimmune disease characterized by muscle weakness associated with acetylcholine receptor (AChR) or muscle-specific receptor tyrosine kinase (MuSK)-antibodies. MuSK-antibodies are predominantly of IgG4 isotype. MuSK associated experimental autoimmune myasthenia gravis (EAMG) model was established to investigate immunoglobulins (Ig) and cytokines related with MuSK associated MG (MuSK-MG).Immunopathological findings of mice immunized with recombinant human MuSK protein and complete Freund’s adjuvant (CFA) (MuSK+CFA, n=8) or only CFA (CFA, n=8) were compared. Muscle weakness was assessed with clinical scoring, inverted screen and motor activity tests. Serum anti-MuSK Ig isotype and complement C3 levels were measured by ELISA, neuromuscular junction (NMJ) Ig, complement deposits, AChR, MuSK levels were measured by immunofluorescence staining and western blot. Levels of cytokines in sera and supernatants of MuSK-stimulated lymph node cells were measured with ELISA and multiplex assays.Seven mice (87.5%) from MuSK+CFA group and no mice from CFA group showed muscle weakness. MuSK+CFA group showed increased anti-MuSK Ig isotype and C3 levels, NMJ Ig isotype, C3 and membrane attack complex (MAC) deposit counts, decreased NMJ AChR and comparable MuSK levels. IgG1 was the dominant anti-MuSK Ig isotype. MuSK+CFA group showed increased IL-4 levels in sera, increased IL-4, IL-10, IFN-γ levels in supernatants. Similar to MuSK-MG patients, non-complement activating IgG1 was the dominant Ig isotype. Nevertheless, NMJ complement deposits suggests that complement-mediated membrane damage might participate in MuSK-MG pathogenesis. Increased IgG1, IL-4, IL-10 levels suggest that Th2-type T-helper cells are involved in MuSK immunity.

Keywords:

Myasthenia gravis, cytokine, MuSK, animal model,

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