Stem cell, T- and B-lymphocyte migration in mouse lines that are high and low reactors to sheep erythrocytes]

Migration of stem cells and B-lymphocytes from the bone marrow and of T-lymphocytes from the thymus was studied on special models in mice of the CBA and C57BL lines, responding to sheep erythrocytes oppositely. Genetically-determined differences in the height of the immune response between the CBA and C57BL mice in immunization with sheep erythrocytes depended to a certain extent on different expression of the process of intensification of migration of the stem cells, T- and B-lymphocytes in response to the antigen administration.     

Changes in the migratory activity of polymorphonuclear leukocytes after administration of Freund’s adjuvant

Spontaneous and chemotactic activity of polymorphonuclear leukocytes, obtained from the peritoneal cavity, was studied after administration of complete and incomplete Freund's adjuvant to rabbits at intervals of 18-26 d after adjuvant injection. Whereas injection of both complete and incomplete adjuvant increased spontaneous migration of rabbit polymorphonuclear leukocytes, the migration directed by the chemotactic signal, induced in fresh serum by E. coli endotoxin, was decreased. The chemotaxigenic activity of two factors, isolated from cell walls of Listeria monocytogenes was also tested: factor Ei (its most potent component is lipopolysaccharide) and a purified phenol extract of lipopeptidopolysaccharide nature.     

Stem cells and T- and B-lymphocytes in acute hypoxia

During the stepped rapid training of mice for hypoxia the number of colony-forming units in the blood and bone marrow increases and that in the spleen falls down. In acute hypoxic hypoxia there is an enhancement of the migration of stem hemopoietic cells and B-lymphocytes from the bone marrow and T-lymphocytes from the thymus.     

The T-cell receptor: just another hypothesis.

Functional activities of T and B lymphocytes and the kinetics of hematopoietic stem cells were studied in mice with inoculated or spontaneous tumors. The development and growth of the tumor inhibited B cells and helper T cells, while the activity of killer T cells and spleen suppressor cells was markedly enhanced. The processes of stem cell migration from the bone marrow were considerably intensified and altered in tumor-bearing mice. Data were obtained suggesting that helper T cells and killer T cells represent nonidentical compartments within the population of thymus-dependent lymphocytes. Immunosuppression during tumor bearing is probably due to an impairment of T lymphocytes cooperating in immune responses, B-lymphocytes and their precursors.     

Induction of delayed hypersensitivity in guinea pigs by staphylococcal cell wall components

The subcutaneous injection of 100 micrograms of staphylococcal cell-wall peptidoglycan in Freund's incomplete adjuvant leads to the development of sensitization which can be detected on days 7-35 by the splenocyte migration inhibition test. Sensitization can be detected with peptidoglycan, as well as with staphylococcal cell walls. Teichoic acids produce no such effect. The sensitization of guinea pigs with whole staphylococcal cells introduced in three injections also leads to the development of delayed hypersensitivity specific to staphylococcal cell walls, peptidoglycan, but not teichoic acids.     

Determinant specific suppression of antigen-induced T cell proliferation in the guinea pig. I. Quantitation of suppressed antigen-specific T cell responses as a consequence of prior exposure to antigen in incomplete Freund's adjuvant

We have asked whether a correlation exists between T cell proliferation and the in vivo suppression of delayed type hypersensitivity observed after administration of antigen in incomplete Freund's adjuvant before exposure to antigen in complete Freund's adjuvant. We find that in vivo suppression is indeed paralleled by diminished in vitro responsiveness to the immunogen. Suppression of T cell proliferation is antigen-specific, dependent upon prior immunization of antigen in IFA, and can be transferred adaptively into unprimed but not primed animals by lymphoid cells from actively suppressed syngeneic donors.     

Semi-synthetic salinomycin analogs exert cytotoxic activity against human colorectal cancer stem cells

Salinomycin, a polyether antibiotic, is a well-known inhibitor of human cancer stem cells. Chemical modification of the allylic C20 hydroxyl of salinomycin has enabled access to synthetic analogs that display increased cytotoxic activity compared to the native structure. The aim of this study was to investigate the activity of a cohort of C20-O-acyl analogs of salinomycin on human colorectal cancer cell lines in vitro. Two human colorectal cancer cell lines (SW480 and SW620) were exposed to three C20-O-acylated analogs and salinomycin. The impact of salinomycin and its analogs on tumor cell number, migration, cell death, and cancer stem cell specifity was analyzed. Exposure of human colorectal cancer cells to the C20-O-acylated analogs of salinomycin resulted in reduced tumor cell number and impaired tumor cell migration at lower concentrations than salinomycin. When used at higher (micromolar) concentrations, these effects were accompanied by induction of apoptotic cell death. Salinomycin analogs further expose improved activity against cancer stem cells compared to salinomycin.     

Lipopeptides: adjuvanticity in conventional and genetic immunization

Synthetic lipopeptides derived from the bacterial cell wall component lipoprotein activate B-lymphocytes and macrophages/monocytes in vitro. In vivo they constitute potent immunoadjuvants for a broad range of different antigens and species comparable or superior to Freund's adjuvant. Here, we demonstrate that P(3)CSK(4), representing a highly active lipopentapeptide derivative in vitro, significantly enhances and accelerates the humoral immune response to tetanus toxoid. P(3)CSK(4) could substitute for up to 90% of the antigen without any decrease in the specific IgG level, and the presence of the lipopeptide resulted in a prolonged production of specific IgG in time. Investigations using P(3)CSK(4) as an adjuvant in genetic immunization confirmed earlier data demonstrating that lipopeptides constitute adjuvants for low-immunogenic DNA constructs and/or for application routes resulting in weak immune responses. We monitored a lipopeptide-dependent shift from a Th1-type to Th2-type response, when DNA immunization was followed by i.p. administration of the lipopeptide adjuvant.     

Hapten-specific T cell response to azobenzenearsonate-N-acetyl-L-tyrosine in the Lewis rat. I. Induction and suppression of delayed-type hypersensitivity and in vitro proliferative responses

Pretreatment of Lewis rats with a single i.p. injection of ABA-N-acetyl-tyrosine in incomplete Freund's adjuvant induced an unresponsiveness for delayed-type hypersensitivity to subsequent immunization with the same antigen in complete Freund's adjuvant. Complete suppression of in vitro antigen-induced proliferative responses required repeated pretreatment. Passive transfer of lymphoid cells from spleen and lymph nodes but not sera from suppressed rats induced unresponsiveness of hapten-specific T cell functions. Nylon wool-nonadherent cells and cells panned on F(ab')2 of rabbit anti-Lewis rat Ig plates suppressed the induction of DTH and in vitro antigen-stimulated proliferation. Adult thymectomy increased DTH and failed to abolish the induction of suppression.     

Role of B-lymphocytes in inactivating allogenic stem cells]

Antisera to membrane antigens of B lumphocytes eliminated the capacity of lymphocytes to inactivate allogenous stem cells by 60%; however, lymphocytes from the lymph nodes of B mice possessed no inactivating capacity. T-lymphocytes were the main criteria inactivating allogenous stem cells. Cooperating with T-lymphocytes, B-lymphocytes probably contributed to inactivation of precursor cells realized by T-lymphocytes. However, the presence of B-lymphocytes in the killer cells population was not a determinant, since T-lymphocytes were capable of inactivating allogenous stem cells without any participation of B-lymphocytes.     

Cellular immune reactions and blood cerebrospinal fluid barrier dysfunction in guinea pigs

Both young and adult strain 13 guinea pigs have been treated with complete Freund's adjuvant (CFA). In young animals CFA induced an increase in the lymphocyte cell count in blood and a reversible blood CSF barrier impairment. Both these effects could be suppressed by the immunosuppressant drug Cyclosporin A. In contrast to the young animals treatment with CFA in adult strain 13 guinea pigs influenced neither the lymphocyte cell count nor blood CSF barrier function. In young and adult CFA-treated animals the number of polymorphonuclear cells in blood were increased and this increase was not reversed by Cyclosporin A. We discuss the influence of a systemic immune stimulation and suppression on blood CSF barrier function for proteins and its relevance to the pathogenesis of inflammatory diseases of the central nervous system.     

Stimulation of the in vivo dinitrophenyl antibody response to the DNP conjugate of L-glutamic acid60-L-alanine30-L-Tyrosine10 (GAT) polymer by a synthetic adjuvant, muramyl dipeptide (MDP): target cells for adjuvant activity and isotypic pattern of MDP-stimulated response

Specific anti-dinitrophenyl (DNP) response to DNP-conjugated L-glutamine60-L-alanine30-L-tyrosine10 (DNP-GAT) was obtained in GAT-responder mice by using synthetic N-acetylmuramyl-L-alanyl-D-isoglutamine (MDP) as adjuvant. Significant levels of anti-DNP antibodies were observed during a secondary response to DNP-GAT, when both antigen and MDP were used for priming. In this system, MDP was able to prime the carrier-specific T cells but not the hapten specific B cells. The study of the isotypic pattern of the anti-DNP response shows that MDP stimulates only the appearance of specific anti-DNP IgG1 plaque-forming cells. Anti-DNP plaque-forming cells were stimulated in animals primed with DNP-GAT in Freund's complete adjuvant or in Maalox-pertussis and used as control IgG1, IgG2a, and IgG2b.     

肝细胞生长因子对人羊水干细胞迁移和黏附能力的影响

目的分离培养及鉴定羊水干细胞（hAFSC）,并研究肝细胞生长因子（HGF）对羊水干细胞迁移、黏附能力的影响。方法使用细胞贴壁法分离培养羊水干细胞,细胞免疫荧光及westernblot鉴定羊水干细胞,Transwell小室分析HGF对羊水干细胞迁移的作用。明胶贴壁法分析HGF对羊水干细胞黏附能力的作用。两组之间数据的比较采用独立样本t检验。结果分离的羊水干细胞均表达特异性标记物Oct-4、c-kit、SSEA-4、CD105。HGF在体外对hAFSC的迁移有趋化作用,对照组和HGF组每个视野的迁移细胞数分别为38±2.5和80±3.2。对黏附能力有促进作用,对照组和HGF组每个视野的黏附细胞数分别为19±1.5和50±2.7,差异均有统计学意义（P〈0.01）。结论 HGF可趋化羊水干细胞的迁移,增强羊水干细胞的黏附能力。     

Immunoregulation in experimental interstitial nephritis: immunization with renal tubular antigen in incomplete Freund's adjuvant induces major histocompatibility complex-restricted, OX8+ suppressor T cells which are antigen-specific and inhibit the expression of disease

We utilized a model of experimental interstitial nephritis induced by renal tubular antigen in complete Freund's adjuvant to examine a mechanism of immunologic tolerance produced by priming immunization with tubular antigen in incomplete Freund's adjuvant. Brown Norway rats primed with tubular antigen in incomplete adjuvant do not develop significant nephritis after challenge with antigen in complete adjuvant, and this tolerance can be transferred to naive recipients with donor T cells. These T cells also specifically suppress a delayed-type hypersensitivity response to soluble tubular antigen in recipients immunized to produce disease. This suppression is MHC-restricted and is mediated by OX8+ T cells which bind antigen and bear idiotypes cross-reactive with those on antibodies eluted from the tubular basement membrane. Despite the suppression of histologic disease, tolerized animals were able to produce significant titers of antibodies to tubular basement membrane. Our findings demonstrate an additional strategy for altering the natural history of immune-mediated renal disease, and further refine the characterization of the suppressive effect produced by incomplete Freund's adjuvant.     

Some electron microscopy characteristics of lung immune response in the rats stimulated by respiratory and parenteral ways with pneumococcal suspensions

Conventional albino rats of either sexes, weighing about 100 grams, were stimulated, one group by inhalation of pulverized suspensions of living type III pneumococci, in 6 exposures, and the second group, by 5 intramusculary injections, one a week, of 0,1 ml, emulision of killed germs, in complete Freund's adjuvant submitted after a week free interval to the inhalation of aerosolized living pneumococci. The tissue fragments sampled at two and respective five weeks after the beginning of the experiment, from central and peripherical sites of lungs, inclusively from a control group, were fixed in glutaraldehide-osmic acid and prepared for electron-microscopical examination. In the thickened, densely cellular alveolar walls there are numerous mature plasma cells, intermediate elements between reticular septal cells and plasmablast and also histiocytic cells which phagocytic inclusions and shape suggests their movement in the interalveolar septa. The swollen free alveolar macrophages contain an increased number of lysosomes and ribosomes. The hyperplastic granular pneumocytes possess enlarged cytosomes excreting osmiophilic material in the air spaces. There are discussed the problems of local plasma cell differentiation, alveolar macrophage physiology and transparietal migration of histiocytes.     

Adjuvant regulation of T cell function.

The effect of complete Freund's adjuvant (CFA) on distinct T cell functions was investigated. Adjuvant was found to suppress the generation of cytolytic T cells in vivo when mixed with allogeneic P815 cells before immunization of C57BL/6 mice. Inoculation of the mice with either adjuvant or adjuvant emulsified with allogeneic cells resulted in whole splenic populations or immunoabsorbent-purified T cells that did not generate cytolytic activity in vitro against allogeneic cells. Mixing T cells from normal and adjuvant-treated mice before in vitro sensitization resulted in suppression of lytic activity. However, memory T cells were not subject to the same suppressive regulation as were precytotoxic T cells since adjuvant had no effect on subsequent boosting of memory.     

Different effect of testosterone on polypotential stem hematopoietic stem cells and immunocompetent B-lymphocytes]

A research was made to study the dynamics of the proliferative, colony-forming and migration capacity of stem hemopoietic cells in (CBA X C57Bl) F1 hybrid mice under the influence of testosterone propionate, 10 mg/100 g, as well as the migration of immunocompetent B lymphocytes from the bone marrow to the spleen and the accumlation of their progeny, antibody-producing cells, in the spleen. The immunodepressive effect of testosterone was manifested by a decrease in the migration of B cells and the number of antibody-producing cells in the spleen. On the contrary, testosterone had a stimulating effect on the functional activity of stem hemopoietic cells, increasing their proliferation and migration. Under conditions of the suppressed erythropoietic differentiation of multipotent stem hemopoietic cells the injection of testosterone resulted in an increase in the number of antibody-producing cells in the spleen. This suggests that the stimulation of erythropoiesis and immunosuppression, induced by testosterone, are interconnected and determined by the direct action of the hormone on the cellular cycle of the stem cells, as well as by their prevailing differentiation towards the erythroid series, resulting in the decrease of their differentiation into B cells.     

DNA content of macrophages in pulmonary granuloma induced in rats by complete Freund's adjuvant

Pulmonary granulomas were induced in rats with complete Freund's adjuvant. Histologic and cytologic alterations and Feulgen-DNA amounts determined by a cytophotometric method were studied. Fourteen (hours after injection, the pulmonary interstitia were thickened by cell infiltration. After 3 days; small granulomatous nodules developed containing macrophages, epithelioid and giant cells. The Feulgen-DNA amounts increased significantly in the course of the first day and reached peak values on the 3rd day. Then they showed a tendency to decrease after the 7th day. The present results confirm the previously obtained data, revealing the stimulating effect of the complete Freund's adjuvant on cell metabolism, DNA synthesis included.     

Effect of in vivo administration of Tetrahymena pyriformis on the in vitro toxoplasmacidal activity of mouse peritoneal macrophages

Tetrahymena pyriformis is a free-living ciliate protozoan in the freshwater system. Experiments were carried out to determine whether intraperitoneal administration of T. pyriformis (GL strain) to mice activates macrophages to be able to kill Toxoplasma gondii tachyzoites in vitro. Mice were also injected intraperitoneally with several synthetic activators; dimethyldioctadecylammonium bromide (DDA), dextran sulfate, complete Freund's adjuvant (CFA) as well as Toxoplasma and Tetrahymena lysates in order to activate mouse peritoneal macrophages. One week after the administration of activators, peritoneal cells were harvested and the adherent macrophages were challenged with Toxoplasma tachyzoites. Macrophage monolayers were then fixed with absolute methanol after washing, and stained with Giemsa solution. The percentage of the adherent cells infected and total number of organisms per 100 macrophages were calculated to make toxoplasmacidal activity of macrophages according to the cultivation time. Peritoneal macrophages from mice administered with Tetrahymena exhibited significant protection against target parasites as compared with those treated with synthetic activators. Among non-biological synthetic activators, DDA was evaluated as an excellent activator.     

Increased efficiency of immunotherapy using irradiated tumor cells

Summary The efficacy of irradiated tumor cells combined with chemotherapy or non-specific immunostimulation with complete Freund's adjuvant was tested in a model of minimal residual tumor-bearing syngeneic mice. Male C57BL/6J mice were innoculated in the right rear leg with live tumor cells from a methylcholanthrene induced fibrosarcoma. The tumor was resected when it reached 0.7 cm in diameter and animals were treated with doses of irradiated tumor cells (XTC) from the primary tumor ranging in number from 1×10³ to 9×10³. Best survival was noted using 5×10³ XTC combined with irradiated tumor cells of liver or pulmonary metastases origin, complete Freund's adjuvant or cytoxan. The combination of irradiated tumor cells of metastatic origin did not enhance the therapeutic effect of XTC alone. Freund's adjuvant was not of benefit in enhancing the efficacy of XTC. However, improved survival was noted when chemotherapy in the form of cytoxan was used to supplement XTC. Our data suggests that XTC is more efficacious as a mode of immunotherapy than are live tumor cells. The dose of XTC used is critical in determining its effect. Chemotherapy appears to enhance the benefit of XTC.Research fellow from Zhejiang Medical University, Hongzhou, ChinaSupported by the Brigham Surgical Group, Inc.