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关键词： HLA-B27   machine learning  

摘要： As the association of human leukocyte antigen B27 (HLA-B27) with spondylarthropathies is widely known, HLA-B27 antigen expression is frequently identified using flow cytometric or other techniques. Because of the possibility of cross-reaction with off target antigens, such as HLA-B7, each flow cytometric technique applies a "gray zone" reserved for equivocal findings. Our aim was to use machine learning (ML) methods to classify such equivocal data as positive or negative. Equivocal samples (n = 99) were selected from samples submitted to our institution for clinical evaluation by HLA-B27 antigen testing. Samples were analyzed by flow cytometry and polymerase chain reaction. Features of histograms generated by flow cytometry were used to train and validate ML methods for classification as logistic regression (LR), decision tree (DT), random forest (RF) and light gradient boost method (GBM). All evaluated ML algorithms performed well, with high accuracy, sensitivity, specificity, as well as negative and positive predictive values. Although, gradient boost approaches are proposed as high performance methods;nevertheless, their effectiveness may be lower for smaller sample sizes. On our relatively smaller sample set, the random forest algorithm performed best (AUC: 0.92), but there was no statistically significant difference between the ML algorithms used. AUC values for light GBM, DT, and LR were 0.88, 0.89, 0.89, respectively. Implementing these algorithms into the process of HLA-B27 testing can reduce the number of uncertain, false negative or false positive cases, especially in laboratories where no genetic testing is available.

关键词： Molecular dynamics  

摘要： Interleukin-37b (IL-37b) has emerged as a potential therapeutic agent for inflammatory diseases. However, IL-37b tends to self-assemble into a dimeric form, which compromises its anti-inflammatory efficacy. Furthermore, the currently active monomeric variants of IL-37b are unstable, limiting their clinical utility. To address these challenges, we employed a series of in silico protein engineering strategies to design IL-37b variants with reduced dimerization propensity and enhanced monomeric stability. We identified and analyzed hot-spot residues critical for dimer formation using a double-alanine scanning approach, and subsequently mutated these residues in search of optimal mutations to block self-dimerization. Further, we expanded computational site-saturation mutagenesis to non-binding-site residues and evaluated their impact on monomer stability. Molecular dynamics (MD) simulations and structural analyses were performed to interpret the effects of these variants on stability. In total, we identified 36 mutations that block IL-37b dimer formation and 10 mutations that enhance monomer stability. The mutational effects of the two lists of mutations were independent, enabling potential multi-mutant combinations into dual-functional variants. The systematic in silico design approach presented for IL-37b in this work also provides a generalizable framework for the rational design of other cytokines and proteins.

关键词： autoimmunity   antigen presentation   B cells  

摘要： T cell tolerance to self can prevent self-reactive B cells from mounting effective autoimmune responses by limiting their available help. However, tolerance may be compromised during infection if small amounts of soluble cross-reactive pathogen antigens containing functional T cell epitopes are released for capture by activated B cells. Here, we assess this scenario and show that na & iuml;ve B cells engaged and activated by membrane-bound antigens lacking T helper epitopes are impaired in their ability to capture and present additional soluble antigens containing T helper epitopes. This limits their ability to acquire help from cognate CD4+ T cells required for effective antibody responses. Failure to capture and present soluble antigen is due to IgM and IgD downregulation when engaged with membrane-bound antigen. Interestingly, IgG+ B cells are not similarly constrained and effectively capture soluble antigens. Our findings suggest control of na & iuml;ve B cell tolerance partially depends on efficient receptor downregulation upon antigen engagement.

关键词： NK cells   KIR2DS4   HLA-C   antigen-specificity   peptides   AlphaFold  

摘要： -The ability of NK cells to establish antigen-specific responses has been demonstrated in various infections. NK cell receptors of the diverse family of Killer-cell Immunoglobulin-like Receptors (KIR) interact with HLA class I molecules, and this interaction is peptide-dependent. The activating receptor KIR2DS4 enables NK cell degranulation following interaction with specific peptides presented within HLA-C*05. However, the mechanism underlying the differential NK cell response depending on a peptide remains poorly understood and lacks explanation based on the structure of ligand-receptor interaction. Using AlphaFold 3, we generated models of KIR2DS4-peptide-HLA-C*05 complexes to analyze the contact interfaces. We confirmed the substantial role of the aromatic ring in the 8th amino acid residue of peptide sequences in mediating interactions with KIR2DS4. Even with the same amino acid residue at position 8, different peptides exhibited variability in polar contacts with KIR2DS4. Our results may contribute to the prediction of KIR-HLA interactions and facilitate the identification of specific peptides capable of activating NK cells.

关键词： AT1-AA   Interleukin-6   BK channel   NF-kappa B nuclear translocation  

摘要： The inflammatory phenotype of vascular smooth muscle cells (VSMCs) is an important factor in triggering vascular disease, and interleukin-6 (IL-6) is one of the earliest inflammatory cytokines upregulated in many inflammatory contexts. Angiotensin II-1 receptor autoantibody (AT1-AA) can promote the phenotypic transformation of VSMCs into macrophage-like cells, then synthesize abundant IL-6 to induce vascular inflammation. Previous studies suggested that abnormal BK channel function on the surface of VSMCs played an important role in the synthesis of IL-6, but the mechanism of abnormal BK channel involving in AT1-AA-induced IL-6 synthesis in VSMCs was unclear. In this study, the agonist NS1619 of the BK channel and the inhibitor Paxilline were used to reverse or exacerbate IL-6 synthesis in AT1-AA-induced VSMCs. It is known that NF-kappa B can enter the nucleus due to increased calcium ion concentration caused by BK channel dysfunction, thereby increasing IL-6 transcription. This study observed that Paxilline pretreatment significantly increased the residence time of AT1-AAinduced NF-kappa B in the nucleus, while NS1619 pretreatment showed the opposite trend. JSH-23 inhibiting NF-kappa B nuclear entry reversed the increase in IL-6 expression in VSMCs induced by AT1-AA. This study found that AT1AA enhanced NF-kappa B nuclear translocation by inhibiting BK channel function, which in turn promoted IL-6 transcription in VSMCs, increased IL-6 synthesis and induced vascular inflammation. This study revealed the importance of BK channel dysfunction in the process of AT1-AA increasing IL-6 synthesis and promoting vascular inflammation, and provided a new idea for alleviating vascular inflammatory diseases from the perspective of improving potassium channel function.