Cytotoxicity mediated by human Fc receptors for IgG

The Fc receptors for IgG(Fc gamma R) play a major role in the removal of antibody-coated infectious agents and may be important molecules for triggering cytotoxicity of tumor cells; they may also serve as an entry for infection of Fc gamma R-bearing cells by viral (including HIV and Dengue), and perhaps other infectious agents. Although central to immune defense, an understanding of the role of these Fc gamma R in cytotoxicity has been complicated in part by the presence of several biochemically distinct types of receptor that have different distributions, specificities, affinities and modes of activation for killing. The development of monoclonal antibodies specific for Fc gamma R on human leukocytes has established the existence of three distinct Fc gamma R and furthermore has helped clarify the function of each of these receptors. In this review, Michael Fanger and colleagues discuss the use of Fc gamma R-specific mAb and the hybridoma cell lines that produce them in examining the ability of each of these unique receptors to mediate killing of tumor and red cell targets. In particular, the use of self-directed hybridoma cells as a model of tumor-cell killing and of bi-specific antibodies to link target cells to effector cells through the different Fc gamma R is discussed. The results of these studies suggest that the ability of a given Fc gamma R to trigger killing is sometimes dependent on the type of Fc gamma R, but is also markedly influenced by the type of target cell and by the nature and state of activation of the effector cell.     

Improved filtration method for red cell deformability measurement

A method which enables reliable measurements of red cell deformability in whole blood to be performed is presented. It is based on the Nuclepore filtration method described byReid et al. (1976). Its reliability was much improved, obtaining an accuracy of within 5% by a technique to remove air bubbles trapped in the filter pores, which caused the poor reproducibility observed before. Simple analyses of the flow characteristics of whole blood and the haematocrit dependence of whole blood passage time were given to eliminate the effect of transient pore blockings of white cells and the contribution of different haematocrit in the whole blood passage time. A mean pore passage time of single red cells is obtainable as a quantitative index of red cell deformability. Furthermore, a good utilisation was achieved by making full use of electrically operated valves and aspiration pumps. It is possible to repeat measurements at intervals of 3 min. This improved filtration method will be useful for diagnostic purposes since it can avoid artefacts caused by artificial treatments added on blood samples and time changes after blood sampling.     

Production of migration inhibition factor (MIF) by purified human T cell subpopulations.

T cells possessing suppressor activity (TG) were isolated from normal human peripheral blood by rosetting with IgG-coated ox red blood cells (OxRBC) as shown by their ability to inhibit pokeweed mitogen (PWM) induced proliferation and Ig synthesis in vitro. The suppressor activity of the intact T cell and Tg cell fractions was enhanced by concanavalin A (Con A) stimulation for 48 hr and supernatants from these Con A-treated cell cultures, after removal of mitogen, contained migration inhibition factor (MIF). Its release appeared to be primarily a function of the TG cells. The bulk of the MIF activity was present in a gel filtration fraction of mol. wt approx. 25,000, similar to that found in previous studies of Con A- or antigen-induced MIF in man.     

A Novel Synthesis of Polyacrolein Microspheres and Their Application for Cell Labeling and Cell Separation

A novel method for the synthesis of polyacrolein microspheres with fluorescent or magnetic properties is described. These microspheres carry reactive aldehyde groups on their surface, which are used for covalent binding of various proteins at physiological pH. Polyacrolein microspheres may be used as a simple tool for cell labeling and cell separation. The feasibility of specific labeling of fresh human red blood cells and of the separation of human red blood cells from turkey red blood cells by means of a magnetic field is discussed.     

红细胞冻干保存中的渗透性损伤实验研究

红细胞长期保存中保护剂添加、洗涤过程会引入红细胞渗透性损伤.在冻干保存研究中,由于多种保护剂同时使用,保护剂的类型和功能一直是研究的重点,但很少有从渗透性损伤角度分析保存方案合理性的报道.目前相关文献、专利中所用的保护剂总渗透压差别很大,细胞保存后回收率差异也较大.文中用NaCl溶液实验模拟红细胞保存中保护剂添加、洗涤过程.结果表明,选择合适的添加、洗涤方法可以在一定程度上减小渗透损伤.就红细胞而言,1.5Osmol/kg左右是保护剂总渗透压的一个重要阈值,总渗透压低于该阈值时,渗透性损伤较小;高于该阈值时,渗透损伤随着渗透压的增大而迅速增大.所以选择保护剂时,首先应该根据总渗透压来排除渗透压过高的保存方案,否则红细胞在添加和洗涤保护剂时已经损伤很大.该研究对其它细胞长期保存中保护剂的选择也具有参考意义.     

The development of unusual B-cell functions in the testosterone-propionate-treated chicken.

Chickens were treated with 4 mg of testosterone propionate on the twelfth day of embryonic life. Bursal remnants of testosterone-treated chickens were very small in size and had very few or no bursal follicles: the lymphoid tissue was replaced substantially by fibrosis. Testosterone-treated chickens formed almost exclusively IgM antibodies to sheep red blood cells and influenza virus, whereas no IgM or IgG response to Brucella abortus or Salmonella pullorum, and no IgG response to sheep red blood cells was demonstrable. Surgical removal of bursal remnants of testosterone-treated chickens at hatching did not significantly affect IgM response to sheep red blood cells. These B-cell functions of testosterone-treated chickens were not improved by addition of T cells, as shown by adoptive cell transfer experiments. Thus, there appears to be an unusual type B-cell development which is independent of the bursa of Fabricius.     

A Novel Synthesis of Polyacrolein Microspheres and Their Application for Cell Labeling and Cell Separation

A novel method for the synthesis of polyacrolein microspheres with fluorescent or magnetic properties is described. These microspheres carry reactive aldehyde groups on their surface, which are used for covalent binding of various proteins at physiological pH. Polyacrolein microspheres may be used as a simple tool for cell labeling and cell separation. The feasibility of specific labeling of fresh human red blood cells and of the separation of human red blood cells from turkey red blood cells by means of a magnetic field is discussed.     

红细胞在保护剂添加和洗涤时的非平衡渗透响应实验研究

在红细胞低温保存或冻干保存中，高浓度保护剂的添加和洗涤会使细胞体积收缩或膨胀，造成溶血损失，但其体积并非单调收缩或膨胀到最后平衡体积，而是要经历更严重的体积变化后再趋于平衡。本研究以NaCl溶液来模拟保护剂的添加与洗涤过程．将38种不同浓度的NaC;溶液，以一步直接法、4步等体积法或4步等摩尔浓度变化法添加到红细胞中，测定其溶血率。若以溶血率10％为标准，红细胞所能承受的最小渗透压在161mOsmol／k附近，而最大渗透压与溶液添加方式有关，等体积添加法效果较好（其值约为5680mOsmol／kg）。研究中还将12％、18％NaCl溶液以4步等体积法加到红细胞中，再用生理盐水以三种方式洗涤，发现等摩尔浓度变化法洗涤效果最好；洗涤后溶血率大大增加，说明很多细胞虽然在保护剂添加时未溶血，但膜脆性已改变，难以恢复到等渗时体积。     

Antigen-presenting cell activation: a link between infection and autoimmunity?

The onset of autoimmune diseases such as type I diabetes and multiple sclerosis is often thought to be associated with infection. This has led to studies of molecular mimicry between infectious agents and the self-antigens associated with autoimmunity. Despite many claims, however, a single causative infectious agent for autoimmunity has not been found. An alternative possibility is that many infectious agents are capable of non-specifically enhancing the likelihood of an autoimmune attack. Here we show how infectious agents may activate antigen-presenting cells leading to the activation of autoreactive T cells by otherwise innocuous antigens. The mechanism of activation involves upregulation of co-stimulatory molecules on the antigen-presenting cell resulting in a lowering of the threshold required for activation. These results help explain how diverse infectious agents could cause autoimmune disease in susceptible individuals.     

Inactivation of infectious pathogens in labile blood components: meeting the challenge.

Substantial improvement in the safety of blood transfusion has been achieved through the addition of new tests, such as nucleic acid tests, yet residual risk associated with transfusion of blood components persists. Transfusion of blood components has been implicated in the transmission of viruses, bacteria, and protozoa. While it is commonly recognized that hepatitis B virus (HBV), hepatitis C virus (HCV), cytomegalovirus (CMV), and the retroviruses, such as human immunodeficiency virus (HIV) and the human lymphotrophic viruses (HTLV) can be transmitted through cellular components, other pathogens are emerging as potentially significant transfusion-associated infectious agents. For example, transmission of protozoan infections due to trypanosomes and babesia have been reported. In addition to viral and protozoal infectious agents, bacterial contamination of platelet and red cell concentrates continues to be reported; and may be an under-reported transfusion complication. More importantly, new infectious agents may periodically enter the donor population before they can be definitively identified and tested for to maintain consistent safety of the blood supply. The paradigm for this possibility is the HIV pandemic, which erupted in 1979. During the past decade a number of methods to inactivate infectious pathogens in labile blood components have been developed and have entered the advanced clinical trial phase.     

外磁驱动轴流式血泵磁场分布及红细胞电磁特性分析

外磁驱动轴流式血泵较强的磁场强度会对血液及周围组织细胞产生影响,因此对血泵及其周围红细胞进行电磁场理论计算和仿真分析。利用ANSYS Electronics Desktop中3D瞬态磁场模块对血泵进行瞬态磁场仿真,用理论方法建立细胞膜磁场分布模型,综合利用3D瞬态电场和磁场模块对红细胞膜及其内外电磁场进行研究。给出了血泵稳定状态时的3D和2D磁感应强度分布云图,得到了细胞膜受到的最大磁感应强度值;通过最大磁感应强度值和血泵工况特点得到红细胞膜电场时域上的分布规律和幅值;综合细胞膜静息电位得到细胞膜电场耦合分布规律;基于以上条件求得细胞膜上感应磁场分布及细胞膜所受最大磁场力。尽管钕铁硼材料剩余磁感应强度很大,但血液和红细胞所受最大磁感应强度值仅为812 mT。由此得到的各项红细胞电磁特性参数值可为红细胞受驱动磁场影响下受到的电磁损伤和血泵的临床应用以及优化设计提供理论基础。     

Control of primary IgM antibody responses to H-2 alloantigens by antigen-bearing live B lymphocytes

Alloantigen-bearing (H-2d+) peripheral red blood cells, but not red cell-depleted H-2d+ spleen cells, induce primary IgM anti-H-2d plaque-forming cell responses. In this study it is reported that the primary antibody responses to H-2d+ peripheral red blood cells can be markedly suppressed by a subpopulation of H-2d+ spleen cells when they are injected simultaneously or a few days before injection of red blood cells. This suppression was antigen (H-2d)-specific, did not depend on T cells of either the donor or the recipient, and strictly required live donor cells. An energy-dependent action of the donor cell cortex and some proliferation of donor cells in the recipient seemed to be involved in the mechanism of suppression. The donor-suppressor cell type was largely present in the spleen but not in the bone marrow and thymus, and was present in the spleen of athymic nude mice. The suppressor cells displayed the properties of B lymphocytes: they adhered to the nylon wool but not to glass, were of relatively low density (rho less than 1.09), and were surface Ig+, Ia+, Fc receptor-positive but Thy-1-. H-2d+ suppressor-donor B lymphocytes might directly signal to antigen-specific recipient B cells competing with the signal provided by H-2d+ red blood cells for the B cell activation.     

红细胞聚集与取向对其悬浮液电阻抗的影响——I.实验研究

本文通过测量具有不同流变特性的红细胞悬浮液电阻抗的动态与静态变化，研究了红细胞聚集与取向对其悬浮液电阻抗的影响。首次得出以下结论：１．红细胞聚集使其悬浮液电阻抗显著减小。２．由于低切变率下红细胞聚集与高切变率下红细胞取向的共同影响，使血液电阻抗随切变率的变化具有双相特性。这增加了对血液电特性的新认识，表明可利用电阻抗测定红细胞聚性。     

Modulation of the immune system in the mouse 1. Drug administration prior to antien sensitization

A model has been developed in the mouse in which metylated bovine serum albumin (MBSA) and sheep red blood cells have been used to produced delayed type hypersensitivity (DTH) and humoral immunity (HI), respectively.The time coruse between antigen sensitization and challenge was chosen such that cyclophosphamide (CY), administered prior to sensitization, produced DTH enhancement and also produced suppression of HI when administered at high doses. A number of other drugs have been examined in this model but only CY-like drugs, viz. alkylating agents, produced similar DTH enhancement. DTH was suppressed in a few cases.The HI response was enhanced or suppressed by a number of unrelated drugs. CY was the only alkylatng agent to suppress the antibody titre.The mechanisms for these drug effects are uncertain. However, a number of drugs elicit effects on either DTH or HI which suggests there is selective removal of certain cell populations rather than non-specific cytotoxicity. Possible theories for the effects observed are discussed.These studies also suggest that CY possesses unique properties in this particular model.     

Cytotoxic Effector Cells from Infectious Mononucleosis Patients in the Acute Phase Do Not Specifically Kill Epstein-Barr Virus Genome-Carrying Lymphoid Cell Lines

We describe a study in which we investigated the cytotoxic activities of thymusderived (T) lymphocytes and natural killer cells against Epstein-Barr virus (EBV) genome-carrying lymphoid cell lines. Purified subpopulations of lymphocytes from eight patients with infectious mononucleosis and six healthy normal EBV-seropositive donors were tested. Enriched T-cells were obtained by passing purified whole blood lymphocyte preparations through human immunoglobulin-anti-immunoglobulin-coated glass bead columns. The cytolytic activity of effector cells was determined by the ability of these cells to lyse human target cells that were internally labeled with (51)Cr. These targets included cells from both EBV genome-carrying and EBV genome-negative lymphoid lines derived from malignant tumors, as well as from lymphocytes transformed in vitro by EBV, and were chosen to represent a wide spectrum of EBV-associated membrane antigens. We found that cytotoxic T-cells from patients with infectious mononucleosis showed no EBV-related specific cell killing per se, although a trend for increased killing of cell lines derived from spontaneous in vivo growing tumors, EBV genome carrying or not, was noted; however, this trend was not observed with cell lines derived from cord blood lymphocytes after EBV infection in vitro. In addition, our data suggest that natural killer cells may play an important role in controlling EBV infection in patients with infectious mononucleosis in the acute phase of the disease, particularly since T-cells (obtained after removal on immunoglobulin-anti-immunoglobulin columns of natural killer cells presumably bearing Fc receptors) were less efficient killers than whole blood lymphocytes; furthermore, lysis by whole blood lymphocytes was also greatest against cell lines derived from malignant tumors (as opposed to in vitro EBV-transformed cord blood lymphoid lines), irrespective of whether these targets were EBV genome positive or negative.     

Removal of parvovirus B19 from hemoglobin solution by nanofiltration

Parvovirus B19 (B19), which may contaminate red cell components for blood transfusion, is known to be resistant to several viral inactivation methods. To increase the safety of hemoglobin solutions as a source of red cell substitutes, we investigated the removal of parvovirus B19 from hemoglobin solution using nanofiltration. The hemoglobin solution spiked with parvovirus B19 was tangentially filtered using the BMM-35 filter (mean pore size of 35 nm) followed by BMM-15. The parvovirus B19 of 10(8.5) PT50 (median PCR titer)/10 microL was not changed after the BMM-35 filtration. However, the BMM-15 filtration decreased the parvovirus B19 from 10(8.3-8.7) PT50 to 10(1.3-2.2) PT50, indicating more than 6 log10 reduction. When the initial parvovirus B19 of 10(6.0) PT50 was subjected to the BMM-15 filtration, the residual virus was 10(-0.3-0.5) PT50 orundetected in some fraction of the filtrate. Hemoglobin recovery was 70.4 +/- 3.4%. The ratio of methemoglobin was not changed during the filtration. These findings indicate that the BMM-15 filtration is a promising approach to prepare a safer hemoglobin solution for red cell substitutes.     

一种新的流感快速诊断法

目的 建立一种新的、简便、敏感、特异和重复性好的流感快速诊断法。方法 采集标本经成片MDCK细胞扩增，使细胞表面受体发生改变，借助植物素与细胞表面受体结合的严格特异性，通过普通光学显微镜来观察红细胞聚集现象，对测定标本做出判断。结果 新法与常规的MDCK细胞分离法符合率为100％，一般20h内就可出结果；其敏感性比常规细胞分离法高100-10000倍或以上；它既可用于临床上的流感快速诊断，又可用于流感监测；测定时最适红细胞浓度为1‰，并测定结果不受人血型和豚鼠个体的影响。结论 新法简便、敏感、特异、重复性好，并具有多用途等优点。     

Photoimmobilized array of panel cells for assay of antibodies

Antibodies in blood are checked with panel blood cells before blood transfusion. In this investigation, for the first time, a panel cell-microarray was prepared by using a photoimmobilization method. Different types of red blood cells were microarrayed on a plate. A water-soluble photoreactive polymer as a matrix was synthesized by the coupling reaction of azidoaniline with poly(2-methacryloyloxyethylphosphorylcholine-co-methacrylic acid). The polymer was mixed with cells and the mixtures were microspotted on substrate and photoirradiated after drying in air. For the antibody assay, monoclonal antibodies or human serum was added to the cell-arrayed plate and adsorbed antibodies were detected by horseradish peroxidase-labeled secondary antibody, which recognized the adsorbed antibodies. Antibodies specifically adsorbed on the immobilized cells as expected. The aggregation method has been available for this type of assay, but extensive experience was needed to apply it correctly. The method using a cell array will be useful for antibody detection.     

Lymphocyte subpopulations. Human red blood cell rosettes.

Human red blood cells (HRBC) even without prior neuraminidase treatment, could form rosettes with human peripheral blood lymphocytes in vitro. The optimum conditions for forming these rosettes were a pH of 7-0 and a medium with 5% bovine serum albumin (BSA). Rosette proportions became much less at a different pH or using lower concentrations of BSA, or replacing BSA with foetal calf sera (FCS) or human sera. Rosette formation was also promoted by prior treatment of HRBC or lymphocytes with neuraminidase. Mixed rosettes of HRBC and sheep red blood cells (SRBC) showed that HRBC receptors were detectable only on lymphocytes that possessed SRBC receptors, suggesting that HRBC rosette-forming cells were probably thymus-derived (T) cells. Next, the properties of human red blood cell (HRBC) and sheep red blood cell (SRBC) rosette-forming cells were investigated by comparing the ability of human peripheral blood lymphocytes to form these two types of rosettes after treatment with various inhibitory reagents. HRBC rosettes were relatively more resistant to inhibition with: (1) proteolytic agents, such as trypsin, alpha-chymotrypsin and pronase; (2) anti-thymocyte serum (ATS); (3) metabolic inhibitors, such as sodium azide and 2,4-dinitrophenol (DNP); (4) cytochalasin B. On further incubation after trypsinization, the lymphocytes recovered some ability to form SRBC rosettes, but continued to lose more of their capability to form HRBC rosettes. All these results were regarded as circumstantial evidence that the HRBC rosettes might represent a subpopulation of human T lymphocytes.     

The spleen in malaria: the role of barrier cells

I believe that my laboratory has developed a construct of the spleen useful in understanding its range of normal and pathologic functions. The elements in the construct include recognition of an anatomically open vasculature with the interposition of reticular cell-reticular fiber filtration beds between terminal arterial vessels and proximal venules. The central function of the spleen, moreover--selective clearance of cells, microbes and other particles from the blood--depends upon these filtration beds. Such functions of the spleen as phagocytosis, immunologic reactivity, hematopoiesis, and blood cell storage derive from its clearance capacities. The reticular filtration beds offer but modest levels of basal clearance. The wide ranges of filtration that characterize the stressed spleen depend upon arming or augmenting the basic reticular filtration beds with responsive cells which can rapidly appear, and rapidly disappear. These include macrophages, salient phagocytic cells of rich repertoire, which have been accorded the major, even exclusive, role in splenic clearance. But other stromal cells participate in splenic clearance. I have identified a system of fibroblastic, contractile, granulated cells which fuse to form complex, branched syncytial sheets which, deployed as diverse barriers, augment the basic reticular filtration beds. Hence, I term these cells barrier cells. Barrier cells effectively interact with macrophages, reticular cells, other stromal and blood cells, contributing to the extraordinary range of splenic clearance capacities. Barrier cells may be elicited by a variety of infectious processes, damaged blood cells and hematopoietic factors. Interleukin-1-alpha evokes a strong barrier cell response, and may be the common denominator in splenic stress, stimulated by activated macrophages.(ABSTRACT TRUNCATED AT 250 WORDS)