Neutrophil migration under spatially-varying chemoattractant gradient profiles.

Chemotaxis plays an important role in biological processes such as cancer metastasis, embryogenesis, wound healing, and immune response. Neutrophils are the frontline defenders against invasion of foreign microorganisms into our bodies. To achieve this important immune function, a neutrophil can sense minute chemoattractant concentration differences across its cell body and effectively migrate toward the chemoattractant source. Furthermore, it has been demonstrated in various studies that neutrophils are highly sensitive to changes in the surrounding chemoattractant environments, suggesting the role of a chemotactic memory for processing the complex spatiotemporal chemical guiding signals. Using a microfluidic device, in the present study we characterized neutrophil migration under spatially varying profiles of interleukine-8 gradients, which consist of three spatially ordered regions of a shallow gradient, a steep gradient and a nearly saturated gradient. This design allowed us to examine how neutrophils migrate under different chemoattractant gradient profiles, and how the migratory response is affected when the cell moves from one gradient profile to another in a single experiment. Our results show robust neutrophil chemotaxis in the shallow and steep gradient, but not the saturated gradient. Furthermore, neutrophils display a transition from chemotaxis to flowtaxis when they migrate across the steep gradient interface, and the relative efficiency of this transition depends on the cell's chemotaxis history. Finally, some neutrophils were observed to adjust their morphology to different gradient profiles.

Effects of Hyriopsis cumingii polysaccharides on angiogenesis, macrophage chemotaxis, proliferation and phagocytosis.

Anti-angiogenic activities of crude Hyriopsis cumingii polysaccharides (HCPS) and its purified fractions (HCPS-1, HCPS-2 and HCPS-3) were evaluated in vivo using the chicken embryo chorioallantoic membrane (CAM) assay. The promoting effects of crude HCPS and its purified fractions on the chemotaxis, proliferation and phagocytosis of peritoneal macrophage were tested by cell model in vitro and cyclophosphamide-induced immuno-suppression animal model in vivo. The results showed that HCPS could significantly suppress the neovascularization of chicken embryo CAM and promote peritoneal macrophage migrating to monocyte chemotactic protein-1 (MCP-1), propagating and devouring sheep red blood cell (SRBC) in a dose-dependent manner. In addition, HCPS-3 showed stronger immunostimulatory activities in vitro than crude HCPS, HCPS-1 and HCPS-2. The beneficial effects of HCPS on the immune system might be, at least in part, attributed to the improvement of chemotaxis, proliferation and phagocytosis of peritoneal macrophage. All these results suggest that HCPS is a potential immunoenhancing and anti-tumor agent.

Usefulness of controlled release of growth factors in investigating the early events of dentin-pulp regeneration.

INTRODUCTION: Little information is yet available on the signals involved in progenitor cell migration that precede reparative dentin synthesis. Our aim was to investigate the effect of the controlled release of fibroblast growth factor (FGF)-2 and transforming growth factor ß1 (TGF-ß1) on permanent teeth pulp cell proliferation and progenitor cell migration. METHODS: FGF-2 and TGF-ß1 were encapsulated into a biodegradable polymer matrix of lactide and glycolide. Human pulp cells were prepared from third molars, and progenitor cells were sorted by STRO-1. The synthesized microsphere toxicity was checked with the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide test. The growth factor release kinetics were checked by an enzyme-linked immunosorbent assay while maintaining their biological activity and were evaluated by investigating their effects on pulp cell proliferation. Their chemotactic potential was investigated on STRO-1-sorted cells in a migration chamber on Matrigel (Cambrex Bio Science, Walkersville, MD). RESULTS: The cell viability was unaffected by the presence of microspheres. The released amount of FGF-2 and TGF-ß1 from the microspheres was maintained after 21 days. Increasing the FGF-2-loaded microsphere concentration or the release period significantly increased dental pulp cell proliferation. TGF-ß1 acted as a potent chemotactic factor of STRO-1-sorted cells. CONCLUSIONS: Encapsulating TGF-ß1 and FGF-2 in a biodegradable polymer of lactide and glycolide microsphere allowed a sustained release of growth factors and provided a protection to their biological activities. Our results clearly show the usefulness of growth factor controlled release in investigating the early events of pulp/dentin regeneration. It provides additional data on the signals required for vital pulp therapy and future tissue engineering.

Evaluation of low density polyethylene and nylon for delivery of synthetic mosquito attractants.

BACKGROUND: Synthetic odour baits present an unexploited potential for sampling, surveillance and control of malaria and other mosquito vectors. However, application of such baits is impeded by the unavailability of robust odour delivery devices that perform reliably under field conditions. In the present study the suitability of low density polyethylene (LDPE) and nylon strips for dispensing synthetic attractants of host-seeking Anopheles gambiae mosquitoes was evaluated. METHODS: Baseline experiments assessed the numbers of An. gambiae mosquitoes caught in response to low density polyethylene (LDPE) sachets filled with attractants, attractant-treated nylon strips, control LDPE sachets, and control nylon strips placed in separate MM-X traps. Residual attraction of An. gambiae to attractant-treated nylon strips was determined subsequently. The effects of sheet thickness and surface area on numbers of mosquitoes caught in MM-X traps containing the synthetic kairomone blend dispensed from LDPE sachets and nylon strips were also evaluated. Various treatments were tested through randomized 4 × 4 Latin Square experimental designs under semi-field conditions in western Kenya. RESULTS: Attractant-treated nylon strips collected 5.6 times more An. gambiae mosquitoes than LDPE sachets filled with the same attractants. The attractant-impregnated nylon strips were consistently more attractive (76.95%; n = 9,120) than sachets containing the same attractants (18.59%; n = 2,203), control nylon strips (2.17%; n = 257) and control LDPE sachets (2.29%; n = 271) up to 40 days post-treatment (P < 0.001). The higher catches of mosquitoes achieved with nylon strips were unrelated to differences in surface area between nylon strips and LDPE sachets. The proportion of mosquitoes trapped when individual components of the attractant were dispensed in LDPE sachets of optimized sheet thicknesses was significantly higher than when 0.03 mm-sachets were used (P < 0.001). CONCLUSION: Nylon strips continuously dispense synthetic mosquito attractants several weeks post treatment. This, added to the superior performance of nylon strips relative to LDPE material in dispensing synthetic mosquito attractants, opens up the opportunity for showcasing the effectiveness of odour-baited devices for sampling, surveillance and control of disease vectors.

Controlling airborne cues to study small animal navigation.

Small animals such as nematodes and insects analyze airborne chemical cues to infer the direction of favorable and noxious locations. In these animals, the study of navigational behavior evoked by airborne cues has been limited by the difficulty of precisely controlling stimuli. We present a system that can be used to deliver gaseous stimuli in defined spatial and temporal patterns to freely moving small animals. We used this apparatus, in combination with machine-vision algorithms, to assess and quantify navigational decision making of Drosophila melanogaster larvae in response to ethyl acetate (a volatile attractant) and carbon dioxide (a gaseous repellant).

Chemotactic cell trapping in controlled alternating gradient fields.

Directed cell migration toward spatio-temporally varying chemotactic stimuli requires rapid cytoskeletal reorganization. Numerous studies provide evidence that actin reorganization is controlled by intracellular redistribution of signaling molecules, such as the PI4,5P2/PI3,4,5P3 gradient. However, exploring underlying mechanisms is difficult and requires careful spatio-temporal control of external chemotactic stimuli. We designed a microfluidic setup to generate alternating chemotactic gradient fields for simultaneous multicell exposure, greatly facilitating statistical analysis. For a quantitative description of intracellular response dynamics, we apply alternating time sequences of spatially homogeneous concentration gradients across 300 µm, reorienting on timescales down to a few seconds. Dictyostelium discoideum amoebae respond to gradient switching rates below 0.02 Hz by readapting their migration direction. For faster switching, cellular repolarization ceases and is completely stalled at 0.1 Hz. In this "chemotactically trapped" cell state, external stimuli alternate faster than intracellular feedback is capable to respond by onset of directed migration. To investigate intracellular actin cortex rearrangement during gradient switching, we correlate migratory cell response with actin repolymerization dynamics, quantified by a fluorescence distribution moment of the GFP fusion protein LimEΔcc. We find two fundamentally different cell polarization types and we could reveal the role of PI3-Kinase for cellular repolarization. In the early aggregation phase, PI3-Kinase enhances the capability of D. discoideum cells to readjust their polarity in response to spatially alternating gradient fields, whereas in aggregation competent cells the effect of PI3-Kinase perturbation becomes less relevant.

Adapt locally and act globally: strategy to maintain high chemoreceptor sensitivity in complex environments.

In bacterial chemotaxis, several types of ligand-specific receptors form mixed clusters, wherein receptor-receptor interactions lead to signal amplification and integration. However, it remains unclear how a mixed receptor cluster adapts to individual stimuli and whether it can differentiate between different types of ligands. Here, we combine theoretical modeling with experiments to reveal the adaptation dynamics of the mixed chemoreceptor cluster in Escherichia coli. We show that adaptation occurs locally and is ligand-specific: only the receptor that binds the external ligand changes its methylation level when the system adapts, whereas other types of receptors change methylation levels transiently. Permanent methylation crosstalk occurs when the system fails to adapt accurately. This local adaptation mechanism enables cells to differentiate individual stimuli by encoding them into the methylation levels of corresponding types of chemoreceptors. It tunes each receptor to its most responsive state to maintain high sensitivity in complex environments and prevents saturation of the cluster by one signal.

Microfluidic monitoring of Pseudomonas aeruginosa chemotaxis under the continuous chemical gradient.

This study presents a microfluidic approach for the rapid analysis of bacterial chemotaxis in response to chemical gradients. The diffusional mixing of laminar flow continuously generates a stable chemical gradient in a microfluidic device. For the proof of concept, we have investigated the effects of the attractant peptone and repellent trichloroethylene (TCE) on chemotactic responses of wild type Pseudomonas aeruginosa PAO1 and chemotactic mutant PC4. The microfluidic method clearly demonstrates that P. aeruginosa PAO1 is attracted to peptone and repelled from TCE, whereas PC4 shows non-chemotactic behavior. In addition, the analysis of PAO1 chemotaxis on 20 amino acids revealed the effective concentration range of each amino acid as a chemoeffector. Not only does the microfluidic approach facilitate the quantitative information of chemotaxis, which gives an insight into understanding the mechanism of P. aeruginosa motility, but it also provides a useful tool for the rapid monitoring of bacterial chemotaxis in a reproducible experimental manner.

Intracapillary leucocyte accumulation as a novel antihaemorrhagic mechanism in acute pancreatitis in mice.

BACKGROUND: Pancreatic infiltration by leucocytes represents a hallmark in acute pancreatitis. Although leucocytes play an active role in the pathophysiology of this disease, the relation between leucocyte activation, microvascular injury and haemorrhage has not been adequately addressed. METHODS: We investigated intrapancreatic leucocyte migration, leucocyte extravasation and pancreatic microperfusion in different models of oedematous and necrotising acute pancreatitis in lys-EGFP-ki mice using fluorescent imaging and time-lapse intravital microscopy. RESULTS: In contrast to the current paradigm of leucocyte recruitment, the initial event of leucocyte activation in acute pancreatitis was represented through a dose- and time-dependent occlusion of pancreatic capillaries by intraluminally migrating leucocytes. Intracapillary leucocyte accumulation (ILA) resulted in dense filling of almost all capillaries close to the area of inflammation and preceded transvenular leucocyte extravasation. ILA was also initiated by isolated exposure of the pancreas to interleukin 8 or fMLP, demonstrating the causal role of chemotactic stimuli in the induction of ILA. The onset of intracapillary leucocyte accumulation was strongly inhibited in LFA-1(-/-) and ICAM-1(-/-) mice, but not in Mac-1(-/-) mice. Moreover, prevention of intracapillary leucocyte accumulation led to the development of massive capillary haemorrhages and transformed mild pancreatitis into lethal haemorrhagic disease. CONCLUSIONS: ILA represents a novel protective and potentially lifesaving mechanism of haemostasis in acute pancreatitis. This process depends on expression of LFA-1 and ICAM-1 and precedes the classical steps of the leucocyte recruitment cascade.

Planar microfluidic chamber for generation of stable and steep chemoattractant gradients.

The extracellular availability of growth factors, hormones, chemokines, and neurotransmitters under gradient conditions is required for directional cellular responses such as migration, axonal pathfinding, and tissue patterning. These responses are, in turn, important in disease and developmental processes. This article addresses critical barriers toward devising a chemotaxis assay that is broadly applicable for different kinds of cancer cells through the design of a microfluidic chamber that produces a steep gradient of chemoattractant. Photolithography was used to create microchannels for chemoattractant delivery, flow diversion barriers/conduits, and small outlets in the form of apertures. The 1-microm apertures were made at the active surface by uncapping a thin (1.5 microm) layer of AZ1518. This process also created a vertical conduit that diverted the flow such that it occurred perpendicularly to the active, experimental surface where the gradients were measured. The other side of the vertical conduit opened to underlying 20-microm deep channels that carried microfluidic flows of tracer dyes/growth factors. Modeled data using computational fluid dynamics produced gradients that were steep along the horizontal, active surface. This simulation mirrors empirically derived gradients obtained from the flow analyses of fluorescent compounds. The open chamber contains a large buffer volume, which prevents chemoattractant saturation and permits easy cell and compound manipulation. The technique obviates the use of membranes or laminar flow that may hinder imaging, rinsing steps, cell seeding, and treatment. The utility of the chamber in the study of cell protrusion, an early step during chemotaxis, was demonstrated by growing cancer cells in the chamber, inducing a chemoattractant gradient using compressed air at 0.7 bar, and performing time-lapse microscopy. Breast cancer cells responded to the rapidly developed and stable gradient of epidermal growth factor by directing centroid positions toward the gradient and by forming a leading edge at a speed of 0.45 microm/min.

Tissue- and stimulus-dependent role of phosphatidylinositol 3-kinase isoforms for neutrophil recruitment induced by chemoattractants in vivo.

PI3K plays a fundamental role in regulating neutrophil recruitment into sites of inflammation but the role of the different isoforms of PI3K remains unclear. In this study, we evaluated the role of PI3Kgamma and PI3Kdelta for neutrophil influx induced by the exogenous administration or the endogenous generation of the chemokine CXCL1. Administration of CXCL1 in PI3Kgamma(-/-) or wild-type (WT) mice induced similar increases in leukocyte rolling, adhesion, and emigration in the cremaster muscle when examined by intravital microscopy. The induction of neutrophil recruitment into the pleural cavity or the tibia-femoral joint induced by the injection of CXCL1 was not significantly different in PI3Kgamma(-/-) or WT mice. Neutrophil influx was not altered by treatment of WT mice with a specific PI3Kdelta inhibitor, IC87114, or a specific PI3Kgamma inhibitor, AS605240. The administration of IC87114 prevented CXCL1-induced neutrophil recruitment only in presence of the PI3Kgamma inhibitor or in PI3Kgamma(-/-) mice. Ag challenge of immunized mice induced CXCR2-dependent neutrophil recruitment that was inhibited by wortmannin or by blockade of and PI3Kdelta in PI3Kgamma(-/-) mice. Neutrophil recruitment to bronchoalveolar lavage induced by exogenously added or endogenous production of CXCL1 was prevented in PI3Kgamma(-/-) mice. The accumulation of the neutrophils in lung tissues was significantly inhibited only in PI3Kgamma(-/-) mice treated with IC87114. Neutrophil recruitment induced by exogenous administration of C5a or fMLP appeared to rely solely on PI3Kgamma. Altogether, our data demonstrate that there is a tissue- and stimulus-dependent role of PI3Kgamma and PI3Kdelta for neutrophil recruitment induced by different chemoattractants in vivo.

An information-theoretic characterization of the optimal gradient sensing response of cells.

Many cellular systems rely on the ability to interpret spatial heterogeneities in chemoattractant concentration to direct cell migration. The accuracy of this process is limited by stochastic fluctuations in the concentration of the external signal and in the internal signaling components. Here we use information theory to determine the optimal scheme to detect the location of an external chemoattractant source in the presence of noise. We compute the minimum amount of mutual information needed between the chemoattractant gradient and the internal signal to achieve a prespecified chemotactic accuracy. We show that more accurate chemotaxis requires greater mutual information. We also demonstrate that a priori information can improve chemotaxis efficiency. We compare the optimal signaling schemes with existing experimental measurements and models of eukaryotic gradient sensing. Remarkably, there is good quantitative agreement between the optimal response when no a priori assumption is made about the location of the existing source, and the observed experimental response of unpolarized Dictyostelium discoideum cells. In contrast, the measured response of polarized D. discoideum cells matches closely the optimal scheme, assuming prior knowledge of the external gradient-for example, through prolonged chemotaxis in a given direction. Our results demonstrate that different observed classes of responses in cells (polarized and unpolarized) are optimal under varying information assumptions.

F2L, a peptide derived from heme-binding protein, inhibits formyl peptide receptor-mediated signaling.

F2L is an acetylated amino-terminal peptide derived from the cleavage of the human heme-binding protein. Very recently, F2L was identified as an endogenous chemoattractant peptide acting specifically through formyl peptide receptor-like (FPRL)2. In the present study, we report that F2L stimulates chemotactic migration in human neutrophils. However, F2L inhibits formyl peptide receptor (FPR) and FPRL1 activities, resulting in the complete inhibition of intracellular calcium increases, and superoxide generation induced by N-formyl-Met-Leu-Phe, MMK-1, or Trp-Lys-Tyr-Met-Val-d-Met (WKYMVm) in human neutrophils. In terms of the inhibitory role of F2L on FPR- and FPRL-mediated signaling, we found that F2L competitively inhibits the binding of (125)I-WKYMVm to its specific receptors, FPR and FPRL1. F2L is the first endogenous molecule that inhibits FPR- and FPRL1-mediated signaling, and is expected to be useful in the study of FPR and FPRL1 signaling and in the development of drugs to treat diseases involving the FPR family of receptors.

A mathematical model for neutrophil gradient sensing and polarization.

Directed cell migration in response to chemical cues, also known as chemotaxis, is an important physiological process involved in wound healing, foraging, and the immune response. Cell migration requires the simultaneous formation of actin polymers at the leading edge and actomyosin complexes at the sides and back of the cell. An unresolved question in eukaryotic chemotaxis is how the same chemoattractant signal determines both the cell's front and back. Recent experimental studies have begun to reveal the biochemical mechanisms necessary for this polarized cellular response. We propose a mathematical model of neutrophil gradient sensing and polarization based on experimentally characterized biochemical mechanisms. The model demonstrates that the known dynamics for Rho GTPase and phosphatidylinositol-3-kinase (PI3K) activation are sufficient for both gradient sensing and polarization. In particular, the model demonstrates that these mechanisms can correctly localize the "front" and "rear" pathways in response to both uniform concentrations and gradients of chemical attractants, including in actin-inhibited cells. Furthermore, the model predictions are robust to the values of many parameters. A key result of the model is the proposed coincidence circuit involving PI3K and Ras that obviates the need for the "global inhibitors" proposed, though never experimentally verified, in many previous mathematical models of eukaryotic chemotaxis. Finally, experiments are proposed to (in)validate this model and further our understanding of neutrophil chemotaxis.

Robustness analysis of the E. coli chemosensory system to perturbations in chemoattractant concentrations.

MOTIVATION: Cells of Escherichia coli sense and move toward chemical attractants. This is done through an intricate sensory system that eventually directs the movements of flagellae which regulate the 'runs' and 'tumbles' of the cells. Under realistic conditions, chemical stimuli often fluctuate due to noise from the environment. The effect of noise on the chemosensory system has been investigated here through the sensitivity coefficients of the concentrations of four key proteins--the phosphorylated forms of CheA, CheB and CheY, and the FliM-CheY-P complex--that govern chemotactic motility. The letter P denotes phosphorylation. RESULTS: All sensitivities increased with time and then stabilized. However, the four sets of sensitivities differed in their magnitudes and the durations of their transient phases before stabilization. CheA-P was the least sensitive and CheY-P the most sensitive. Moreover, while the sensitivities of CheA-P, CheB-P and CheY-P increased with chemoattractant concentration, that of the FliM complex decreased. These differences have been interpreted in terms of the mechanism of the chemosensory system and they have important implications for practical applications of chemotaxis.

Effects of adaptation in maintaining high sensitivity over a wide range of backgrounds for Escherichia coli chemotaxis.

An allosteric model is developed to study the cooperative kinase response of wild-type (wt) Escherichia coli cells to the chemoattractant MeAsp in different ambient MeAsp concentrations. The model, together with wt dose response data, reveals the underlying mechanism for E. coli's ability to maintain high sensitivity over a wide range of backgrounds. We find: 1), Adaptation tunes the system to the steepest part of the dose response curve, where the sensitivity to a given type of stimulus is amplified by the number of corresponding receptors in the (mixed) functional receptor complex. A lower bound on the number of Tar receptor dimers (Na) in the complex Na>approximately 6 is obtained from the measured sensitivity. 2), Accurate adaptation synchronizes the kinase activities from different (uncoupled) receptor complexes in a single cell and is crucial in maintaining the high Hill coefficient in the (population averaged) kinase response curve. 3), The wide dynamic range of the high sensitivity can be explained in our model by either having a very small ratio between ligand dissociation constants of the inactive and the active receptors C=0.006, Na=6, and a (methylation level independent) dissociation constant for the inactive Tar receptor K=18.2 microM or by having K and/or Na increase with receptor methylation level together with a larger value of C>0.01. Specific experiments are suggested to distinguish these two scenarios. 4), The receptor occupancy in a wt cell should also adapt and exhibit a slow (approximately logarithmic) dependence on the ligand concentration in the adapted state; this general prediction can be tested experimentally to verify/falsify our model.

Progesterone at the picomolar range is a chemoattractant for mammalian spermatozoa.

By means of a videomicroscopy system and a computer image analysis, we performed chemotaxis assays to detect true chemotaxis in human spermatozoa, in parallel to immunohistochemistry detection of progesterone inside the cumulus cells. Progesterone indeed chemotactically guides mammalian spermatozoa at very low hormone concentrations, and the cumulus oophorus could be a potential place for sperm chemotaxis mediated by progesterone in vivo.

Stem cell factor stimulates the chemotaxis, integrin upregulation, and survival of human basophils.

BACKGROUND: Little is known about the mechanisms that regulate the selective recruitment of basophils to sites of allergic inflammation. OBJECTIVE: Here we examine the role of stem cell factor (SCF) in the regulation of basophil function. METHODS: Human basophils were isolated from peripheral blood, and their migration was investigated in chemotaxis assays. Apoptosis was detected by means of annexin V and propidium iodide staining. The expression of cell-surface molecules was measured by means of flow cytometry. RESULTS: SCF amplified the chemotactic responsiveness of human peripheral blood basophils to the chemoattractants eotaxin, monocyte chemotactic protein 2 and macrophage inflammatory protein 1alpha, and C5a, without being chemotactic or chemokinetic by itself. SCF synergized with chemoattractants in causing basophil upregulation of the integrin CD11b, and this effect was inhibited by a c-kit antibody, the tyrosine kinase inhibitor imatinib mesylate (STI-571), and a phosphatidylinositol 3 kinase inhibitor but not by inhibitors of p38 mitogen-activated protein kinase or mitogen-activated protein kinase/extracellular signal-regulated kinase kinase. Basophils bound fluorescence-labeled SCF and expressed its receptor, c-kit, which was markedly upregulated in culture for 24 to 48 hours in the presence of IL-3. Moreover, SCF prolonged basophil survival in concert with IL-3 by delaying apoptosis. These effects of SCF were selective for basophils because chemotaxis and CD11b upregulation of eosinophils or neutrophils were unchanged. CONCLUSION: SCF might be an important selective modulator of basophil function through a phosphatidylinositol 3 kinase-dependent pathway.

Directed cell migration via chemoattractants released from degradable microspheres.

Chemotaxis, cell migration directed by spatial concentration gradients of chemoattractant molecules, is critical for proper function of the immune system. Materials capable of generating defined chemoattractant gradients via controlled release may be useful for the design of improved vaccines and immunotherapies that draw specific cells to an immunization site. To this end, we encapsulated formyl-Nle-Leu-Phe-Nle-Tyr-Lys (fN'LFN'YK) peptides or macrophage inflammatory protein-3alpha (MIP-3alpha or CCL20) in degradable poly(lactide-co-glycolide) microspheres that provided sustained release for more than 2 weeks in vitro. fN'LFN'YK and MIP-3alpha chemoattract dendritic cells (DCs), the key antigen-presenting cells involved in generation of primary immune responses, and their precursors, monocytes. Using an in vitro videomicroscopy migration assay, we detected strong chemotaxis of human monocytes and monocyte-derived DCs through 3D collagen gels toward microspheres releasing fN'LFN'YK. Similarly, microparticles releasing MIP-3alpha were able to attract mouse bone marrow-derived dendritic cells. Strikingly, prolonged attraction of DCs from distances up to 500 microm from the source to the point of contact with individual microspheres was observed. Such microspheres could be of general interest for the design of vaccines that promote adaptive immunity and as a platform for studying the biology of chemotaxis in vitro and in vivo.

Proinflammatory activities of S100: proteins S100A8, S100A9, and S100A8/A9 induce neutrophil chemotaxis and adhesion.

S100A8 and S100A9 are small calcium-binding proteins that are highly expressed in neutrophil and monocyte cytosol and are found at high levels in the extracellular milieu during inflammatory conditions. Although reports have proposed a proinflammatory role for these proteins, their extracellular activity remains controversial. In this study, we report that S100A8, S100A9, and S100A8/A9 caused neutrophil chemotaxis at concentrations of 10(-12)-10(-9) M. S100A8, S100A9, and S100A8/A9 stimulated shedding of L-selectin, up-regulated and activated Mac-1, and induced neutrophil adhesion to fibrinogen in vitro. Neutralization with Ab showed that this adhesion was mediated by Mac-1. Neutrophil adhesion was also associated with an increase in intracellular calcium levels. However, neutrophil activation by S100A8, S100A9, and S100A8/A9 did not induce actin polymerization. Finally, injection of S100A8, S100A9, or S100A8/A9 into a murine air pouch model led to rapid, transient accumulation of neutrophils confirming their activities in vivo. These studies 1) show that S100A8, S100A9, and S100A8/A9 are potent stimulators of neutrophils and 2) strongly suggest that these proteins are involved in neutrophil migration to inflammatory sites.