胃癌组织HLA－I类和DR分子免疫组化研究

本文应用免疫组化法对６４例胃癌、癌旁组织和６例胃溃疡大致正常胃粘膜冰冻和石蜡切片进行了染色．结果表明，正常胃粘膜和癌旁胃粘膜上皮细胞ＨＬＡ－Ｉ类分子表达阳性，其着色较均一，ＨＬＡ－ＤＲ染色均阴性．胃癌细胞Ｉ类分子表达缺失（２７／６４例），与癌旁上皮比较差异显著（Ｐ＜０．０１）。粘液细胞癌和低分化癌Ｉ类分子缺失率显著高于高分化癌（Ｐ＜０．０２５）．此外，发生肿瘤转移的病例Ｉ类分子缺失率（１２／１５例）显著高于无转移组（１／５例，Ｐ＜０．０２５）．ＤＲ分子在癌组织表达阳性，其阳性率高达５３．１％（３４／６４例）．低分化癌ＤＲ分子阳性率亦显著高于高分化癌和中分化癌，未分化癌ＤＲ分子阳性率亦显著高于高分化癌（Ｐ＜０．０１～０．０５）．提示（１）ＨＬＡ－Ｉ类分子表达缺失可能与癌细胞逃避宿主免疫监视发生润浸生长和转移有关；（２）分化程度不同的癌组织ＨＬＡ－Ｉ类分子表达差异显著，提示癌细胞分化可能影响Ｉ、Ⅱ类分子表达和肿癌抗原呈递；（３）ＨＬＡ－Ｉ类和ＤＲ分子表达异常可能是上皮恶性转变的标志之一．     

Pretransplant Exposure to Donor HLA-DR Antigen in Random Transfusion Units and the Development of Donor Antigen-Specific Hyporeactivity

Our previous studies have shown that the in vitro assay of donor antigen-specific hyporeactivity is a useful marker for identifying solid organ transplant recipients (kidney, lung and heart) at low risk for immunologic complications (i.e., late acute rejection episodes and chronic rejection). Donor antigen-specific hyporeactivity is defined as a significantly decreased post- vs. pretransplant proliferative response to donor antigens while response to third-party controls remains unchanged. We analyzed whether exposure to the same HLA-DR antigen pretransplant via random blood transfusion and posttransplant via the transplanted organ influenced the development of hyporeactivity. Thirty previously nontransfused recipients, each receiving two 150 ml pretransplant random blood transfusions, were assessed for hyporeactivity at 1 year posttransplant. Of the 12 recipients with pretransplant exposure to kidney HLA-DR via transfusions, 6 (50%) developed hyporesponsiveness; in contrast, of the 18 recipients who were not preexposed, only 3 (15%) exhibited this form of immunomodulation. Of interest, 2 of the 3 hyporesponsive recipients who were not preexposed, received units containing HLA-DR antigens previously shown to share crossreactive epitopes with the kidney HLA-DR. In conclusion, these results suggest a increased incidence in the development of hyporeactivity in patients receiving pretransplant transfusions which share an HLA-DR antigen with the transplanted kidney.     

Tissue typing in support of unrelated hematopoietic cell transplantation

The success of unrelated hematopoietic cell transplantation (HCT) for the treatment of hematologic malignancies has closely paralleled development of robust typing methods for comprehensive and precise donor-recipient matching. The application of molecular methods in clinical research has led to a more complete understanding of the immunogenetic barriers involving host-vs-graft (HVG) and graft-vs-host (GVH) reactions. Along with the development of less toxic transplant regimens, advances in the prevention and treatment of graft-vs-host disease (GVHD) and in the supportive care of the transplant recipient, improved HLA matching of potential unrelated donors has led to clinical results that begin to compare favorably with that of HLA-identical sibling transplants.     

Increased soluble CD14 serum levels and altered CD14 expression of peripheral blood monocytes in HIV-infected patients.

Serum levels of soluble CD14 were elevated in HIV-infected asymptomatic patients or those with lymphadenopathy (CDC II/III) 2.9 +/- 0.8 mg/l compared with normal controls with 2.2 +/- 0.47 mg/l, P < 0.001. A further rise was seen in patients with ARC (CDC IVA) 3.8 +/- 1.1 mg/l, P < 0.01 and patients with AIDS (CDC IVB-D) 5.7 +/- 2.5 mg/l, P < 0.01. Although absolute numbers of CD14+ cells decrease in the AIDS group, the percentage of CD14+ monocytes did not change. In contrast, levels of soluble T cell antigens sCD4 and sCD8, which are higher in HIV-infected patients compared with normal subjects, showed no increase with disease progression. Serum levels of sCD14 were correlated positively with beta 2-microglobulin levels (rs = 0.63, P < 0.0001). Whereas the percentage of CD14+ monocytes did not change, an increase in monocytic CD14 expression in HIV-infected patients was observed (P < 0.01). The percentage of a monocyte subset expressing both CD14 and CD16 increased from 6% in normal healthy persons to 13% in HIV-infected patients (P < 0.001), and did not vary between the HIV patient groups. Incubation of cultured peripheral blood monocytes with azidothymidine had no effect on either normal or LPS-induced or IL-4-inhibited sCD14 release in vitro. Therefore, an effect of AZT on sCD14 serum values in vivo is considered to be unlikely. Our data further provide evidence that monocytes/macrophages are engaged in HIV infection.     

HIV感染与HLAⅠ类基因相关性的研究

目的　研究HIV感染与HLAⅠ类基因相关性 ,分析HLAⅠ类基因与HIV易感性及HIV感染者疾病进程的关系。方法　应用聚合酶链反应 序列特异性引物技术 (PCR SSP)检测 2 9例HIV感染者HLAⅠ类等位基因的特异性 ,从而确定HIV感染者的HLA分型。结果　在 2 9例HIV感染者中 ,HLA B35的等位基因频率较对照组明显增高 (B35 :Pc<0 .0 5 ,RR =5 .83)。结论　HIV感染者HLA B35的等位基因频率明显高于正常人 ,可能与HIV感染的易感性有关。     

Increased level of serum HLA class I antigens in HIV infection correlation with disease progression

Analysis of (sHLA-I) antigens in a large number of HIV-positive subjects found a significant increase of their level, but did not detect any change in their molecular profile. Monitoring at yearly intervals for four years of the sHLA-I antigen level in 14 HIV-positive subjects with a normal sHLA-I antigen level at study entry showed a significant correlation between progressive increase of sHLA-I antigen level and disease progression. Furthermore, a Kaplan-Meier plot of the frequency of development of AIDS in 34 patients whose cases were followed for 7 years showed that sHLA-I antigen level is a strong predictor of progression to AIDS. Its predictive value is comparable to that of serum β₂-μ level, greater than that of serum neopterin, and lower than that of CD4 + T-cell percentage. The predictive value of sHLA-I antigen level in combination with serum β₂-μ level, neopterin level, or CD4 + T-cell percentage is greater than that of each individual variable. These results suggest that measurement of the sHLA-I antigen level may provide useful prognostic information in HIV-positive subjects. Human Immunology 40, 259–266 (1994)     

Allogeneic immunity clears latent virus following allogeneic stem cell transplantation in SIV-infected ART-suppressed macaques

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人类免疫缺陷病毒感染细胞毒性T细胞逃逸突变的研究进展

人类免疫缺陷病毒（HIV）-1感染者体内活化的细胞毒性T细胞（CTL）反应不足以清除病毒,这是由于HIV。1在HLA限制的CTL压力下经常发生逃逸突变。但是,部分CTL逃逸突变会造成HIV病毒适应性的下降。近年来,人们对HIV-1感染中CTL逃逸突变的特征以及不同位点的逃逸突变在疾病进程中的作用进行了比较深入的研究,因而探讨CTL压力下HIV逃逸突变的规律有助于了解HIV-1自然感染中的免疫保护机制,并为开发有效的HIV疫苗提供依据。     

Human platelet alloantigen (HPA)-5a/b mismatch decreases disease-free survival in unrelated bone marrow transplantation

Matching of human platelet alloantigen (HPA) systems 2-6 was retrospectively investigated in 715 unrelated bone marrow transplantations. Of the five HPA systems studied, HPA-5 mismatching was found to have a significant effect on the disease-free survival rate of recipients following transplantation in the HLA-A, -B, -C, and -DR allele-matched donor-recipient pairs. The effect of the HPA-5 mismatch was most significant in the recipient group possessing the HLA haplotype A*2402-B*5201, which is a highly frequent haplotype among the Japanese population. However, the probability of development of acute graft-versus-host disease (GVHD) was not increased significantly by the HPA-5 mismatching. These findings suggest that the HPA-5 mismatching decreases the recipient's survival by a mechanism different from that in the case of mismatching of minor antigens found often in transplant recipients developing GVHD.     

Immunogenetics of HLA null alleles: implications for blood stem cell transplantation

The transplantation of haematopoietic stem cells is a potentially curative therapy for a variety of haematological and non-haematological diseases. Matching of donor and recipient for human leucocyte antigens (HLA) is pivotal for the success of blood stem cell transplantation. HLA null alleles are characterized by the lack of a serologically detectable product. Because serological HLA diagnostics are increasingly replaced by DNA-based typing methods considering only small regions of the genes, null alleles may be misdiagnosed as normally expressed variants. The failure to identify an HLA null allele as a non-expressed variant in the stem cell transplantation setting may result in an HLA mismatch that is highly likely to stimulate allogeneic T cells and to trigger graft-vs-host disease. For some HLA null alleles, the translation into a truncated polypeptide chain seems possible, which thus might act as minor histocompatibility antigens. Because the prevalence of HLA null alleles may be around 0.3% or even higher, a screening strategy for HLA null alleles should, therefore, be implemented in the clinical laboratory. It may consist of the combination of serology and standard molecular typing techniques. As the standard molecular techniques are sometimes troublesome especially for characterizing the cytosine island at the 5' end of HLA class I exon 4 and need continuously be updated, an alternative approach may consist of sequencing all samples from genomic DNA for exons 2-3 or 4 (class I) or exon 2 (class II), including the adjacent intron splicing sites. This approach will detect 36/40 so far known non-expressed variants and has the potential to easily uncover novel variants, thus essentially minimizing the risk of overlooking these challenging variants.     

Behavior of non-classical soluble HLA class G antigens in human immunodeficiency virus 1-infected patients before and after HAART: Comparison with classical soluble HLA-A, -B, -C antigens and potential role in immune-reconstitution

We have reported that serum level of soluble HLA-A, -B, -C (sHLA-A,-B,-C) antigens is elevated in HIV-infected subjects and decreases after antiretroviral therapy (HAART). In this study, we measured the levels of soluble HLA-G (sHLA-G) antigens in a cohort of HIV-infected patients before and during HAART. sHLA-G and sHLA-A, -B, -C levels were significantly elevated in HIV-infected subjects as compared with controls before antiretroviral treatment and significantly decreased after 36 months of HAART. sHLA-G levels were correlated with sHLA-A, -B, -C levels, the decrease of plasma HIV-RNA level, the increase of CD4+ T-lymphocyte number and the decrease of CD8+ T-lymphocyte number. These results suggest that the measurement of sHLA-G and sHLA-A, -B, -C antigen serum levels might represent a useful surrogate marker to monitor virological response and immune reconstitution in HIV-positive subjects undergoing HAART treatment.     

Impact of fetal-maternal tolerance in hematopoietic stem cell transplantation

Allogeneic hematopoietic stem cell transplantation (HSCT) is known to cure various hematological disorders; however, its widespread use is limited due to a lack of histocompatible donors. Reciprocal cell traffic between the mother and fetus during pregnancy gives rise to postpartum fetal-maternal lymphohematopoietic microchimerism, which is frequently detected in the blood or tissue of healthy individuals. Studies in clinical and experimental transplantation provide evidence that exposure to non-inherited maternal antigens (NIMAs) during pregnancy may result in long-lasting fetomaternal microchimerism and tolerance induction. Studies of HLA-mismatched HSCT have suggested a relatively lower incidence of severe graft-versus-host disease (GVHD) after transplantation from a NIMA-mismatched donor. Studies using a mouse model have also demonstrated a “child-to-mother” bone marrow transplantation from an NIMA-exposed donor to reduce the morbidity and mortality of GVHD in an antigen-specific manner while preserving the graft-versus-leukemia effects and favoring the immune reconstitution, thus resulting in a marked improvement in outcome after HSCT. Prospective clinical studies are therefore warranted to confirm these beneficial effects of fetal-maternal tolerance in allogeneic HSCT.     

Correlation of serum HIV antigen and antibody with clinical status in HIV-infected patients

An enzyme immunoassay (EIA) has been developed which detects antigen(s) (Ag) of the human immunodeficiency virus (HIV) in the serum of patients with the acquired immunodeficiency syndrome (AIDS), AIDS-related complex (ARC), and patients at high risk for HIV infection. The test has a sensitivity of approximately 50 pg/ml of HIV protein. The specificity of the assay was determined with various virus infected cell lines, normal human sera/plasma, and serum from patients not known to be at risk for HIV infection. No false-positive HIV-Ag results were seen. Sera from 69% of patients with AIDS were positive for HIV-Ag as were 46% of patients with ARC and 19% of asymptomatic, HIV-antibody-positive individuals. There were significant associations between the stage of HIV infection--ie, AIDS vs ARC vs asymptomatic--and the detection of HIV-Ag in serum (p less than 0.0001) and the lack of detection of antibody to HIV core Ag (p less than 0.0001). HIV-Ag was also found in the serum of two asymptomatic antibody-negative individuals who were at high risk for AIDS and who later developed HIV antibody. The presence of HIV-Ag in sera was confirmed by an inhibition procedure. Thus, HIV-Ag can be detected in the serum of infected individuals prior to antibody production and correlates with the clinical stage of HIV infection.     

Elevated serum levels of soluble tumour necrosis factor receptors (sTNF-R) in patients with HIV infection.

Serum levels of the soluble form of tumour necrosis factor receptor type II (p75) (sTNF-R) were determined in HIV-infected individuals and risk groups and were then correlated with the course of infection and prognosis. sTNF-R levels were determined by an ELISA with MoAbs and polyclonal antibodies to urine-derived sTNF-R proteins. The mean +/- s.e. levels of sTNF-R in the sera of 49 HIV+ male homosexuals, 34 HIV- male homosexuals and 44 matched controls were 6.1 +/- 0.3 ng/ml, 4.4 +/- 0.3 ng/ml and 3.4 +/- 0.2 ng/ml, respectively. All these values were significantly different between each of the groups (P less than 0.001-0.05). Sequential studies of sTNF-R revealed higher levels following seroconversion in 5/8 individuals, remained persistently high during the asymptomatic phase of the infection and became even more elevated in some ARC and AIDS patients. At the same time TNF-alpha was undetectable in sera obtained from HIV+ male homosexuals and from healthy controls. This was independent of stage of HIV infection, serum sTNF-R level and type of ELISA kit used. These findings suggest that TNF-alpha/TNF-R system is turned on before and during HIV infection and raise the possibility that sTNF-R, the natural inhibitor of TNF, may be of importance in determining the course and probably prognosis of the disease.     

HLA-DR antigens and phenotypes in Dutch coeliac children and their families

In a study on the HLA-DR antigens and phenotypes in a series of Dutch coeliac children and their first-degree relatives, the B-cell antigens of 36 unrelated coeliac children, 110 first-degree relatives of 33 of them, and 201 controls were typed with the two-colour fluorescence test. The most frequent antigen was HLA-DR3 (69%), followed by DR7 (36%). The distribution of DR phenotypes showed that the most frequent was DR3/other DR (25%), followed by DR3/DR7 (17%), DR3/DR4 (14%), and DR3/DR3 (14%). However, due to the frequency of certain antigens in the controls, only phenotypes DR3/DR3 (relative risk = 6.2), DR3/DR7 (relative risk = 6.4), and DR3/DR4 (relative risk = 6.2) were significantly associated with CD. The family study confirmed the segregation of the disease with phenotypes DR3/DR3 and DR3/DR7. The present results show that the association between CD and phenotypes DR3/DR3 and DR3/DR7 is not an exclusive characteristic of Southern coeliac children.     

Increased beta2-microglobulin-free HLA class I heavy chain serum levels in the course of immune responses to viral antigens and to mismatched HLA antigens

Besides being present in serum in association with beta2-mu, HLA class I heavy chains are also present in serum as beta2-micro-free moieties. The increase in serum levels of beta2-micro-associated HLA class I heavy chains in conditions associated with an activation of the immune system have prompted us to measure the serum levels of beta2-mu-free HLA class I heavy chains in the course of immune responses to viral antigens and to mismatched histocompatibility antigens. The serum level of beta2-mu-free HLA class I heavy chains, like that of beta2-mu-associated HLA class I heavy chains was significantly increased in patients affected by advanced HIV-1 infection or by chronic hepatitis C (CHC). In the latter group of patients an association was found between a reduction in the beta2-mu-free HLA class I heavy chain serum level and response to therapy with interferon alpha and ribavirin. Moreover, the beta2-mu-free HLA class I heavy chain serum level was increased more than that of beta2-mu-associated HLA class I heavy chains during episodes of liver ischemia following liver transplantation and in the course of acute graft rejection and of acute graft-versus-host-disease (GVHD) after allogeneic bone marrow transplantation (BMT). These results suggest that the serum levels of beta2-mu-free and beta2-mu-associated HLA class I heavy chains are independently regulated. Furthermore, beta2-mu-free HLA class I heavy chain serum level may be a useful marker to monitor response to therapy in CHC patients and the clinical course of liver and bone marrow grafts.     

Osteocalcin serum levels in patients following renal transplantation

Summary Osteocalcin serum levels reflect bone turnover. In renal insufficiency secondary hyperparathyroidism and reduced renal clearance might be responsible for elevated serum levels of osteocalcin. Renal transplantation might improve renal osteodystrophy and therefore could influence osteocalcin serum levels.We determined the influence of renal transplantation on osteocalcin levels in 37 consecutive patients (25m/12f) by RIA. Blood samples were collected prior to, 3 days, 28 days, 6 months and 12 months after renal transplantation.Prior to renal transplantation osteocalcin levels were significantly elevated (x±s: 23.4±12.8 ng/ml) compared to healthy volunteers (4.1±1.4 ng/ml). Following renal transplantation osteocalcin decreased significantly (9.4±8.9 ng/ml) 3 days and (7.1±7.8 ng/ml) 28 days. However, 6 and 12 months following renal transplantation the mean osteocalcin level increased again (8.3±5.7 ng/ml, 12.1±15.4 ng/ml). At 6 months 11 and at 12 months only 6 of 37 patients had osteocalcin levels in the normal range. 12 months following renal transplantation 21 out of 37 patients with elevated osteocalcin levels had parathyroid hormone levels above the normal range. Additionally to increased osteocalcin levels patients prior to renal transplantation had elevated alkaline phosphatase. Alkaline phosphatase had following renal transplantation a similar pattern as osteocalcin with initial decrease and secondary increase 6 and 12 months after renal transplantation. Parathyroid hormone was elevated in all patients before renal transplantation. Following renal transplantation mean parathyroid hormone levels fell significantly, however remained above normal range in 57% of these 37 patients. Osteocalcin serum levels correlated positively with alkaline phosphatase (rs=0.43–0.62;p<0.011) and parathyroid hormone (0.3–0.66p<0.07). This correlation of osteocalcin with alkaline phosphatase and parathyroid hormone prior to and after renal transplantation suggests that osteocalcin may be a confirmative parameter in renal osteodystrophy in patients on chronic intermittent hemodialysis and following renal transplantation.Abbreviations RIA radio immuno assay - OC Osteocalcin - rtx renal transplantation - GLA gammacarboxyglutamic acid - PTH parathyroid hormone - AP alkaline phosphatase - GLA gamma-carboxyglutamic acid     

Soluble HLA class I and class II antigens in patients with multiple sclerosis

Abstract: Soluble HLA class I (sHLA-I) and soluble HLA class II (sHLA-II) antigen levels during different stages of disease were investigated in paired serum and cerebrospinal fluid (CSF) samples from 37 patients with multiple sclerosis (MS) using ELISA and Western blot analysis. Soluble HLA-II antigens in the serum of untreated patients with the relapsing-remitting type of MS (RRMS) were found to be significantly elevated in acute relapse as compared to values obtained from patients under steroid treatment, in remission or healthy controls. No significant differences in circulating sHLA-I levels could be detected. In contrast, a trend towards increased intrathecal production of sHLA-I molecules in the CSF was observed in untreated RRMS patients in acute relapse, whereas the levels of soluble HLA-II antigens in the CSF were below the detection limit of the ELISA method. Our observations underline the presence of systemic immune activation in MS patients, as reflected in elevated serum sHLA-II antigen levels, and reveal a dichotomy between sHLA class I and II antigen production in the peripheral blood versus CSF in acute MS. Serial measurements of sHLA-II antigen levels might represent a non-invasive method to assess disease activity in MS patients.     

Cell-mediated immune responses to mycobacterial antigens in patients with pulmonary tuberculosis and HIV infection

Lymphocyte proliferation and cytokine responses induced by a panel of mycobacterial antigens were compared in Portuguese donors with pulmonary tuberculosis (TB) with or without HIV co-infection, HIV⁺ patients and healthy Mantoux-positive controls. Control donors showed stronger proliferative responses than any of the patient groups, with secreted antigens (Mycobacterium tuberculosis (Mtb) 30 kD and short-term culture filtrate proteins (ST-CFP)), purified protein derivative (PPD) and Mtb H37Rv Sonicate (MtbS) inducing the strongest proliferation. Patients with pulmonary TB showed lower proliferation to PPD or to the 30-kD antigen. Responses to all the antigens (PPD, ST-CFP, MtbS, 70 kD, 65 kD, 38 kD, 30 kD and 10 kD) were higher in TB/HIV patients with CD4 counts 200 CD4⁺ T cells/mm³ compared with HIV alone (CD4 200 T cells/mm³), but were lost in both TB/HIV and HIV patients when CD4 counts fell below 200 T cells/mm³. Measurements of interferon-gamma (IFN-γ) in culture supernatants revealed that PPD, 30 kD, MtbS and ST-CFP induced the strongest Th1 response. Analysis of mRNA for IFN-γ, IL-4 and IL-10 confirmed that IFN-γ production was maintained in patients with pulmonary TB without any concomitant increase in IL-4 or IL-10 mRNA expression, although expression of IL-10 mRNA was increased if HIV infection was present. These results reveal that IFN-γ production is retained in pulmonary TB patients to a broad range of mycobacterial antigens, and that no switch to IL-4 production is seen even with HIV infection. Secreted antigens, and in particular ST-CFP, were the best inducers of IFN-γ secretion, confirming their role in protective responses to Mtb.