scholarly journals Actomyosin dynamics, Bmp and Notch signaling pathways drive apical extrusion of proepicardial cells

2018 ◽  
Author(s):  
Laura Andrés-Delgado ◽  
Alexander Ernst ◽  
María Galardi-Castilla ◽  
David Bazaga ◽  
Marina Peralta ◽  
...  
Keyword(s):  
Cell Viability ◽  
Notch Signaling ◽  
Heart Development ◽  
Zebrafish Model ◽  
Pharmacological Agents ◽  
Apical Extrusion ◽  
A Cell ◽  
Cell Extrusion ◽  
Unique Mechanism

ABSTRACTThe epicardium, the outer mesothelial layer enclosing the myocardium, plays key roles in heart development and regeneration. During embryogenesis it arises from the proepicardium (PE), a cell cluster that appears in the dorsal pericardium close to the venous pole of the heart. Little is known about how the PE emerges from the pericardial mesothelium. Using the zebrafish model and a combination of genetic tools, pharmacological agents and quantitative in vivo imaging we reveal that a coordinated collective movement of the dorsal pericardium drives PE formation. We found that PE cells are apically extruded in response to actomyosin activity. Our results reveal that the coordinated action of Notch/Bmp pathways is critically needed for apical extrusion of PE cells. More generally, by comparison to cell extrusion for the elimination of unfit cells from epithelia, our results describe a unique mechanism where extruded cell viability is maintained.

scholarly journals High-resolution cardiovascular function confirms functional orthology of myocardial contractility pathways in zebrafish

2010 ◽  
Vol 42 (2) ◽  
pp. 300-309 ◽  
Author(s):  
Jordan T. Shin ◽  
Eugene V. Pomerantsev ◽  
John D. Mably ◽  
Calum A. MacRae
Keyword(s):  
High Speed ◽  
Dynamic Range ◽  
Model Organism ◽  
Cardiovascular Physiology ◽  
Zebrafish Embryos ◽  
Cardiovascular Development ◽  
Zebrafish Model ◽  
Ventricular Performance ◽  
Pharmacological Agents

Phenotype-driven screens in larval zebrafish have transformed our understanding of the molecular basis of cardiovascular development. Screens to define the genetic determinants of physiological phenotypes have been slow to materialize as a result of the limited number of validated in vivo assays with relevant dynamic range. To enable rigorous assessment of cardiovascular physiology in living zebrafish embryos, we developed a suite of software tools for the analysis of high-speed video microscopic images and validated these, using established cardiomyopathy models in zebrafish as well as modulation of the nitric oxide (NO) pathway. Quantitative analysis in wild-type fish exposed to NO or in a zebrafish model of dilated cardiomyopathy demonstrated that these tools detect significant differences in ventricular chamber size, ventricular performance, and aortic flow velocity in zebrafish embryos across a large dynamic range. These methods also were able to establish the effects of the classic pharmacological agents isoproterenol, ouabain, and verapamil on cardiovascular physiology in zebrafish embryos. Sequence conservation between zebrafish and mammals of key amino acids in the pharmacological targets of these agents correlated with the functional orthology of the physiological response. These data provide evidence that the quantitative evaluation of subtle physiological differences in zebrafish can be accomplished at a resolution and with a dynamic range comparable to those achieved in mammals and provides a mechanism for genetic and small-molecule dissection of functional pathways in this model organism.

Using in vivo zebrafish models to understand the biochemical basis of neutrophilic respiratory disease

2009 ◽  
Vol 37 (4) ◽  
pp. 830-837 ◽  
Author(s):  
Jane S. Martin ◽  
Stephen A. Renshaw
Keyword(s):  
Respiratory Disease ◽  
Fluorescent Protein ◽  
Reverse Genetic ◽  
Zebrafish Model ◽  
Pharmacological Agents ◽  
Biochemical Basis ◽  
Neutrophilic Inflammation ◽  
Green Fluorescent ◽  
Inflammation Resolution

Neutrophilic inflammation in the lung protects against infectious disease, and usually resolves spontaneously after removal of the inflammatory stimulus. However, much lung disease is caused by a failure of resolution of neutrophilic inflammation. Our laboratory is seeking an understanding of the biochemical basis of inflammation resolution, using the zebrafish model system. Zebrafish larvae are transparent, allowing visualization of GFP (green fluorescent protein)-labelled leucocytes during inflammation in vivo, and they can be readily manipulated by a range of forward and reverse genetic techniques. This combination of advantages makes zebrafish a powerful tool for the study of in vivo inflammatory processes. Using this model, we have visualized the process of inflammation resolution in vivo, and identified a role for apoptosis in this process. In addition, we have performed a forward genetic screen for mutants with defective resolution of inflammation, and reverse genetic experiments examining the influence of candidate genes on inflammation resolution. We have established a platform for screening for compounds with anti-inflammatory activity, which has yielded a number of interesting leads. Looking forward to succeed in the future, we are working at combining mutants, transgenes and pharmacological agents to dissect the biochemical basis of inflammation resolution, and to identify compounds that might be used to treat patients with respiratory disease.

scholarly journals Small-molecule synergist of the Wnt/β-catenin signaling pathway

2007 ◽  
Vol 104 (18) ◽  
pp. 7444-7448 ◽  
Author(s):  
Qisheng Zhang ◽  
Michael B. Major ◽  
Shinichi Takanashi ◽  
Nathan D. Camp ◽  
Naoyuki Nishiya ◽  
...  
Keyword(s):  
Wnt Signaling ◽  
Small Molecules ◽  
Signaling Pathway ◽  
Cell Fate ◽  
Protein Trafficking ◽  
Purine Derivative ◽  
Pharmacological Agents ◽  
A Cell ◽  
And Behavior

The Wnt/β-catenin signaling pathway regulates cell fate and behavior during embryogenesis, adult tissue homeostasis, and regeneration. When inappropriately activated, the pathway has been linked to colorectal cancer and melanoma, and when attenuated it may contribute to Alzheimer's disease and osteoporosis. Small molecules that modulate Wnt signaling will likely provide new insights into the regulation of this key developmental pathway and ultimately provide pharmacological agents to control Wnt signaling in vivo. To this end, we screened a library of 100,000 small molecules for activity in a cell-based assay of Wnt/β-catenin signaling and discovered a purine derivative, QS11, that synergizes with Wnt-3a ligand in the activation of Wnt/β-catenin signal transduction. Through affinity chromatography and subsequent functional assays, we showed that QS11 binds and inhibits the GTPase activating protein of ADP-ribosylation factor 1 (ARFGAP1), suggesting that QS11 modulates Wnt/β-catenin signaling through an effect on protein trafficking. Consistent with its function as an ARFGAP inhibitor, QS11 inhibits migration of ARFGAP overexpressing breast cancer cells.

Notch signaling represses the glial fate in fly PNS

Development ◽  
2001 ◽  
Vol 128 (8) ◽  
pp. 1381-1390 ◽  
Author(s):  
V. Van De Bor ◽  
A. Giangrande
Keyword(s):  
Nervous System ◽  
Notch Signaling ◽  
Glial Cell ◽  
Glial Cells ◽  
Genetic Switch ◽  
Organ Type ◽  
A Cell ◽  
Glial Fate ◽  
Glial Precursor

By using gain-of-function mutations it has been proposed that vertebrate Notch promotes the glial fate. We show in vivo that glial cells are produced at the expense of neurons in the peripheral nervous system of flies lacking Notch and that constitutively activated Notch produces the opposite phenotype. Notch acts as a genetic switch between neuronal and glial fates by negatively regulating glial cell deficient/glial cells missing, the gene required in the glial precursor to induce gliogenesis. Moreover, Notch represses neurogenesis or gliogenesis, depending on the sensory organ type. Numb, which is asymmetrically localized in the multipotent cell that produces the glial precursor, induces glial cells at the expense of neurons. Thus, a cell-autonomous mechanism inhibits Notch signaling.

scholarly journals A Ca2+ wave generates a force during cell extrusion in zebrafish

2021 ◽  
Author(s):  
Sohei Yamada ◽  
Yasumasa Bessho ◽  
Yasuyuki Fijita ◽  
Yoichiroh Hosokawa ◽  
Takaaki Matsui
Keyword(s):  
Epithelial Cells ◽  
Force Measurement ◽  
Gene Knockout ◽  
Ip3 Receptor ◽  
Mammalian Cell Cultures ◽  
Apical Extrusion ◽  
Epithelial Sheet ◽  
Wave Induced ◽  
Cell Extrusion

When oncogenic transformed or damaged cells appear within an epithelial sheet, they are apically extruded by surrounding cells. Recently, using cultured mammalian epithelial cells and zebrafish embryonic epithelial cells, we found that a calcium (Ca2+) wave propagates from RasV12-transformed cells and laser-irradiated damaged cells to surrounding cells and promotes apical extrusion by inducing polarized movements of the surrounding cells. In mammalian cell cultures, we reported that the inositol trisphosphate (IP3) receptor, gap junctions, and the mechanosensitive Ca2+ channel TRPC1 are involved in Ca2+ wave-mediated polarized movements. However, which molecules regulate Ca2+ wave-mediated polarized movements in zebrafish and whether the Ca2+ wave can generate a force remain unknown. In this study, we aimed to answer these questions. By performing pharmacological and gene knockout experiments, we showed that a Ca2+ wave induced by the IP3 receptor and trpc1 led to formation of cryptic-lamellipodia and polarized movements of surrounding cells toward extruding cells in zebrafish. By using an in vivo force measurement method, we found that the Ca2+ wave generated approximately 1 kPa of force toward extruding cells. Our results reveal a previously unidentified molecular mechanism underlying the Ca2+ wave in zebrafish and demonstrate that the Ca2+ wave generates a force during cell extrusion.

scholarly journals Overexpression of Notch Signaling Induces Hyperosteogeny in Zebrafish

2019 ◽  
Vol 20 (15) ◽  
pp. 3613 ◽  
Author(s):  
Sung-Tzu Liang ◽  
Jung-Ren Chen ◽  
Jhih-Jie Tsai ◽  
Yu-Heng Lai ◽  
Chung-Der Hsiao
Keyword(s):  
Notch Signaling ◽  
Skeletal Development ◽  
Transgenic Zebrafish ◽  
Bone Homeostasis ◽  
Zebrafish Model ◽  
Therapeutic Drugs ◽  
Evolutionarily Conserved ◽  
Multicellular Organisms

Notch signaling is one of the evolutionarily conserved signaling pathways in multicellular organisms. It plays an important role in embryonic development. During skeletal development of vertebrates, it regulates bone homeostasis by manipulating both osteoblastogenesis and osteoclastogenesis through different mechanisms. However, due to the different nature of Notch signaling in mesenchymal stem cell and osteoblast, regulation of Notch signaling in bone-related diseases remains unsettled. Previous studies by cell culture and mouse models showed contradictory results regarding the role of Notch signaling in bone homeostasis. To clarify the role of Notch signaling in osteogenesis, we established a zebrafish model, in which Notch1a intracellular domain (N1aICD) was specifically expressed in the osteoblasts. We found that overexpression of N1aICD in osteoblasts caused hyperosteogeny in the column region of zebrafish with the morphology of narrowed neural/hemal canals. Moreover, increased metabolic activity of osteoblasts instead of augmenting osteoblast number led to hyperosteogeny in N1aICD-overexpressed zebrafish. In summary, we successfully established a transgenic zebrafish line overexpressing N1aICD to clarify the in-vivo function of Notch signaling during osteoblastogenesis. In the future, this fish line can serve as a valuable tool to test the therapeutic drugs for hyperosteogeny.

Ultrastructure of melanocyte-keratinocyte interactions and pigment transfer in vitro

1978 ◽  
Vol 36 (2) ◽  
pp. 650-651
Author(s):  
Raul I. Garcia ◽  
Evelyn A. Flynn ◽  
George Szabo
Keyword(s):  
Direct Injection ◽  
Skin Pigmentation ◽  
Freeze Fracture ◽  
Pigment Granule ◽  
Time Lapse ◽  
Ultrastructural Level ◽  
Lanthanum Tracer ◽  
A Cell

Skin pigmentation in mammals involves the interaction of epidermal melanocytes and keratinocytes in the structural and functional unit known as the Epidermal Melanin Unit. Melanocytes(M) synthesize melanin within specialized membrane-bound organelles, the melanosome or pigment granule. These are subsequently transferred by way of M dendrites to keratinocytes(K) by a mechanism still to be clearly defined. Three different, though not necessarily mutually exclusive, mechanisms of melanosome transfer have been proposed: cytophagocytosis by K of M dendrite tips containing melanosomes, direct injection of melanosomes into the K cytoplasm through a cell-to-cell pore or communicating channel formed by localized fusion of M and K cell membranes, release of melanosomes into the extracellular space(ECS) by exocytosis followed by K uptake using conventional phagocytosis. Variability in methods of transfer has been noted both in vivo and in vitro and there is evidence in support of each transfer mechanism. We Have previously studied M-K interactions in vitro using time-lapse cinemicrography and in vivo at the ultrastructural level using lanthanum tracer and freeze-fracture.

Analysis of the Dynamics of the Electric Capacitance of a Cell Monolayer at the Late Stages of Caco-2 cell Differentiation

2019 ◽  
Vol 35 (6) ◽  
pp. 87-90
Author(s):  
S.V. Nikulin ◽  
V.A. Petrov ◽  
D.A. Sakharov
Keyword(s):  
Impedance Spectroscopy ◽  
Cell Monolayer ◽  
Microfluidic Chips ◽  
Russian Science ◽  
Electric Capacitance ◽  
A Cell ◽  
Static Conditions ◽  
Late Stages

The real-time monitoring of electric capacitance (impedance spectroscopy) allowed obtaining evidence that structures which look like intestinal villi can be formed during the cultivation under static conditions as well as during the cultivation in microfluidic chips. It was shown in this work via transcriptome analysis that the Hh signaling pathway is involved in the formation of villus-like structures in vitro, which was previously shown for their formation in vivo. impedance spectroscopy, intestine, villi, electric capacitance, Hh The study was funded by the Russian Science Foundation (Project 16-19-10597).

scholarly journals Group III phospholipase A2 downregulation attenuated survival and metastasis in ovarian cancer and promotes chemo-sensitization

2021 ◽  
Vol 40 (1) ◽  
Author(s):  
Upasana Ray ◽  
Debarshi Roy ◽  
Ling Jin ◽  
Prabhu Thirusangu ◽  
Julie Staub ◽  
...  
Keyword(s):  
Ovarian Cancer ◽  
Cell Viability ◽  
Phospholipase A2 ◽  
Mtt Assay ◽  
Metabolic Inhibitor ◽  
Autophagic Flux ◽  
Ciliated Cells ◽  
Group Iii

Abstract Background Aberrant lipogenicity and deregulated autophagy are common in most advanced human cancer and therapeutic strategies to exploit these pathways are currently under consideration. Group III Phospholipase A2 (sPLA2-III/PLA2G3), an atypical secretory PLA2, is recognized as a regulator of lipid metabolism associated with oncogenesis. Though recent studies reveal that high PLA2G3 expression significantly correlates with poor prognosis in several cancers, however, role of PLA2G3 in ovarian cancer (OC) pathogenesis is still undetermined. Methods CRISPR-Cas9 and shRNA mediated knockout and knockdown of PLA2G3 in OC cells were used to evaluate lipid droplet (LD) biogenesis by confocal and Transmission electron microscopy analysis, and the cell viability and sensitization of the cells to platinum-mediated cytotoxicity by MTT assay. Regulation of primary ciliation by PLA2G3 downregulation both genetically and by metabolic inhibitor PFK-158 induced autophagy was assessed by immunofluorescence-based confocal analysis and immunoblot. Transient transfection with GFP-RFP-LC3B and confocal analysis was used to assess the autophagic flux in OC cells. PLA2G3 knockout OVCAR5 xenograft in combination with carboplatin on tumor growth and metastasis was assessed in vivo. Efficacy of PFK158 alone and with platinum drugs was determined in patient-derived primary ascites cultures expressing PLA2G3 by MTT assay and immunoblot analysis. Results Downregulation of PLA2G3 in OVCAR8 and 5 cells inhibited LD biogenesis, decreased growth and sensitized cells to platinum drug mediated cytotoxicity in vitro and in in vivo OVCAR5 xenograft. PLA2G3 knockdown in HeyA8MDR-resistant cells showed sensitivity to carboplatin treatment. We found that both PFK158 inhibitor-mediated and genetic downregulation of PLA2G3 resulted in increased number of percent ciliated cells and inhibited cancer progression. Mechanistically, we found that PFK158-induced autophagy targeted PLA2G3 to restore primary cilia in OC cells. Of clinical relevance, PFK158 also induces percent ciliated cells in human-derived primary ascites cells and reduces cell viability with sensitization to chemotherapy. Conclusions Taken together, our study for the first time emphasizes the role of PLA2G3 in regulating the OC metastasis. This study further suggests the therapeutic potential of targeting phospholipases and/or restoration of PC for future OC treatment and the critical role of PLA2G3 in regulating ciliary function by coordinating interface between lipogenesis and metastasis.

Sign in / Sign up

Export Citation Format

Share Document