BACKGROUND: Nanoparticles (NPs) prepared from biodegradable polymers, such as poly (D,L-lactic acid-co-glycolic acid) (PLGA), have been studied as vehicles for the delivery of antigens to phagocytes. This paper describes the preparation of antigen-loaded PLGA-NPs for efficient cross-priming. METHODS: NPs containing a similar amount of ovalbumin (OVA) but different sizes were produced using a micromixer-based W/O/W solvent evaporation procedure, and the efficiency of the NPs to induce the cross-presentation of OVA peptides were examined in dendritic cells (DCs). Cellular uptake and biodistribution studies were performed using fluorescein isothiocyanate (FITC)-loaded NPs in mice. RESULTS: The NPs in the range of 1.1~1.4microm in size were the most and almost equally efficient in inducing the cross-presentation of OVA peptides via H-2Kb molecules. Cellular uptake and biodistribution studies showed that opsonization of the NPs with mouse IgG greatly increased the percentage of FITC-positive cells in the spleen and lymph nodes. The major cell type of FITC-positive cells in the spleen was macrophages, whereas that of lymph nodes was DCs. CONCLUSION: These results show that IgG-opsonized PLGA-NPs with a mean size of 1.1microm would be the choice of biodegradable carriers for the targeted-delivery of protein antigens for cross-priming in vivo.
Animals ; Cross-Priming ; Dendritic Cells ; Fluorescein ; Immunoglobulin G ; Isothiocyanates ; Lactic Acid ; Lymph Nodes ; Macrophages ; Mice ; Nanoparticles ; Ovalbumin ; Ovum ; Peptides ; Phagocytes ; Polyglycolic Acid ; Polymers ; Spleen

OBJECTIVES: To quantitate apoptosis and Fas antigen expression of T lymphocytes by activation in aplastic anemia (AA) and compare with that of normal controls and completely-recovered AA, and to investigate the apoptotic sensitivity to anti-fas antibody of activated T lymphocytes in AA. METHODS: We studied the expression of Fas antigen on fresh T lymphocytes of twenty patients with AA [13 newly diagnosed, 7 recorvered AA after immunosuppressive therapy (IST)], and investigated the activation-induced cell death (AICD) and Fas expression by activation [interleukin-2 (200 U/ml) and phytohemagglutinin (50 micrograms/ml)] in 5 newly-diagnosed AA, 5 normal controls and 5 AA in complete response (CR). Apoptotic sensitivity to anti-Fas antibody was assessed by the time-course kinetics of induction of cell death by anti-Fas antibody (500 ng/ml). RESULTS: There was no significant difference of Fas antigen expression on freshly-isolated T lymphocytes among newly-diagnosed severe AA, normal control s and patients with AA in CR after IST. In normal controls, T lymphocytes death was greatly increased at 3 days of activation, and Fas antigen expression on T lymphocytes was increased above baseline at day 1 of activation. In contrast, in newly-diagnosed AA, T lymphocytes showed delayed cell death, which correlated with a slowed increase of Fas antigen expression by activation. Also, anti-Fa s antibody sensitivity of activated T lymphocytes was decreased in newly-diagnosed AA. In completely recovered AA, these abnormal AICD and Fas antigen expressions by activation were recovered to normal range. CONCLUSIONS: Abnormal AICD plays a role in the immune pathophysiology of AA, and defective Fas system is involved in this process.
Anemia, Aplastic/pathology ; Anemia, Aplastic/immunology* ; Antigens, CD95/blood ; Apoptosis ; Case-Control Studies ; Human ; In Vitro ; Lymphocyte Transformation ; T-Lymphocytes/pathology* ; T-Lymphocytes/immunology* ; Time Factors

BACKGROUND: EY-6 is one of the newly synthesized indoledione derivatives to induce tumor cell-specific cell death. In this study, we investigated the mechanism of immunological death induced by EY-6 at mouse colon cancer cell as well as at the normal immune cell represented by dendritic cell. METHODS: C57BL/6 mouse syngeneic colon cancer cell MC38 was treated with EY-6, and analyzed by MTT for viability test, flow cytometry for confirming surface expressing molecules and ELISA for detection of cytokine secretion. Normal myeloid-dendritic cell (DC) was ex vivo cultured from bone marrow hematopoietic stem cells of C57BL/6 mice with GM-CSF and IL-4 to analyze the DC uptake of dead tumor cells and to observe the effect of EY-6 on the normal DC. RESULTS: EY-6 killed the MC38 tumor cells in a dose dependent manner (25, 50 and 100 microM) with carleticulin induction. And EY-6 induced the secretion of IFN-gamma but not of TNF-alpha from the MC38 tumor cells. EY-6 did not kill the ex-vivo cultured DCs at the dose killing tumor cells and did slightly but not significantly induced the DC maturation. The OVA-specific cross-presentation ability of DC was not induced by chemical treatment (both MHC II and MHC I-restricted antigen presentation). CONCLUSION: Data indicate that the EY-6 induced tumor cell specific and immunological cell death by modulation of tumor cell phenotype and cytokine secretion favoring induction of specific immunity eliminating tumor cells.
Animals ; Bone Marrow ; Cell Death ; Colonic Neoplasms ; Cross-Priming ; Dendritic Cells ; Enzyme-Linked Immunosorbent Assay ; Flow Cytometry ; Granulocyte-Macrophage Colony-Stimulating Factor ; Hematopoietic Stem Cells ; Homicide ; Interleukin-4 ; Mice ; Phenotype ; Tumor Necrosis Factor-alpha

For those with allergy, vaccination with a specific allergen has often been used as a major therapeutic measure. However, the universal application of this technique in clinics have been restricted due to its low success rates and the risk of active systemic anaphylactic shock (ASAS). In this regard, we constructed a fusion protein (OVA-DT), ovalbumin (OVA) fused with diphtheria toxin protein (DT), which may exert a specific cytotoxicity to cells bearing OVA-specific IgE. Its therapeutic effect was evaluated in mice (BALB/c) sensitized with OVA (Os-mice). OVA challenges to the OVA-sensitized mice (Os-mice) caused ASAS to death within 30 min, but OVA-DT treatment afforded mice complete protection. When OVA-DT was treated to the Os-mice, none showed the signs of ASAS when re-challenged 48 h after the treatment. OVA-DT itself was not found to be toxic or allergenic in normal mice. The effect of OVA-DT on the biological functions of mast cells was also studied. Binding of OVA-DT to OVA-specific IgE bearing mast cells and the inhibition of histamine release from these cells were observed. In addition, OVA-DT treatment inhibited the proliferation of OVA-specific B cells in mice. In Os-mice treated with OVA-DT, levels of anti-OVA IgG2a in serum and the production of IFN-gamma by splenic lymphocytes were found to increase, but the production of IL-4 by these cells decreased. Re-direction of cytokine profiles from OVA-specific Th2 to OVA-specific Thl is suggested. These results indicate that OVA-DT can protect Os-mice from ASAS due to OVA challenge, because it inactivates OVA-specific IgE-expressing cells, including mast cells and B cells.
Anaphylaxis/prevention | control* ; Animal ; B-Lymphocytes/immunology ; Female ; Histamine Release/drug effects ; IgE/metabolism ; Interferon Type II/biosynthesis ; Interleukin-4/biosynthesis ; Lymphocyte Transformation/drug effects ; Mast Cells/metabolism ; Mice ; Mice, Inbred BALB C ; Ovalbumin/immunology* ; Recombinant Fusion Proteins/therapeutic use*

This study is aimed to investigate the activity of lymphocytes of recurrent spontaneous aborters, their response to phytohemagglutinin (PHA) mitogen was studied by comparing with that of normal pregnancy and normal non-pregnancy women. The results are as follows: there is no or a significant decrease of suppression of lymphoblast transformation in recurrent spontaneous abortion comparing to that of normal pregnancy women. It should be considered all of three parameters: absolute counts of OD with PHA, non-PHA and stimulation index as a general analysis for lymphoblast transformation.
Lymphocyte Activation ; Abortion, Spontaneous

The prevalence of peanut allergy in Korea is lower than in America. Peanut extract allergens between the two countries have not been standardized. This study was performed to compare the allergenicity of raw Korean and American peanuts with that of roasted peanuts. We prepared the peanut extracts in Korean raw (KP) and roasted peanuts (KRP), and also in American raw (AP) and roasted (ARP) peanuts. We compared the peanut extract allergens of KP, KRP, AP and ARP in vitro with sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) followed by immunoblotting, T-cell proliferation assay and skin prick test with sera from peanut-allergic patients. SDS-PAGE and Western blotting demonstrated four allergenic extracts, numerous bands that displayed a high prevalence of IgE binding. IgE-binding bands were at 64, 36 and 17 kDa. Western blot inhibition revealed that either KP or AP could almost completely inhibit the reactivity of the other extract. There were no differences between T-cell proliferation assay and skin prick test. In conclusion, this investigation showed no different allergic components in both raw and roast extracts of Korean and American peanuts.
Allergens/immunology ; Allergens/analysis+ACo- ; Allergens/adverse effects ; Blotting, Western ; Cells, Cultured ; Comparative Study ; Electrophoresis, Polyacrylamide Gel ; Food Hypersensitivity/immunology ; Heat ; Human ; Hybridization ; Korea ; Lymphocyte Transformation ; North America ; Peanuts/immunology ; Peanuts/classification ; Peanuts/chemistry+ACo- ; Plant Extracts/immunology ; Plant Proteins/immunology ; Plant Proteins/analysis+ACo- ; Plant Proteins/adverse effects ; Skin Tests ; Species Specificity ; T-Lymphocytes/immunology ; T-Lymphocytes/drug effects

Inflammation and activation of immune cells have important roles in the pathogenesis of atherosclerosis. We analyzed the plasma levels of inflammatory markers and the degree of activation of peripheral blood monocytes and T-lymphocytes isolated from 12 unstable angina, 12 stable angina, and 12 normal subjects. In 20%-33% of patients, monocytes expressed high basal levels of IL-8, tissue factor, IL-1beta, and monocyte chemoattractant protein-1 mRNA. Furthermore, basal mRNA levels of these cytokines showed strong correlation with each other (p < 0.01 in all combination) but not with tumor necrosis factor-alpha or transforming growth factor-beta1. Plasma level of C-reactive protein was highest in the unstable angina patients (1.63+/-0.70 mg/l) and lowest in the control subjects (0.22+/-0.08 mg/l) (P = 0.03). We also observed a high correlation between C-reactive protein level and the occurrence of minor and major coronary events during 6 months of follow-up. Activation status of T-cells, assessed by the percentage of HLA-DR positive cells, was highest in the unstable angina patients (26.8+/-1.4%) compared with that in the control (14.7+/-1.2%) (P = 0.0053). Our data represent the first case showing that the circulating monocytes in angina patients are activated to a state express numerous proatherogenic cytokines. These results may help to diagnose angina patients according to the inflammatory markers and evaluate the prognosis of the disease.
Aged ; Angina Pectoris/immunology* ; Angina Pectoris/diagnosis ; Angina, Unstable/immunology* ; Angina, Unstable/diagnosis ; Biological Markers/blood ; C-Reactive Protein/analysis ; Cytokines/blood* ; Female ; HLA-DR Antigens/immunology ; Human ; Interleukins/blood ; Lymphocyte Transformation ; Male ; Middle Age ; Monocyte Chemoattractant Protein-1/blood ; Monocytes/metabolism* ; RNA, Messenger/metabolism ; T-Lymphocytes/metabolism* ; Transforming Growth Factor beta/analysis ; Tumor Necrosis Factor/analysis

OBJECTIVETo analyze the clonal relationship between transformed and non-transformed components in mucosa-associated lymphoid tissue (MALT) lymphoma.

METHODSSix cases of MALT lymphoma with high-grade transformation were studied. Immunohistochemical study was carried out by EliVision using bcl-10 antibody. Reverse transcription-polymerase chain reaction was used to detect the presence of API2-MALT1 fusion gene transcripts. The target cells were selected by laser microdissection and studied by polymerase chain reaction and sequence analysis for rearrangement of immunoglobulin heavy chain gene.

RESULTSIn the 6 cases of MALT lymphoma with high-grade transformation, nuclear and cytoplasmic expression of bcl-10 was demonstrated in 1 case, while API2-MALT1 fusion gene was present in 2 cases. Identical fragments of rearranged immunoglobulin heavy chain gene were detected in both transformed and non-transformed components in each of the 6 cases, except for the differences at 2 nucleotide positions in N or D regions in 2 cases.

CONCLUSIONThe tumor cells from both transformed and non-transformed components in MALT lymphoma derive from the same clone.

Adaptor Proteins, Signal Transducing ; metabolism ; Adult ; Aged ; B-Cell CLL-Lymphoma 10 Protein ; Base Sequence ; Cell Nucleus ; metabolism ; Cell Transformation, Neoplastic ; Cloning, Molecular ; Cytoplasm ; metabolism ; Female ; Gene Rearrangement, B-Lymphocyte, Heavy Chain ; genetics ; Humans ; Lymphoma, B-Cell, Marginal Zone ; genetics ; metabolism ; pathology ; Male ; Middle Aged ; Oncogene Proteins, Fusion ; metabolism ; Polymerase Chain Reaction

60 breast cancer patients in the II and III phrase were treated on the regime: Surgery with after radiotherapy at the II radio Department, K Hospital in Ha Noi. These breast cancer patients after radiotherapy, Vitexin was used within continuous 2 months and 30 patients in the control group with placebo of 50mg Vitamine C. The number of lymphocytes were significantly increased after radiotherapy. In the presence of phytohemaglutinin, Vitexin also restores the number of lumphocytes to the level before radiotherapy significantly.
Breast Neoplasms ; Lymphocyte Activation ; Diseases ; Apigenin ; Radiotherapy

Human T lymphocytes are divided into αβT cells and γδT cells on the basis of the different expressions of T cell receptors. In recent years, the studies of the regulation of T cell activation and tolerance by co-stimulatory molecules and their signaling pathways in αβT cell have made remarkable progress; however, relatively fewer investigations have been performed on γδT cells. A clearer understanding of the roles of co-stimulatory molecules and their signaling pathways in the positive/negative regulation of γδT cells at different stages will provides new insights for the treatment of viral infections, cancer, autoimmune diseases, transplant rejection, and other conditions.
Humans ; Lymphocyte Activation ; physiology ; Signal Transduction ; T-Lymphocyte Subsets ; physiology