HLA-B27与强直性脊柱炎免疫学机制的研究进展

强直性脊柱炎是与HLA-B27有明确相关性的自身免疫性疾病,其发病过程包括了微生物和宿主的相互作用、免疫细胞对MHC-I类分子的识别以及细胞因子网络的失平衡等方面.未折叠蛋白应答反应参与了强直性脊柱炎发病过程,并且下游IL-23/IL-17轴的激活可能在发病过程中起重要作用.未折叠蛋白反应和IL-23/IL-17轴是研究强直性脊柱炎发病机制和防治措施的新靶点.     

人白细胞抗原HLA-B27在强直性脊柱炎中的临床意义

目的:探讨人白细胞抗原HLA-B27在强直性脊柱炎(AS)中的临床意义,及联合检测HLA-B27、C反应蛋白(CRP)和红细胞沉降率(ESR)的意义。方法:对18例AS患者、24例腰背痛患者和21健康体检者的血液标本进行HLA-B27、CRP、ESR检测,并比较其HLA-B27阳性率。结果:与健康对照组比较,AS组的3项指标均高于对照组(P0.05),且AS组HLA-B27阳性率均高于另外2组(P0.05)。结论:HLA-B27在强直性脊柱炎诊断中具有重要意义,与其具有高度疾病相关性,联合检测HLA-B27及CRP比联合检测HLA-B27及ESR更有利于疾病诊断。     

Characterization of a Proteasome and TAP-independent Presentation of Intracellular Epitopes by HLA-B27 Molecules

Nascent HLA-class I molecules are stabilized by proteasome-derived peptides in the ER and the new complexes proceed to the cell surface through the post-ER vesicles. It has been shown, however, that less stable complexes can exchange peptides in the Trans Golgi Network (TGN). HLA-B27 are the most studied HLA-class I molecules due to their association with Ankylosing Spondylitis (AS). Chimeric proteins driven by TAT of HIV have been exploited by us to deliver viral epitopes, whose cross-presentation by the HLA-B27 molecules was proteasome and TAP-independent and not restricted to Antigen-Presenting Cells (APC). Here, using these chimeric proteins as epitope suppliers, we compared with each other and with the HLA-A2 molecules, the two HLA-B*2705 and B*2709 alleles differing at residue 116 (D116H) and differentially associated with AS. We found that the antigen presentation by the two HLA-B27 molecules was proteasome-, TAP-, and APC-independent whereas the presentation by the HLA-A2 molecules required proteasome, TAP and professional APC. Assuming that such difference could be due to the unpaired, highly reactive Cys-67 distinguishing the HLA-B27 molecules, C67S mutants in HLA-B*2705 and B*2709 and V67C mutant in HLA-A*0201 were also analyzed. The results showed that this mutation did not influence the HLA-A2-restricted antigen presentation while it drastically affected the HLA-B27-restricted presentation with, however, remarkable differences between B*2705 and B*2709. The data, together with the occurrence on the cell surface of unfolded molecules in the case of C67S-B*2705 mutant but not in that of C67S-B*2709 mutant, indicates that Cys-67 has a more critical role in stabilizing the B*2705 rather than the B*2709 complexes.     

Complexity among constituents of the HLA-B*1501 peptide motif

 Analysis of peptides derived from HLA class I molecules indicates that thousands of unique peptides are bound by a single molecular type, and sequence examination of the pooled constituents yields a motif which collectively defines the peptides bound by a given class I molecule. Motifs resulting from pooled sequencing are then used to infer whether particular viral and tumor protein fragments might serve as class I-presented peptide therapeutics. Still undetermined from a pooled motif is the breadth or range of peptides in the population which are brought together to form the pooled motif, and it is therefore not yet known how representative of the population a pooled motif is. By employing hollow fiber bioreactors for large-scale production of HLA class I molecules, sufficient peptides are produced to investigate individual subsets of peptides comprising a motif. Edman sequencing and mass spectrometric analysis of peptides eluted from HLA-B^*1501 reveal that many peptide sequences fail to align with either the N- or C-terminal anchors predicted for the B^*1501 peptide motif through whole pool sequencing. These analyses further reveal auxiliary anchors not previously detected and peptides significantly larger and smaller than the predicted nonamer, ranging from 6 to 12 amino acids in length. These results demonstrate that constituents of the B^*1501 peptide pool vary markedly in comparison with one another and therefore in comparison with previously established B^*1501 motifs, and such complexity indicates that many of the peptide ligands presented to CTL cannot be predicted using class I consensus motifs as search criteria. Received: 7 October 1997 / Revised: 10 December 1997     

Mutational analysis reveals a complex interplay of peptide binding and multiple biological features of HLA-B27

Molecular polymorphism influences the strong association of HLA-B27 with ankylosing spondylitis through an unknown mechanism. Natural subtypes and site-directed mutants were used to analyze the effect of altering the peptide-binding site of this molecule on its stability, interaction with tapasin, folding, and export. The disease-associated subtypes B*2705, B*2702, and B*2704 showed higher thermostability at 50 °C than all other subtypes and mutants, except some mimicking B*2702 polymorphism. The lowest values were found among pocket B mutants, most of which interacted strongly with tapasin, but otherwise there was no correlation between thermostability and tapasin interaction. Mutants resulting in increased hydrophobicity frequently acquired their maximal thermostability faster than those with increased polarity, suggesting that this process is largely driven by the thermodynamics of peptide binding. Folding, export, and tendency to misfold were influenced by polymorphism all along the peptide-binding site and were not specifically dependent on any particular region or structural feature. Frequent uncoupling of thermostability, folding/misfolding, and export can be explained by the distinct effect of mutations on the acquisition of a folded conformation, the optimization rate of B27-peptide complexes, and their quality control in the endoplasmic reticulum, all of which largely depend on the ways in which mutations alter peptide binding, without excluding additional effects on interactions with tapasin or other proteins involved in folding and export. The similarity of the generally disease-associated B*2707 to nondisease-associated subtypes in all the features analyzed suggests that molecular properties other than antigen presentation may not currently explain the relationship between HLA-B27 polymorphism and ankylosing spondylitis.     

HLA-B27分子及其与强直性脊柱炎关联的研究进展简

HLA-B27是MHCI类分子,因而具有MHCI类分子的相关生物学特性。在正常免疫状态下,HLA-B27分子执行对内源性抗原如肿瘤及病毒的免疫监视及杀伤功能;但在机体免疫状态出现异常的情况下,它又可能成为引发自身免疫性疾病的主要相关因素,其中最为著名的是其与强制性脊柱炎（As）的关联研究。我们首先概述了HLA—B27分子正常免疫学功能的研究进展,然后对其在AS相关性研究中的进展进行了总结和分析。通过对上述两方面的综述与分析,全方位展示HLA-B27在执行免疫学功能和引发免疫病理性改变方面的作用,为全面认识和深入研究HLA-B27分子的生理与病理学作用提供坚实的基础。     

HLA-B27: natural function and pathogenic role in spondyloarthritis

The human leukocyte antigen HLA-B27 is strongly associated with development of a group of inflammatory arthritides collectively known as the spondyloarthritides. We have set out to define the natural immunological function of HLA-B27, and then to apply this knowledge to understand its pathogenic role. Human leukocyte antigen class 1 molecules bind antigenic peptides for cell surface presentation to cytotoxic T lymphocytes. HLA-B27 binds and presents peptides from influenza, HIV, Epstein-Barr virus, and other viruses. This leads to vigorous and specific cytotoxic T lymphocyte responses, which play an important role in the body's immune response to these viruses. HLA-B27 thus carries out its natural function highly effectively. Although many theories have been proposed to explain the role of HLA-B27 in the pathogenesis of spondyloarthropathy, we favour those postulating that the pathogenic role of HLA-B27 stems from its natural function. For example, the 'arthritogenic' peptide hypothesis suggests that disease results from the ability of HLA-B27 to bind a unique peptide or a set of antigenic peptides. Additionally, a number of lines of evidence from our laboratory and other laboratories have suggested that HLA-B27 has unusual cell biology. We have recently demonstrated that HLA-B27 is capable of forming disulfide-bonded homodimers. These homodimers are expressed on the cell surface and are ligands for a number of natural killer and related immunoreceptors, expressed on a variety of cell types including natural killer cells, T lymphocytes and B lymphocytes, and members of the monocyte/macrophage lineage. We are currently investigating the possibility that such interactions could be involved in disease pathogenesis.     

Thermodynamic and structural equivalence of two HLA-B27 subtypes complexed with a self-peptide

The F pocket of major histocompatibility complex (in humans HLA) class I molecules accommodates the C terminus of the bound peptide. Residues forming this pocket exhibit considerable polymorphism, and a single difference (Asp116 in HLA-B*2705 and His116 in HLA-B*2709 heavy chains) confers differential association of these two HLA-B27 subtypes to the autoimmune disease ankylosing spondylitis. As peptide presentation by HLA molecules is of central importance for immune responses, we performed thermodynamic (circular dichroism, differential scanning calorimetry, fluorescence polarization) and X-ray crystallographic analyses of both HLA-B27 subtypes complexed with the epidermal growth factor response factor 1-derived self-peptide TIS (RRLPIFSRL) to understand the impact of the Asp116His exchange on peptide display. This peptide is known to be presented in vivo by both subtypes, and as expected for a self-peptide, TIS-reactive cytotoxic T lymphocytes are absent in the respective individuals. The thermodynamic analyses reveal that both HLA-B27:TIS complexes exhibit comparable, relatively high thermostability (Tm approximately 60 degrees C) and undergo multi-step unfolding reactions, with dissociation of the peptide in the first step. As shown by X-ray crystallography, only subtle structural differences between the subtypes were observed regarding the architecture of their F pockets, including the presence of distinct networks of water molecules. However, no consistent structural differences were found between the peptide presentation modes. In contrast to other peptides displayed by the two HLA-subtypes which show either structural or dynamical differences in their peptide presentation modes, the TIS-complexed HLA-B*2705 and HLA-B*2709 subtypes are an example for thermodynamic and structural equivalence, in agreement with functional data.     

Strong memory CD8+ T cell responses against immunodominant and three new subdominant HLA-B27-restricted influenza A CTL epitopes following secondary infection of HLA-B27 transgenic mice

We previously showed that the known HLA-B27-restricted influenza A epitope identified from human studies, NP.383-391, was recognized by CTLs following influenza A infection of transgenic (Tg) HLA-B27/H2 class I-deficient (H2 DKO) mice. Here, we examined the kinetics of the primary NP.383-391-specific response in Tg HLA-B27/H2 DKO mice at the site of respiratory infection, along with the profile of additional influenza A epitopes recognized. While the temporal kinetics of the Tg HLA-B27/NP.383-391-specific CD8+ T cell response paralleled the H2-D(b)/NP.366-374-specific response of non-Tg H2b mice, the magnitude was less. Using epitope prediction programs, we identified three novel B27-restricted influenza A epitopes, PB2.702-710, PB1.571-579, and PB2.368-376, recognized during both the primary and secondary response to infection. Although the secondary NP.383-391-specific response was dominant, PB1.571-579 and PB2.368-376 stimulated stronger proliferative expansion in memory T cells. Our results indicate a broader B27/influenza A CTL repertoire than previously known. Together with results for other HLA class I alleles, this information will become important in improving vaccine strategies for influenza A and other human pathogens.     

TNFalpha blockade prevents the development of inflammatory bowel disease in HLA-B27 transgenic rats

Rats transgenic for HLA-B27 and human β2microglobulin (B27TR) develop a multi-systemic disease resembling inflammatory bowel disease (IBD) and spondyloarthritis. TNFα has a crucial role in chronic inflammation. Our objective was to evaluate the effect of anti-TNFα treatment on spontaneous IBD in B27TR. Nine-week-old B27TR received monoclonal anti-TNFα or an isotypic IgG2a,k up to age of 18 weeks. A second group was monitored up to 18 weeks and then randomly assigned to anti-TNFα or IgG2 a,k treatment. Each rat was monitored for clinical IBD manifestations. After sacrifice, the colon was examined for pathological changes. TNFα receptors (TNF-R1, TNF-R2), Fas/Fas-L expression and apoptosis were evaluated. IgG2a,k-treated and untreated B27TR presented signs of IBD at 11 weeks, whereas in anti-TNFα-treated B27TR no IBD signs were detected. In the late treatment, IBD signs improved after 1 week. Histopathological analysis of IgG2a,k-treated B27TR colon showed inflammatory signs that were widely prevented by early anti-TNFα treatment. Late treatment did not significantly reduce inflammation. TNF-R1 was weakly expressed in intestinal epithelial cells of IgG2a,k-treated B27TR, while it was comparable to controls in anti-TNFα-treated animals. TNF-R2 immunopositivity was strongly evident in IgG2a,k-treated B27TR, whereas was absent in anti-TNFα-treated rats. RT-PCR confirmed these results. IgG2a,k-treated B27TR showed, at 18 weeks, few Fas-positive cells and an increase of Fas-L-positive cells. At 27 weeks, Fas-/Fas-L-positive cell number was significantly low. Anti-TNFα treatment increased Fas-L expression, whereas Fas increased only with the early treatment. TNFα blockade is effective in preventing inflammation in early phase of IBD, maintaining the homeostatic balance of apoptosis.     

Class I and class II MHC bind self peptide sets that are strikingly different in their evolutionary characteristics

 Comparison of peptides eluted from human class I and class II major histocompatibility complex (MHC) molecules and the proteins from which they are derived (source proteins) revealed that class I MHC bind peptides derived from proteins that are highly conserved, hydrophilic, and universally expressed, while the peptides themselves are hydrophobic and even more conserved than their source proteins. In contrast, source proteins for class II-bound peptides were not significantly more conserved than a random sample of proteins. Class II-bound peptides were generally more conserved than their source proteins but were significantly less conserved than class I-bound peptides. The characteristics of class I-bound peptides can probably be explained by the selectivity of processing and transport of peptides for binding by class I, while the relative lack of selectivity of peptide binding for class II may explain the high incidence of autoimmune diseases associated with alleles of these molecules. Received: 17 May 1999 / Revised: 5 August 1999     

Highly conserved antigen-presenting function of CD1d molecules

 The lack of polymorphism of nonclassical antigen-presenting molecules has led to the proposal that they may carry out some conserved and essential antigen-presenting function. Although this is a plausible hypothesis, the major histocompatibility complex has undergone dramatic expansions and contractions through evolution, and there is surprisingly little evidence for interspecies conservation of nonclassical class I molecules. The CD1d molecule, by contrast, shows an extremely high degree of functional conservation between mice and humans, with regard to its interaction with the relatively invariant TCRs that are expressed by NK T cells. This conservation for CD1d recognition is observed either in the absence of exogenous Ag or together with a lipoglycan antigen. The close functional and phenotypic conservation of NK T cells, in mammalian species separated by approximately 50 million years, suggests an essential role in the immune system for CD1d recognition by NK T cells.