Epithelial-mesenchymal transformation during palatal fusion: carboxyfluorescein traces cells at light and electron microscopic levels

Development ◽  
1992 ◽  
Vol 116 (4) ◽  
pp. 1087-1099 ◽  
Author(s):  
C.M. Griffith ◽  
E.D. Hay
Keyword(s):  
Electron Microscopy ◽  
Epithelial Cells ◽  
Basal Layer ◽  
Subsequent Development ◽  
Electron Microscopic ◽  
Medial Edge ◽  
Mesenchymal Transformation ◽  
First Time

During the fusion of rodent embryo palatal shelves, the cells of the outer epithelial layer slough off, allowing the cells of the medial edge basal layer to form a midline seam that undergoes epithelial-mesenchymal transformation, as judged by electron microscopy and immunohistochemistry. In this study, we analyze the fate of the transformed cells using a lipid soluble dye to label the medial edge epithelium in situ. Prefusion E14 mouse palates were exposed in vitro or in vivo to a fluoresceinated lipid soluble marker, carboxydichlorofluorescein diacetate succinimidyl ester (CCFSE), which localizes in epithelia as a lipid insoluble compound that does not pass into the connective tissue compartment. The midline seam that formed after 24 hours contained labelled epithelial cells that were replaced by individually labelled mesenchymal cells where the seam transformed. By light microscopy, the labelled cells were seen to contain intensely fluorescent bodies that do not react for acid phosphatase. We were able for the first time to identify these structures by electron microscopy as CCFSE isolation bodies. The cells with isolation bodies are clearly healthy and able to participate in subsequent development of the palate. At 4 days after labelling, individual CCFSE containing cells present in the palate mesenchyme occupy both midline and lateral areas and can clearly be classified as fibroblasts by electron microscopy. CCFSE is a far more useful marker than another lipid soluble marker, DiI, for following cells, because the cells can be fixed and identified both at the light and electron microscope levels. Interestingly, if labelled palatal shelves are not allowed to fuse in vitro, the basal epithelial cells do not form mesenchyme after sloughing, indicating that formation of the epithelial midline seam is necessary to trigger its epithelial-mesenchymal transformation.

scholarly journals Quercetin inhibits intrahepatic cholangiocarcinoma by inducing ferroptosis and inhibiting epithelial-mesenchymal transformation via the NF-κB pathway

2021 ◽  
Author(s):  
Yinghui Song ◽  
Zhihua Zhang ◽  
Qin Chai ◽  
GuoYi Xia ◽  
Zhangtao Yu ◽  
...  
Keyword(s):  
Intrahepatic Cholangiocarcinoma ◽  
Clonogenic Assay ◽  
Direct Interaction ◽  
Rna Seq ◽  
Wound Healing Assay ◽  
Tumor Bearing ◽  
Mesenchymal Transformation ◽  
First Time

Abstract Intrahepatic cholangiocarcinoma (ICC) is a rare high-fatal hepatobiliary malignancy, the treatment option of ICC is very limited, and the prognosis is also poor. Recently, emerging evidence has shown the potential of quercetin (QE) for cancer therapy. We explored the effect and mechanism of QE on ICC in vitro and in vivo. CCK-8 assay and Clonogenic assay showed that QE could inhibit ICC cells proliferation and survival. PI staining suggested QE could induce ICC cells arrest in G1 phase. AV/PI staining suggested QE could promote ICC cells apoptosis. Wound Healing Assay and Transwell chamber experiment suggested QE could inhibit ICC cells EMT. RNA-seq, the changes in the structure of mitochondria by electron microscopy and the key markers of ferroptosis (free iron ions, MDA, SOD, GPX4) were supported QE could promote ferroptosis in ICC cells. Molecular docking showed that QE had direct interaction with NF-κB and GPX4. In vivo, treatment with QE inhibited tumor growth and prolonged survival time of tumor-bearing nude mice. Our data for the first time suggest that QE is a new ferroptosis inducer and combinative treatment of inhibiting NF-κB in ICC cells by inducing ferroptosis and inhibiting EMT, which will hopefully provide a prospective strategy for ICC patients.

The fate of medial edge epithelial cells during palatal fusion in vitro: an analysis by DiI labelling and confocal microscopy

Development ◽  
1992 ◽  
Vol 114 (2) ◽  
pp. 379-388 ◽  
Author(s):  
M.J. Carette ◽  
M.W. Ferguson
Keyword(s):  
Laser Scanning ◽  
Time Course ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Palatal Fusion ◽  
Morphological Criteria ◽  
Medial Edge ◽  
Mesenchymal Transformation

Fusion of bilateral shelves, to form the definitive mammalian secondary palate, is critically dependent on removal of the medial edge cells that constitute the midline epithelial seam. Conflicting views suggest that programmed apoptotic death or epithelial-mesenchymal transformation of these cells is predominantly involved. Due in part to the potentially ambiguous interpretation of static images and the notable absence of fate mapping studies, the process by which this is achieved has, however, remained mechanistically equivocal. Using an in vitro mouse model, we have selectively labelled palatal epithelia with DiI and examined the fate of medial edge epithelial (MEE) cells during palatal fusion by localisation using a combination of conventional histology and confocal laser scanning microscopy (CLSM). In dynamic studies using CLSM, we have made repetitive observations of the same palatal cultures in time-course investigations. Our results concurred with the established morphological criteria of seam degeneration; however, they provided no evidence of MEE cell death or transformation. Instead we report that MEE cells migrate nasally and orally out of the seam and are recruited into, and constitute, epithelial triangles on both the oral and nasal aspects of the palate. Subsequently these cells become incorporated into the oral and nasal epithelia on the surface of the palate. We hypothesize an alternative method of seam degeneration in vivo which largely conserves the MEE population by recruiting it into the nasal and oral epithelia.

Morphometrical Ultrastructural Studies of the Rat's Hepatocytes Under Cooling

1997 ◽  
Vol 3 (S2) ◽  
pp. 245-246
Author(s):  
A.S. Kaprelyants ◽  
A.A. Kaprelyants ◽  
A.N. Reylan ◽  
R.K. Migunova
Keyword(s):  
Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Electron Microscopic ◽  
Rat Hepatocyte ◽  
Ultrastructural Studies ◽  
Transmission Electron ◽  
Transmission Electron Microscopic ◽  
Morphometric Methods

The aim of given investigation is to study the effect of cooling upon rat hepatocyte structure using transmission electron microscopic and computer morphometric methods. Ultrastructural and morphometrical characteristics of hepatocytes under liver cooling for various levels under in vivo and in vitro conditions were investigated. Vistar rats of 180-250 g were used in the experiment. Liver cooling (in vivo) was performed by means of original cryoapplicator with different probe temperature (1,2). Liver tissue for transmission electron microscopy was fixed in glutaraldehyde fixator on cocadylate buffer and OsO4. Dehydration was completed on acetone (3). Tissue embedding was done into the mixture of Epon/Araldite epoxy rasin. Ultrathin slices were contrasted by the method of Reinolds. Cell viewing and imaging were accomplished by electron microscope at accelerating power of 75kV.Morphometrical and stereometrical analysis was performed using the “Morpho-Tools” original computer system (c) 1994-1996 A.S. Kaprelyants, A.A. Kaprelyants, A.N. Reylan .

TGFbeta3 promotes transformation of chicken palate medial edge epithelium to mesenchyme in vitro

Development ◽  
1998 ◽  
Vol 125 (1) ◽  
pp. 95-105
Author(s):  
D. Sun ◽  
C.R. Vanderburg ◽  
G.S. Odierna ◽  
E.D. Hay
Keyword(s):  
Close Contact ◽  
Defined Medium ◽  
Physical Contact ◽  
Developmental Potential ◽  
Agar Gels ◽  
Membrane Bound ◽  
Medial Edge ◽  
Mesenchymal Transformation

Epithelial-mesenchymal transformation plays an important role in the disappearance of the midline line epithelial seam in rodent palate, leading to confluence of the palate. The aim of this study was to test the potential of the naturally cleft chicken palate to become confluent under the influence of growth factors, such as TGFbeta3, which are known to promote epithelial-mesenchymal transformation. After labeling medial edge epithelia with carboxyfluorescein, palatal shelves (E8-9) with or without beak were dissected and cultured on agar gels. TGFbeta1, TGFbeta2 or TGFbeta3 was added to the chemically defined medium. By 24 hours in culture, medial edge epithelia form adherent midline seams in all paired groups without intact beaks. After 72 hours, seams in the TGFbeta3 groups disappear and palates become confluent due to epithelial-mesenchymal transformation, while seams remain mainly epithelial in control, TGFbeta1 and TGFbeta2 groups. Epithelium-derived mesenchymal cells are identified by carboxyfluorescein fluorescence with confocal microscopy and by membrane-bound carboxyfluorescein isolation bodies with electron microscopy. Labeled fibroblasts completely replace the labeled epithelia of origin in TGFbeta3-treated palates without beaks. Single palates are unable to undergo transformation, and paired palatal shelves with intact beaks do not adhere or undergo transformation, even when treated with TGFbeta3. Thus, physical contact of medial edge epithelia and formation of the midline seam are necessary for epithelial-mesenchymal transformation to be triggered. We conclude that there may be no fundamental difference in developmental potential of the medial edge epithelium for transformation to mesenchyme among reptiles, birds and mammals. The bird differs from other amniotes in having developed a beak and associated craniofacial structures that seemingly keep palatal processes separated in vivo. Even control medial edge epithelia partly transform to mesenchyme if placed in close contact. However, exogenous TGFbeta3 is required to achieve complete confluence of the chicken palate.

scholarly journals Epithelia suspended in collagen gels can lose polarity and express characteristics of migrating mesenchymal cells.

1982 ◽  
Vol 95 (1) ◽  
pp. 333-339 ◽  
Author(s):  
G Greenburg ◽  
E D Hay
Keyword(s):  
Epithelial Cells ◽  
Cell Polarity ◽  
Three Dimensional ◽  
Collagen Gel ◽  
Mesenchymal Cells ◽  
Apical Surface ◽  
Collagen Gels ◽  
Mesenchymal Transformation

This study of epithelial-mesenchymal transformation and epithelial cell polarity in vitro reveals that environmental conditions can have a profound effect on the epithelial phenotype, cell shape, and polarity as expressed by the presence of apical and basal surfaces. A number of different adult and embryonic epithelia were suspended within native collagen gels. Under these conditions, cells elongate, detach from the explants, and migrate as individual cells within the three-dimensional lattice, a previously unknown property of well-differentiated epithelia. Epithelial cells from adult and embryonic anterior lens were studied in detail. Elongated cells derived from the apical surface develop pseudopodia and filopodia characteristic of migratory cells and acquire a morphology and ultrastructure virtually indistinguishable from that of mesenchymal cells in vivo. It is concluded from these experiments that the three-dimensional collagen gel can promote dissociation, migration, and acquisition of secretory organelles by differentiated epithelial cells, and can abolish the apical-basal cell polarity characteristic of the original epithelium.

scholarly journals Cell Wall-associated Protein A as a Tool for Immunolocalization of Staphylococcus aureus in Infected Human Airway Epithelium

2000 ◽  
Vol 48 (4) ◽  
pp. 523-533 ◽  
Author(s):  
Emmanuel Mongodin ◽  
Odile Bajolet ◽  
Jocelyne Hinnrasky ◽  
Edith Puchelle ◽  
Sophie de Bentzmann
Keyword(s):  
Electron Microscopy ◽  
Staphylococcus Aureus ◽  
Epithelial Cells ◽  
Airway Epithelium ◽  
Protein A ◽  
Staphylococcal Protein A ◽  
Basal Cells ◽  
Bronchial Diseases

Staphylococcus aureus is a common human pathogen involved in non-bronchial diseases and in genetic and acquired bronchial diseases. In this study, we applied an immunolabeling approach for in vivo and in vitro detection of S. aureus, based on the affinity of staphylococcal protein A (SpA) for the Fc region of immunoglobulins, especially IgG. Most strains of S. aureus, including clinical strains, can be detected with this labeling technique. The approach can be used for detection and localization with transmission electron microscopy or light-fluorescence microscopy of S. aureus in infected tissues such as human bronchial tissue from cystic fibrosis (CF) patients. The methodology can also be applied to cell culture models with the aim of characterizing bacterial adherence to epithelial cells in backscattered electron imaging with scanning electron microscopy. Application to the study of S. aureus adherence to airway epithelium showed that the bacteria did not adhere in vivo to intact airway epithelium. In contrast, bacteria adhered to the basolateral plasma membrane of columnar cells, to basal cells, to the basement membrane and were identified beneath the lamina propria when the epithelium was injured and remodeled, or in vitro when the epithelial cells were dedifferentiated.

scholarly journals Schwann Cell Autophagy and Necrosis as Mechanisms of Cell Death by Acanthamoeba

Pathogens ◽  
2020 ◽  
Vol 9 (6) ◽  
pp. 458
Author(s):  
Ismael Castelan-Ramírez ◽  
Lizbeth Salazar-Villatoro ◽  
Bibiana Chávez-Munguía ◽  
Citlaltepetl Salinas-Lara ◽  
Carlos Sánchez-Garibay ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Confocal Microscopy ◽  
Rattus Norvegicus ◽  
Multilamellar Bodies ◽  
Transmission Electron ◽  
Granulomatous Amoebic Encephalitis ◽  
Etiological Agents ◽  
First Time

Amoebae of the genus Acanthamoeba are etiological agents of granulomatous amoebic encephalitis (GAE). Recently, through an in vivo GAE model, Acanthamoeba trophozoites were immunolocalized in contact with the peripheral nervous system (PNS) cells—Schwann cells (SC). In this study, we analyzed in greater detail the in vitro early morphological events (1, 2, 3, and 4 h) during the interaction of A. culbertsoni trophozoites (ATCC 30171) with SC from Rattus norvegicus (ATCC CRL-2941). Samples were processed for scanning and transmission electron microscopy as well as confocal microscopy. After 1 h of interaction, amoebae were observed to be adhered to the SC cultures, emitting sucker-like structures associated with micro-phagocytic channels. In addition, evidence of necrosis was identified since edematous organelles as well as multivesicular and multilamellar bodies characteristics of autophagy were detected. At 2 h, trophozoites migrated beneath the SC culture in which necrosis and autophagy persisted. By 3 and 4 h, extensive lytic zones were observed. SC necrosis was confirmed by confocal microscopy. We reported for the first time the induction of autophagic and necrotic processes in PNS cells, associated in part with the contact-dependent pathogenic mechanisms of A. culbertsoni trophozoites.

Steps of nuclear pore complex disassembly and reassembly during mitosis in earlyDrosophilaembryos

2001 ◽  
Vol 114 (20) ◽  
pp. 3607-3618 ◽  
Author(s):  
Elena Kiseleva ◽  
Sandra Rutherford ◽  
Laura M. Cotter ◽  
Terence D. Allen ◽  
Martin W. Goldberg
Keyword(s):  
Electron Microscopy ◽  
Nuclear Pore Complex ◽  
Drosophila Embryo ◽  
Nuclear Pore ◽  
Polar Regions ◽  
Transmission Electron ◽  
Vitro Model ◽  
First Time

The mechanisms of nuclear pore complex (NPC) assembly and disassembly during mitosis in vivo are not well defined. To address this and to identify the steps of the NPC disassembly and assembly, we investigated Drosophila embryo nuclear structure at the syncytial stage of early development using field emission scanning electron microscopy (FESEM), a high resolution surface imaging technique, and transmission electron microscopy. Nuclear division in syncytial embryos is characterized by semi-closed mitosis, during which the nuclear membranes are ruptured only at the polar regions and are arranged into an inner double membrane surrounded by an additional ‘spindle envelope’. FESEM analysis of the steps of this process as viewed on the surface of the dividing nucleus confirm our previous in vitro model for the assembly of the NPCs via a series of structural intermediates, showing for the first time a temporal progression from one intermediate to the next. Nascent NPCs initially appear to form at the site of fusion between the mitotic nuclear envelope and the overlying spindle membrane. A model for NPC disassembly is offered that starts with the release of the central transporter and the removal of the cytoplasmic ring subunits before the star ring.

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