Eosinophil and neutrophil chemotactic activities of adult worm extracts of Schistosoma japonicum in vivo and in vitro

Large numbers of eosinophils and neutrophils attracted to the soluble extract of Schistosoma japonicum adult worms (SjAW-ext) were detected at the injection site of normal guinea pig skin. Eosinophil and neutrophil chemotactic activities were also confirmed in in vitro assay by using blind-well chambers with Millipore filters in dose-dependent fashion. Two components of SjAW-ext showed eosinophil chemotactic activity; one was in the high molecular weight fraction (JAE-H), estimated to be more than 440,000 daltons, the other in the low molecular weight fraction (JAE-L) obtained by Sephadex G-200 gel filtration. High neutrophil chemotactic activity was detected in the JAE-L. These eosinophil and neutrophil chemotactic activities were also detected in culture fluid of S. japonicum adult worms. Eosinophil chemotactic factor (ECF) of JAE-H was stable to heating (100 C, 30 min) and pronase digestion, but completely destroyed by periodate oxidation. It is suggested that the ECF of JAE-H is a glycoprotein. JAE-L was also stable to heating (56 and 100 C, 30 min) and pronase digestion for eosinophil chemotaxis. Possible roles of those activities in schistosome infections are discussed.     

Cellular changes in bone marrow of malaria-infected mice. III. Chemotaxis of granulocytes

The number of bone marrow cells and their chemotactic activity was studied during malaria infection. Two days after infection of Balb/c mice with Plasmodium berghei, an increase in granulocyte number was observed in the blood. A modified Boyden chamber chemotaxis assay was employed to investigate the mechanism of granulocyte accumulation in the blood. Bone marrow cells from normal mice, from mice during a primary lethal infection and from immune mice after challenge were compared. The complement factor C5a showed chemotactic activity for bone marrow cells; a significant decrease of chemotaxis was only observed after 6 days of primary infection. Extracts of spleen, liver and infected erythrocytes lacked chemotactic activity, or caused inhibition of cell migration. Serum from mice with a 2-day primary infection contained chemotactic activity. The active component was heat labile, protease sensitive and had an estimated molecular weight of 250,000.     

Study of chemotactic activity developed by neutrophils from rheumatoid arthritis patients

Neutrophils are the predominant cells accumulated in the synovial fluid (SF) of rheumatoid arthritis (RA) patients. Accumulation of neutrophils may be regarded as a possible way by which neutrophils exert cytotoxic functions. The aim of the present study was to analyze the chemotactic response of neutrophils (PMNs) isolated from the peripheral blood or SF of patients with RA by performing the chemotaxis assay, in which N-formyl-methionyl-leucyl-phenylalanine (FMLP) was used as chemotactic agent. Our results showed that FMLP induced response of peripheral blood neutrophils from 12 patients with RA was similar with the response of 15 healthy controls. A decreased chemotactic response to FMLP was, however, observed in PMNs isolated from the SF of RA patients as comlipared with peripheral blood cells. Therefore, this defective chemotactic ability of neutrophil, was inversely correlated with the number of infiltrating cells in SF. These results indicate that chemotactic ability of neutrophils may be reduced after migration to the SF. Because PMNs chemotaxis in vivo has likely occurred in the presence of serum or SF, we tried to simulate the same conditions in vitro. Therefore, we analyzed the effect of serum or SF on the RA-PMNs chemotaxis. Heat-inactivated serum produced a marked reduction of chemotactic activity developed by PMNs isolated from patients with RA. Notably, a significant increase of chemotactic activity was observed when FMLP and serum stimuli were used together, as compared with the same stimuli used alone. The results suggested that complement activation might interfere with neutrophils chemotaxis. SF amplifies the chemotactic activity of PMNs isolated from peripheral blood of RA patients, but does not affect the chemotaxis developed by PMNs isolated from SF. The data might suggest that several components of SF (IL-8, leukotrien B4, thrombin, platelet-activating factor, etc.) could serve as a potent stimulus for recruitment of neutrophils from periphery into the RA joint. In conclusion, serum or SF components seem to contribute to chemotaxis of neutrophils and play a role in differential killing of PMNs and incidence of infection.     

Monocyte chemotaxis in patients with nonseminomatous testicular carcinoma

Summary The chemotactic responsiveness of blood monocytes was tested in 16 patients with nonseminomatous testicular carcinoma before, during, and after chemotherapy. All the patients initially had monocyte chemotaxis within the normal range. No correlation with the histology of the tumor, the clinical stage, or the presence in serum of -fetoprotein and human chorionic gonadotropin was observed. Plasma from the patients did not inhibit the chemotaxis of normal monocytes, and serum from the patients contained no chemotactic factor inhibitor. During intensive chemotherapy with cis-platinum, bleomycin, and vinblastine a reversible defect in chemotaxis occurred without correlation to the development of fever. Two months after the completion of chemotherapy the chemotactic responsiveness was unchanged compared with pretreatment values. In conclusion, this study shows normal monocyte chemotaxis in patients with testicular carcinoma, which is in contrast to reports on a variety of other solid tumors.     

A quantitative and qualitative study of blood monocytes in patients with bronchogenic carcinoma

Summary Absolute circulating number and functions of blood monocytes (i.e., pinocytosis, phagocytosis, and chemotaxis) were studied in 25 patients with untreated bronchogenic carcinoma and in 28 control subjects. The absolute circulating monocyte count was increased in 20 (80%) of the patients. There was no difference in the pinocytic and phagocytic activity of patient and control monocytes. In contrast, patient monocytes showed depressed chemotactic responsiveness. This defect was more severe in small cell anaplastic carcinoma than in the other histologic types of bronchogenic carcinoma (P=0.001), and may explain the difference in macrophage infiltration seen in solid tumours of the lung. There was no correlation between chemotaxis and clinical stage. Depressed chemotaxis may be related to a plasma factor, since patient plasma inhibited the chemotaxis of control monocytes as well as the activity of chemotactic agents. The defective chemotaxis and the presence of plasma inhibitory activity may interfere with the ability of blood monocytes to accumulate as macrophages in tumour sites. Abbreviations used in this paper are: MCR, monocyte chemotactic response; SAC, small cell anaplastic bronchogenic carcinoma; OBC, non-small cell bronchogenic carcinoma MEM, Eagle's minimal essential medium; CFI, chemotactic factor inhibitor(s); HSA, human serum albumin     

Improved quality of 1H NMR spectroscopic data for enhanced metabolic profiling of low molecular weight metabolites in human serum

NMR based metabolic profiling of blood samples in epidemiological studies can be used for molecular phenotyping and biomarker discovery. Often metabolic changes in blood are more subtle and demand a high quality spectrum especially when looking at low molecular weight compounds. In order to improve ¹H NMR spectroscopic data we compared different serum sample preparation methods. Application of phosphate buffer reduces chemical shift variation, enhances resolution of signal multiplicity, facilitates visual inspection of NMR spectra and annotation of signals compared to traditionally used saline. For analysis of low molecular weight compounds we found that standard 1D spectra of ultrafiltrated serum samples show enhanced spectral quality of small metabolites as compared to transverse relaxation edited spectra (also called Carr–Purcell–Meiboom–Gill, CPMG) spectra of unfiltered serum samples due to improved signal-to-noise ratio. Thus, NMR signals attributable to different amino acids and other small metabolites could readily be detected in spectra of ultrafiltrated serum, but remained invisible in the corresponding CPMG spectra. An OPLS model of fasting blood glucose showed an increase of Q² when using spectra from ultrafiltrated serum (Q² = 0.261) compared to using CPMG spectra (Q² = 0.173). Similar results were observed for OPLS models of BMI (Q² = 0.253 and Q² = 0.216, respectively). Furthermore, a reduction in model dimensionality was observed when using ultrafiltrated serum data. In conclusion we recommend sample preparation of serum samples in phosphate buffer instead of saline. Ultrafiltration of serum samples prior to NMR analysis is beneficial especially for low concentrated small metabolites.     

THE REGULATORY ROLE OF DIVALENT CATIONS IN HUMAN GRANULOCYTE CHEMOTAXIS : Evidence for an Association between Calcium Exchanges and Microtubule Assembly

Optimal human granulocyte chemotaxis has been shown to require both calcium and magnesium. Exposure of granulocytes to three different chemotactic factors (C5a, kallikrein, and dialyzable transfer factor) yielded a rapid calcium release, depressed calcium uptake, and was associated with a shift of calcium out of the cytoplasm and into a granule fraction. Colchicine, sodium azide, and cytochalasin B, in concentrations that inhibited chemotaxis, also inhibited calcium release while low concentrations of cytochalasin B, which enhanced chemotaxis, also enhanced calcium release. Microtubule assembly was visualized both in cells suspended in C5a without a chemotactic gradient and in cells actively migrating through a Micropore filter. The data suggest microtubule assembly is regulated, at least, in part, by the level of cytoplasmic calcium. It is proposed that asymmetric assembly of microtubules may be instrumental in imparting the net vector of motion during chemotaxis.     

A methyl-accepting protein is involved in benzoate taxis in Pseudomonas putida.

Pseudomonas putida is attracted to at least two groups of aromatic acids: a benzoate group and a benzoylformate group. Members of the benzoate group of chemoattractants stimulated the methylation of a P. putida polypeptide with an apparent molecular weight of 60,000 in sodium dodecyl sulfate-polyacrylamide gels. This polypeptide is presumed to be a methyl-accepting chemotaxis protein for several reasons: its molecular weight is similar to the molecular weights of Escherichia coli methyl-accepting chemotaxis proteins, the amount of time required to attain maximal methylation correlated with the time needed for behavioral adaptation of P. putida cells to benzoate, and methylation was stimulated by benzoate only in cells induced for chemotaxis to benzoate. Also, a mutant specifically defective in benzoate taxis failed to show any stimulation of methylation upon addition of benzoate. Benzoylformate did not stimulate protein methylation in cells induced for benzoylformate chemotaxis, suggesting that sensory input from this second group of aromatic-acid attractants is processed through a different kind of chemosensory pathway. The chemotactic responses of P. putida cells to benzoate and benzoylformate were not sensitive to external pH over a range (6.2 to 7.7) which would vary the protonated forms of these weak acids by a factor of about 30. This indicates that detection of cytoplasmic pH is not the basis for aromatic-acid taxis in P. putida.     

Identification of a ubiquitin family protein as a novel neutrophil chemotactic factor

Neutrophil chemotaxis is a process that is essential for the recruitment of neutrophils to an inflamed site. In the present study, we found a remarkable increase in neutrophil chemotactic activity in the lysate of red blood cells (RBC) of mice infected with murine malaria, Plasmodium yoelii. A neutrophil chemotactic factor with an apparent molecular weight of 17 kDa (IP17) was isolated from RBC by a combination of anion-exchange chromatography on DE52 and cation-exchange chromatography on Mono S. A comprehensive GenBank database search of N-terminal amino acid sequences and MALDI-TOF mass analysis of IP17 revealed that IP17 is identical to a murine homologue of ISG15/UCRP, a member of the ubiquitin family of proteins that are inducible by interferon-beta. Recombinant mouse ISG15 showed neutrophil chemotactic activity comparable to that of natural IP17. IP17 showed specific chemotactic activity forward neutrophils and activated neutrophils to induce the release of eosinophil chemotactic factors. These results suggest that the ubiquitin family protein ISG15/UCRP has novel functions in neutrophil-mediated immune mechanisms.     

Characterization of two different factors chemotactic for cancer cells from tumor tissue.

Two different factors chemotactic for cancer cells were extracted in pseudoglobulin fraction of rat ascites hepatoma transplanted tissue. After chromatography on Sephadex G-50 and CM-sephadex, these factors were separated by gel filtration on Sephadex G-100. The factor a was further fractionated by immunoadsorbent chromatography with goat antirat gamma-globulin antibody and then with rabbit antirat hemoglobin antibody; it was a protein with a molecular weight of about 78,000, resembling a chemotactic factor previously reported, and its activity was thermolabile. The previously undescribed factor b was also a protein with a molecular weight of about 14,000 and its activity was thermostable. Intradermal injection of these factors at low concentrations induced an extravascular migration of circulating tumor cells and formation of metastatic secondary tumors; and little difference in the in vivo effect between these factors was observed. It was thus assumed that the combined action of these two factors may be involved in malignant invasion.     

Selective chemotaxis of subsets of B lymphocytes from gut-associated lymphoid tissue and its implications for the recruitment of mucosal plasma cells

As they differentiate, precursor cells from the gut-associated lymphoid tissue are known to travel via the lymphatic system to the blood and then preferentially to home to various mucosal and exocrine sites such as the lamina propria of the gut and the lactating mammary gland, where they give rise to IgA-secreting plasma cells. The present study, directed at the mechanism by which the circulating precursors of mucosal IgA plasma cells selectively lodge in characteristic locations, explored the hypothesis that such homing is due to a locally produced chemotactic factor and that milk might be a source of such a factor. Subsets of lymphocytes bearing particular surface markers and purified by panning from lymph nodes of mice were examined in a micropore chemotaxis assay to search for the presence of chemotactic activity in mouse milk. The globulin fraction of whey was shown to contain a nondialyzable factor that is chemotactic for IgA (and also IgG)-positive lymphocytes when these are obtained from mesenteric lymph nodes as a source of mucosal-associated lymphoid tissue. Lymphocytes from peripheral lymph nodes, nonmucosal associated, were unaffected as were surface IgM-positive lymphocytes and T lymphocytes obtained from mesenteric nodes. Chemotactic activity for IgA lymphocytes was undetectable in mouse serum. The data are consistent with the idea that precursors of mucosal IgA plasma cells home to mucosal and exocrine sites in response to a specific chemotactic factor elaborated by local differentiated epithelial cells.     

Chemotaxis of Pseudomonas aeruginosa: involvement of methylation.

The involvement of a protein methyl transfer system in the chemotaxis of Pseudomonas aeruginosa was investigated. When a methionine auxotroph of P. aeruginosa was starved for methionine, chemotaxis toward serine, measured by a quantitative capillary assay, was reduced 80%, whereas background motility was unaffected or increased. When unstarved bacteria were labeled with L-[methyl-3H]methionine, a labeled species of 73,000 molecular weight which was methylated in response to stimulation by L-serine was identified. Under appropriate electrophoretic conditions, the 73,000 molecular weight species was resolved into two bands, both of which responded to stimulation by L-serine, L-arginine, and alpha-aminoisobutyrate (AIB) with an increased incorporation of methyl label. Arginine, which elicited the strongest chemotactic response in the capillary assay, also stimulated the greatest methylation response. Methylation of the 73,000 molecular weight species reached a maximum 10 min after stimulation by AIB and returned to the unstimulated level upon removal of the AIB. In vitro labeling of cell extracts with S-adenosyl[methyl-3H]methionine indicated that the 73,000 molecular weight species are methylated by an S-adenosylmethionine-mediated reaction. These results indicate that chemotaxis of P. aeruginosa toward amino acids is mediated by dynamic methylation and demethylation of methyl-accepting chemotaxis proteins analogous to those of the enteric bacteria.     

卵巢激素对肺泡巨噬细胞趋化活性的影响

为探讨卵巢激素对非性器官肺脏的防御功能有无影响,本研究以肺泡巨噬细胞（ＡＭ）趋化活性为指标,观察了卵巢激素对成年雄性大鼠离体ＡＭ的趋化活性的作用。结果显示：不同浓度的酵母多糖激活血清与ＡＭ在体外培养３．５ｈ,对ＡＭ趋化性有良好的线性关系。雌二醇能抑制ＡＭ的趋化活性,量效关系显著（ｒ＝－０．９２８０,Ｐ＜０．０１）;而孕酮则促进ＡＭ的趋化活性,亦具有剂量依从性（ｒ＝０．９９７５,Ｐ＜０．０１）。提示：卵巢激素除参与性器官功能的调节外,对于非性器官肺脏的防御功能亦具有一定的调控作用。     

Mammalian sperm chemotaxis and its association with capacitation

Much progress has been made in recent years in establishing mammalian sperm chemotaxis and understanding sperm capacitation. Thus far, chemotaxis to follicular fluid has been established by a variety of means in human and mouse spermatozoa. It was found that only a small fraction of a given sperm population (averaging around 10%) is chemotactically responsive and that this fraction constitutes capacitated (ripe) spermatozoa. Both the chemotactic responsiveness and the capacitated state are transient (with a lifetime of 50 min to 4 h) and they occur only once in the sperm's lifetime. It has been proposed that the role of sperm chemotaxis in mammals (at least in humans) is selective recruitment of capacitated spermatozoa for fertilizing the egg, and that the role of the continuous replacement of chemotactic/capacitated spermatozoa is to prolong the time during which capacitated spermatozoa are available in the female reproductive tract. The sperm chemoattractants have not been identified, but they appear to be heat‐stable peptides. Although the molecular mechanism and the in vivo location of sperm chemotaxis are not known, a number of possible mechanisms and locations are discussed. Dev. Genet. 25:87–94, 1999. © 1999 Wiley‐Liss, Inc.     

Myelin basic protein inhibits formyl-peptide induced chemotaxis in human neutrophils

Myelin basic protein, one of the major membrane protein component of the central nervous system, was used to probe the molecular mechanism of cellular activation. Pre-treatment of human neutrophils with myelin basic protein selectively inhibits the formyl-peptide-induced chemotaxis, without affecting chemotaxis evoked by casein and activated serum. Furthermore, both the degranulation and superoxide anion production stimulated by the chemotactic peptide are not modified by the prior treatment of the neutrophils with myelin basic protein.     

河蚌血细胞对细菌趋化移动的初步研究

采用改进的毛细管法 ,研究了圆背角无齿蚌 (Anodontawoodianapacifica)和三角帆蚌 (Hyriopsiscum ingii)两种淡水河蚌离体血细胞对两种水体中常见病原细菌的趋化移动作用 ,及血清对其的影响。结果显示 ,两种河蚌的离体血细胞对细菌都具有趋化移动作用 ,产生趋化移动的血细胞数量都显著高于无细菌的对照组 (P <0 0 5 )。在有血清时 ,血细胞对荧光极毛杆菌 (Pseudomonasfluorescens)的趋化移动活性略高于肠型点状气单孢菌 (Aeromonaspunctataf.intestinalis) ,圆背角无齿蚌离体血细胞的趋化移动能力显著高于三角帆蚌 (P <0 0 5 )。血清对河蚌离体血细胞的趋化移动作用有显著的促进作用 (P <0 0 5 )。     

Ostertagia ostertagi: excretory secretory chemotactic substance from infective larvae as cause of eosinophil locomotion

The infective (third stage) larva of Ostertagia ostertagi produced an excretory secretory substance that is chemotactic for bovine eosinophils. The eosinophil chemotactic substance was present in supernatants of larval cultures within 6 hr of incubation in Eagle's Minimum Essential Medium containing 25 mM N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid. The substance responsible for eosinophil chemotaxis has activity in small amounts, a molecular weight (MW) greater than 2000, and is heat labile at 100 C. Bovine eosinophils appeared to have a receptor for the chemotactic excretory secretory substance which is either identical or structurally similar to the previously described eosinophil chemotactic substance present in soluble third stage larval extracts. The larval substance may cause eosinophil accumulation in the abomasal tissue of cattle with ostertagiasis.     

Specific inhibitor of complement (C5)-derived chemotactic activity in systemic lupus erythematosus related antigenically to the Bb fragment of human factor B

Serum and plasma from patients with active systemic lupus erythematosus contain a specific inhibitor of complement (C5)-derived chemotactic activity. We found that the inhibitor is antigenically related to the Bb fragment of complement factor B. Lupus plasma and purified inhibitor significantly reduced the chemotactic activity of zymosan-treated normal serum, an effect that was abolished by antibodies to factor B. Similar results were obtained when purified Bb was used. Neither purified inhibitor nor Bb inhibited the chemotactic activity of purified human C5a or C5a des Arg. As reported previously, the chemotactic activity of C5a des Arg was enhanced significantly by the addition of an anionic polypeptide (cochemotaxin) present in normal serum and plasma. Interestingly, both purified lupus inhibitor and Bb inhibited the chemotactic activity exhibited by mixtures of C5a des Arg and its cochemotaxin. This effect was due, most likely, to their ability to neutralize the enhancing effect of the cochemotaxin on the chemotactic activity of C5a des Arg. Immunoelectrophoresis and western blots revealed that the purified inhibitor reacted with anti-factor B and exhibited a similar charge and molecular weight as purified Bb.     

The combined action of hyaluronic acid and fibronectin stimulates neutrophil migration

Previous investigations have demonstrated that the chemotactic response of polymorphonuclear leukocytes (PMN) was stimulated by hyaluronic acid (HA) when serum was present. The aim of the present investigation was to identify the serum factor necessary for the stimulation of PMN chemotaxis by HA. By means of gel filtration, the m.w. of the serum component was shown to be greater than 350,000. Immunoprecipitation of serum with anti-fibronectin, but not with anti-IgM and anti-alpha 2-macroglobulin, inhibited the stimulation of PMN chemotaxis by HA. Preincubation of PMN with HA (10 to 500 micrograms/L) and isolated fibronectin (0.1 to 100 mg/L) significantly stimulated the chemotactic response of PMN. Also, random migration of PMN was significantly increased by preincubation of the cells with HA (10 to 500 micrograms/L) and isolated fibronectin (50 to 200 mg/L). Additionally, PMN preincubated with HA (10 to 50 micrograms/L) and with fibronectin (10 to 50 mg/L) added afterwards, and PMN preincubated with fibronectin (10 mg/L) and with HA (5 to 10 micrograms/L) added after the preincubation, showed a significant stimulation of the chemotactic response. PMN preincubated with serum and chondroitin sulfate, or with fibrinogen and HA, demonstrated no stimulation of the chemotactic response. The present investigation suggests that the combined action of HA and fibronectin, which probably takes place at the cellular membrane, is a major mechanism in the HA-mediated stimulation of PMN migration.     

Appearance of chemotactic responsiveness to elastin peptides by developing fetal bovine ligament fibroblasts parallels the onset of elastin production

We studied chemotaxis to elastin peptides by bovine ligamentum nuchae fibroblasts to determine whether there is a developmental association between chemotactic responsiveness to elastin and expression of the elastin phenotype. Undifferentiated ligament cells demonstrate chemotactic responsiveness to platelet-derived growth factor and fibronectin, known chemoattractants for fibroblasts, but do not show chemotaxis to elastin peptides. After matrix-induced differentiation, however, young cells display a positive chemotactic response to elastin that persists even after the cells are removed from the matrix substratum. Matrix-induced chemotaxis to elastin could be inhibited selectively by incorporation of bromodeoxyuridine into DNA of undifferentiated cells before (but not after) contact with inducing matrix. These results show that the appearance of chemotaxis to elastin peptides parallels the onset of elastin synthesis and suggests that the acquisition of chemotactic responsiveness to elastin and expression of the elastin phenotype are affected by the same inducing elements or processes and may be closely coupled in development.