Cellular mRNA expression of interferon-gamma (IFN-γ), IL-4 and IL-10 relates to resistance to experimental autoimmune myasthenia gravis (EAMG) in young Lewis rats

Age-related alterations in the immune system, including changes in lymphocyte subset composition, result in changes of cytokine patterns and might thereby influence the incidence and severity of autoimmune diseases. To investigate the age-related resistance to EAMG, an animal model for human MG, young (4-week-old) and adult (8–10-week-old) female Lewis rats were immunized with Torpedo acetylcholine receptor (AChR) and Freund's complete adjuvant (FCA). Adult Lewis rats showed severe weight loss and progressive muscular weakness after immunization, while young rats developed minor clinical signs of EAMG after a prolonged interval post-immunization. By comparison with adult rats, the young had lower AChR-specific T and B cells responses, and less muscle AChR loss. In situ hybridization performed on mononuclear cells (MNC) from lymph nodes revealed that young rats had lower levels of AChR-specific IFN-γ, IL-4 and IL-10 mRNA-expressing cells compared with adult rats. Since IFN-γ, IL-4 and IL-10 promote the development of EAMG, the low expression of these cytokines might contribute to EAMG resistance in young Lewis rats.     

丙型肝炎病毒核心蛋白在患者外周血单个核细胞内的核表达检测

目的　检测和研究丙型肝炎病毒 (hepatitisCvirus,HCV)核心蛋白在患者外周血单个核细胞 (peripheralbloodmononuclearcells ,PBMC)内核表达的意义 ,并探讨其与临床的关系。方法　对 6 6例慢性丙型肝炎患者PBMC标本进行免疫组化检测 ,并将HCV蛋白抗原定位分布情况与患者临床状况进行比较分析 ,对其中 2 7例患者PBMC进行HCVRNA和HCVAg的平行检测和分析。结果　免疫组化结果显示 ,慢性丙型肝炎患者PBMCHCVAg(core +NS3)阳性检出率为 77 2 7% (5 1 6 6 )。结果还证实 ,HCV核心蛋白均定位于胞核内 ,且呈强表达 ;NS3蛋白主要定位于胞质内 ,呈弱表达。当进行HCVAg在PBMC内定位情况与患者临床状况比较分析时显示 ,病情较重患者PBMC内核心蛋白表达阳性率 (35 2 9% )明显高于病情较轻者 (5 88% ) (P <0 0 0 1)。结论　HCV核心蛋白在PBMC内核表达与患者临床状况相关 ,提示其可能是丙型肝炎慢性化的一个指标 ,并可能在肝硬化和肝癌发生上起一定作用     

Simultaneous activation of natural killer T cells and autoantibody production in mice injected with denatured syngeneic liver tissue

Denatured syngeneic liver tissue prepared by mechanical procedures was intraperitoneally injected into adult C57BL/6 mice. In parallel with a decrease in the total number of lymphocytes in the liver, spleen, and thymus from days 1-7 after the injection, the proportion of the CD4+NK1.1+CD3(int) subset of these cells (i.e. natural killer T or NKT cells) increased in the liver. Even the absolute number of these NKT cells increased in the liver on days 14 and 21. In response to the injection of denatured liver tissue, tissue damage was induced in the liver, as shown by elevated levels of serum transaminases and hepatocyte degeneration observed by electron microscopy. Sera obtained on days 7 and 14 contained autoantibodies including anti-DNA antibodies. The proportion of CD1d(high)B cells in the liver was found to decrease on days 1-7. In other words, denatured liver tissue stimulated both NKT cells and certain B cells in the liver. These results suggest that liver lymphocytes might contain not only autoreactive T cells (e.g. CD3(int) or NKT cells) but also some B cells (e.g. B-1 cells) which produce autoantibodies and that the denatured tissue had the potential to stimulate these lymphocytes and to evoke an autoimmune-like state.     

Evidence for the presence of a histaminergic neuron system in the rat brain: an immunohistochemical analysis

The binding of calcium antagonists in the rat hippocampal formation was studied using autoradiography. Hippocampal slices were labeled in vitro with [3H]PN 200-110. High densities of binding sites for calcium antagonists were found in the molecular layer of the dentate gyrus and in the CA3 subfield of the hippocampus. After ablation of the granule cells by local injection of colchicine a marked decrease in the number of [3H]PN 200-110 binding sites density was observed on these areas, while binding to other parts of the hippocampal formation and brain was spared. These results strongly suggest the localization of high densities of calcium channels to the granule cells of the dentate gyrus.     

B-cell function in patients with chronic lymphocytic leukemia

Summary Using a reverse hemolytic protein A plaque assay, spontaneous and pokeweed mitogen (PWM)-induced immunoglobulin (Ig) secretion was determined in peripheral blood from 22 patients with B₁-chronic lymphocytic leukemia (CLL), one patient with B₂-CLL, and one patient with suppressor T-CLL. Diagnoses were established by cytological and histological criteria as well as several marker analyses. Lymphocytes from B₁- and B₂-CLL patients were unable to secrete Ig either spontaneously or after PWM stimulation. In T-CLL lymphocytes, spontaneous Ig secretion was suppressed very probably by the OKT-8-positive leukemic population, since, after cultivation with PWM, a normal Ig secretion could be demonstrated which was paralleled by a decrease in the OKT-8-positive cells. Cocultivation experiments with freshly isolated, unseparated lymphocytes from normal subjects and lymphocytes from patients were of no informational value, since isolated normal B-cells alone already showed a high rate of Ig secretion. However, coculture experiments with separated subpopulations after PWM stimulation revealed an intrinsic B-cell defect in lymphocytes from B₁-CLL patients, whereas their T-lymphocytes were found to be normal helper cells.

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Abbreviations CLL Chronic lymphocytic leukemia - PWM Pokeweed mitogen - ISC Immunoglobulin-secreting cells - Ig Immunoglobulin(s) Supported by the Deutsche Forschungsgemeinschaft (Ru 215/2)     

A method for the accurate determination of anti-hapten cytotoxic T lymphocyte precursors: correction for apparent 'anti-self' reactivity

A method is described for accurately determining the frequency of precursors of hapten specific cytotoxic T cells. The method is based on a standard Poisson analysis of limit dilution cultures, but makes a correction of 'anti-self' reacting clones and for spontaneously arising clones that recognise modified self. These corrections are shown to be especially important when low hapten densities are used, where there may be more than a 10-fold difference between the corrected and uncorrected frequency estimates. Determined levels of antigen specificity and of H-2 restriction are significantly enhanced by application of this method.     

Acquired α2 macroglobulin on the surface of human lymphoblastoid cells

The molecular nature of the membrane antigen that is acquired from FCS-containing media by human lymphoblastoid cells has been investigated. The presence of bovine α₂, macroglobulin on the surface of Namalva cells was demonstrated by radioimmunoassay using specific antisera. Alternatively, cell-bound bovine α₂,M could be detected by the more sensitive heterophile rosette assay described previously. Namalva cells grown in NHS-containing media acquired bovine α₂ M upon subsequent incubation with the purified protein in a dose- and time-dependent way. Acquisition of α₂ M was demonstrated using both viable and formaldehyde-fixed cells. Purified fetuin, which carries a heterophile epitope shared with bovine α₂ M as well as with other glycoproteins, failed to bind with the membrane of Namalva cells. The possible role of acquired α₂ macroglobulin on cells has been discussed.     

The thymus is a site of mast cell development in chicken embryos

Thymic mast cells were studied by light and transmission electron microscopy in chicken embryos during organogenesis. Mast cells made their first appearance at day 15. At days 16 and 17, there was a burst of mast cell development with a peak of 278 ± 54 cells/mm² at day 16. Then, mast cell density decreased until hatching. During the whole embryonic period, about 80% of mast cells localized to the thymic medulla. In the cortex, they were less numerous, and some rare mast cells could be identified in the capsule and septa. Thymic mast cells could be recognized in association with hematopoietic foci, but frequently they grew independently from areas of hematopoiesis and appeared as single cells interspersed among thymocytes, thymic epithelial cells, and interdigitating cells. They were often recognized in close relationship with the scanty and delicate extracellular matrix of the developing gland. Viewed by electron microscopy, mast cells were relatively small cells, with a few secretory granules. Exocytosis was never seen, but, notably, granules emptied in a piecemeal degranulation fashion. This study demonstrates that the chicken thymus is a site of mast cell development during embryogenesis. The high mast cell density we found suggests a possible role for these cells during thymus organogenesis.     

The inhibitory NK cell receptor CD94/NKG2A and the activating receptor CD94/NKG2C bind the top of HLA-E through mostly shared but partly distinct sets of HLA-E residues

The human non-classical MHC class I molecule HLA-E is a ligand for both an inhibitory NK cell receptor (CD94/NKG2A) and an activating receptor (CD94/NKG2C). To identify HLA-E surface recognized by both receptors, especially to determine if both receptors recognize the same epitope, we made a series of individually Ala-substituted HLA-E proteins and analyzed their binding to CD94/NKG2A orCD94/NKG2C. Eight HLA-E mutations that significantly impaired HLA-E binding to CD94/NKG2A are all found in the top of alpha1/alpha2 domain of HLA-E. These results suggest that CD94/NKG2A binds a HLA-E surface equivalent to a NKG2D binding site on MICA. Of the eight mutations that impaired HLA-E binding to CD94/NKG2A, six significantly impaired HLA-E binding to CD94/NKG2C suggesting that CD94/NKG2C also binds a similar surface of HLA-E. Unexpectedly, the two HLA-E mutations (D69A and H155A) selectively abrogated HLA-E binding to CD94/NKG2A, not largely affected CD94/NKG2C. These results indicate that a mostly shared, but partly distinct set of HLA-E residues is discriminated by the two receptors.     

人嗜铬细胞瘤细胞的原代培养及鉴定

本研究拟建立人嗜铬细胞瘤细胞的原代培养方法。采用连续分次胶原酶消化法分离培养人嗜铬细胞瘤细胞，采用细胞培养液中儿茶酚胺水平检测、多聚甲醛诱发荧光及细胞嗜铬粒蛋白A(CgA)和神经元特异性烯醇化酶(NSE)的免疫组化染色等方法进行细胞性质和功能鉴定，并用噻唑兰(MTT)法观察原代培养的人嗜铬细胞瘤细胞的生长状况。结果表明，人嗜铬细胞瘤细胞在培养3～7天时生长较快，7天后细胞开始分化。经检测细胞培养液中的儿茶酚胺浓度、多聚甲醛诱发荧光等，证明该细胞有合成和分泌儿茶酚胺的功能。并且培养的细胞CgA和NSE免疫组化染色阳性。因此，本研究成功建立了人嗜铬细胞瘤细胞的原代培养方法，并鉴定其具有嗜铬细胞瘤的分泌和表达功能，国内尚未见报告。