Migratory Mechanisms Research Articles

Tunable motile sperm separation based on sperm persistence in migrating through shear barriers.

Rheotaxis is one of the major migratory mechanisms used in autonomous swimmers such as sperms and bacteria. Here, we present a microfluidic chip using joint rheotaxis and boundary-following behavior that selects sperms based on the motility and persistence. The proposed device consists of a channel decorated with diamond-shaped pillars that create spots of increased velocity field and shear rate. These spots are supposed as hydrodynamic barriers that impede the passage of less motile sperms through the channels, while highly motile sperms were able to overcome the generated barrier and swim through the structures. The proposed device was able to populate the chamber with sorted sperms that were fully viable and motile. The experimental results validated the separation of highly motile sperms with enhanced motility parameters compared with the initial sample. Our device was able to improve linear straight velocity, curvilinear velocity, and average path velocity of the sorted population surpassing 35%, compared with the raw semen. The processing time was also reduced to 20 min.

Triple threat: how diabetes results in worsened bacterial infections.

Diabetes mellitus, characterized by impaired insulin signaling, is associated with increased incidence and severity of infections. Various diabetes-related complications contribute to exacerbated bacterial infections, including hyperglycemia, innate immune cell dysfunction, and infection with antibiotic-resistant bacterial strains. One defining symptom of diabetes is hyperglycemia, resulting in elevated blood and tissue glucose concentrations. Glucose is the preferred carbon source of several bacterial pathogens, and hyperglycemia escalates bacterial growth and virulence. Hyperglycemia promotes specific mechanisms of bacterial virulence known to contribute to infection chronicity, including tissue adherence and biofilm formation. Foot infections are a significant source of morbidity in individuals with diabetes and consist of biofilm-associated polymicrobial communities. Bacteria perform complex interspecies behaviors conducive to their growth and virulence within biofilms, including metabolic cross-feeding and altered phenotypes more tolerant to antibiotic therapeutics. Moreover, the metabolic dysfunction caused by diabetes compromises immune cell function, resulting in immune suppression. Impaired insulin signaling induces aberrations in phagocytic cells, which are crucial mediators for controlling and resolving bacterial infections. These aberrancies encompass altered cytokine profiles, the migratory and chemotactic mechanisms of neutrophils, and the metabolic reprogramming required for the oxidative burst and subsequent generation of bactericidal free radicals. Furthermore, the immune suppression caused by diabetes and the polymicrobial nature of the diabetic infection microenvironment may promote the emergence of novel strains of multidrug-resistant bacterial pathogens. This review focuses on the "triple threat" linked to worsened bacterial infections in individuals with diabetes: (i) altered nutritional availability in diabetic tissues, (ii) diabetes-associated immune suppression, and (iii) antibiotic treatment failure.

The multifaceted role of aquaporins in physiological cell migration.

Cell migration is an essential process that underlies many physiological processes, including the immune response, organogenesis in the embryo, and angiogenesis, as well as pathological processes such as cancer metastasis. Cells have at their disposal a variety of migratory behaviors and mechanisms that seem to be specific to cell type and the microenvironment. Research over the past two decades has elucidated the water channel protein family of aquaporins (AQPs) as a regulator of many cell migration-related processes, from physical phenomena to biological signaling pathways. The roles that AQPs play in cell migration are both cell type- and isoform-specific; thus, a large swath of information has accumulated as researchers seek to identify the responses across these distinct variables. There does not seem to be a universal role that AQPs play in cell migration; the complex interplay between AQPs and cell volume management, signaling pathway activation, and in a few identified circumstances, gene expression regulation, has shown the intricate, and perhaps paradoxical, role of AQPs in cell migration. The objective of this review is to provide an organized and integrated collection of recent work that has elucidated the many mechanisms by which AQPs regulate cell migration.NEW & NOTEWORTHY Research has elucidated the water channel protein family of aquaporins (AQPs) as a regulator of many cell migration-related processes, from physical phenomena to biological signaling pathways. The roles that AQPs play in cell migration are both cell type- and isoform-specific; thus, a large swath of information has accumulated as researchers seek to identify the responses across these distinct variables. This review compiles insights into the recent findings linking AQPs to physiological cell migration.

Cell contractility drives mechanical memory of oral squamous cell carcinoma.

Matrix stiffening is ubiquitous in solid tumors and can direct epithelial-mesenchymal transition (EMT) and cancer cell migration. Stiffened niche can even cause poorly invasive oral squamous cell carcinoma (OSCC) cell lines to acquire a less adherent, more migratory phenotype, but mechanisms and durability of this acquired "mechanical memory" are unclear. Here, we observed that contractility and its downstream signals could underlie memory acquisition; invasive SSC25 cells overexpress myosin II (vs. noninvasive Cal27 cells) consistent with OSCC. However, prolonged exposure of Cal27 cells to a stiff niche or contractile agonists up-regulated myosin and EMT markers and enabled them to migrate as fast as SCC25 cells, which persisted even when the niche softened and indicated "memory" of their prior niche. Stiffness-mediated mesenchymal phenotype acquisition required AKT signaling and was also observed in patient samples, whereas phenotype recall on soft substrates required focal adhesion kinase (FAK) activity. Phenotype durability was further observed in transcriptomic differences between preconditioned Cal27 cells cultured without or with FAK or AKT antagonists, and such transcriptional differences corresponded to discrepant patient outcomes. These data suggest that mechanical memory, mediated by contractility via distinct kinase signaling, may be necessary for OSCC to disseminate.

The mammalian target of rapamycin contributes to synovial fibroblast pathogenicity in rheumatoid arthritis.

The mammalian target of Rapamycin (mTOR) is a metabolic master regulator of both innate and adaptive immunity; however, its exact role in stromal cell biology is unknown. In this study we explored the role of the mTOR pathway on Rheumatoid Arthritis synovial fibroblast (RASF) metabolism and activation and determined if crosstalk with the Hippo-YAP pathway mediates their effects. Primary RA synovial fibroblasts (RASF) were cultured with TNFα alone or in combination with the mTOR inhibitor Rapamycin or YAP inhibitor Verteporfin. Chemokine production, matrix metalloproteinase (MMP) production, and adhesion marker expression were quantified by real-time PCR, ELISA, and/or Flow Cytometry. Invasion assays were performed using Transwell invasion chambers, while wound repair assays were used to assess RASF migration. Cellular bioenergetics was assessed using the Seahorse XFe96 Analyzer. Key metabolic genes (GLUT-1, HK2, G6PD) were measured using real-time PCR. Reanalysis of RNA-Seq analysis was performed on RA (n = 151) and healthy control (HC) (n = 28) synovial tissue biopsies to detect differential gene and pathway expression. The expression of YAP was measured by Western Blot. Transcriptomic analysis of healthy donor and RA synovial tissue revealed dysregulated expression of several key components of the mTOR pathway in RA. Moreover, the expression of phospho-ribosomal protein S6 (pS6), the major downstream target of mTOR is specifically increased in RA synovial fibroblasts compared to healthy tissue. In the presence of TNFα, RASF display heightened phosphorylation of S6 and are responsive to mTOR inhibition via Rapamycin. Rapamycin effectively alters RASF cellular bioenergetics by inhibiting glycolysis and the expression of rate limiting glycolytic enzymes. Furthermore, we demonstrate a key role for mTOR signaling in uniquely mediating RASF migratory and invasive mechanisms, which are significantly abrogated in the presence of Rapamycin. Finally, we report a significant upregulation in several genes involved in the Hippo-YAP pathway in RA synovial tissue, which are predicted to converge with the mTOR pathway. We demonstrate crosstalk between the mTOR and YAP pathways in mediating RASF invasive mechanism whereby Rapamycin significantly abrogates YAP expression and YAP inhibition significantly inhibits RASF invasiveness. mTOR drives pathogenic mechanisms in RASF an effect which is in part mediated via crosstalk with the Hippo-YAP pathway.

Migratory regulation by MTA homologous genes is essential for the uniform distribution of planarian adult pluripotent stem cells.

The migration of adult stem cells in vivo is an important issue, but the complex tissue structures involved, and limited accessibility of the cells hinder a detailed investigation. To overcome these problems, the freshwater planarian Dugesia japonica was used because it has a simple body plan and abundant adult pluripotent stem cells (neoblasts) distributed uniformly throughout its body. To investigate the migratory mechanisms of neoblasts, two planarian homologous genes of metastatic tumor antigen (MTA-A and MTA-B), a protein involved in cancer metastasis that functions through histone deacetylation, were identified, and their function was analyzed using RNA interference (RNAi). MTA-A or MTA-B knockdown disrupted homeostatic tissue turnover and regeneration in planarians. Whereas neoblasts in MTA-A (RNAi) and MTA-B (RNAi) animals were maintained, neoblast differentiation was inhibited. Furthermore, the normal uniform neoblast distribution pattern changed to a branch-like pattern in MTA-A (RNAi) and MTA-B (RNAi) animals. To examine the neoblast migratory ability, a partial X-ray irradiation assay was performed in D. japonica. Using this assay system, the MTA-A knockdown neoblasts migrated collectively in a branch-like pattern, and the MTA-B knockdown neoblasts were not able to migrate. These results indicated that MTA-A was required for the exit of neoblasts from the branch-like region, and that MTA-B was required for neoblast migration. Thus, the migration mediated by MTA-A and MTA-B enabled uniform neoblast distribution and was required for neoblast differentiation to achieve tissue homeostasis and regeneration.

RNA localization in confined cells depends on cellular mechanical activity and contributes to confined migration

SummaryCancer cells experience mechanical confining forces during metastasis and, consequently, can alter their migratory mechanisms. Localization of numerous mRNAs to cell protrusions contributes to cell polarization and migration and is controlled by proteins that can bind RNA and/or cytoskeletal elements, such as the adenomatous polyposis coli (APC). Here, we demonstrate that peripheral localization of APC-dependent RNAs in cells within confined microchannels is cell type dependent. This varying phenotype is determined by the levels of a detyrosinated tubulin network. We show that this network is regulated by mechanoactivity and that cells with mechanosensitive ion channels and increased myosin II activity direct peripheral localization of the RAB13 APC-dependent RNA. Through specific mislocalization of the RAB13 RNA, we show that peripheral RNA localization contributes to confined cell migration. Our results indicate that a cell’s mechanical activity determines its ability to peripherally target RNAs and utilize them for movement in confinement.

Nonmuscle Myosin II in cancer cell migration and mechanotransduction.

Cell migration is a key step of cancer metastasis, immune-cell navigation, homing of stem cells and development. What adds complexity to it is the heterogeneity of the tissue environment that gives rise to a vast diversity of migratory mechanisms utilized by cells. A majority of cell motility mechanisms reported elsewhere largely converge in depicting the importance of the activity and complexity of actomyosin networks in the cell. In this review, we highlight the less discussed functional diversity of these actomyosin complexes and describe in detail how the major cellular actin-binding molecular motor proteins, nonmuscle myosin IIs are regulated and how they participate and mechanically reciprocate to changes in the microenvironment during cancer cell migration and tumor progression. Understanding the role of nonmuscle myosin IIs in the cancer cell is important for designing efficient therapeutic strategies to prevent cancer metastasis.

SATB2 overexpression promotes oral squamous cell carcinoma progression by up-regulating NOX4

While atypical expression of special AT-rich sequence-binding protein 2 (SATB2) has been approved associated with tumor progression, metastasis and unfavourable prognosis in various carcinomas. However, in oral squamous cell carcinoma (OSCC), both the expressive state and associated functions of SATB2's are still undefined. Here we show that, in clinical samples from a retrospective cohort of 58 OSCC patients, high expression of SATB2 is associated with poor prognosis of OSCC patients. In this study, we investigated SATB2 is highly expressed in OSCC tissues and cell lines, which can promote OSCC cells' proliferation, migration, invasion and tumor growth. According to sequencing results based on previous literature, we identified NOX4 is a bona fide downstream target of SATB2, when it was knockdown, OSCC's proliferation can be partially suppressed. Furthermore, NOX4 knockdown inhibits tumorigenicity, which can be rescued partially by ectopic expression of SATB2 in HNSCC cell line, and vice versa. Collectively, our findings not only indicate overexpression of SATB2 triggers the proliferative, migratory and invasive mechanisms which are important in the malignant phenotype of OSCC, but also identify NOX4 as the downstream gene for SATB2. These findings indicate that SATB2 may play a key role in OSCC tumorigenicity and may be a future target for the development of new therapeutic regimens.

RNA Localization in Confined Cells Depends on Cellular Mechanical Activity and Contributes to Confined Migration

Cancer cells experience mechanical confining forces during metastasis and consequently, can alter their migratory mechanisms. Localization of numerous mRNAs to cell protrusions contributes to cell polarization and migration and is controlled by proteins that can bind RNA and/or cytoskeletal elements, such as the Adenomatous Polyposis Coli (APC). Here, we demonstrate that peripheral localization of APC-dependent RNAs in cells within confined microchannels is cell type dependent. This varying phenotype is determined by the presence or absence of a fibrillar detyrosinated tubulin network. We show that this network is regulated by mechanoactivity, and that cells with mechanosensitive ion channels and increased myosin II activity direct peripheral localization of the RAB13 APC-dependent RNA. Through specific mislocalization of the RAB13 RNA, we show that peripheral RNA localization contributes to confined cell migration. Our results indicate that a cell’s mechanical activity determines its ability to peripherally target RNAs and utilize them for movement in confinement.

Stem Cell-Derived Exosomes and Nanovesicles: Promotion of Cell Proliferation, Migration, and Anti-Senescence for Treatment of Wound Damage and Skin Ageing.

Extracellular vesicles (EVs), such as exosomes, are nano-sized vesicles derived from endocytic membranes and contain biomolecules such as proteins, lipids, RNAs, and DNAs for the transfer of signals to recipient cells, playing significant roles in cell-to-cell communication. Discovery of exosomes has attracted attention for possible use as next generation therapies in clinical applications; however, several studies suggest that cells secrete exosomes that perform as mediators in the tumor niche and play several roles in tumorigenesis, angiogenesis, and metastasis. Recently, stem cell-derived exosomes have been suggested as a desirable source for regenerative medicine due to their roles in the promotion of angiogenesis via migratory and proliferative mechanisms. This review is aimed at demonstrating the present knowledge of stem cell-derived exosomes and cell-engineered nanovesicles (CNVs) as proliferative, migratory, and anti-senescent therapeutic biomaterial for use in tissue regeneration; wound healing and anti-ageing are explained. We conclude this review by discussing the future perspectives of stem cell-derived exosomes and CNVs as a platform in therapeutic strategies for treatment of wound damage and skin aging.

Chronic Pancreatitis and the Development of Pancreatic Cancer.

Pancreatitis is a fibro-inflammatory disorder of the pancreas that can occur acutely or chronically as a result of the activation of digestive enzymes that damage pancreatic cells, which promotes inflammation. Chronic pancreatitis with persistent fibro-inflammation of the pancreas progresses to pancreatic cancer, which is the fourth leading cause of cancer deaths across the globe. Pancreatic cancer involves cross-talk of inflammatory, proliferative, migratory, and fibrotic mechanisms. In this review, we discuss the role of cytokines in the inflammatory cell storm in pancreatitis and pancreatic cancer and their role in the activation of SDF1α/CXCR4, SOCS3, inflammasome, and NF-κB signaling. The aberrant immune reactions contribute to pathological damage of acinar and ductal cells, and the activation of pancreatic stellate cells to a myofibroblast-like phenotype. We summarize several aspects involved in the promotion of pancreatic cancer by inflammation and include a number of regulatory molecules that inhibit that process.

Mechanisms of 3D cell migration.

Cell migration is essential for physiological processes as diverse as development, immune defence and wound healing. It is also a hallmark of cancer malignancy. Thousands of publications have elucidated detailed molecular and biophysical mechanisms of cultured cells migrating on flat, 2D substrates of glass and plastic. However, much less is known about how cells successfully navigate the complex 3D environments of living tissues. In these more complex, native environments, cells use multiple modes of migration, including mesenchymal, amoeboid, lobopodial and collective, and these are governed by the local extracellular microenvironment, specific modalities of Rho GTPase signalling and non-muscle myosin contractility. Migration through 3D environments is challenging because it requires the cell to squeeze through complex or dense extracellular structures. Doing so requires specific cellular adaptations to mechanical features of the extracellular matrix (ECM) or its remodelling. In addition, besides navigating through diverse ECM environments and overcoming extracellular barriers, cells often interact with neighbouring cells and tissues through physical and signalling interactions. Accordingly, cells need to call on an impressively wide diversity of mechanisms to meet these challenges. This Review examines how cells use both classical and novel mechanisms of locomotion as they traverse challenging 3D matrices and cellular environments. It focuses on principles rather than details of migratory mechanisms and draws comparisons between 1D, 2D and 3D migration.

Overlapping migratory mechanisms between neural progenitor cells and brain tumor stem cells.

Neural stem cells present in the subventricular zone (SVZ), the largest neurogenic niche of the mammalian brain, are able to self-renew as well as generate neural progenitor cells (NPCs). NPCs are highly migratory and traverse the rostral migratory stream (RMS) to the olfactory bulb, where they terminally differentiate into mature interneurons. NPCs from the SVZ are some of the few cells in the CNS that migrate long distances during adulthood. The migratory process of NPCs is highly regulated by intracellular pathway activation and signaling from the surrounding microenvironment. It involves modulation of cell volume, cytoskeletal rearrangement, and isolation from compact extracellular matrix. In malignant brain tumors including high-grade gliomas, there are cells called brain tumor stem cells (BTSCs) with similar stem cell characteristics to NPCs but with uncontrolled cell proliferation and contribute to tumor initiation capacity, tumor progression, invasion, and tumor maintenance. These BTSCs are resistant to chemotherapy and radiotherapy, and their presence is believed to lead to tumor recurrence at distal sites from the original tumor location, principally due to their high migratory capacity. BTSCs are able to invade the brain parenchyma by utilizing many of the migratory mechanisms used by NPCs. However, they have an increased ability to infiltrate the tight brain parenchyma and utilize brain structures such as myelin tracts and blood vessels as migratory paths. In this article, we summarize recent findings on the mechanisms of cellular migration that overlap between NPCs and BTSCs. A better understanding of the intersection between NPCs and BTSCs will to provide a better comprehension of the BTSCs' invasive capacity and the molecular mechanisms that govern their migration and eventually lead to the development of new therapies to improve the prognosis of patients with malignant gliomas.

A combination of coarse-grain molecular dynamics to investigate the effects of sodium dodecyl sulfate on grafted reaction of starch-based adhesive

Graft copolymerization is a challenging step in preparation of starch-based adhesive due to the complexity and instantaneity. A combination of both experimental and simulation methodology has been employed to investigate the process at microscopic level. Through a series of characterizations of adhesives and copolymers with different SDS (sodium dodecyl sulfate) contents, 2% (w/w, 2g SDS/100 g starch) SDS demonstrated outstanding balance between the starch grafted percentage and interfacial properties. The coarse-grain molecular dynamics (CGMD) was utilized to reveal the molecular distribution and migratory mechanisms during the reaction by calculating radius distribution function (RDF) and mean square displacement (MSD). Starch chains covering the monomers surface was found to exhibit longer radius of gyration (Rg). Furthermore, the interfacial models were constructed in this study, and interfacial tension between water and VAc beads was calculated to confirm the improvement in interfacial properties and the rationality of simulation with the addition of SDS.

Abstract 2017: Phenotypic plasticity and class switching in ovarian cancer

Abstract The multifaceted evolution of cancer involves differential responses of tumor cell populations to intrinsic and extrinsic stimuli. Numerous reports have examined the role of epithelial-mesenchymal transition (EMT) in the progression of high-grade serous ovarian carcinoma (HGSC). An earlier study from our group successfully resolved molecular subclasses in HGSC wherein metastasis was mediated by cooperative cell migration in the epithelial subclass and EMT in the mesenchymal subclass. We now explore the transcriptional circuitry in expression datasets and cell lines regulating these states. This led to the identification that Tcf21 and Slug expression correlated with the epithelial and mesenchymal phenotypes, respectively, and inversely correlated with each other. Probing the differential altered localization and expression of TCF21 and Slug further resolved a spectrum of intermediary phenotypic states along the epithelial to mesenchymal spectrum, each of which is associated with the expression of specific markers. Further, functional assays reveal the inherent plasticity of each phenotype with migratory mechanisms emerging as a differential parameter. Rigidity and high degree of differentiation of the epithelial and mesenchymal state is evident from a lack of responsiveness to specific microenvironmental conditions. Importantly, such cellular plasticity induced shifts from a steady-state phenotype following modulation of the microenvironment only in the intermediate phenotypes to exhibit flexibility to transit towards either the two ends of the spectrum. Specifically, a tendency to acquire an epithelial phenotype on growth factor deprivation was noted. The effects of BMP7 and TGFβ to induce epithelial or mesenchymal phenotypes, respectively, were also noted exclusively within the intermediary states. The stress generated by chemotherapeutic agents like paclitaxel led to enrichment of and dominant effects of the epithelial state. We further queried if this plasticity is also a feature of HGSC tumors. Marker expression (Tcf21, E-cadherin, Parp1, Slug, Anxa2 and Hyaluronan) was scored based on their frequency, intensity and location in tumors to derive a definitive scoring system comprising biomarker and tumor class indices. Application of such scoring could effectively stratifiy 95 human HGSC cases comprising primary and metastatic tumors into epithelial and mesenchymal classes, and further resolve a third class likely to represent the intermediate states. A salient feature noted was that metastases and chemotherapy could lead to class switching. Our study thus provides a comprehensive view of the intrinsic and extrinsic molecular events governing cellular plasticity in HGSC. Citation Format: Sharmila A. Bapat, Sagar Varankar, Swapnil Kamble. Phenotypic plasticity and class switching in ovarian cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2017.

Gas6/AXL Modulates Increased Inflammatory Cytokines During Preeclampsia in Rats

Preeclampsia (PE) is a complicated obstetric complication characterized by increased blood pressure and the production of inflammatory cytokines. The growth arrest‐specific 6 (Gas6) protein is known to induce dynamic cellular responses and is elevated in PE. Gas6 binds to the AXL tyrosine kinase receptor and AXL‐mediated signaling is implicated in proliferation and migratory mechanisms in several tissues. Our laboratory utilized Gas6 to induce preeclamptic conditions in pregnant rats. Our objective was to determine the role of Gas6 and AXL signaling in the release of cytokines during the development of PE in the rat. Briefly, pregnant rats were divided into 3 groups that received daily ip injections of PBS, Gas6, or Gas6 + R428 (an AXL inhibitor). Treatment was done from embryonic day 7.5 (E7.5) to E18.5, at which time they were euthanized and serum was harvested for Multiplex cytokine determination. Fetal and placental weights were recorded. Blood pressure and proteinuria were determined. We observed: increased proteinuria and elevated blood pressure in Gas6 treated animals, but each were inhibited by R428; a 12‐fold increase of serum interferon gamma was reduced by AXL inhibition; a 12‐fold increase in serum IL1‐alpha was inhibited by AXL inhibition; a 45‐fold increase in serum IL‐2 was reduced by AXL inhibition; a 10‐fold increase in serum IL1‐beta was reduced by AXL inhibition; a 29‐fold increase in serum IL4 was decreased by AXL inhibition; a 13‐fold increase in serum IL5 was reduced by AXL inhibition; an 24‐fold increase in serum IL6 was reduced by AXL inhibition; and a 20‐fold increase in IL12p70 was reduced by AXL inhibition. These data suggest roles for Gas6/AXL in cytokine elaboration coincident with PE and offer possible treatment avenues that may alleviate PE symptoms.Support or Funding InformationThis work was supported by a grant from the Flight Attendant's Medical Research Institute (FAMRI, PRR and JAA) and a BYU Mentoring Environment Grant (JAA and PRR).This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Semi‐Synthetic Glycosaminoglycan Ethers Decrease Receptors for Advanced Glycation End‐Products and Increase AXL Receptor in the Lung from Secondhand Smoke Treated Mice

Tobacco exposure is one of the top three global health risks leading to the development of chronic obstructive pulmonary disease (COPD). Although there is extensive research into the effects of cigarette smoke, the effect of secondhand smoke (SHS) in the lung remains limited. SHS induces Receptors for advanced glycation end‐products (RAGE) and an inflammatory response that leads to the development of COPD characteristics. Semi‐synthetic glycosaminoglycan ethers (SAGEs) are sulfated polysaccharides derived from hyaluronic acid (HA) that inhibit RAGE signaling. The growth arrest‐specific 6 (Gas6) protein is known to induce dynamic cellular responses and is correlated with cell function. Gas6 binds to the AXL tyrosine kinase receptor and AXL‐mediated signaling is implicated in proliferation and migratory mechanisms in several tissues. This project's purpose was to study the correlation between RAGE, AXL, and Gas6 during SHS exposure in the lung. C57/Bl6 mice were exposed to SHS alone or SHS + SAGEs for 4 weeks and compared to control animals exposed to room air (RA). At the time of necropsy lungs were extracted, frozen for mRNA studies or embedded for immunohistological analysis. Compared to controls we observed: 1) increased RAGE mRNA expression in SHS‐exposed lungs which was decreased by SAGEs; 2) no difference in Gas6 or AXL expression in SHS lungs and expression was significantly increased with SAGEs; 3) increased RAGE protein expression in SHS lungs that was decreased by SAGEs; and 4) decreased AXL protein in SHS animals and increased AXL protein in animals treated with SAGEs. Our results suggest that there is a direct correlation between RAGE and AXL during SHS exposure. These studies provide insight into tobacco‐mediated effects in the lung and clarify possible avenues for alleviating complications that could arise during SHS exposure such as those observed during COPD.Support or Funding InformationThis work was supported by a grant from the Flight Attendant's Medical Research Institute (FAMRI, PRR and JAA) and a BYU Mentoring Environment Grant (JAA and PRR).This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Cdc42 Activation by Endothelin Regulates Neural Crest Cell Migration in the Cardiac Outflow Tract

Congenital cardiovascular malformations are the most common birth defects affecting children. Several of these defects occur in structures developing from neural crest cells (NCC) that migrate ventrally to populate the pharyngeal arches. Cardiac NCCs (CNCC) participate in the asymmetric remodeling of the pharyngeal arch arteries (PAA) into the great vessels and the septation of the cardiac outflow tract (COFT) into the pulmonary and aortic outflows. One of the key signaling pathways regulating CNCC development involves the Endothelin‐A receptor (Ednra). The absence of Ednra signaling in the mouse causes severe cardiovascular defects, including patent ductus arteriosus, coarctation of the aorta, persistent truncus arteriosus and ventricular septal defects. However, the exact function of Ednra signaling in CNCCs is unknown.CNCC fate mapping in the Ednra mouse revealed that the migration of these cells is aberrant in the COFT of the mutant embryos, but not in the pharyngeal arches. This premature arrest of CNCC migration appeared to be independent of CNCC proliferation and apoptosis changes. These results suggest that Ednra has a patterning function in the pharyngeal arches and controls CNCC migration in the COFT. Based on these results, we tested whether CNCC migration was arrested prematurely because CNCCs fail to activate specific cell migratory mechanisms by looking at migratory markers of the Rho family of GTPases. In the mutant embryos, Cdc42 fails to localize at the migratory front of the CNCCs migrating in the COFT. Cultured E9.25 whole embryos treated with the Cdc42 inhibitor ML141 recapitulated the phenotype. To confirm that Ednra and Cdc42 were acting in the same pathway, primary NCC from the cardiac area and mandibular pharyngeal arch were isolated for culture and treated with the Ednra ligand Edn1, ML141 and Ednra antagonist TBC3214. Western blot analysis revealed that Edn1 stimulated the phosphorylation of the Cdc42 downstream target ACK only in CNCCs and the addition of either inhibitor prevented ACK phosphorylation. Erk1/2 phosphorylation was observed in the mandibular NCCs upon Edn1 treatment but this was not prevented by the addition of the Cdc42 inhibitor. Wound healing scratch assay showed that the addition of Edn1 stimulated CNCC migration, which was repressed by the addition of either the Ednra or Cdc42 inhibitor. However, no such stimulation or inhibition was observed with the mandibular NCCs.In conclusion, these results demonstrate that endothelin signaling controls the migration of NCCs in the COFT via the specific activation of Cdc42 and this pathway is not necessary for the migration of NCC in the pharyngeal arches and the rearrangement of the pharyngeal arch arteries.Support or Funding InformationAmerican Heart Association 11BGIA 5670010NIH T32 DE018380Baylor Oral Health FoundationThis abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Redirecting adult mesenchymal stromal cells to the brain: a new approach for treating CNS autoimmunity and neuroinflammation?

Mesenchymal stromal cells or stem cells (MSCs) have been shown to participate in tissue repair and are immunomodulatory in neuropathological settings. Given this, their potential use in developing a new generation of personalized therapies for autoimmune and inflammatory diseases of the central nervous system (CNS) will be explored. To effectively exert these effector functions, MSCs must first gain entry into damaged neural tissues, a process that has been demonstrated to be a limiting factor in their therapeutic efficacy. In this review, we discuss approaches to maximize the therapeutic efficacy of MSCs by altering their intrinsic trafficking programs to effectively enter neuropathological sites. To this end, we explore the significant role of chemokine receptors and adhesion molecules in directing cellular traffic to the inflamed CNS and the capacity of MSCs to adopt these molecular mechanisms to gain entry to this site. We postulate that understanding and exploiting these migratory mechanisms may be key to the development of cell-based therapies tailored to respond to the migratory cues unique to the nature and stage of progression of individual CNS disorders.