Isolation and characterization of new human carrier peptides from two important vaccine immunogens

In the preparation of glycoconjugate vaccines in clinical practice, two highly immunogenic carrier proteins, CRM₁₉₇ and tetanus toxoid (TT), are predominantly conjugated with the capsular polysaccharides (CPSs) of bacterial pathogens. In addition, TT has long been used as an effective vaccine to prevent tetanus. While these carrier proteins play an important role in immunogenicity and vaccine design alike, their defined human major histocompatibility complex class II (MHCII) T cell epitopes are inadequately characterized. In this current work, we use mass spectrometry to identify the peptides from these carrier proteins that are naturally processed and presented by human B cells via MHCII pathway. The MHCII-presented peptides are screened for their T cell stimulation using primary CD4+ T cells from four healthy adult donors. These combined methods reveal a subset of eleven CD4+ T cell epitopes that proliferate and stimulate human T cells with diverse MHCII allelic repertoire. Six of these peptides stand out as potential immunodominant epitopes by responding in three or more donors. Additionally, we provide evidence of these natural epitopes eliciting more significant T cell responses in donors than previously published TT peptides selected from T cell epitope screening. This study serves toward understanding carrier protein immune responses and thus enables the use of these peptides in developing novel knowledge-based vaccines to combat persisting problems in glycoconjugate vaccine design.     

自体骨髓移植联合MHC单倍体相合淋巴细胞治疗急性髓性白血病的临床疗效

目的:自体骨髓移植联合MHC单倍体相合淋巴细胞治疗急性髓性白血病的治疗效果和安全性。方法:以40例急性髓性白血病患者作为研究对象，随机分为两组各20例，研究组患者使用自体骨髓移植联合MHC单倍体相合淋巴细胞进行治疗，对照组单用自体骨髓移植进行治疗。结果:两组患者的造血系统都得到重建，重建时间以及并发症的发生均没有显著差异。而研究组患者复发率显著降低，且复发时间明显地长于对照组患者。研究组患者3年累积无病生存率明显地高于对照组患者，且差异具有统计学意义。结论:自体骨髓移植联合MHC单倍体相合淋巴细胞治疗能够有效控制急性髓性白血病病情，降低其复发，延长患者生存期。     

Antigen Presentation By Class I Major Histocompatibility Complex Molecules: A Context for Thinking About HLA-G

PROBLEM: There is substantial data that support the efficacy of paternal leukocyte immunization (PLI) for the treatment of alloimmune mediated miscarriage; however, there is confusion regarding the laboratory test that should be performed to determine levels of maternal anti-paternal leukocyte antibodies (MAPLA). METHOD: Popular methodologies employed include: 1) microcytotoxicity (MCX), 2) mixed lymphocyte culture (MLC), and 3) cell flow cytometry crossmatch (FCXM). Cell flow cytometry crossmatch correlates well with the more difficult MLC assay although the former proves the more sensitive study. This work compares the MCX assays with FCXM. The study group consisted of ten women who had a history of three or more spontaneous abortions (SABs). All ten had very low levels (<10%) of MAPLA as measured by FCXM. Following PLI all subjects demonstrated elevated levels (>50%) of MAPLA by FCXM. At 12 weeks gestation, sera were simultaneously measured for MAPLA by MCX and FCXM. RESULTS: Although all ten patients had very high levels of MAPLA by FCXM during pregnancy, five of ten had antibodies to HLA Class I and two of ten had antibodies to HLA Class II paternal antigens by MCX. Furthermore, all patients who were positive by MCX to paternal Class I antigens were also positive to Class I antigens not seen in either parent. Both patients who were positive by MCX to paternal Class II antigens were also positive to maternal Class II antigens. Notable is that all ten women eventually delivered healthy infants. CONCLUSION: Based on this preliminary study, the MCX assay is neither sensitive or reliable enough to determine the need and/or to monitor the effectiveness of PLI. Flow cytometry should be the modality of choice when determining the need for alloimmunotherapy and to monitor the effectiveness of treatment.     

逆转录病毒载体介导的外源性主要组织相容性抗原复合体基因转移及其在T细胞的表达

目的 建立一个在胸腺内表达外源性主要组织相容性抗原复合体(MHC)抗原的实验模型,为下一步研究作准备。方法 应用逆转录病毒载体介导的基因转移技术,首次将外源性MHC基因转移到T淋巴表达并应用聚合酶链反应(PCR)及反转录聚合酶链反应(RT-PCR)检测转染T细胞DNA及mRNA。结果 外源性MHC基因已整合到靶细胞染色体DNA并有效地转录;单克隆抗体免疫荧光染色流式细胞仪检测显示在转染T细胞膜有外源性MHC分子表达;转染效率为46.2％。结论 外源性MHC基因可以转移到T细胞稳定表达,为今后的研究提供了经验。     

Regulation of major histocompatibility complex (MHC) products in fresh and cultured human fetal pancreata aged 9–16 gestational weeks by gamma-interferon

Immunological data on the human fetal pancreas (HFP) are mainly confined to its constitutive expression of the MHC antigens. However, cytokines, such as gamma-interferon (g-IFN), released by lymphocytes during immune reactions, can induce or upregulate the expression of MHC products in allografts and alter their immunological behaviour. We investigated the effects of g-IFN on fresh and cultured HFPs aged 9–16 gestational weeks (gw). Following g-IFN stimulation of fresh HFPs, there was class I hyperexpression by the ductal cells, and some of the ductal, endothelial and islet cells also became class II⁺. Conventional tissue culture (5% CO₂ in air at 37°C) reduced the number of interstitial class II⁺ cells within the HFP after 1 week but was associated with de novo class I expression by some of the ductal cells. Remarkably, the changes in major histocompatibility complex (MHC) antigen expression by the ductal cells occurred earlier and were markedly enhanced when the HFPs were cultured beforehand. The number of interstitial class II⁺ cells in fresh and cultured HFPs was not influenced by g-IFN. The significance of these observations with regard to clinical HFP transplantation is discussed.     

Lesions of T-Cell-Mediated Kidney Allograft Rejection in Mice Do Not Require Perforin or Granzymes A and B

Organ allograft rejection is strongly associated with the presence of alloreactive cytotoxic T cells but the role of cytotoxicity in the pathologic lesions is unclear. Previous studies showed that the principal lesions of kidney rejection - interstitial infiltration, tubulitis, and endothelial arteritis - are T-cell-dependent and antibody-independent. We studied the role of cytotoxic granule components perforin and granzymes A and B in the evolution of the T-cell-mediated lesions of mouse kidney transplant rejection. By real-time RT-PCR, allografts rejecting in wild-type hosts at days 5, 7, 21, and 42 showed massively elevated and persistent expression of perforin and granzymes A and B, but evolution of tubulitis and arteritis did not correlate with increasing granzyme or perforin expression. Allografts transplanted into hosts with disrupted genes for perforin or granzymes A and B showed no change in tubulitis, arteritis, or MHC induction. Thus the development of the histologic lesions diagnostic of T-cell-mediated kidney transplant rejection are associated with but not mediated by perforin or granzyme A or B. Together with previous graft survival studies, these results indicate that the granule-associated cytotoxic mechanisms of T cells are not the effectors of T-cell-mediated allograft rejection.     

Genetic polymorphism of MHC class III and GLO in Chinese Yao nationality

Yao nationality is one of the minority nationalities living mainly in South China (Guangxi Province). The purpose of this study was to provide data of MHC classI and GLO in Chinese Yao nationality and the different genetic background of Yao and Han nationality, the latter representing the major nationality in China. The genetic polymorphism of MHC classI and GLO in Chinese Yao nationality was determined. Previously the Japanese were considered to have the lowest C*₂C frequencies (0.9386), but now we ascertained that the Yao have the lowest C2*C frequencies (0.9336). The data concerning gene frequencies of Yao are presented. They were also compared with the available data of Han.     

Phenotypes and spontaneous cell cytotoxicity of mononuclear cells from patients with seronegative spondyloarthropathies: Ankylosing spondylitis,psoriatic arthropathy and pauciarticular juvenile chronic arthritis — Analysis of mononuclear cells from peripheral blood,synovial fluid and synovial membranes

Summary T-cell subpopulations and natural killer (NK) cells from peripheral blood, synovial fluid and synovial membranes from patients with seronegative spondyloarthropathies were investigated. Thirty-four patients with ankylosing spondylitis, sixteen patients with psoriatic arthropathy and six patients with pauciarticular juvenile chronic arthritis were studied. All the patient groups had normal proportions of T4⁺ and T8⁺ cells as well as normal T4/T8 ratios in peripheral blood. In the synovial fluids the T4/T8 ratios were reduced in ankylosing spondylitis and psoriatic arthropathy (p<0.05). Although both the T4 and T8 subpopulations were reduced, the T4/T8 ratios in the synovial membranes of patients with these two disorders tended to be within the normal range of that of peripheral blood. Increased numbers of T-cells in the synovial fluid from patients with ankylosing spondylitis expressed class II MHC antigens. The natural killer cell activity was normal in peripheral blood and synovial fluids of patients with ankylosing spondylitis and psoriatic arthropathy while it tended to be reduced, although not significantly, in pauciarticular juvenile chronic arthritis. Synovial membranes were almost devoid of NK cell activity. The number of Leu 7⁺ cells were reduced in synovial fluid of patients with psoriatic arthropathy (p<0.04), but not as significantly as in the two other patient groups.