Publication: Ankilozan spondilit’in moleküler ve immün patogenezinin incelenmesi
Abstract
Amaç: Ankilozan Spondilit (AS) ile İnsan Lökosit Antijen (HLA)-B27 arasındaki ilişki, artritojenik peptid teorisi ve hücre yüzeyinde sunulan peptid repertuvarının değişiminin tetiklediği süreçler dikkate alınarak, hastalığın immün mekanizmalarının aydınlatılması ve immünojenik/ artritojenik peptid adaylarının belirlenmesi amaçlanmıştır. Gereç ve Yöntem: Bu çalışma kapsamında, hasta ve kontrollerin HLA-B tiplendirilmesi dört basamaklı SeCore SSO XR yöntemi ile, ERAP1 ve ERAP2 haplotipleri ise gerçek zamanlı polimeraz zincir reaksiyonu (RT-PCR) ile belirlenmiştir. in silico veriler ve genotip-fenotip ilişkileri göz önüne alınarak seçilen ikinci basamak çalışma grubunun periferik kan mononükleer ve sinoviyal sıvı mononükleer hücrelerinden elde edilen peptidler, sıvı kromatografi kütle spektrometresi (LC-MS/ MS) yöntemi ile tanımlanmış ve peptidlerin olası artritojenik/ immünojenik potansiyelleri yazılımlar ve in silico çalışmalarla değerlendirilmiştir. Bulgular: ERAP1 ve ERAP2 haplotiplerinin incelendiği popülasyonda, hasta ve kontrol grupları arasındaki dağılımlar farklılık göstermezken haplotip 8, hasta popülasyonunda en sık rastlanan haplotip olarak saptanmıştır. ERAP haplotipleri ve HLA-B alt tipleri bilinen hasta ve kontrollerden oluşturulan immünopeptidom havuzunda, GGQGGAAPV, QVSTHEQ, GSTLGGLL, KSAALLL, NSGPIII, KLAADNK, KRATVAP, TNAAEIP ve QSAALEP peptidleri, bakteri ve fungal dizilerle olan benzerlikleri nedeniyle immünojenik ve artritojenik aday peptidler olarak tanımlanmıştır. Sonuç: Bu çalışmada, antijen işleme ve sunumundaki rolleri göz önüne alındığında, ERAP1 ve ERAP2’deki değişimlerin enzim aktivitesini ve dolayısıyla peptid kesimini etkilediği gösterilmiş; HLA-B27 ile hastalık ilişkisi ve artrit gelişimine katkıda bulunan peptid teorisi temelinde teoriyi destekler nitelikte, hastalıkla ilişkili olabilecek peptidler tanımlanmıştır. Bu peptidler yapılacak ileri analizlerle yeni ilaç hedefleri olarak kullanılabilir.
Objective: This study aims to elucidate the immune mechanisms underlying Ankylosing Spondylitis (AS) by investigating the relationship between Human Leukocyte Antigen (HLA)-B27 and the disease. The focus is on the arthritogenic peptide hypothesis and the processes triggered by changes in the peptide repertoire presented on the cell surface, with the goal of identifying potential immunogenic and arthritogenic peptide candidates. Materials and Methods: HLA-B typing of patients and controls was performed using the SeCore SSO XR method with four-digit resolution, while ERAP1 and ERAP2 haplotypes were determined using real-time polymerase chain reaction (RT-PCR). Peptides isolated from peripheral blood mononuclear cells (PBMCs) and synovial fluid mononuclear cells (SFMCs) from a second-stage study group, selected based on in silico data and genotype-phenotype correlations, were identified using liquid chromatography-mass spectrometry (LC-MS/ MS). The immunogenic and arthritogenic potential of the peptides was evaluated through in silico analyses and bioinformatic software tools. Results: Although the distribution of ERAP1 and ERAP2 haplotypes did not differ between patients and controls, haplotype 8 was the most frequent in the patient population. In the immunopeptidome pool generated from known patients and controls, peptides GGQGGAAPV, QVSTHEQ, GSTLGGLL, KSAALLL, NSGPIII, KLAADNK, KRATVAP, TNAAEIP, and QSAALEP were identified as immunogenic and arthritogenic candidates due to their similarities with bacterial and fungal sequences. Conclusion: This study demonstrates that variations in ERAP1 and ERAP2, considering their roles in antigen processing and presentation, affect enzyme activity and peptide trimming. Peptides potentially associated with the disease were identified, supporting the arthritogenic peptide hypothesis, which underpins the link between HLA-B27 and AS development. These peptides may serve as novel therapeutic targets in future analyses.
Objective: This study aims to elucidate the immune mechanisms underlying Ankylosing Spondylitis (AS) by investigating the relationship between Human Leukocyte Antigen (HLA)-B27 and the disease. The focus is on the arthritogenic peptide hypothesis and the processes triggered by changes in the peptide repertoire presented on the cell surface, with the goal of identifying potential immunogenic and arthritogenic peptide candidates. Materials and Methods: HLA-B typing of patients and controls was performed using the SeCore SSO XR method with four-digit resolution, while ERAP1 and ERAP2 haplotypes were determined using real-time polymerase chain reaction (RT-PCR). Peptides isolated from peripheral blood mononuclear cells (PBMCs) and synovial fluid mononuclear cells (SFMCs) from a second-stage study group, selected based on in silico data and genotype-phenotype correlations, were identified using liquid chromatography-mass spectrometry (LC-MS/ MS). The immunogenic and arthritogenic potential of the peptides was evaluated through in silico analyses and bioinformatic software tools. Results: Although the distribution of ERAP1 and ERAP2 haplotypes did not differ between patients and controls, haplotype 8 was the most frequent in the patient population. In the immunopeptidome pool generated from known patients and controls, peptides GGQGGAAPV, QVSTHEQ, GSTLGGLL, KSAALLL, NSGPIII, KLAADNK, KRATVAP, TNAAEIP, and QSAALEP were identified as immunogenic and arthritogenic candidates due to their similarities with bacterial and fungal sequences. Conclusion: This study demonstrates that variations in ERAP1 and ERAP2, considering their roles in antigen processing and presentation, affect enzyme activity and peptide trimming. Peptides potentially associated with the disease were identified, supporting the arthritogenic peptide hypothesis, which underpins the link between HLA-B27 and AS development. These peptides may serve as novel therapeutic targets in future analyses.