Chips-on-a-plate device for monitoring cellular migration in a microchannel-based intestinal follicle-associated epithelium model

Abstract Background The first visible signs of Crohn’s disease (CD) are microscopic erosions over the follicle-associated epithelium (FAE). The aim of the study was to investigate the effects of human α-defensin 5 (HD5) on adherent-invasive Escherichia coli LF82 translocation and HD5 secretion after LF82 exposure in an in vitro model of human FAE and in human FAE ex vivo. Methods An in vitro FAE-model was set up by the coculture of Raji B cells and Caco-2-cl1 cells. Ileal FAE from patients with CD and controls were mounted in Ussing chambers. The effect of HD5 on LF82 translocation was studied by LF82 exposure to the cells or tissues with or without incubation with HD5. The HD5 secretion was measured in human FAE exposed to LF82 or Salmonella typhimurium. The HD5 levels were evaluated by immunofluorescence, immunoblotting, and ELISA. Results There was an increased LF82 translocation across the FAE-model compared with Caco-2-cl1 (P < 0.05). Incubation of cell/tissues with HD5 before LF82 exposure reduced bacterial passage in both models. Human FAE showed increased LF82 translocation in CD compared with controls and attenuated passage after incubation with sublethal HD5 in both CD and controls (P < 0.05). LF82 exposure resulted in a lower HD5 secretion in CD FAE compared with controls (P < 0.05), whereas Salmonella exposure caused equal secretion on CD and controls. There were significantly lower HD5 levels in CD tissues compared with controls. Conclusions Sublethal HD5 reduces the ability of LF82 to translocate through FAE. The HD5 is secreted less in CD in response to LF82, despite a normal response to Salmonella. This further implicates the integrated role of antimicrobial factors and barrier function in CD pathogenesis.

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Identification of PIM1 substrates reveals a role for NDRG1 phosphorylation in prostate cancer cellular migration and invasion

Communications Biology ◽

10.1038/s42003-020-01528-6 ◽

2021 ◽

Vol 4 (1) ◽

Author(s):

Russell J. Ledet ◽

Sophie E. Ruff ◽

Yu Wang ◽

Shruti Nayak ◽

Jeffrey A. Schneider ◽

...

Keyword(s):

Prostate Cancer ◽

Cellular Migration ◽

Migration And Invasion ◽

Prostate Cancer Cells ◽

Invasion And Metastasis ◽

Serine Threonine Kinase ◽

Prostate Tumorigenesis ◽

Threonine Kinase ◽

Chemical Genetic ◽

Protein Levels

AbstractPIM1 is a serine/threonine kinase that promotes and maintains prostate tumorigenesis. While PIM1 protein levels are elevated in prostate cancer relative to local disease, the mechanisms by which PIM1 contributes to oncogenesis have not been fully elucidated. Here, we performed a direct, unbiased chemical genetic screen to identify PIM1 substrates in prostate cancer cells. The PIM1 substrates we identified were involved in a variety of oncogenic processes, and included N-Myc Downstream-Regulated Gene 1 (NDRG1), which has reported roles in suppressing cancer cell invasion and metastasis. NDRG1 is phosphorylated by PIM1 at serine 330 (pS330), and the level of NDRG1 pS330 is associated higher grade prostate tumors. We have shown that PIM1 phosphorylation of NDRG1 at S330 reduced its stability, nuclear localization, and interaction with AR, resulting in enhanced cell migration and invasion.

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Effect of Inflammation on Gingival Mesenchymal Stem/Progenitor Cells’ Proliferation and Migration through Microperforated Membranes: An In Vitro Study

Stem Cells International ◽

10.1155/2020/5373418 ◽

2020 ◽

Vol 2020 ◽

pp. 1-10 ◽

Cited By ~ 3

Author(s):

M. Al Bahrawy ◽

K. Ghaffar ◽

A. Gamal ◽

K. El-Sayed ◽

V. Iacono

Keyword(s):

Pore Size ◽

Progenitor Cells ◽

Cellular Migration ◽

Gingival Tissue ◽

Population Doubling ◽

Population Doubling Time ◽

Electron Microscopic ◽

Healthy Human ◽

Membrane Pores ◽

Scanning Electron

Background. In the field of periodontal guided tissue regeneration, microperforated membranes have recently proved to be very promising periodontal regenerative tissue engineering tools. Regenerative periodontal approaches, employing gingival mesenchymal stem/progenitor cells in combination with these novel membranes, would occur mostly in inflamed microenvironmental conditions intraorally. This in turn entails the investigation into how inflammation would affect the proliferation as well as the migration dynamics of gingival mesenchymal stem/progenitor cells. Materials and Methods. Clones of human gingival mesenchymal stem/progenitor cells (GMSCs) from inflamed gingival tissues were characterized for stem/progenitor cells’ characteristics and compared to clones of healthy human GMSCs (n=3), to be subsequently seeded on perforated collagen-coated poly-tetra-floro-ethylene (PTFE) membranes with a pore size 0.4 and 3 microns and polycarbonic acid membranes of 8 microns pore size in Transwell systems. The population doubling time and the MTT test of both populations were determined. Fetal bovine serum (FBS) was used as a chemoattractant in the culturing systems, and both groups were compared to their negative controls without FBS. Following 24 hours of incubation period, migrating cells were determined on the undersurface of microperforated membranes and the membrane-seeded cells were examined by scanning electron microscopy. Results. GMSCs demonstrated all predefined stem/progenitor cell characteristics. GMSCs from inflamed gingival tissues showed significantly shorter population doubling times. GMSCs of inflamed and healthy tissues did not show significant differences in their migration abilities towards the chemoattractant, with no cellular migration observed in the absence of FBS. GMSCs from healthy gingival tissue migrated significantly better through larger micropores (8 microns). Scanning electron microscopic images proved the migratory activity of the cells through the membrane pores. Conclusions. Inflammation appears to boost the proliferative abilities of GMSCs. In terms of migration through membrane pores, GMSCs from healthy as well as inflamed gingival tissues do not demonstrate a difference in their migration abilities through smaller pore sizes, whereas GMSCs from healthy gingival tissues appear to migrate significantly better through larger micropores.

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Studies on cellular migration in developing cerebral wall using explant culture system

Neuroscience Research Supplements ◽

10.1016/0921-8696(89)90623-3 ◽

1989 ◽

Vol 9 ◽

pp. 54

Author(s):

Noriyuki Morita ◽

Kazuhiro Ikenaka ◽

Masaharu Ogawa ◽

Katsuhiko Mikoshiba

Keyword(s):

Culture System ◽

Explant Culture ◽

Cellular Migration

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The Foot-and-Mouth Disease Carrier State Divergence in Cattle

Journal of Virology ◽

10.1128/jvi.00388-16 ◽

2016 ◽

Vol 90 (14) ◽

pp. 6344-6364 ◽

Cited By ~ 51

Author(s):

Carolina Stenfeldt ◽

Michael Eschbaumer ◽

Steven I. Rekant ◽

Juan M. Pacheco ◽

George R. Smoliga ◽

...

Keyword(s):

Foot And Mouth Disease ◽

Antiviral Response ◽

Carrier State ◽

Gamma Interferon ◽

Mouth Disease ◽

Fmdv Infection ◽

Mouth Disease Virus ◽

Follicle Associated Epithelium ◽

Foot And Mouth ◽

Host Antiviral Response

ABSTRACTThe pathogenesis of persistent foot-and-mouth disease virus (FMDV) infection was investigated in 46 cattle that were either naive or had been vaccinated using a recombinant, adenovirus-vectored vaccine 2 weeks before challenge. The prevalence of FMDV persistence was similar in both groups (62% in vaccinated cattle, 67% in nonvaccinated cattle), despite vaccinated cattle having been protected from clinical disease. Analysis of antemortem infection dynamics demonstrated that the subclinical divergence between FMDV carriers and animals that cleared the infection had occurred by 10 days postinfection (dpi) in vaccinated cattle and by 21 dpi in nonvaccinated animals. The anatomic distribution of virus in subclinically infected, vaccinated cattle was restricted to the pharynx throughout both the early and the persistent phases of infection. In nonvaccinated cattle, systemically disseminated virus was cleared from peripheral sites by 10 dpi, while virus selectively persisted within the nasopharynx of a subset of animals. The quantities of viral RNA shed in oropharyngeal fluid during FMDV persistence were similar in vaccinated and nonvaccinated cattle. FMDV structural and nonstructural proteins were localized to follicle-associated epithelium of the dorsal soft palate and dorsal nasopharynx in persistently infected cattle. Host transcriptome analysis of tissue samples processed by laser capture microdissection indicated suppression of antiviral host factors (interferon regulatory factor 7, CXCL10 [gamma interferon-inducible protein 10], gamma interferon, and lambda interferon) in association with persistent FMDV. In contrast, during the transitional phase of infection, the level of expression of IFN-λ mRNA was higher in follicle-associated epithelium of animals that had cleared the infection. This work provides novel insights into the intricate mechanisms of FMDV persistence and contributes to further understanding of this critical aspect of FMDV pathogenesis.IMPORTANCEThe existence of a prolonged, asymptomatic carrier state is a political impediment for control and potential eradication of foot-and-mouth disease (FMD). When FMD outbreaks occur, they are often extinguished by massive depopulation of livestock due to the fear that some animals may have undiagnosed subclinical infection, despite uncertainty over the biological relevance of FMD virus (FMDV) persistence. The work described here elucidates aspects of the FMDV carrier state in cattle which may facilitate identification and/or abrogation of asymptomatic FMDV infection. The divergence between animals that clear infection and those that develop persistent infection was demonstrated to occur earlier than previously established. The host antiviral response in tissues maintaining persistent FMDV was downregulated, whereas upregulation of IFN-λ mRNA was found in the epithelium of cattle that had recently cleared the infection. This suggests that the clearing of FMDV infection is associated with an enhanced mucosal antiviral response, whereas FMDV persistence is associated with suppression of the host antiviral response.

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The Role of Src Kinase in Macrophage-Mediated Inflammatory Responses

Mediators of Inflammation ◽

10.1155/2012/512926 ◽

2012 ◽

Vol 2012 ◽

pp. 1-18 ◽

Cited By ~ 115

Author(s):

Se Eun Byeon ◽

Young-Su Yi ◽

Jueun Oh ◽

Byong Chul Yoo ◽

Sungyoul Hong ◽

...

Keyword(s):

Rheumatoid Arthritis ◽

Immune Responses ◽

Inflammatory Responses ◽

Inflammatory Diseases ◽

Src Kinase ◽

Multiple Roles ◽

Cellular Migration ◽

Pharmaceutical Drugs ◽

Tyrosine Protein Kinase

Src kinase (Src) is a tyrosine protein kinase that regulates cellular metabolism, survival, and proliferation. Many studies have shown that Src plays multiple roles in macrophage-mediated innate immunity, such as phagocytosis, the production of inflammatory cytokines/mediators, and the induction of cellular migration, which strongly implies that Src plays a pivotal role in the functional activation of macrophages. Macrophages are involved in a variety of immune responses and in inflammatory diseases including rheumatoid arthritis, atherosclerosis, diabetes, obesity, cancer, and osteoporosis. Previous studies have suggested roles for Src in macrophage-mediated inflammatory responses; however, recently, new functions for Src have been reported, implying that Src functions in macrophage-mediated inflammatory responses that have not been described. In this paper, we discuss recent studies regarding a number of these newly defined functions of Src in macrophage-mediated inflammatory responses. Moreover, we discuss the feasibility of Src as a target for the development of new pharmaceutical drugs to treat macrophage-mediated inflammatory diseases. We provide insights into recent reports regarding new functions for Src that are related to macrophage-related inflammatory responses and the development of novel Src inhibitors with strong immunosuppressive and anti-inflammatory properties, which could be applied to various macrophage-mediated inflammatory diseases.

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Improved biocompatibility and efficient labeling of neural stem cells with poly(L-lysine)-coated maghemite nanoparticles

Beilstein Journal of Nanotechnology ◽

10.3762/bjnano.7.84 ◽

2016 ◽

Vol 7 ◽

pp. 926-936 ◽

Cited By ~ 15

Author(s):

Igor M Pongrac ◽

Marina Dobrivojević ◽

Lada Brkić Ahmed ◽

Michal Babič ◽

Miroslav Šlouf ◽

...

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Neural Stem Cells ◽

Cellular Uptake ◽

Cell Tracking ◽

Cellular Migration ◽

Cell Labeling ◽

Blue Staining ◽

Maghemite Nanoparticles ◽

Acetoxymethyl Ester

Background: Cell tracking is a powerful tool to understand cellular migration, dynamics, homing and function of stem cell transplants. Nanoparticles represent possible stem cell tracers, but they differ in cellular uptake and side effects. Their properties can be modified by coating with different biocompatible polymers. To test if a coating polymer, poly(L-lysine), can improve the biocompatibility of nanoparticles applied to neural stem cells, poly(L-lysine)-coated maghemite nanoparticles were prepared and characterized. We evaluated their cellular uptake, the mechanism of internalization, cytotoxicity, viability and proliferation of neural stem cells, and compared them to the commercially available dextran-coated nanomag®-D-spio nanoparticles. Results: Light microscopy of Prussian blue staining revealed a concentration-dependent intracellular uptake of iron oxide in neural stem cells. The methyl thiazolyl tetrazolium assay and the calcein acetoxymethyl ester/propidium iodide assay demonstrated that poly(L-lysine)-coated maghemite nanoparticles scored better than nanomag®-D-spio in cell labeling efficiency, viability and proliferation of neural stem cells. Cytochalasine D blocked the cellular uptake of nanoparticles indicating an actin-dependent process, such as macropinocytosis, to be the internalization mechanism for both nanoparticle types. Finally, immunocytochemistry analysis of neural stem cells after treatment with poly(L-lysine)-coated maghemite and nanomag®-D-spio nanoparticles showed that they preserve their identity as neural stem cells and their potential to differentiate into all three major neural cell types (neurons, astrocytes and oligodendrocytes). Conclusion: Improved biocompatibility and efficient cell labeling makes poly(L-lysine)-coated maghemite nanoparticles appropriate candidates for future neural stem cell in vivo tracking studies.

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