BACKGROUNDLittle is known of the effects of hydrocortisone on cell adhesion molecules such as intercellular adhesion molecule-1 (ICAM-1) and its counterreceptors (LFA-1, Mac-1) in acute pancreatitis (AP). We investigated the effects of prior treatment with hydrocortisone on the production of ICAM-1 and its counterreceptors (LFA-1 and Mac-1) in AP of rats to clarify the effect of hydrocortisone on induced acute pancreatitis.

METHODSAcute pancreatitis was induced by infusion of 5% chenodeoxycholic acid into the pancreatic duct, followed by ligation of pancreatic duct. Before induction of acute pancreatitis, rats were treated with hydrocortisone (n = 20) or 0.9% saline (n = 20). Blood and specimens from pancreas and lung were obtained from 5 rats from each treatment euthanized at 1 hour or 3 hours, 6 hours, 12 hours. Expression of ICAM-1 was assessed by immunohistochemistry and Western blot analysis of pancreas and lungs. The expression of LFA-1 and Mac-1 on neutrophils was detected by flow cytometer. The therapeutic effect of hydrocortisone was assessed from injuries to pancreas and lung.

RESULTSICAM-1 expression in the pancreas of hydrocortisone group was significantly less than in control group at 3 hours and 6 hours. In the lungs of hydrocortisone group, ICAM-1 expression was significantly less than in control group at 3 hours, 6 hours and 12 hours. The expression of LFA-1 and Mac-1 on neutrophils in blood increased significantly in control group over hydrocortisone group. Increased expression of ICAM-1, LFA-1 and Mac-1 preceded leukocyte infiltration. Compared to untreated animals with acute pancreatitis, rats pretreated with hydrocortisone had significantly reduced histological lung injury and output of ascitic fluid.

CONCLUSIONSPrior treatment with hydrocortisone before the induction of acute pancreatitis ameliorates pulmonary injury and the output of ascitic fluid and reduces the expression of ICAM-1 and its counterreceptors (LFA-1, Mac-1) in acute pancreatitis.

Acute Disease ; Amylases ; blood ; Animals ; Hydrocortisone ; pharmacology ; therapeutic use ; Intercellular Adhesion Molecule-1 ; analysis ; Lymphocyte Function-Associated Antigen-1 ; blood ; Macrophage-1 Antigen ; blood ; Pancreatitis ; blood ; drug therapy ; Rats ; Rats, Sprague-Dawley

Prostaglandin E2(PGE2) has been implicated as an immunosuppressive agent and plasma levels of PGE2 are elevated in patients sustaining thermal injury. We examined the effect of 10(-7) M prostaglandin E2(PGE2) on human polymorphonuclear leukocytes (PMN) to determine whether it directly inhibits stimulated responses of these cells. At this concentration, PGE2 alone was incapable of stimulating PMN intracellular hydrogen peroxide production (indirectly assayed by fluorescence of 2',7'ichlorofluorescin) or expression of the PMN CD11b/CD16 surface glycoproteins. PMN incubated in the presence of the soluble stimul phorbol myristate acetate(PMA, 100 ng/ml) or recombinant human C5a(rHC5a, 10(-8) M) generated significant amounts of hydrogen peroxide, increased their CD11b expression and decreased their CD16 expression. Pre-incubation of cells with PGE2 caused significant inhibition of all the observed changes stimulated by rHC5a. In contrast, events stimulated by PMA were not affected by preincubation of cells with PGE2. We conclude that PGE2, in concentrations identical to those found in the plasma of patients with burn injuries, is capable of selectively inhibiting some stimulated events and phenotypic expression of PMN in vitro study.
Dinoprostone/*pharmacology ; Dose-Response Relationship, Drug ; Human ; Hydrogen Peroxide/metabolism ; Immunosuppressive Agents/*pharmacology ; Macrophage-1 Antigen/biosynthesis ; Neutrophils/*drug effects/immunology ; Receptors, IgG/biosynthesis ; Support, Non-U.S. Gov't ; Temperature ; Tetradecanoylphorbol Acetate/pharmacology ; Time Factors

To determine whether gamma irradiation influences phenotype and function of human dendritic cells (DC) in vitro, dendritic cells were induced from the peripheral blood mononuclear cells of multiple sclerosis patients with RPMI 1640 medium containing recombinant human GM-CSF (rhGM-CSF, 800 U/ml) and recombinant human IL-4 (rhIL-4, 500 U/ml). Phenotypic changes were monitored by light microscopy. Lipopolysaccharide at a concentration of 5 micro g/ml was added into the cultures after 6 days of growth for DC complete maturation, and the cells were cultured for another 24 hours. The harvested DC on day 7 were divided equally into several parts. One part was used as non-irradiated DC (naive DC) while the other parts were irradiated by gamma ray at a dose of 25 Gy and 30 Gy respectively. Cell surface molecules were analyzed by flow cytometry. The capability of DC to stimulated autologous T cell proliferation were determined. The results showed that gamma irradiation reduced expression of CD86, CD80 and HLA-DR molecules on dendritic cells, especially CD86 molecules. Dendritic cells effectively stimulated autologous T cells proliferation while irradiated DC in all groups showed profound decrease of capability to promote T cells proliferation. It is concluded that gamma irradiation of dendritic cells not only influenced phenotype of DC but also altered their function as stimulator cells in mixed lymphocyte reaction.
Antigens, CD ; analysis ; B7-1 Antigen ; analysis ; B7-2 Antigen ; Cell Division ; immunology ; Dendritic Cells ; drug effects ; immunology ; radiation effects ; Flow Cytometry ; Gamma Rays ; Granulocyte-Macrophage Colony-Stimulating Factor ; pharmacology ; HLA-DR Antigens ; analysis ; Humans ; Immunophenotyping ; Interleukin-4 ; pharmacology ; Membrane Glycoproteins ; analysis ; Multiple Sclerosis ; blood ; Recombinant Proteins ; pharmacology ; T-Lymphocytes ; cytology ; immunology

The present study was aimed to investigate the pathways, by which IL-27 regulates the expression of adherent molecule Mac-1, chemotactic factor receptor fMLP-R and pro-inflammatory cytokine IL-1beta in human neutrophils. Highly purified human neutrophils were isolated from peripheral blood using Ficoll-Hypaque gradients centrifugation and erythrocyte lysis. The mRNA expression of IL-27 receptor components (WSX-1/TCCR and gp130) in human neutrophils was detected by reverse transcription polymerase chain reaction (RT-PCR). After incubation with IL-27 and specific inhibitors (p38 MAPK inhibitor SB203580, PI3K inhibitor LY294002 and ERK inhibitor U0126), the mRNA levels of fMLP-R and IL-1beta were determined by real time RT-PCR, and the adherent molecule Mac-1 expression in human neutrophils was determined by flow cytometry. The IL-1beta level in culture supernatant of human neutrophils was assayed by radioimmunoassay. The results showed that IL-27 receptor components (WSX-1/TCCR and gp130) were constitutively expressed in human neutrophils. IL-27 down-regulated Mac-1 expression in human neutrophils (p<0.05). After incubation with specific inhibitors, SB203580, not LY294002 and U0126, inhibited the down-regulation of Mac-1 expression by IL-27. However, IL-27 up-regulated the mRNA expression of fMLP-R and IL-1beta, and increased the release of IL-1beta (p<0.05). Interestingly, LY294002, not SB203580 and U0126, inhibited the up-regulation of fMLP-R and IL-1beta by IL-27. It is concluded that the IL-27 may regulate the expression of Mac-1, fMLP-R and IL-1beta in human neutrophils through p38 MAPK and PI3K signal pathways.
Butadienes ; pharmacology ; Chromones ; pharmacology ; Down-Regulation ; Humans ; Imidazoles ; pharmacology ; Interleukin-1beta ; metabolism ; Interleukins ; metabolism ; Macrophage-1 Antigen ; metabolism ; Morpholines ; pharmacology ; Neutrophils ; metabolism ; Nitriles ; pharmacology ; Phosphatidylinositol 3-Kinases ; metabolism ; Pyridines ; pharmacology ; Receptors, Formyl Peptide ; metabolism ; Signal Transduction ; Up-Regulation ; p38 Mitogen-Activated Protein Kinases ; metabolism

This study was aimed to investigate and compare the anti-leukemic effect mediated by dendritic cells (DC) derived from multidrug resistant (MDR) leukemia K562/A02 cells with high expression of p-glycoprotein (P-gp) and sensitive K562 cells. Multidrug resistant K562/A02 cell line and sensitive K562 cell line from chronic myeloid leukemia (CML) were induced for differentiating to DC in complete RPMI 1640 culture medium supplemented with GM-CSF (1 000 U/ml), IL-4 (500 U/ml) and TNF-alpha (100 ng/ml) for 14 days. The morphologic features of DC were observed by means of optical microscopy and the phenotype of DC was detected by flow cytometry. T-cell stimulating activity was determined by allogeneic lymphocyte reaction (allo-MLR). Cytotoxic activity was measured by MTT assay. The results indicated that DC derived from K562/A02 cells and K562 cells similarly showed the typical morphology of dendritic cell and expressed the surface differentiation antigens and costimulatory molecules CD1a, CD83, HLA-DR, CD80 and CD86 of DC. In allo-MLR, K562/A02-DC had a higher capacity to induce lymphocyte proliferation, compared with K562-DC (P < 0.05). K562/A02-DC and K562-DC could similarly generate specific cytotoxic activity against K562/A02 cells or K562 cells respectively, but low reactivity against HL-60 cells. More importantly, the cytotoxic activity mediated by K562/A02-DC was stronger than that by K562-DC against K562/A02 cells or HL-60/VCR cells (P < 0.01, respectively). It is concluded that functional DC can be differentiated from multidrug resistant leukemia K562/A02 cells as well as sensitive K562 cells in the presence of GM-CSF, IL-4 and TNF-alpha. Especially, DC derived from K562/A02 cells can induced a p-glycoprotein specific anti-leukemic immunity.
ATP-Binding Cassette, Sub-Family B, Member 1 ; biosynthesis ; Antigens, CD ; analysis ; Antigens, CD1 ; analysis ; B7-1 Antigen ; analysis ; B7-2 Antigen ; analysis ; Cell Differentiation ; drug effects ; immunology ; Cytotoxicity, Immunologic ; Dendritic Cells ; cytology ; immunology ; metabolism ; Doxorubicin ; pharmacology ; Drug Resistance, Multiple ; Drug Resistance, Neoplasm ; Flow Cytometry ; Granulocyte-Macrophage Colony-Stimulating Factor ; pharmacology ; Humans ; Immunoglobulins ; analysis ; Interleukin-4 ; pharmacology ; K562 Cells ; Leukemia ; immunology ; pathology ; Membrane Glycoproteins ; analysis ; Tumor Necrosis Factor-alpha ; pharmacology

BACKGROUNDDendritic cells (DCs) are one of the most important antigen presenting cells in the human body, and DCs at various stages of maturation possess different or even opposite functions. The aim of this study was to investigate the influence of growth hormones on the functional status of cord blood-derived DCs encompassing immunophenotype, ability to excrete interleukin (IL)-12 and provoke autologous leukomonocyte.

METHODSMononuclear cells were isolated from fresh cord blood, with IL-4 and granulocyte-macrophage colony-stimulating factor (GM-CSF) used to induce and stimulate the mononuclear cells. Growth hormone at different concentrations was used to modify DCs, and then DCs morphology, number and growth status were observed. The immunophenotype of DCs was detected with a flow cytometer. The concentration of IL-12 in the DCs supernatant was determined by enzyme linked immunosorbent assay (ELISA) and DCs functional status was evaluated by autologous mixed lymphocyte reactions.

RESULTSMononuclear cells from cord blood can be differentiated into DCs by cytokine induction and growth hormone modification. With the increase in growth hormone concentrations (5 - 100 microg/L), the expression of DCs HLA-DR, CD1alpha, CD80 and CD83 were significantly increased (P < 0.05). The ability of DCs to secrete IL-12 was significantly improved (P < 0.05), and the ability of DCs to activate autologous lymphocytes was significantly enhanced (P < 0.05). Pegvisomant was able to ablate the effects of growth hormone on DCs.

CONCLUSIONSGrowth hormone may facilitate DCs induction and maturation, and improve the reproductive activity of autologous lymphocytes in a dose-dependent manner. Growth hormone may serve as a factor of modifying DCs to achieving maturity.

Antigens, CD ; metabolism ; B7-1 Antigen ; metabolism ; Cells, Cultured ; Dendritic Cells ; drug effects ; metabolism ; Enzyme-Linked Immunosorbent Assay ; Flow Cytometry ; Granulocyte-Macrophage Colony-Stimulating Factor ; metabolism ; Growth Hormone ; pharmacology ; HLA-DR Antigens ; metabolism ; Humans ; Immunoglobulins ; metabolism ; Interleukin-12 ; metabolism ; Membrane Glycoproteins ; metabolism

OBJECTIVETo investigate the phosphorylation intensity of MAPK pathway molecular Erk1/2 and the proliferation of prostate cancer cell line PC-3M.

METHODSFlow cytometry and RT-PCR were employed to study the ratio of different cell cycles and phases, respectively, before and after GM-CSF stimulation. Erk1/2 phosphorylation intensity was examined by Western blot simultaneously.

RESULTSThe rate of PC-3M cells at S and G2/M stages and the expression intensity of Ki-67 increased after GM-CSF incubation in a dose-dependent manner. The phosphorylation intensity of Erk1/2 increased remarkably after stimulation with GM-CSF.

CONCLUSIONThe intensification of Erk1/2 phosphorylation is one important molecular mechanism of the proliferation of hormone-independent prostate cancer.

Cell Line, Tumor ; Cell Proliferation ; Granulocyte-Macrophage Colony-Stimulating Factor ; pharmacology ; Humans ; Ki-67 Antigen ; biosynthesis ; Male ; Mitogen-Activated Protein Kinase 1 ; metabolism ; Mitogen-Activated Protein Kinase Kinases ; metabolism ; physiology ; Neoplasms, Hormone-Dependent ; metabolism ; pathology ; physiopathology ; Phosphorylation ; Prostatic Neoplasms ; metabolism ; pathology ; physiopathology ; Signal Transduction ; drug effects

Rac1 and Rac2 are essential for the control of oxidative burst catalyzed by NADPH oxidase. It was also documented that Rho is associated with the superoxide burst reaction during phagocytosis of serum- (SOZ) and IgG-opsonized zymosan particles (IOZ). In this study, we attempted to reveal the signal pathway components in the superoxide formation regulated by Rho GTPase. Tat-C3 blocked superoxide production, suggesting that RhoA is essentially involved in superoxide formation during phagocytosis of SOZ. Conversely SOZ activated both RhoA and Rac1/2. Inhibition of RhoA-activated kinase (ROCK), an important downstream effector of RhoA, by Y27632 and myosin light chain kinase (MLCK) by ML-7 abrogated superoxide production by SOZ. Extracellular signaling-regulated kinase (ERK)1/2 and p38 mitogen-activated protein kinase (MAPK) were activated during phagocytosis of SOZ, and Tat-C3 and SB203580 reduced ERK1/2 and p38 MAPK activation, suggesting that RhoA and p38 MAPK may be upstream regulators of ERK1/2. Inhibition of ERK1/2, p38 MAPK, phosphatidyl inositol 3-kinase did not block translocation of RhoA to membranes, suggesting that RhoA is upstream to these kinases. Inhibition of RhoA by Tat-C3 blocked phosphorylation of p47 PHOX. Taken together, RhoA, ROCK, p38MAPK, ERK1/2, and p47 PHOX may be subsequently activated, leading to activation of NADPH oxidase to produce superoxide.
Animals ; Cell Line ; Cell Membrane ; Cytosol ; Enzyme Inhibitors/pharmacology ; Extracellular Signal-Regulated MAP Kinases/metabolism ; Macrophage-1 Antigen/pharmacology ; Macrophages/drug effects/*metabolism/ultrastructure ; Mice ; Myosin-Light-Chain Kinase/metabolism ; Opsonin Proteins/blood/*metabolism ; *Phagocytosis ; Protein Transport ; Protein-Serine-Threonine Kinases/metabolism ; Research Support, Non-U.S. Gov't ; *Signal Transduction ; Superoxides/*metabolism ; Tetradecanoylphorbol Acetate/pharmacology ; Zymosan/*blood ; p38 Mitogen-Activated Protein Kinases/metabolism ; rhoA GTP-Binding Protein/antagonists & inhibitors/*metabolism