Chemotaxis Ability Research Articles

The ageing immune system and its clinical implications

SummaryAgeing is associated with multiple changes in many different components of the immune system. A healthy immune system exists in a state of balance between efficient effector responses against pathogens and tolerance to self antigens. This balance is changed with age; functions such as antigen recognition, phagocytosis, antigen presentation, chemotaxis, cytokine secretion and killing ability are all compromised. Aberrant cellular responses lead to an altered cytokine network with increases in inflammatory cytokines and decreases in anti-inflammatory cytokines leading to a pro-inflammatory state. Consequently older patients require extra care in diagnosis of infections as symptoms may be perturbed, resulting in unusual presentations of common conditions. The defects in immunity due to immunosenescence also mean that older patients require more care and screening than other patients in the same disease cohort. Though it is generally understood by clinicians that older patients are more at risk from multiple infections, the wider clinical effects of immunosenescence are less understood. The immune system is involved in several neurodegenerative conditions and the inflammatory conditions of immunosenescence may be a key factor in pathogenesis. Similarly, there is reason to believe that immunosenescence might be a key factor explaining the increased incidence of cancer in older age. With increasing understanding of the immune system's involvement in many of these pathological processes, and the contribution that immunosenescence makes to these, more efficient vaccines and novel therapies may be developed to prevent/treat these conditions.

A hybrid two-component system protein from Azospirillum brasilense Sp7 was involved in chemotaxis

We here report the sequence and functional analysis of org35 of Azospirillum brasilense Sp7, which was originally identified to be able to interact with NifA in yeast-two-hybrid system. The org35 encodes a hybrid two-component system protein, including N-terminal PAS domains, a histidine kinase (HPK) domain and a response regulator (RR) domain in C-terminal. To determine the function of the Org35, a deletion–insertion mutant in PAS domain [named Sp7353] and a complemental strain Sp7353C were constructed. The mutant had reduced chemotaxis ability compared to that of wild-type, and the complemental strain was similar to the wild-type strain. These data suggested that the A. brasilense org35 played a key role in chemotaxis. Variants containing different domains of the org35 were expressed, and the functions of these domains were studied in vitro. Phosphorylation assays in vitro demonstrated that the HPK domain of Org35 possessed the autokinase activity and that the phosphorylated HPK was able to transfer phosphate groups to the RR domain. The result indicated Org35 was a phosphorylation-communicating protein.

Lipopolysaccharide triggers macrophage activation of inflammatory cytokine expression, chemotaxis, phagocytosis, and oxidative ability via a toll-like receptor 4-dependent pathway: Validated by RNA interference

RNA interference has been extensively used to knock-down the translation of certain genes. Toll-like receptor 4 (TLR4) produced by macrophages can be activated in response to endotoxin stimulation. This study used the RNA interference technique to evaluate the roles of TLR4 in lipopolysaccharide (LPS)-stimulated activation of macrophages from the aspects of cytokine production, chemotaxis, phagocytosis, and oxidative ability. Exposure of macrophages to 1, 25, 50, 100ng/mL LPS for 1, 6, and 24h did not affect cell viability. Meanwhile, treatment with 100ng/mL LPS induced interleukin (IL)-1β protein and mRNA syntheses in a time-dependent manner. Application of TLR4 small interference (si)RNA into macrophages decreased the levels of this receptor, and simultaneously ameliorated LPS-induced IL-1β and IL-6 mRNA production. Transwell analysis showed that LPS increased chemotactic activity of macrophages, but application of TLR4 siRNA reduced such an effect. Phagocytic activities of macrophages were significantly augmented following LPS treatment. However, knocking-down the translation of TLR4 mRNA using RNA interference lowered the LPS-enhanced phagocytic activity. Analysis of flow cytometry revealed that LPS increased oxidative ability of macrophages, but TLR4 siRNA inhibited such development. This study used RNA interference techniques to show that TLR4 can mediate LPS-induced macrophage activations of IL-1β and IL-6 gene expression, chemotaxis, phagocytosis, and oxidative ability.

Nitric Oxide and Carbon Monoxide mediate the anti-inflammatory effects of Remote Ischemic Preconditioning (RIPC) (94.12)

Abstract Remote ischemic preconditioning(RIPC) consists of short periods of ischemia followed by reperfusion, which induces protective anti-inflammatory effect on distance organs. Studies have shown that inflammatory parameters such as leukocyte migration are significantly reduced in pre-conditioned animals.However, the mechanism involved in this effect is not yet well established. This study evaluated if NO and CO is mediating the anti-inflammatory activity of RIPC.RIPC model is based on turnstile in right leg of a mouse for 10 minutes followed by 30 of reperfusion. Participation of NO and CO was investigated using inhibitors of iNOS (1400W; 3mg/kg or Aminoguanide; 50mg/kg) and HO-1 (ZnPPIX;10mg/kg) as pre-treatment 30 minutes before RIPC. Afterwards, inflammatory reaction was induced by intraperitoneal administration of Carragenin(500µg/cav).Four hours after the peritoneal cavities were washed with saline and number of migrated leukocytes was evaluated. Chemotaxis ability of neutrophils obtained from mice under RIPC treated or not with the inhibitors was analyzed in Boyden Chamber. Expression of CXCR2 and GRK2 in neutrophils was determined by flow cytometry and immunofluorescence, respectively.CO and NO inhibitors prevented the inhibitory effect of RIPC upon Cg-induced neutrophil migration into peritoneal cavity of animals, as well as, upon KC induced neutrophils chemotaxis in Boyden Chamber. The reduction of neutrophil chemotaxis by RICP correlated with CXCR2 internalization and increase of GRK2 expression in the neutrophils.CO and NO production prevented these effects of RICP.Thus,inhibitors of HO-1/CO and NOSi/NO pathways seem to mediate the anti-inflammatory effect of RICP.

Patterns of expression and function of the p75 NGFR protein in pancreatic cancer cells and tumours

Backgrounds/aims Pancreatic carcinoma is one of the most aggressive human malignancies. The aggressive and highly metastatic behaviour of pancreatic carcinoma may partly be attributable to the autocrine and/or paracrine interactions involving altered expression of neurotrophin growth factors and their corresponding receptors. The aim of the present study is to investigate the expression pattern and function of the p75 NGFR protein in pancreatic cancer cell lines and tumours to explain the phenomenon of perineural invasion in pancreatic cancer. Methods The expression of p75 NGFR in 137 pancreatic adenocarcinoma samples and the corresponding adjacent pancreatic samples was examined immunohistochemically using the EnVision Plus System. Then we examined the in vitro chemotaxis behaviour of cancer cells transfected with p75 NGFR plasmid to nerve growth factor (NGF). Results Immunostaining for p75 NGFR was weak or absent in both normal pancreata and pancreatic carcinoma tissues; however, the immunostaining was relatively weaker in the pancreatic carcinoma tissues than in the normal pancreata. It is interesting to note that p75 NGFR expression in the cancer tissues was positively correlated with the degree of perineural invasion (χ 2 = 32.94, P < 0.01). The chemotaxis ability of the p75 NGFR-transfected pancreatic cancer cells to NGF was significantly stronger than that of the non-transfected or vacant vector transfected cells ( P < 0.01). Conclusions Our findings indicate that p75 NGFR expression may be involved in the perineural invasion of pancreatic cancer cells, and the mechanism might be through mediating the chemoattraction of cancer cells for neural tissues.

Streptococcus pyogenes induces epithelial inflammatory responses through NF-κB/MAPK signaling pathways

Innate immunity involves a cascade of inflammatory events, resulting in the secretion of chemokines and cytokines to recruit mediator cells in adaptive immunity. To study epithelial inflammatory responses initiated by Streptococcus pyogenes infection, we investigated chemotaxis ability in the supernatant of infected human respiratory epithelial HEp-2 cells. Our results showed that these supernatants showed significantly increased ability to attract monocytes, implying the release of inflammatory chemoattractants into the medium. Expression of interleukin (IL)-8 and IL-6 in HEp-2 cells was significantly increased at both the mRNA and protein levels after infection with S. pyogenes. Electrophoretic mobility shift and reporter-gene assays demonstrated that the transcription factors NF-κB and AP-1, regulated by mitogen-activated protein (MAP) kinase, were activated after streptococcal infection. The increases in mRNAs for IL-8 and IL-6 were abrogated by addition of NF-κB and MAP kinase inhibitors, suggesting that the upregulation of IL-8 and IL-6 is mediated through NF-κB and MAP kinase signaling pathways. Taken together, our results indicate that S. pyogenes infection of epithelial cells induces the secretion of pro-inflammatory chemokines/cytokines through activation of NF-κB and MAP kinase signaling pathways. These early innate responses initiated by S. pyogenes-infected respiratory epithelial cells may recruit immune cells to the airway and induce inflammation.

Enhancement of Metastatic and Invasive Capacity of Gastric Cancer Cells by Transforming Growth Factor-&amp;beta;1

Transforming growth factor-beta (TGF-beta), a multifunctional cytokine, exerts contradictory roles in different kinds of cells. A number of studies have revealed its involvement in the progression of many types of tumors. To investigate the effect of TGF-beta on gastric carcinoma, SGC7901, BGC823 and MKN28 (a TGF-beta-resistant cell line) adenocarcinoma clones were used. After pretreatment in serum-free medium with or without 10 ng/ml TGF-beta1, their experimental metastatic potential, chemotaxis, and invasive and adhesive ability were measured. Furthermore, zymography for gelatinase was processed. Liver colonies were also measured 4 weeks after inoculation of SGC7901, BGC823 and MKN28 in Balb/c nude mice, and an increase in the number of surface liver metastases was seen in SGC7901 (from 11.0+/-3.0 to 53.3+/-3.3) and BGC823 (from 9.3+/-2.5 to 60.0+/-2.8) groups, whereas there was no difference between MKN28 groups (from 35.2+/-3.8 to 38.5+/-2.7). In vitro experiments showed that TGF-beta1 increased the adhesion capacity of SGC7901 and BGC823 cells to immobilized reconstituted basement membrane/fibronectin matrices and promoted their penetration through reconstituted basement membrane barriers. Zymography demonstrated that enhanced invasive potential was partly due to the increased type IV collagenolytic (gelatinolytic) activity, but there was no difference in type IV collagenolytic activity and other biological behaviors between MKN28 groups. These results suggested that TGF-beta1 might modulate the metastatic potential of gastric cancer cells by promoting their ability to break down and penetrate basement membrane barriers and their adhesive and motile activities. We speculated that TGF-beta1 might act as a progression-enhancing factor in gastric cancer. Therefore blockage of TGF-beta or TGF-beta signaling might prevent gastric cancer cells from invading and metastasizing.

Partial characterization of phagocytic activity in neutrophils of the nine-banded armadillo Dasypus novemcinctus

Though the armadillo is important as a research model in leprosy studies, the activity of armadillo's neutrophils is an aspect of little research. The aim of this study was carried out to partially characterize the chemotaxis, endocytosis and bacteriocidal ability of the neutrophils found in the nine-banded armadillo ( Dasypus novemcinctus) . Results showed that the chemotactic activity of the neutrophils, evaluated by the movement of the neutrophils through a nitrocellulose membrane (5 μm) in response to a chemo-attractive substance, was greater towards the armadillo serum (5.16 ± 1.35 migration index, p < 0.05) than towards the formil methionyl leucil phenylalanine (fMLP, 1.43 ± 0.18 migration index) or human serum (0.56 ± 0.18 migration index). Regarding endocytic capacity of the neutrophils and the monocytes against Escherichia coli was evaluated by a flow cytometry and using opsonized and non-opsonized E. coli-FITC at the following incubation times: 5, 10, 20, 30 and 60 min. The largest percentage of endocytosis by the neutrophils was 92.32 ± 0.12% with opsonized bacteria and 77.73 ± 14.33% with non-opsonized bacteria at 10 min incubation time, while the largest percentage of endocytosis by monocytes was 89.94 ± 1.40% with opsonized bacteria and 73.07 ± 15.6% with non-opsonized bacteria at 20 min incubation time. Evaluation of the bacteriocidal capacity of neutrophils using the methyl-thiazol-tetrazolium salts (MTT) reduction color-measurement assay showed an 89.0 ± 10% mortality rate of non-opsonized E. coli and 89.0 + 10% of opsonized E. coli. In conclusion, the armadillo neutrophils show a good phagocytosis and bacteriocidal activity; however, a deficiency in the migration towards the fMLP was observed. This deficiency could be a cause so that the armadillo neutrophils do not respond quickly to invading microorganism.

Chemotaxis-mediated biodegradation of cyclic nitramine explosives RDX, HMX, and CL-20 by Clostridium sp. EDB2

Cyclic nitramine explosives, RDX, HMX, and CL-20 are hydrophobic pollutants with very little aqueous solubility. In sediment and soil environments, they are often attached to solid surfaces and/or trapped in pores and distribute heterogeneously in aqueous environments. For efficient bioremediation of these explosives, the microorganism(s) must access them by chemotaxis ability. In the present study, we isolated an obligate anaerobic bacterium Clostridium sp. strain EDB2 from a marine sediment. Strain EDB2, motile with numerous peritrichous flagella, demonstrated chemotactic response towards RDX, HMX, CL-20, and NO 2 −. The three explosives were biotransformed by strain EDB2 via N-denitration with concomitant release of NO 2 −. Biotransformation rates of RDX, HMX, and CL-20 by the resting cells of strain EDB2 were 1.8 ± 0.2, 1.1 ± 0.1, and 2.6 ± 0.2 nmol h −1 mg wet biomass −1 (mean ± SD; n=3), respectively. We found that commonly seen RDX metabolites such as TNX, methylenedinitramine, and 4-nitro-2,4-diazabutanal neither produced NO 2 − during reaction with strain EDB2 nor they elicited chemotaxis response in strain EDB2. The above data suggested that NO 2 − released from explosives during their biotransformation might have elicited chemotaxis response in the bacterium. Biodegradation and chemotactic ability of strain EDB2 renders it useful in accelerating the bioremediation of explosives under in situ conditions.

Effects of aflatoxin and carotenoids on growth performance and immune response in mule ducklings

The purpose of this study was to investigate if carotenoids could alleviate the adverse effects caused by aflatoxin with respect to growth performance and immune response. In two experiments, a total of 320 mule ducklings were assigned to 5 treatments, i.e. control, aflatoxin B 1 (AFB 1) 200 ppb, AFB 1 +β-carotene (BC) 200 ppm, AFB 1+BC 400 ppm, and AFB 1+astaxanthin (AS) 200 ppm. In experiment 1, the addition of β-carotene or astaxanthin in the diet containing AFB 1 200 ppb resulted in a significant decrease in average daily gain as compared with the control. AFB 1 200 ppb alone and the addition of BC or AS on top of AFB 1 resulted in a significantly lower daily feed intake than for the control group. There were no significant differences in relative organ weights among treatment groups. Both treatments of BC 400 ppm and AS 200 ppm had significantly more macrophages harvested per duck than the control and AFB 1 200 ppb treatments. However, there were no significant differences among treatments in percentages of phagocytotic macrophages and number of Candida albican phagocytized by phagocytotic macrophages. In experiment 2, blood biochemical parameters and antibody titers were evaluated. There were no significant differences among treatments in total bilirubin content and alkaline phosphatase activity in the serum or in antibody titers against fowl cholera. However, AFB 1 treatment had the highest activities of AST and ALT in the serum. The addition of BC 400 ppm on top of AFB 1 significantly reduced ALT activity as compared with the AFB 1 200 ppb treatment. These results suggest that carotenoids could provide a slightly toxic alleviating effect on growth performance, enhance the chemotaxis ability of macrophages, and reduce ALT activity elevated by AFB 1.

Neither motility nor chemotaxis plays a role in the ability of Escherichia coli F-18 to colonize the streptomycin-treated mouse large intestine

Escherichia coli F-18, isolated from the feces of a healthy human in 1977, is an excellent colonizer of the streptomycin-treated mouse large intestine and displays normal motility and chemotaxis ability. A chemotaxis-defective derivative of E. coli F-18, E, coli F-18 CheA-, and a nonflagellated derivative, E. coli F-18 Fla-, were constructed. These strains were found to colonize the streptomycin-treated mouse large intestine as well as E. coli F-18 when mice were fed both E. coli F-18 and either the CheA- or Fla- derivative at high levels (10(10) CFU of each strain per mouse) or low levels (10(4) CFU of each strain per mouse). Furthermore, E. coli F-18 lost motility and chemotaxis ability when grown in colonic or cecal mucus in vitro despite retaining the ability to synthesize flagella. Thus, it appears that neither motility nor chemotaxis plays a role in the ability of E. coli F-18 to colonize because this strain becomes functionally nonmotile upon growth in the streptomycin-treated mouse large intestine.

S-adenosylmethionine may not be essential for signal transduction during bacterial chemotaxis.

We previously showed that a mutant strain of Salmonella typhimurium completely deficient in both the chemoreceptor methylating (CheR) and demethylating (CheB) enzymes can still exhibit chemotaxis to aspartate and other attractants (J. Stock, A. Borczuk, F. Chiou, and J. E. B. Burchenal, Proc. Natl. Acad. Sci. USA 82:8364-8368, 1985). We used this cheR cheB mutant to examine the possibility of an additional requirement for S-adenosylmethionine in chemotaxis besides its role in chemoreceptor methylation. A metE mutation was transduced into a cheR cheB double mutant, and the cells were starved for methionine. Despite the fact that intracellular S-adenosylmethionine dropped from approximately 100 microM to less than 0.2 microM, chemotaxis was largely unaffected. In contrast, a corresponding cheR+ cheB+ metE mutant completely lost its chemotaxis ability after being starved for methionine. We conclude from this observation that the primary requirement for S-adenosylmethionine during bacterial chemotaxis is in the methylation of receptor proteins.

Identification of the tip-encoded receptor in bacterial sensing.

A chemotaxis gene encoding a protein with receptorlike properties has been identified in Salmonella typhimurium and termed tip for taxis-involved protein. Based on the stringency of DNA hybridization, the tip gene has about 75% homology with a region of the tar gene encoding the cytoplasmic domain of the aspartate receptor. Introduction of the tip gene into a smooth-swimming Escherichia coli receptor mutant (tar tsr tap) restored both chemotaxis ability on soft-agar-tryptone plates and a wild-type swimming phenotype. We have shown, by overexpressing the CheY protein, that shifting of the mutant swimming bias in the absence of receptors is insufficient to restore chemotaxis ability. This suggests that in addition to resetting the swimming bias, the tip gene product functions as a receptor. By functional criteria, we found that Tip is not a duplicate aspartate (Tar) or serine (Tsr) receptor gene. Based on behavioral properties, the S. typhimurium Tip receptor provides functional features similar to those of the E. coli Tap receptor.

Expression of macrophage functions in hybrids of a myeloma cell line with inflammatory macrophages: evidence for negative control mechanisms in the expression of macrophage functions.

Mouse inflammatory macrophages from C57BL/6N mice were fused with BALB/c mouse-derived myeloma cells (the CANS series). The hybrids in the early period after cell fusion (8 weeks) showed no macrophage functions (chemotaxis, EA and EAC rosette-forming abilities, phagocytosis or lysozyme production). EA rosette-forming ability was observed when these hybrids were treated with trypsin, whereas other macrophage functions were not. After prolonged culture, the hybrids (12 clones of 13 randomly selected) showed all the macrophage functions along with chromosome loss. Myeloma cell functions (kappa light chain production) were found in the young hybrids soon after cell fusion but were absent in the aged hybrids. These results indicated that reexpression of macrophage properties, except for EA rosette-forming abilities, takes place after the loss of chromosomes or genes repressing the expression of macrophage functions.

Granulocyte function during lithium therapy

Random migration, chemotaxis, phagocytosis, and bactericidal ability of neutrophils from 5 patients receiving lithium carbonate were compared with those of neutrophils from healthy donors. These cells functioned normally in all respects. Neither sera from patients receiving lithium carbonate nor the addition of lithium chloride to control cells in vitro significantly altered their functional capacity. These findings suggest that neutrophil function in patients receiving lithium therapy is preserved, and they support the potential utility of this drug as a leukopoietic agent in neutropenic states.

Granulocyte Function During Lithium Therapy

AbstractRandom migration, chemotaxis, phagocytosis, and bactericidal ability of neutrophils from 5 patients receiving lithium carbonate were compared with those of neutrophils from healthy donors. These cells functioned normally in all respects. Neither sera from patients receiving lithium carbonate nor the addition of lithium chloride to control cells in vitro significantly altered their functional capacity. These findings suggest that neutrophil function in patients receiving lithium therapy is preserved, and they support the potential utility of this drug as a leukopoietic agent in neutropenic states.