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Cell migration is central to development, wound healing, tissue regeneration, and immunity. Despite extensive knowledge of muscle regeneration, myoblast migration during regeneration is not well understood. C2C12 mouse myoblast migration and morphology were investigated using a triple-docking polydimethylsiloxane-based microfluidic device in which cells moved under gravity-driven laminar flow on uniform (=) collagen (CN=), fibronectin (FN=), or opposing gradients (CN-FN or FN-CN). In haptotaxis experiments, migration was faster on FN= than on CN=. At 10 h, cells were more elongated on FN-CN and migration was faster than on the CN-FN substrate. Net migration distance on FN-CN at 10 h was greater than on CN-FN, as cells rapidly entered the channel as a larger population (bulk-cell movement, wave 1 ). Hepatocyte growth factor (HGF) stimulated rapid chemotaxis on FN= but not CN=, increasing migration speed at 10 h early in the channel at low HGF in a steep HGF gradient. HGF accelerated migration on FN= and bulk-cell movement on both uniform substrates. An HGF gradient also slowed cells in wave 2 moving on FN-CN, not CN-FN. Both opposing-gradient substrates affected the shape, speed, and net distance of migrating cells. Gradient and uniform configurations of HGF and substrate differentially influenced migration behavior. Therefore, haptotaxis substrate configuration potently modifies myoblast chemotaxis by HGF. Innovative microfluidic experiments advance our understanding of intricate complexities of myoblast migration. Findings can be leveraged to engineer muscle-tissue volumes for transplantation after serious injury. New analytical approaches may generate broader insights into cell migration.

Chemotaxis plays a fundamental role in immune defense and cancer metastasis. Microfluidic devices are increasingly applied to studying chemotaxis, owing to their advantages of reduced reagent consumption, ability to control chemical gradients, tracking of individual cells, and quantification of chemotaxis. Many existing microfluidic chemotaxis devices suffer from limited throughput and complex operation. Here, we describe a microfluidic device with a radial channel design which allows for simultaneous chemotaxis tests of different cell types and different gradient conditions. This radial microfluidic device was capable of stand-alone stable gradient generation using passive pumping and pressure-balancing strategies. The device was validated by testing the migration of fast-migrating human neutrophils and two slower-migrating human breast cancer cell lines, MDA-MB-231 and MCF-7 cells. Furthermore, this radial microfluidic device was useful in studying the influence of the nuclear chromatin binding protein high mobility group A2 (HMGA2) on the migration of the human triple negative breast cancer cell line MDA-MB-231.

Chemotaxis is a classic mechanism for guiding cell migration and an important topic in both fundamental cell biology and health sciences. Neutrophils are a widely used model to study eukaryotic cell migration and neutrophil chemotaxis itself can lead to protective or harmful immune actions to the body. While much has been learnt from past research about how neutrophils effectively navigate through a chemoattractant gradient, many interesting questions remain unclear. For example, while it is tempting to model neutrophil chemotaxis using the well-established biased random walk theory, the experimental proof was challenged by the cell's highly persistent migrating nature. A special experimental design is required to test the key predictions from the random walk model. Another question that has interested the cell migration community for decades concerns the existence of chemotactic memory and its underlying mechanism. Although chemotactic memory has been suggested in various studies, a clear quantitative experimental demonstration will improve our understanding of the migratory memory effect. Motivated by these questions, we developed a microfluidic cell migration assay (so-called dual-docking chip or D 2 -Chip) that can test both the biased random walk model and the memory effect for neutrophil chemotaxis on a single chip enabled by multi-region gradient generation and dual-region cell alignment. Our results provide experimental support for the biased random walk model and chemotactic memory for neutrophil chemotaxis. Quantitative data analyses provide new insights into neutrophil chemotaxis and memory by making connections to entropic disorder, cell morphology and oscillating migratory response.

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.

Microfluidic systems can better control cellular microenvironments and therefore are increasingly used for cell migration research. However, most existing systems are impractical to use without specialized facilities and researchers. Toward removing this barrier, we developed a compact USB microscope-based Microfluidic Chemotaxis Analysis System (UMCAS). This system integrates microfluidic devices, live cell imaging, environmental control and data analysis to provide a solution for rapid microfluidic cell migration and chemotaxis experiments with real-time result reporting. This developed system was successfully validated by testing neutrophil chemotaxis.

Directed cell migration plays important roles in physiological processes such as host defense, wound healing, cancer metastasis and embryogenesis. Many organisms are capable of directional migration, which can be guided by diverse cellular factors including chemical and electrical cues. Recently, microfluidic devices that consist of small channels with micrometer dimensions are being developed for cell migration studies. These devices can precisely configure and flexibly manipulate chemical concentration gradients and electric fields, and thus can be used to study the complex guiding mechanisms for cell migration. In this paper we highlight recent applications of microfluidic devices for cell migration research, with a focus on electric field-directed cell migration, to provide important and timely updates of this rapidly developing research field., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

Directed cell migration mediates physiological and pathological processes. In particular, immune cell trafficking in tissues is crucial for inducing immune responses and is coordinated by multiple environmental cues such as chemoattractant gradients. Although the chemotaxis mechanism has been extensively studied, how cells integrate multiple chemotactic signals for effective trafficking and positioning in tissues is not clearly defined. Results from previous neutrophil chemotaxis experiments and modeling studies suggested that ligand-induced homologous receptor desensitization may provide an important mechanism for cell migration in competing chemoattractant gradients. However, the previous mathematical model is oversimplified to cell gradient sensing in one-dimensional (1-D) environment. To better understand the receptor desensitization mechanism for chemotactic navigation, we further developed the model to test the role of homologous receptor desensitization in regulating both cell gradient sensing and migration in different configurations of chemoattractant fields in two-dimension (2-D). Our results show that cells expressing normal desensitizable receptors preferentially orient and migrate toward the distant gradient in the presence of a second local competing gradient, which are consistent with the experimentally observed preferential migration of cells toward the distant attractant source and confirm the requirement of receptor desensitization for such migratory behaviors. Furthermore, our results are in qualitative agreement with the experimentally observed cell migration patterns in different configurations of competing chemoattractant fields.

Growth factor-induced chemotaxis of cancer cells is believed to play a critical role in metastasis, directing the spread of cancer from the primary tumor to secondary sites in the body. Understanding the mechanistic and quantitative behavior of cancer cell migration in growth factor gradients would greatly help in future treatment of metastatic cancers. Using a novel microfluidic chemotaxis chamber capable of simultaneously generating multiple growth factor gradients, we examined the migration of the human metastatic breast cancer cell line MDA-MB-231 in various conditions. First, we quantified and compared the migration in two gradients of epidermal growth factor (EGF) spanning different concentrations: 0-50 ng/ml and 0.1-6 ng/ml. Cells showed a stronger response in the 0-50 ng/ml gradient. However, the fact that even a shallow gradient of EGF can induce chemotaxis, and that EGF can direct migration over a large dynamic range of gradients, confirms the potency of EGF as a chemoattractant. Second, we investigated the effect of antibody against the EGF receptor (EGFR) on MDA-MB-231 chemotaxis. Quantitative analysis indicated that anti-EGFR antibody impaired both motility and directional orientation (CI = 0.03, speed = 0.71 microm/min), indicating that cell motility was induced by the activation of EGFR. The ability to compare, in terms of quantitative parameters, the effects of different pharmaceutical inhibitors, as well as subtle differences in experimental conditions, will aid in our understanding of mechanisms that drive metastasis. The microfluidic chamber described in this work will provide a platform for cell-based assays that can be used to compare the effectiveness of different pharmaceutical compounds targeting cell migration and metastasis.

Chemotaxis, directed cell migration in a gradient of chemoattractant, is an important biological phenomenon that plays pivotal roles in cancer metastasis. Newly developed microfluidic chemotaxis chambers (MCC) were used to study chemotaxis of metastatic breast cancer cells, MDA-MB-231, in EGF gradients of well-defined profiles. Migration behaviors of MDA-MB-231 cells in uniform concentrations of EGF (0, 25, 50, and 100 ng/ml) and EGF (0-25, 0-50, and 0-100 ng/ml) with linear and nonlinear polynomial profiles were investigated. MDA-MB-231 cells exhibited increased speed and directionality upon stimulation with uniform concentrations of EGF. The cells were viable and motile for over 24 h, confirming the compatibility of MCC with cancer cells. Linear concentration gradients of different ranges were not effective in inducing chemotactic movement as compared to nonlinear gradients. MDA-MB-231 cells migrating in EGF gradient of 0-50 ng/ml nonlinear polynomial profile exhibited marked directional movement toward higher EGF concentration. This result suggests that MDA-MB-231 cancer cell chemotaxis depends on the shape of gradient profile as well as on the range of EGF concentrations.

Chronic obstructive pulmonary disease (COPD) is a persistent and progressive respiratory disorder characterized by expiratory airflow limitation caused by chronic inflammation. Evidence has shown that COPD is correlated with neutrophil chemotaxis towards the airways, resulting in neutrophilic airway inflammation. This study aimed to evaluate neutrophil chemotaxis in bronchoalveolar lavage fluid (BALF) from COPD patients using a high-throughput nine-unit microfluidic platform and explore the possible correlations between neutrophil migratory dynamics and COPD development. The results showed that BALF from COPD patients induced stronger neutrophil chemotaxis than the Control BALF. Our results also showed that the chemotactic migration of neutrophils isolated from the blood of COPD patients was not significantly different from neutrophils from healthy controls, and neutrophil migration in three known chemoattractants (fMLP, IL-8, and LTB4) was not affected by glucocorticoid treatment. Moreover, comparison with clinical data showed a trend of a negative relationship between neutrophil migration chemotactic index (C. I.) in COPD BALF and patient's spirometry data, suggesting a potential correlation between neutrophil migration and the severity of COPD. The present study demonstrated the feasibility of using the microfluidic platform to assess neutrophil chemotaxis in COPD pathogenesis, and it may serve as a potential marker for COPD evaluation in the future. [ABSTRACT FROM AUTHOR]

Immune surveillance is dependent on lymphocyte migration and targeted recruitment. This can involve different modes of cell motility ranging from random walk to highly directional environment-guided migration driven by chemotaxis. This study protocol describes a flow-based microfluidic device to perform quantitative multiplex cell migration assays with the potential to investigate in real time the migratory response of T cells at the population or single-cell level. The device also allows for subsequent in situ fixation and direct fluorescence analysis of the cells in the microchannel. [ABSTRACT FROM AUTHOR]

Chronic obstructive pulmonary disease (COPD) is a common lung disease characterized by narrowed airways, resulting in serious breathing difficulty. Previous studies have demonstrated that inflammatory infiltration of leukocytes in the airway is associated with the pathogenesis of COPD. In the present study, we employed a microfluidic approach to assess the effect of COPD sputum on activated human peripheral blood T cell migration and chemotaxis under well‐controlled gradient conditions. Our results showed considerable basal migration of T cells derived from peripheral blood of COPD patients and healthy controls in the medium control groups. By contrast, the migration of T cells from COPD patients and healthy controls was significantly inhibited in the presence of a gradient of sputum supernatant from COPD patients. Furthermore, chemotaxis of T cells from COPD patients or healthy subjects toward an SDF‐1α gradient was clearly inhibited by sputum samples from the COPD patients. The inhibition effect revealed by the microfluidic cell migration experiments provides new information about the complex involvement of T cell trafficking in COPD. [ABSTRACT FROM AUTHOR]

Chronic Obstructive Pulmonary Disease (COPD) is a common lung disease characterized by breathing difficulty as a consequence of narrowed airways. Previous studies have shown that COPD is correlated with neutrophil infiltration into the airways through chemotactic migration. However, whether neutrophil chemotaxis can be used to characterize and diagnose COPD is not well established. In the present study, we developed a microfluidic platform for evaluating neutrophil chemotaxis to sputum samples from COPD patients. Our results show increased neutrophil chemotaxis to COPD sputum compared to control sputum from healthy individuals. The level of COPD sputum induced neutrophil chemotaxis was correlated with the patient’s spirometry data. The cell morphology of neutrophils in a COPD sputum gradient is similar to the morphology displayed by neutrophils exposed to an IL-8 gradient, but not a fMLP gradient. In competing gradients of COPD sputum and fMLP, neutrophils chemotaxis and cell morphology are dominated by fMLP. [ABSTRACT FROM AUTHOR]

Migration and localization of NK cells into peripheral tissues are tightly regulated under normal and pathological conditions. The physiological importance of NK cell-DC crosstalk has been well documented. However, the ways in which DCs regulate the migratory properties of NK cells (such as chemotaxis, chemokinesis, chemo-repulsion) are not fully defined in vitro. Here, we employed a microfluidic platform to examine, at the single-cell level, C57BL/6 NK-cell migrations in a stable chemical gradient. We observed that soluble factors released by the immature and LPS-activated mature DCs induced a high level of chemotactic movement of IL-2-activated NK cells in vitro. We confirmed these findings in a standard trans-well migration assay, and identified CXCR3 as a key receptor on the NK cells that mediated the migration. More interestingly, we revealed a novel function of granulocyte macrophage colony-stimulating factor in repulsing NK-cell migrations. The future uses of such microfluidic device in the systematic evaluations of NK-cell migratory responses in NK cell-DC crosstalk will provide new insights into the development of DC-based NK-cell therapies against tumor and infections. [ABSTRACT FROM AUTHOR]

Microfluidic devices can better control cellular microenvironments compared to conventional cell migration assays. Over the past few years, microfluidics-based chemotaxis studies showed a rapid growth. New strategies were developed to explore cell migration in manipulated chemical gradients. In addition to expanding the use of microfluidic devices for a broader range of cell types, microfluidic devices were used to study cell migration and chemotaxis in complex environments. Furthermore, high-throughput microfluidic chemotaxis devices and integrated microfluidic chemotaxis systems were developed for medical and commercial applications. In this article, we review recent developments in microfluidics-based chemotaxis studies and discuss the new trends in this field observed over the past few years. [ABSTRACT FROM AUTHOR]

Abstract: Directed cell migration in tissues mediates various physiological processes and is guided by complex cellular factors such as chemoattractant gradients and electric fields. Direct current (DC) electric fields can be generated in physiological settings and the electric field guided migration of various cell types (i.e., electrotaxis) has been demonstrated both in vitro and in vivo. Although several mechanisms have been proposed for electrotaxis, there are so far very few quantitative models. Furthermore, because chemoattractant gradients and electric fields co-exist in tissues, it is important to understand how chemotaxis and electrotaxis interact for mediating cell migration and trafficking. In this study, we developed a mathematical model to investigate the role of electromigration of cell surface chemoattractant receptors in mediating electrochemical sensing and migration of cells. Our results show that electromigration of chemoattractant receptors enables cell electrotactic sensing and migration in the presence of a uniform chemoattractant field. Furthermore, in the physiologically-relevant range of receptor electromigration rates, application of electric fields overcomes chemical guiding signals for directional sensing and migration of cells in co-existing competing electric fields and chemoattractant gradients. [Copyright &y& Elsevier]

Directed cell migration mediates physiological and pathological processes. In particular, immune cell trafficking in tissues is crucial for inducing immune responses and is coordinated by multiple environmental cues such as chemoattractant gradients. Although the chemotaxis mechanism has been extensively studied, how cells integrate multiple chemotactic signals for effective trafficking and positioning in tissues is not clearly defined. Results from previous neutrophil chemotaxis experiments and modeling studies suggested that ligand-induced homologous receptor desensitization may provide an important mechanism for cell migration in competing chemoattractant gradients. However, the previous mathematical model is oversimplified to cell gradient sensing in one-dimensional (1-D) environment. To better understand the receptor desensitization mechanism for chemotactic navigation, we further developed the model to test the role of homologous receptor desensitization in regulating both cell gradient sensing and migration in different configurations of chemoattractant fields in two-dimension (2-D). Our results show that cells expressing normal desensitizable receptors preferentially orient and migrate toward the distant gradient in the presence of a second local competing gradient, which are consistent with the experimentally observed preferential migration of cells toward the distant attractant source and confirm the requirement of receptor desensitization for such migratory behaviors. Furthermore, our results are in qualitative agreement with the experimentally observed cell migration patterns in different configurations of competing chemoattractant fields. [ABSTRACT FROM AUTHOR]

Neutrophils need to correctly interpret gradients of chemotactic factors (CFs) such as interleukin 8 (IL-8) to migrate to the site of infection and perform immune functions. Because diffusion-based chemotaxis assays used in previous studies suffer from temporally changing gradients, it is difficult to distinguish the influence of CF gradient steepness from mean CF concentration on chemotaxis. To better understand the roles of mean CF concentration and CF gradient steepness, we developed a microfluidic device that can maintain stable IL-8 gradients. We report that the random motility of neutrophils is a biphasic function of IL-8 concentration and its magnitude plays a decisive role in effective chemotaxis, a quantitative measure of migration. We show that the concentrations for the optimum chemotaxis in linear IL-8 gradients and for the maximum random motility in uniform IL-8 coincide. In contrast, we find that the steepness of IL-8 gradients has no significant effect on effective chemotaxis. [Copyright &y& Elsevier]

Cell migration is involved in physiological processes such as wound healing, host defense, and cancer metastasis. The movement of various cell types can be directed by chemical gradients (i.e., chemotaxis). In addition to chemotaxis, many cell types can respond to direct current electric fields (dcEF) by migrating to either the cathode or the anode of the field (i.e., electrotaxis). In tissues, physiological chemical gradients and dcEF can potentially co-exist and the two guiding mechanisms may direct cell migration in a coordinated manner. Recently, microfluidic devices that can precisely configure chemical gradients or dcEF have been increasingly developed and used for chemotaxis and electrotaxis studies. However, a microfluidic device that can configure controlled co-existing chemical gradients and dcEF that would allow quantitative cell migration analysis in complex electrochemical guiding environments is not available. In this study, we developed a polydimethylsiloxane-based microfluidic device that can generate better controlled single or co-existing chemical gradients and dcEF. Using this device, we showed chemotactic migration of T cells toward a chemokine CCL19 gradient or electrotactic migration toward the cathode of an applied dcEF. Furthermore, T cells migrated more strongly toward the cathode of a dcEF in the presence of a competing CCL19 gradient, suggesting the higher electrotactic attraction. Taken together, the developed microfluidic device offers a new experimental tool for studying chemical and electrical guidance for cell migration, and our current results with T cells provide interesting new insights of immune cell migration in complex guiding environments. [ABSTRACT FROM AUTHOR]

M obile s ensing based on the integration of m icrofluidic device and s martphone, so-called MS 2 technology, has enabled many applications over recent years, and continues to stimulate growing interest in both research communities and industries. In particular, it has been envisioned that MS 2 technology can be developed for various cell functional assays to enable basic research and clinical applications. Toward this direction, in this paper, we describe the development of a MS 2 -based cell functional assay for testing cell migration (the M kit ). The system is constructed as an integrated test kit, which includes microfluidic chips, a smartphone-based imaging platform, the phone apps for image capturing and data analysis, and a set of reagent and accessories for performing the cell migration assay. We demonstrated that the M kit can effectively measure purified neutrophil and cancer cell chemotaxis. Furthermore, neutrophil chemotaxis can be tested from a drop of whole blood using the M kit with red blood cell (RBC) lysis. The effects of chemoattractant dose and gradient profile on neutrophil chemotaxis were also tested using the M kit . In addition to research applications, we demonstrated the effective use of the M kit for on-site test at the hospital and for testing clinical samples from chronic obstructive pulmonary disease patient. Thus, this developed M kit provides an easy and integrated experimental platform for cell migration related research and potential medical diagnostic applications. [ABSTRACT FROM AUTHOR]

T lymphocyte migration is crucial for adaptive immunity. Manipulation of signaling molecules controlling cell migration combined with in-vitro cell migration analysis provides a powerful research approach. Microfluidic devices, which can precisely configure chemoattractant gradients and allow quantitative single cell analysis, have been increasingly applied to cell migration and chemotaxis studies. However, there are a very limited number of published studies involving microfluidic migration analysis of genetically manipulated immune cells. In this study, we describe a simple microfluidic method for quantitative analysis of T cells expressing transfected chemokine receptors and other cell migration signaling probes. Using this method, we demonstrated chemotaxis of Jurkat transfectants expressing wild-type or C-terminus mutated CCR7 within a gradient of chemokine CCL19, and characterized the difference in transfectant migration mediated by wild-type and mutant CCR7. The EGFP-tagged CCR7 allows identification of CCR7-expressing transfectants in cell migration analysis and microscopy assessment of CCR7 dynamics. Collectively, our study demonstrated the effective use of the microfluidic method for studying CCR7 mediated T cell transfectant migration. We envision this developed method will provide a useful platform to functionally test various signaling mechanisms at the cell migration level. [ABSTRACT FROM AUTHOR]