Search

Bu çalışma, anti-asetilkolin reseptörü (AChR) ve kasa özgü kinaza (MuSK) karşı otoantikor pozitif myastenia gravis (MG) hastalarında regülatör T hücrelerinin (Treg) değerlendirilmesini hedeflemektedir. Treg hücrelerinin süpresyon kapasitesi ve MG patogenezindeki rolü ve 1,25-dihidroksi vitamin D3 (VitD3) ile interlökin 21 (İL-21)'in Treg aracılığıyla meydana gelen baskılamadaki etkisi araştırılmıştır .Çalışmaya 66 AChR-MG, 27 MuSK-MG ve 53 sağlıklı donör (KON) dâhil edildi. Treg hücrelerinde `programmed cell death-1` (PD-1), `PD-1 ligand-1` (PD-L1) `glucocorticoid induced tumor necrosis factor related receptor` (GITR) ve hücre içi `forkhead box P3` (FOXP3) ekspresyonu akım sitometrisi ile analiz edildi. Treg ve CD4+CD25- (standart T hücre, STH) hücreler ayrıldı ve Treg süpresyon yüzdesi rIL-21 ve VitD3'ün varlığı ya da yokluğunda ölçüldü. AChR-MG'de Treg hücrelerin oranında KON'a göre anlamlı artış olmasına rağmen (p= 0,04) FOXP3 ekspresyonu AChR-MG ve MuSK-MG'de KON grubu ile kıyaslandığında düşüktü (p< 0,001 ve p< 0,001). Treg yüzeyinde GITR oranında AChR-MG grubunda azalma gözlemlendi (p< 0,001). AChR-MG (p= 0,02) ve MuSK-MG (p= 0,005) gruplarında kontrol grubuna göre PD-L1 ekspresyonu açısından azalma görülürken, PD-1 ekspresyonu farklı değildi.Treg süpresyon yüzdesi AChR-MG'de KON grubuna göre düşük bulundu (p< 0,001). VitD3 Treg hücrelerin süpresyon gücünü, AChR-MG (p= 0,001) ve KON'da (p< 0,001) anlamlı derecede arttırdı. Ancak VitD3'ün etkisi AChR-MG ve KON arasında anlamlı bir fark oluşturmadı. rİL-21 hem AChR-MG grubunda (p< 0,001), hem kontrol grubunda (p= 0,002) Treg hücrelerin süpresyon gücünü anlamlı derecede baskıladı, bu baskılama MG grubunda daha güçlüydü (p= 0,05). AChR-MG grubunda Treg sayısı artış göstermesine rağmen bu hücrelerde fonksiyon bozukluğu gözlemlendi. Treg fonksiyonunda önemli rol oynayan FOXP3, PD-L1 ve GITR ekspresyonlarında bir düşüş görüldü. AChR-MG'de VitD3'ün Treg süpresyon gücünü arttırması nedeniyle, VitD3 kullanımının AChR-MG hastalarında Treg fonksiyonlarının arttırılmasında yararlı olabileceği düşünülmektedir. İn vitro ortamda İL-21'in Treg süpresyon gücünü azaltması, bu sitokinin Treg üzerine bir rolü olduğunu düşündürmektedir. The purpose of this study is to evaluate peripheral blood regulatory T cells (Tregs) in anti-acetylcholine receptor (AChR) and muscle specific kinase (MuSK) antibody positive myasthenia gravis (MG) and their suppressive capacity with respect to pathogenesis of the disease. In addition, the role of 1,25-dihydroxyvitamin D3 (VitD3) and IL-21 on Tregs suppression were investigated.Tregs from 66 AChR-MG and 27 MuSK-MG patients and 53 healthy controls (HC) were analyzed for programmed cell death-1 (PD-1), PD-1 ligand-1 (PD-L1) glucocorticoid induced tumor necrosis factor related receptor (GITR) and intracellular forkhead box P3 (FOXP3) expression by multicolor flow cytometry. Tregs and CD4+CD25- (conventional T cell, Tconv) cells were sorted and suppression assays were performed with/without IL-21 and VitD3.Although the frequencies of Tregs were increased in AChR-MG patients (p= 0.04), FOXP3 expression in Tregs was significantly decreased both in AChR-MG and MuSK-MG compared to HC (p< 0.001 and p< 0.001). We observed a significant decrease of GITR+ Tregs in AChR-MG (p< 0.001). PD-L1 expression on Tregs was significantly decreased in AChR-MG (p= 0.02) and MuSK-MG (p=0.005) compared to HC, whereas PD-1 expression on Tregs was not different.A strong suppressive effect of Tregs was demonstrated in HC and this suppressive effect was attenuated in AChR-MG (p< 0.001). VitD3 significantly increased the suppressive capacity of Tregs both in AChR-MG (p=0.001) and HC (p< 0.001). However, the increase of suppression was not significantly different between AChR-MG and HC. IL-21 significantly decreased Tregs suppression both in AChR-MG (p< 0.001) and HC (p=0.002) with stronger effect in patients (p= 0.05). A higher number of Tregs in AChR-MG, but a severe functional defect in their regulatory activity was found together with a decreased expression of the FOXP3, PD-L1 and GITR which have a role for T cell regulatory function. Treg-mediated suppression of Tconv was impaired in AChR-MG and the addition of VitD3 significantly increased Tconv suppression by inhibiting Tconv proliferation. The use of VitD3 could be considered for the improvement of Tregs function in AChR-MG. Moreover, a possible regulatory role of IL-21 on suppression of Tregs can be assumed, as IL-21 caused an attenuation of the suppression in vitro. 73

Susceptibility to myasthenia gravis (MG) has been demonstrated with several HLA in different disease subgroups. HLA-DR14, -DR16 and -DQ5 were reported as predisposing factors in muscle-specific kinase antibody positive MG (MuSK-MG). These markers were evaluated in MG subgroups from Turkey. Among 164 generalized MG patients, 116 had antibodies against anti-acetylcholine receptor (AChR-MG) and 48 had MuSK-MG. Only HLA-DRB1 and DQB1 allele groups reported to be associated with MuSK-MG were compared with 250 healthy controls (HC). Highly significant associations of both DRB1*16 and DRB1*14 were found with MuSK-MG compared to HC (p = 1.9 x 10(-5), OR: 4.95 and p = 0.0028, OR: 3.1). On the contrary, HLA-DRB1*03 was less frequent in MuSK-MG (p = 0.006, OR: 0.09). DQB1*05 was also associated with MuSK-MG (p = 2.5 x 10(-6) OR: 4.8). This study provides a replication of the highly significant associations of both HLA-DRB1*16,-DRB1*14 and -DQB1*05 with MuSK-MG. Moreover, HLA-DRB1*03 appears to have a distinguishing role for this disease subgroup compared to early-onset and AChR-MG. (C) 2013 American Society for Histocompatibility and Immunogenetics. Published by Elsevier Inc. All rights reserved.