Sepsis remains one of the leading causes of death globally, in spite of advanced developments in intensive care and better understandings of pathophysiology related to sepsis. There is no special treatment or drug available for sepsis, currently. Under normal circumstances, neutrophil is a major player in acute infection control. However, during sepsis, the migration abilities and antimicrobial functions of neutrophils are impaired, resulting in a dysregulated immune response. Recent studies have indeed demonstrated that blocking or reversing neutrophil migration and impaired antibacterial function can improve the outcomes in septic animal models. This article systemically synthesized information regarding related factors and signaling involved in the functions of neutrophils in sepsis. This review also discussed the possibility that neutrophils be used as a marker for specific diagnosis and/or prediction of the outcomes of sepsis.
Animals ; Neutrophils/physiology* ; Chemotaxis ; Chemotaxis, Leukocyte ; Sepsis ; Cell Movement

To investigate the effects of cigarette smoking in different manners on acute lung injury in rats.The commercially available cigarettes with tar of 1,5, 11 mg were smoked in Canada depth smoking (health canada method, HCM) manner, and those with tar of 11 mg were also smoked in international standard (ISO) smoking manner. Rats were fixed and exposed to mainstream in a manner of nose-mouth exposure. After 28 days, the bronchoalveolar lavage fluids from left lung were collected for counting and classification of inflammatory cells and determination of pro-inflammatory cytokines IL-1β and TNF-α. The right lungs were subjected to histological examination and determination of myeloperoxidase (MPO) and superoxide dismutase (SOD) activities and glutathione, reactive oxygen species (ROS) and malondialdehyde (MDA) levels.In both HCM and ISO manners, the degree of lung injury was closely related to the tar content of cigarettes, and significant decrease in the body weight of rats was observed after smoking for one week. In a HCM manner, smoking with cigarette of 11 mg tar resulted in robust infiltration of macrophages, lymphocytes and neutrophils into lungs, significant increase in IL-1β and TNF-α levels and MPO activities, and significant decrease in GSH levels and SOD activities and increase in ROS and MDA levels (all<0.05). Smoking with cigarette of 5 mg tar led to moderate increase in IL-1β and TNF-α levels, and MPO activities (all<0.05), and moderate decrease in GSH levels and SOD activities and increase of ROS and MDA levels (all<0.05). However, smoking with cigarette of 1 mg tar affected neither inflammatory cell infiltration nor IL-1β and TNF-α levels.Cigarette smoking in nose-mouth exposure manner can induce acute lung injury in rats; and the degree of lung injury is closely related to the content of tar and other hazards in cigarettes.
Acute Lung Injury ; etiology ; pathology ; physiopathology ; Animals ; Bronchoalveolar Lavage Fluid ; chemistry ; cytology ; Chemotaxis, Leukocyte ; drug effects ; Glutathione ; analysis ; drug effects ; Interleukin-1beta ; analysis ; drug effects ; Lung ; chemistry ; pathology ; Lymphocytes ; drug effects ; pathology ; Macrophages ; drug effects ; pathology ; Male ; Malondialdehyde ; analysis ; Neutrophil Infiltration ; drug effects ; Neutrophils ; drug effects ; pathology ; Peroxidase ; analysis ; drug effects ; Rats ; Reactive Oxygen Species ; analysis ; Smoking ; adverse effects ; Superoxide Dismutase ; analysis ; drug effects ; Tobacco Products ; adverse effects ; classification ; Tumor Necrosis Factor-alpha ; analysis ; drug effects ; Weight Loss ; drug effects

Neutrophils play a key role in innate immunity, and the identification of new stimuli that stimulate neutrophil activity is a very important issue. In this study, we identified three novel peptides by screening a synthetic hexapeptide combinatorial library. The identified peptides GMMWAI, MMHWAM, and MMHWFM caused an increase in intracellular Ca2+ in a concentration-dependent manner via phospholipase C activity in human neutrophils. The three peptides acted specifically on neutrophils and monocytes and not on other non-leukocytic cells. As a physiological characteristic of the peptides, we observed that the three peptides induced chemotactic migration of neutrophils as well as stimulated superoxide anion production. Studying receptor specificity, we observed that two of the peptides (GMMWAI and MMHWFM) acted on formyl peptide receptor (FPR)1 while the other peptide (MMHWAM) acted on FPR2. Since the three novel peptides were specific agonists for FPR1 or FPR2, they might be useful tools to study FPR1- or FPR2-mediated immune response and signaling.
Animals ; Calcium/metabolism ; Cell Line ; Cells, Cultured ; Chemotaxis, Leukocyte/drug effects ; Humans ; Mice ; NIH 3T3 Cells ; Neutrophils/*cytology/*drug effects ; PC12 Cells ; Peptides/*pharmacology ; Rats ; Receptors, Formyl Peptide/agonists

Secondary lymphoid tissue chemokine (SLC), which is expressed in T cell zones of secondary lymphoid organs, including the spleen and lymph nodes, strongly recruits both T lymphocytes and mature dendritic cells. As appropriate interaction of tumor-specific T cells and mature dendritic cells, equipped with tumor antigens, is a prerequisite for effective T cell immunity against established tumors, we mobilized lymphocytes and dendritic cells to tumor sites by intratumoral injection of secondary lymphoid tissue chemokine-Fc (SLC-Fc) fusion protein using the B16F10 murine melanoma model. Activation of dendritic cells, another prerequisite for the effective activation of naive tumor-specific T cells, was achieved by the addition of immunostimulatory cytosine-phosphorothioate-guanine oligodeoxynucleotide (CpG-ODN) into the tumor site. Intratumoral administration of SLC-Fc or CpG-ODN revealed antitumor effects against B16F10 murine melanoma grown in the subcutaneous space. Co-treatment of SLC-Fc and CpG-ODN displayed synergistic effects in reducing the tumor size. The synergistic antitumor effect in co-treatment group was correlated with the synergistic/additive increase in the infiltration of CD4+ T cells and CD11c+ dendritic cells in the tumor mass compared to the single treatment groups. These results suggest that the combined use of chemokines and adjuvant molecules may be a possible strategy in clinical tumor immunotherapy.
Animals ; Antigens, CD11c/immunology ; CD4-Positive T-Lymphocytes/immunology ; Cell Line, Tumor ; Cell Proliferation/drug effects ; Chemokine CCL21/*administration & dosage/pharmacology ; Chemotaxis, Leukocyte ; Dendritic Cells/immunology/metabolism ; Immunotherapy ; Injections, Intralesional ; Melanoma, Experimental/*immunology/*therapy ; Mice ; Mice, Inbred C57BL ; Oligodeoxyribonucleotides/*administration & dosage/pharmacology ; T-Lymphocytes/immunology/metabolism

OBJECTIVETo study the influence of transcranial high-voltage electrical burn (HEB) on rheological changes of leukocytes in mesentery capillary in rats and the therapeutic effects of ulinastatin.

METHODSForty-five SD rats were divided into control (C), electrical burns (EB), and ulinastatin treatment (UT) groups according to the random number table, with 15 rats in each group. Model of HEB was reproduced in rats of EB and UT groups with voltage regulator and experimental transformer, and then rats in EB group was intraperitoneally injected with 2 mL isotonic saline while rats in UT group was intraperitoneally injected with 2 mL ulinastatin (2 x 10(4) U/kg). Rats in C group received sham burn with the same treatment as used in EB group but without electric current. Rheological changes of leukocytes in mesentery capillary were observed with Bradford microscope at 15 minutes before HEB and 5 minutes, 1, 2, 4, 8 hour (s) after HEB (PHM or PHH), including counting the number of rolling leukocytes, leukocytes rolling speed, the number of leukocytes adherent to mesentery capillary, total leukocyte-endothelium contact time (TLECT). Data were processed with t test.

RESULTS(1) The number of rolling leukocytes from PHM 5 to PHH 8 was increased in EB group and UT group as compared with that at 15 minutes before HEB, especially at PHM 5 [(51.4 +/- 3.2), (24.6 +/- 1.9) cells/min, respectively] which were higher than that in C group [( 1.1 +/- 0.7) cells/min, with t value respectively 59.28, 44.99, P values all below 0.05]. The number in UT group at each time point after burn was less than those in EB group, especially at PHM 5 (t = 27.97, P < 0.05). (2) Compared with that at 15 minutes before HEB, the rolling speed of leukocytes from PHM 5 to PHH 8 was slow in EB group and UT group, especially at PHM 5 [(90 +/- 9), (175 +/- 13) microm/s, respectively] which were slower than that in C group [(277 +/- 12) microm/s, with t value respectively 47.97, 21.59, P values all below 0.05]. The rolling speed in UT group from PHM 5 to PHH 8 was faster than that in EB group, especially at PHM 5 (t = 20.55, P < 0.05). (3) Compared with that at 15 minutes before HEB, the number of leukocytes per 100 micrometer capillary from PHM 5 to PHH 8 was increased in EB group and UT group, especially at PHM 5 (23.27 +/- 3.20, 5.80 +/- 1.61, respectively) which were higher than that in C group (0, with t value respectively 28.16, 13.95, P values all below 0.05). The number of adhered leukocytes in UT group at each time point after burn was less than that in EB group, especially at PHM 5 ( t = 18.89, P < 0.05). (4) Compared with that at 15 minutes before HEB, TLECT from PHM 5 to PHH 8 was increased in EB group and UT group, especially at PHM 5 [(14.45 +/- 1.99), (3.66 +/- 0.96) s/min, respectively] which were longer than that in C group (0 s/min, with t value respectively 28.12, 14.77, P values all below 0.05). TLECT in UT group from PHM 5 to PHH 8 was shorter than that in EB group, especially at PHM 5 (t = 18.91, P < 0.05). (5) No rolling leukocyte or wall-adherent leukocyte was found in blood flow of arterioles or capillaries of rats in three groups at each time point.

CONCLUSIONSTranscranial HEB can lead to abnormal rheological changes of leukocytes in mesentery capillary in rats, and the changes can be ameliorated by ulinastatin.

Animals ; Burns, Electric ; physiopathology ; Capillaries ; Glycoproteins ; pharmacology ; Leukocyte Rolling ; Leukocytes ; drug effects ; Male ; Mesentery ; blood supply ; drug effects ; Microcirculation ; Rats ; Rats, Sprague-Dawley

OBJECTIVETo construct the murine CCL21 eukaryotic expression plasmid, and to investigate the chemotactic function of its products.

METHODSMurine CCL21 cDNA was amplified by RT-PCR from murine total RNA, and was inserted into eukaryotic expression plasmid pcDNA3.1 after confirmation of sequencing. The recombinant CCL21 plasmid was transferred into mouse forestomach carcinoma (MFC) cells and the chemotactic function of expressed products was detected by chemotaxis assay.

RESULTGene sequencing, gel electrophoresis of PCR products and restrictive digestion proved the successful construction of CCL21, and its expression was confirmed by Western Blot. The transfected tumor cells had a significant chemotactic function to DC.

CONCLUSIONThe recombinant murine CCL21 eukaryotic expression plasmid has been successfully constructed, and its expression products in tumor cells have a marked chemotactic function to DC.

Animals ; Base Sequence ; Chemokine CCL21 ; biosynthesis ; genetics ; Chemotaxis, Leukocyte ; Cloning, Molecular ; DNA, Complementary ; genetics ; Dendritic Cells ; drug effects ; immunology ; Genetic Vectors ; genetics ; Mice ; Mice, Inbred C57BL ; Molecular Sequence Data ; Plasmids ; genetics ; Recombinant Proteins ; biosynthesis ; genetics ; immunology ; Stomach Neoplasms ; metabolism ; pathology ; Transfection ; Tumor Cells, Cultured

Serum amyloid A (SAA) has been regarded as an important mediator of inflammatory responses. The effect of several formyl peptide receptor-like 1 (FPRL1) ligands on the production of IL-8 by SAA was investigated in human neutrophils. Among the ligands tested, LL-37 was found to specifically inhibit SAA-induced IL-8 production in transcriptional and post-transcriptional levels. Since SAA stimulated IL-8 production via ERK and p38 MAPK in human neutrophils, we tested the effect of LL-37 on SAA induction for these two MAPKs. LL-37 caused a dramatic inhibition of ERK and p38 MAPK activity, which is induced by SAA. LL-37 was also found to inhibit SAA-stimulated neutrophil chemotactic migration. Further, the LL-37-induced inhibitory effect was mediated by FPRL1. Our findings indicate that LL-37 is expected to be useful in the inhibition of SAA signaling and for the development of drugs against SAA-related inflammatory diseases.
Animals ; Antimicrobial Cationic Peptides/*pharmacology ; Cell Line, Tumor ; Cell Movement ; Chemotaxis, Leukocyte ; Humans ; Interleukin-8/*biosynthesis ; MAP Kinase Kinase Kinases/metabolism ; Neutrophils/drug effects/*immunology ; Proto-Oncogene Proteins/metabolism ; Rats ; Receptors, Formyl Peptide/metabolism ; Receptors, Lipoxin/metabolism ; Serum Amyloid A Protein/*antagonists & inhibitors ; Signal Transduction ; Transcription, Genetic

Serum amyloid A (SAA) has been regarded as an important mediator of inflammatory responses. The effect of several formyl peptide receptor-like 1 (FPRL1) ligands on the production of IL-8 by SAA was investigated in human neutrophils. Among the ligands tested, LL-37 was found to specifically inhibit SAA-induced IL-8 production in transcriptional and post-transcriptional levels. Since SAA stimulated IL-8 production via ERK and p38 MAPK in human neutrophils, we tested the effect of LL-37 on SAA induction for these two MAPKs. LL-37 caused a dramatic inhibition of ERK and p38 MAPK activity, which is induced by SAA. LL-37 was also found to inhibit SAA-stimulated neutrophil chemotactic migration. Further, the LL-37-induced inhibitory effect was mediated by FPRL1. Our findings indicate that LL-37 is expected to be useful in the inhibition of SAA signaling and for the development of drugs against SAA-related inflammatory diseases.
Animals ; Antimicrobial Cationic Peptides/*pharmacology ; Cell Line, Tumor ; Cell Movement ; Chemotaxis, Leukocyte ; Humans ; Interleukin-8/*biosynthesis ; MAP Kinase Kinase Kinases/metabolism ; Neutrophils/drug effects/*immunology ; Proto-Oncogene Proteins/metabolism ; Rats ; Receptors, Formyl Peptide/metabolism ; Receptors, Lipoxin/metabolism ; Serum Amyloid A Protein/*antagonists & inhibitors ; Signal Transduction ; Transcription, Genetic

In this study, we observed that lysophosphatidylglycerol (LPG) completely inhibited a formyl peptide receptor like-1 (FPRL1) agonist (MMK-1)-stimulated chemotactic migration in human phagocytes, such as neutrophils and monocytes. LPG also dramatically inhibited IL-1beta production by another FPRL1 agonist serum amyloid A (SAA) in human phagocytes. However, LPG itself induced intracellular calcium increase and superoxide anion production in human phagocytes. Keeping in mind that phagocytes migration and IL-1beta production by FPRL1 are important for the induction of inflammatory response, our data suggest that LPG can be regarded as a useful material for the modulation of inflammatory response induced by FPRL1 activation.
Chemotaxis, Leukocyte/*drug effects ; Humans ; Interleukin-1beta/*biosynthesis ; Lysophospholipids/*pharmacology ; Monocytes/drug effects/immunology/metabolism/physiology ; Neutrophils/drug effects/immunology/metabolism/physiology ; Peptides/metabolism/pharmacology ; *Phagocytes/drug effects/immunology/metabolism/physiology ; Receptors, Formyl Peptide/*metabolism ; Receptors, Lipoxin/*metabolism ; Serum Amyloid A Protein/metabolism/pharmacology

UNLABELLEDOBJECTIVE To explore the effects of human macrophage inflammatory protein-1 beta (hMIP-1beta) modification on the in vivo tumorigenicity and vaccine efficacy of tumor cells.

METHODSMurine colorectal adenocarcinoma CT26 cells were transfected with a recombinant adenovirus carring the hMIP-1beta gene (AdhMIP-1beta). The efficacy of gene transfection was tested by X-gal staining. The hMIP-1beta level in the supernatant of hMIP-1beta gene-modified CT26 cells was assayed by ELISA, and the chemotactic activity for CD4+ T cells, CD8+ T cells, NK cells and immature dendritic cells (imDCs) were assayed by a transwell chamber. The changes of growth characteristics and in vivo tumorigenicity of hMIP-1beta gene-modified CT26 cells were also assessed. BALB/c mice were immunized with hMIP-1beta gene-modified CT26 tumor vaccine and the antitumor effect was evaluated.

RESULTShMIP-1beta gene could be transfected into CT26 cells by AdhMIP-1beta with an efficiency over 95%. The level of hMIP-1beta in the culture supernatant of hMIP-1beta gene-modified CT26 cells was 980 pg/ml and the supernatant displayed ramarkable chemotactic activity to CD4+ T cells, CD8+ T cells, NK cells and imDCs compared with LacZ gene-modified CT26 cells and control. When the hMIP-1beta gene-modifited CT26 cells were subcutaneously inoculated in BALB/c mice, the tumorigencity was delayed and suppressed, and overt necrosis and lymphocyte infiltration were observed in the tumor tissue, but not in those inoculated with LacZ gene-modified CT26 cells or parental CT26 cells. The mice immunized with hMIP-1beta gene-modified CT26 tumor vaccine could induce tumor specific CTL activity and nonspecific NK activity, and exhibited resistance to later challenge with wild-type CT26 cells.

CONCLUSIONhMIP-1beta gene-modified CT26 cells exhibit decreased tumorigenicity, and hMIP-1beta gene-modified tumor vaccine may induce a powerful specific and nonspecific antitumor response. The data suggested that hMIP-1beta gene-modified tumor vaccine may play a potent role in prevention of metastasis and recurrence of malignant tumors.

Adenocarcinoma ; metabolism ; pathology ; Adenoviridae ; genetics ; Animals ; CD4-Positive T-Lymphocytes ; immunology ; CD8-Positive T-Lymphocytes ; immunology ; Cancer Vaccines ; Cell Line, Tumor ; Chemokine CCL4 ; genetics ; metabolism ; Chemotaxis, Leukocyte ; Colonic Neoplasms ; metabolism ; pathology ; Cytotoxicity, Immunologic ; Dendritic Cells ; immunology ; Female ; Genetic Vectors ; Humans ; Killer Cells, Natural ; immunology ; Mice ; Mice, Inbred BALB C ; Neoplasm Transplantation ; Random Allocation ; Recombinant Proteins ; genetics ; metabolism ; Transfection ; Tumor Burden