Biomaterial Database

Quantifying Polarized Extracellular Matrix Secretion in Cultured Endothelial Cells. 2021

Fabiana Clapero, and Dora Tortarolo, and Donatella Valdembri, and Guido Serini

Candiolo Cancer Institute, FPO-IRCCS, Candiolo, Italy.

In endothelial cells (ECs), the onset of apicobasal polarity is primarily regulated by the interaction of integrins with the surrounding extracellular matrix (ECM). ECs secrete and polymerize fibronectin (FN), a unique, permissive substrate that allows for vascular morphogenesis and lumen formation. We previously identified a signaling pathway that, under the control of the adhesion site adaptor protein PPFIA1, integrates the polarized secretion of freshly synthesized FN with the recycling of conformationally active α5β1 integrin, the main FN receptor in ECs. To characterize the functional role of PPFIA1-dependent signaling in ECs, we set up a Transwell-based assay to quantify the polarized secretion of ECM proteins. To this aim, we allowed ECs to form a confluent monolayer on the Transwell membrane and checked its integrity by measuring transendothelial electric resistance and controlling the stability of tight junctions over time by fluorescent confocal microscope analysis. Finally, we quantified apical and basolateral FN secretion in control and PPFIA1-silenced EC culture medium by western blot analysis coupled to spike-in normalization.

UI	MeSH Term	Description	Entries
D008856	Microscopy, Fluorescence	Microscopy of specimens stained with fluorescent dye (usually fluorescein isothiocyanate) or of naturally fluorescent materials, which emit light when exposed to ultraviolet or blue light. Immunofluorescence microscopy utilizes antibodies that are labeled with fluorescent dye.	Fluorescence Microscopy,Immunofluorescence Microscopy,Microscopy, Immunofluorescence,Fluorescence Microscopies,Immunofluorescence Microscopies,Microscopies, Fluorescence,Microscopies, Immunofluorescence
D005109	Extracellular Matrix	A meshwork-like substance found within the extracellular space and in association with the basement membrane of the cell surface. It promotes cellular proliferation and provides a supporting structure to which cells or cell lysates in culture dishes adhere.	Matrix, Extracellular,Extracellular Matrices,Matrices, Extracellular
D005353	Fibronectins	Glycoproteins found on the surfaces of cells, particularly in fibrillar structures. The proteins are lost or reduced when these cells undergo viral or chemical transformation. They are highly susceptible to proteolysis and are substrates for activated blood coagulation factor VIII. The forms present in plasma are called cold-insoluble globulins.	Cold-Insoluble Globulins,LETS Proteins,Fibronectin,Opsonic Glycoprotein,Opsonic alpha(2)SB Glycoprotein,alpha 2-Surface Binding Glycoprotein,Cold Insoluble Globulins,Globulins, Cold-Insoluble,Glycoprotein, Opsonic,Proteins, LETS,alpha 2 Surface Binding Glycoprotein
D005786	Gene Expression Regulation	Any of the processes by which nuclear, cytoplasmic, or intercellular factors influence the differential control (induction or repression) of gene action at the level of transcription or translation.	Gene Action Regulation,Regulation of Gene Expression,Expression Regulation, Gene,Regulation, Gene Action,Regulation, Gene Expression
D006056	Golgi Apparatus	A stack of flattened vesicles that functions in posttranslational processing and sorting of proteins, receiving them from the rough ENDOPLASMIC RETICULUM and directing them to secretory vesicles, LYSOSOMES, or the CELL MEMBRANE. The movement of proteins takes place by transfer vesicles that bud off from the rough endoplasmic reticulum or Golgi apparatus and fuse with the Golgi, lysosomes or cell membrane. (From Glick, Glossary of Biochemistry and Molecular Biology, 1990)	Golgi Complex,Apparatus, Golgi,Complex, Golgi
D006801	Humans	Members of the species Homo sapiens.	Homo sapiens,Man (Taxonomy),Human,Man, Modern,Modern Man
D001692	Biological Transport	The movement of materials (including biochemical substances and drugs) through a biological system at the cellular level. The transport can be across cell membranes and epithelial layers. It also can occur within intracellular compartments and extracellular compartments.	Transport, Biological,Biologic Transport,Transport, Biologic
D015194	Diffusion Chambers, Culture	Devices used in a technique by which cells or tissues are grown in vitro or, by implantation, in vivo within chambers permeable to diffusion of solutes across the chamber walls. The chambers are used for studies of drug effects, osmotic responses, cytogenic and immunologic phenomena, metabolism, etc., and include tissue cages.	Culture Diffusion Chambers,Semi-Permeable Chambers,Tissue Cages,Semipermeable Chambers,Cage, Tissue,Cages, Tissue,Chamber, Culture Diffusion,Chamber, Semi-Permeable,Chamber, Semipermeable,Chambers, Culture Diffusion,Chambers, Semi-Permeable,Chambers, Semipermeable,Culture Diffusion Chamber,Diffusion Chamber, Culture,Semi Permeable Chambers,Semi-Permeable Chamber,Semipermeable Chamber,Tissue Cage
D015398	Signal Transduction	The intracellular transfer of information (biological activation/inhibition) through a signal pathway. In each signal transduction system, an activation/inhibition signal from a biologically active molecule (hormone, neurotransmitter) is mediated via the coupling of a receptor/enzyme to a second messenger system or to an ion channel. Signal transduction plays an important role in activating cellular functions, cell differentiation, and cell proliferation. Examples of signal transduction systems are the GAMMA-AMINOBUTYRIC ACID-postsynaptic receptor-calcium ion channel system, the receptor-mediated T-cell activation pathway, and the receptor-mediated activation of phospholipases. Those coupled to membrane depolarization or intracellular release of calcium include the receptor-mediated activation of cytotoxic functions in granulocytes and the synaptic potentiation of protein kinase activation. Some signal transduction pathways may be part of larger signal transduction pathways; for example, protein kinase activation is part of the platelet activation signal pathway.	Cell Signaling,Receptor-Mediated Signal Transduction,Signal Pathways,Receptor Mediated Signal Transduction,Signal Transduction Pathways,Signal Transduction Systems,Pathway, Signal,Pathway, Signal Transduction,Pathways, Signal,Pathways, Signal Transduction,Receptor-Mediated Signal Transductions,Signal Pathway,Signal Transduction Pathway,Signal Transduction System,Signal Transduction, Receptor-Mediated,Signal Transductions,Signal Transductions, Receptor-Mediated,System, Signal Transduction,Systems, Signal Transduction,Transduction, Signal,Transductions, Signal
D016764	Cell Polarity	Orientation of intracellular structures especially with respect to the apical and basolateral domains of the plasma membrane. Polarized cells must direct proteins from the Golgi apparatus to the appropriate domain since tight junctions prevent proteins from diffusing between the two domains.	Cell Polarities,Polarities, Cell,Polarity, Cell