A Monoclonal Antibody Specific to Embryonic Trunk-Lateral Cells of the Ascidian Halocynthia roretzi Stains Coelomic Cells of Juvenile and Adult Basophilic Blood Cells

In spite of extensive knowledge on the structure and function of ascidian blood cells, little is known about their embryological origin. In the present investigation, the developmental fate of trunk lateral cells (TLCs) was explored using a specific monoclonal antibody. TLCs comprise a group of undefined embryonic cells of the ascidian Halocynthia roretzi , which arise from the A7.6 blastomeres of a 64-cell embryo. The antigenicity first appeared at the middle tailbud stage in a pair of TLC-clusters situated lateral to the brain stem of the bilaterally symmetrical embryo. The position and number of stained cells did not change during later embryogenesis until hatching. After hatching, the stained cells were found in the entire trunk region of the swimming larva. After metamorphosis, cells that expressed the antigen were present within the coelom and within the tunic layer of the juvenile. In addition, the antibody stained adult basophilic blood cells. These observations suggest a relationship of this group of embryonic cells with the prospective blood forming mesenchymal cells.     

Production and characterization of monoclonal antibodies with P1 specificity

Four monoclonal antibodies, with P1 specificity were obtained after fusion of myeloma cells and spleen cells from mice immunized with turtle dove ovomucoid. Immediately after the fusion, the culture supernatants were studied for specificity with panels of erythrocytes and red blood cells sharing rare phenotypes (P1K, P2K, p) in the P system. After cloning, four monoclonal antibodies were produced, these antibodies strongly agglutinate P1 red blood cells, specially when they are used with 3% of dextran or with a 350 mmol/l concentration of sodium.     

Human leukaemia-associated antigens expressed by acute myelocytic leukaemia cells and their detection by heterologous antisera.

Antisera against human acute myelocytic leukaemias were tested in complement-dependent in-vitro cytotocity tests against leukaemia cells and normal cells as targets. After absorption with erythrocytes and spleen cells from allogeneous donors the antisera reacted with leukaemia cells, but not with leukocytes from bone marrow and the peripheral blood of children in remission, lymphocytes from healthy donors, enriched B-lymphocytes, enriched T-lymphocytes, PHA-induced blasts and cord blood lymphocytes. Extensive cross reactions were obtained in the tests against leukaemia cells. The antisera reacted not only with AML cells, but also with ALL, CLL, and CML cells. It was possible to remove the cross-reactivity with ALL cells through absorption with ALL cells or with fetal tissue, and to remove the cross reactivity with CLL cells through absorption with CLL. A complete absorption of the anti-AML sera was possible with AML and CML cells. After absorption with fetal tissue and CLL cells the antisera showed exclusively specificity for myelocytic leukaemias. Thus, AML cells contain three leukaemia-associated membrane antigen components: an antigen of fetal origin, a "CLL-specific" antigen, and an antigen that occurs on myelocytic leukaemias.     

A new method for fish leucocyte counting and partial differentiation by flow cytometry

A simple and rapid method for analysis of fish blood cells is presented. Carp (Cyprinus carpio) blood was diluted 200 times with Hanks' solution containing 1 microg/ml of DiOC6(3) which is a fluorescent, lipophilic dye. After staining for 10 min, the blood cells were measured by a flow cytometer (FACS). Several blood cell populations were identified by different FL-1 (green fluorescence), FSC (forward scatter), and SSC (side scatter) properties. FL-1 v. SSC or FSC v. SSC dot-plot of stained blood cells displayed five separate cell populations: erythrocytes: a mixture of thrombocytes plus lymphocytes; monocytes; neutrophils; and basophils. The number of each type of blood cell counted by the FACS was in good agreement with those counted microscopically.     

Morphologic alterations in cultured human synovial fibroblasts induced by blood mononuclear cells

Effects of peripheral blood mononuclear cells on cultured synovial fibroblasts were studied. When mononuclear cells from normal or rheumatoid blood were incubated on synovial fibroblast cultures, a part of the cells adhered to the fibroblasts. They were mainly T lymphocytes but also some B lymphocytes and monocytes. After a 10-hour incubation, adhered mononuclear cells induced morphologic alterations to synovial fibroblasts: appearance of stellate cells and thinning and branching of fibroblasts. No changes were seen when the cells were incubated in the presence of indomethacin. Cytotoxicity of peripheral blood mononuclear cells from 8 rheumatoid patients was also tested against three rheumatoid and three normal synovial fibroblast strains. Only 2 out of 48 combinations were cytotoxicity. The potentially cytotoxic mononuclear cells were bound equally well to rheumatoid and control synovial fibroblast cultures.     

Ontogeny of T Cells in the Pig Fetus

Mononuclear cells isolated from thymus, spleen and cord blood of pig fetuses ranging in age from 48 to 112 days were examined for the presence of sheep red blood cell rosette-forming cells (SRBC-RFC). After an initial increase from 77 % (mean) at 48 days of gestation to 88 % at 60 days, the proportion of SRBC-RFC in thymus remained constant throughout the gestational period. In spleen and cord blood, the proportion of SRBC-RFC increased with age, from occasional rosette-forming cells at 48 days of gestation to 21 % and 30 %, respectively, at 112 days. The demonstrated development of SRBC-RFC in the thymus, spleen and cord blood is considered to reflect the ontogeny of T cells in these fetal pig tissues.     

Abnormal flow properties of white blood cells in patients with severe ischaemia of the leg

The possible role of white blood cells in tissue ischaemia has attracted recent interest. White blood cells can block narrow vessels, particularly if perfusion pressure is reduced or if the cells become activated. To investigate the role of white blood cells in ischaemia microfiltration techniques were used to measure the flow properties of these cells in patients with severe leg ischaemia before and after amputation. Compared with controls white blood cells from patients showed impaired ability to flow through 8 μm and 5 μm pore filters. This applied to fractionated granulocytes and mononuclear cells as well as to unfractionated mixed white blood cells. White blood cells from blood draining the ischaemic leg had worse filterability than those from arm blood of the same patients. After amputation of the ischaemic leg there was definite improvement, the flow properties of the cells being no longer significantly different from controls.These abnormalities detected in white blood cells probably reflect activation of the cells by factors released in the ischaemic tissue. As activation alters the mechanical and adhesive properties of white blood cells, a vicious cycle of microcirculatory trapping at low perfusion pressure, activation, tissue damage, and further activation and trapping might contribute to the progressive worsening of tissue ischaemia.     

体外培养牛白细胞中环形泰勒焦虫(Theileria annulata)的增殖、释放与感染

在体外培养的牛外周血白细胞中,环形泰勒焦虫裂殖子与裂殖体寄生于宿主细胞的细胞质中,并且随着宿主细胞的分裂而分到两个子细胞中。焦虫染色质粒的分裂方式为二分裂,随着焦虫颗粒的不断增殖,逐渐发育为成熟的裂殖体。体外培养感染焦虫的牛白细胞可通过伪足与细胞裂解两种途径向培养液中释放焦虫颗粒。释放到培养液中的焦虫颗粒对体外培养的健康牛外周血白细胞具有感染能力,感染细胞能在体外连续传代培养。     

Effect of the interferon inducer tilorone in inbred CBA mice.

Tilorone hydrochloride (200 mg/kg) was administered intragastrically to inbred CBA mice. After 5, 18 and 48 hr the number of circulating leucocytes and peritoneal cells as well as the migration of unstimulated peritoneal cells, the blood corticosteroid level and interferon production were investigated. In spite of the considerable decrease of the number of mononuclear cells in the blood and polynuclear ones in the peritoneal exudate, the drug induced production of circulating interferon and stimulated its synthesis by peritoneal cells. The blood corticosteroid level and the mast cell count in the peritoneal cavity were significantly elevated, but the migration of peritoneal cells in antigen-free medium decreased.     

Erythrocyte senescence and haematological changes induced by starvation in the neotropical fish traíra, Hoplias malabaricus (Characiformes, Erythrinidae)

Adult specimens of traira (Hoplias malabaricus Bloch) were subjected to long-term starvation (30 to 240 days) and re-fed for 30 days after 90 and 240 days of food deprivation. Counting of immature erythrocytes in peripheral blood showed that erythropoiesis decreased significantly during the first 30 days of food deprivation. The results suggest that a process of senescence takes place in the pre-existent red blood cells and that the cells are not replaced during starvation. After 240 days of starvation, H. malabaricus had a significantly reduced number of red blood cells, causing changes in hematocrit and blood indices (mean corpuscular volume, mean corpuscular haemoglobin and mean corpuscular haemoglobin concentration). Furthermore, during this period, the fish presented leukopenia (lymphocytopenia) and thrombocytopenia. After re-feeding, the number of leukocytes and thrombocytes recovered, but the red blood cell number remained reduced and there was a significant increase in abnormal red cell nuclei.     

Fetal cells in maternal blood: recovery by charge flow separation

Fetal blood cells can be recovered from the maternal circulation by charge flow separation (CFS), a method that obviates the risks associated with amniocentesis and chorionic villus sampling. By CFS, we processed blood samples from 13 women carrying male fetuses, 2 carrying fetuses with trisomy 21, and 1 who had delivered a stillborn infant with trisomy 18. On average more than 2000 fetal nucleated red blood cells were recovered per 20-ml sample of maternal blood. Recovery of fetal cells was confirmed by fluorescence in situ hybridization with probes for chromosomes Y, 18 and 21. After culturing of CFS-processed cells, amplification by the polymerase chain reaction revealed Y-chromosomal DNA in clones from four of six women bearing male fetuses, but not in clones from three women bearing female fetuses. Received: 8 January 1996 / Revised: 22 March 1996     

Changes in T-cell subpopulations in mice during prolonged experimental secondary infection withEchinococcus granulosus

Balb/c mice were infected intraperitoneally with protoscoleces ofEchinococcus granulosus. After 15 months of infection, and by means of flow cytometry, the expression of T-cell markers CD3, CD4, and CDS on T cells from peripheral blood, spleen, and thymus was analyzed and compared with that of age-matched controls. Infected mice had higher percentages of CD3+, and CD4+ cells in peripheral blood, and higher percentages of CD8+ cells in the spleen, when compared with control mice. CD4+ and CD8+ cells in peripheral blood and CD8+ cells in thymus also showed higher percentages of expression of interleukin-2 receptor. The results infer a role for interleukin-2 in experimental secondary echinococcosis.     

Effects of 4-nonylphenol on blood cells of the African catfish Clarias gariepinus (Burchell, 1822)

In the present work, the destructive effects of the 4-nonylphenol on one of the most economically important Nile fishes, namely African catfish (Clarias gariepinus) were studied. Apoptosis, erythrocytes alterations, micronucleus test and blood parameters count were used as biological indicators to detect those effects. After exposure to sublethal concentrations of 4-nonylphenol (0, 0.05, 0.08 and 0.1 mg/l), apoptotic red blood cells with many malformations and micronucleated erythrocytes were recorded. Decrease in the blood parameters such as red blood cells (RBCs), hemoglobin (Hb), package cell volume (PCV), mean corpuscular hemoglobin concentration (MCHC), platelets, white blood cells (WBCs), lymphocytes, basophils, monocytes and increase in mean corpuscular volume (MCV), mean corpuscular hemoglobin (MCH), neutrophils, eosinophils indicated the negative effects of 4-nonylphenol. It was concluded that, the 4-nonylphenol caused genotoxicity in erythrocytes with many malformations in shape and number indicated with other blood parameters.     

Tissue engineering of blood vessels with endothelial cells differentiated from mouse embryonic stem cells

Endothelial cells (TEC3 cells) derived from mouse embryonic stem (ES) cells were used as seed cells to construct blood vessels. Tissue engineered blood vessels were made by seeding 8 X 106 smooth muscle cells (SMCs) ob-tained from rabbit arteries onto a sheet of nonwoven polyglycolic acid (PGA) fibers, which was used as a biode-gradable polymer scaffold. After being cultured in DMEM medium for 7 days in vitro, SMCs grew well on the PGA fibers, and the cell-PGA sheet was then wrapped around a silicon tube, and implanted subcutaneously into nude mice. After 6～8 weeks, the silicon tube was replaced with another silicon tube in smaller diameter, and then the TEC3 cells (endothelial cells differentiated from mouse ES cells) were injected inside the engineered vessel tube as the test group. In the control group only culture medium was injected. Five days later, the engineered vessels were harvested for gross observation, histological and immunohistochemical analysis. The preliminary results demonstrated that the SMC-PGA construct could form a tubular structure in 6-8 weeks and PGA fibers were completely degraded. Histological and immunohistochemical analysis of the newly formed tissue revealed a typical blood vessel structure, including a lining of endothelial cells (ECs) on the lumimal surface and the presence of SMC and collagen in the wall. No EC lining was found in the tubes of control group. Therefore, the ECs differentiated from mouse ES cells can serve as seed cells for endothelium lining in tissue engineered blood vessels.     

Activation and migration of allo-peptide specific TCR transgenic T cells in cardiac allograft rejection

T cells from TCR transgenic mice, expressing receptors specific for an allogeneic MHC class I peptide, were used to track T cell activation and migration in normal adoptive recipients that were subsequently transplanted with heterotopic hearts that were syngeneic except for a transgenic MHC class I antigen. T cells rapidly disappeared from the blood into the lymphoid tissues where they were activated within one day after transplantation. T cells initially formed discrete clusters in the spleen and lymph nodes. After proliferating for 2-4 days in lymphoid tissues, T cells reappeared in the blood and migrated to the heart and the intestines. The T cells underwent another round of proliferation in the heart, but not the intestines, and induced cardiac rejection uniformly on 6 day.     

Neutrophil aggregation is beta 2-integrin- and L-selectin-dependent in blood and isolated cells.

Neutrophil aggregation in response to formyl peptide was analyzed in blood and isolated cells by fluorescence flow cytometry. The isolated leukocyte aggregates and the leukocytes in blood were identified with the vital nucleic acid stain LDS-751. This method enabled us to discriminate nucleated cells from other blood cells and to detect granulocyte aggregates without isolation or E lysis. Cells isolated in the absence of endotoxin retained the characteristics of cells in blood and exhibited similar aggregation kinetics and dose-response to formyl peptide. We show that it is possible to analyze epitope expression in blood with homogeneous flow cytometric assays and that carefully isolated neutrophils retain the expression characteristics of those in blood. The expression of CD18 was at its lowest levels in unstimulated cells, while the rate of formyl peptide stimulated aggregation was most rapid in these cells. Aggregation in isolated cells as well as blood preceded an increase in receptor expression. After stimulation, L-selectin expression decreased in both blood and isolated cells over a time frame similar to disaggregation. The aggregation response in blood was blocked by pretreatment with antibody to CD18 over a concentration range consistent with the amount of antibody bound. Aggregation was also blocked in isolated cells and blood by antibodies DREG-200 and DREG-56 to L-selectin, but not by isotype controls or anti-LFA-1. The results are discussed in terms of the roles of adhesive receptor expression and recognition in neutrophil aggregation. The methods validated here permit linkage between isolated cells and in vivo studies.     

蝮蛇抗栓酶对高血压病者甲襞微循环的影响(附50例分析)

用蝮蛇抗栓酶治疗高血压病５０例,用药前后对比甲襞微循环管袢轮廊、管袢口径、管袢内红细胞聚集等指标均有非常显著性差异。治疗后患者血压明显下降     

Separation of proximal tubule cells from suspensions of rat kidney cells by free-flow electrophoresis

Suspensions of rat kidney cells obtained by disaggregation of the kidney with 0.25% trypsin were separated by electrophoresis. Previously, we found a correlation between cells with histochemically demonstrable alkaline phosphatase (HDAP) and cells with brush borders which established that HDAP is a useful marker for rat proximal tubule cells (Kreisberg et al., '77). The starting suspension of cells for electrophoresis consisted of 38.4 +/- 5.7% nucleated cells with HDAP, 39.8 +/- 5.7% nucleated cells without HDAP, and 20.8 +/- 9.2% red blood cells. After electrophoresis, the purest fraction contained 85.8 +/- 3.5% nucleated cells with HDAP, 8.4 +/- 2.2% nucleated cells lacking HDAP, and 5.8 +/- 2.8% red blood cells; 91.9 +/- 2.4% of the nucleated cells in the purest fractions had HDAP.     

X-ray induction of micronuclei in human lymphocyte subpopulations differentiated by immunoperoxidase staining.

In this study we sought to confirm the radiosensitivity of human peripheral blood lymphocyte subpopulations using a micronucleus assay. Mononucleated cells isolated from peripheral blood were irradiated with X rays. After being cultured for 3 days, cells were fixed and stained using the immunoperoxidase staining technique. Lymphocyte subpopulations were characterized by means of the monoclonal antibodies Leu4 (CD3), Leu2a (CD8) and Leu19 (CD56). Dose-response curves were obtained by scoring the number of micronuclei in binucleated cells that reacted with a specific antibody and were then stained. The dose response of CD8+ (suppressor/cytotoxic) cells was quite similar to that of CD3+ (pan T) cells. In comparison, CD56+ (natural killer) cells were significantly less sensitive, although scorable binucleated CD56+ cells made up less than 4% of the total number of binucleated cells.     

脐血干细胞移植治疗中间型脊髓性肌萎缩症1 例并文献复习

目的:探讨脐血干细胞移植治疗中间型脊髓性肌萎缩症的临床治疗可行性及效果。方法:已确诊的中间型脊髓性肌萎缩症患儿,采用脐血干细胞移植治疗,4次为一疗程,移植途径采用静脉输注(1次)加蛛网膜下腔注射(3次)的方法,治疗前和治疗后半年均需完善神经系统体检、实验室检查、FIM评分、肌电图等。结果:移植治疗后患儿神经系统症状明显改善,FIM评分提高,实验室检查肌酶下降,肌电图提示重收缩每10.0ms所检肌运动单位较前增加。随访10月患儿未出现副反应。结论:应用脐血干细胞移植治疗中间型脊髓性肌萎缩症是有效安全的,可以改善患儿神经功能。