Wachstumsfaktorenkonzentration im Kulturüberstand von Keratozyten mit humanem Serum in vitro

2017 ◽  
Vol 235 (07) ◽  
pp. 840-845
Author(s):  
Nóra Szentmáry ◽  
Tanja Stachon ◽  
Ming-Feng Wu ◽  
Mona Bischoff ◽  
Manuela Huber ◽  
...  
Keyword(s):  
Growth Factor ◽  
Fibroblast Growth Factor ◽  
Transforming Growth Factor ◽  
Keratinocyte Growth Factor ◽  
Transforming Growth Factor Β1 ◽  
Fibroblast Growth ◽  
Fibroblast Growth Factor Basic ◽  
Hepatocyte Growth ◽  
Tgf Β1

Zusammenfassung Hintergrund Die Anwendung von Serumaugentropfen (AS) stellt eine alternative Behandlungsmethode für therapieresistente korneale Epitheldefekte dar. Bei persistierenden Epitheldefekten könnten Zytokine, die von Keratozyten im Stroma produziert werden, eine entscheidende Rolle bei der epithelialen Wundheilung spielen. Ziel dieser Studie ist es, Transforming Growth Factor β1 (TGF-β1), Keratinocyte Growth Factor (KGF), Hepatocyte Growth Factor (HGF) und Fibroblast Growth Factor basic (FGFb) im Kulturüberstand von Keratozyten mit humanem Serum (HS) in vitro zu untersuchen. Material und Methoden Serumaugentropfen von 10 Patienten wurden nach der Standardmethode der LIONS Hornhautbank Saar-Lor-Lux präpariert und bei − 80 °C tiefgefroren. Primäre humane Keratozyten wurden durch enzymatische Behandlung mit Kollagenase A (1 mg/ml) aus humanen Korneoskleralscheiben isoliert (n = 1) und in DMEM/Hamʼs Kulturmedium, versetzt mit 5% fetalem Kälberserum (FCS), kultiviert. Für den Testansatz wurden die Keratozyten mit 15 oder 30% HS (in DMEM/F14 ohne FCS) inkubiert und nach 24 h die Konzentration von TGF-β1, KGF, HGF und FGFb mittels Enzyme-linked Immunoabsorbent Assay (ELISA) aus dem Kulturüberstand bestimmt. Als Kontrolle wurden 15 oder 30% HS ohne Keratozyten nach 24 h Inkubationszeit (unter den gleichen Bedingungen wie die Keratozyten) verwendet. Ergebnisse Die HGF-Konzentration mit beiden HS-Konzentrationen war im Kulturüberstand von Keratozyten signifikant höher, im Vergleich zur HS-Kontrolle (ohne Keratozyten) nach 24 h Inkubationszeit (p < 0,01). Die FGFb-Konzentration war im Kulturüberstand mit 30% HS signifikant höher im Vergleich zur Kontrollgruppe ohne Keratozyten nach 24 Stunden Inkubationszeit (p < 0,01). Die TGF-β1- und KGF-Konzentrationen im Kulturüberstand blieben durch die Keratozyten unverändert. Schlussfolgerungen Durch die Anwesenheit von Keratozyten steigt die Konzentration von HGF und FGFb im Kulturmedium mit humanem Serum innerhalb von 24 Stunden an. Diese Konzentrationsänderungen könnten die Wundheilung bei Epitheldefekten beeinflussen.

scholarly journals Transforming Growth Factor β1 (TGF-β1) Promotes Endothelial Cell Survival during In Vitro Angiogenesis via an Autocrine Mechanism Implicating TGF-α Signaling

2001 ◽  
Vol 21 (21) ◽  
pp. 7218-7230 ◽  
Author(s):  
Francesc Viñals ◽  
Jacques Pouysségur
Keyword(s):  
Endothelial Cells ◽  
Growth Factor ◽  
Cell Survival ◽  
Network Formation ◽  
Egf Receptor ◽  
Transforming Growth Factor ◽  
Transforming Growth Factor Β1 ◽  
In Vitro Angiogenesis ◽  
Tgf Β1

ABSTRACT Mouse capillary endothelial cells (1G11 cell line) embedded in type I collagen gels undergo in vitro angiogenesis. Cells rapidly reorganize and form capillary-like structures when stimulated with serum. Transforming growth factor β1 (TGF-β1) alone can substitute for serum and induce cell survival and tubular network formation. This TGF-β1-mediated angiogenic activity depends on phosphatidylinositol 3-kinase (PI3K) and p42/p44 mitogen-activated protein kinase (MAPK) signaling. We showed that specific inhibitors of either pathway (wortmannin, LY-294002, and PD-98059) all suppressed TGF-β1-induced angiogenesis mainly by compromising cell survival. We established that TGF-β1 stimulated the expression of TGF-α mRNA and protein, the tyrosine phosphorylation of a 170-kDa membrane protein representing the epidermal growth factor (EGF) receptor, and the delayed activation of PI3K/Akt and p42/p44 MAPK. Moreover, we showed that all these TGF-β1-mediated signaling events, including tubular network formation, were suppressed by incubating TGF-β1-stimulated endothelial cells with a soluble form of an EGF receptor (ErbB-1) or tyrphostin AG1478, a specific blocker of EGF receptor tyrosine kinase. Finally, addition of TGF-α alone poorly stimulated angiogenesis; however, by reducing cell death, it strongly potentiated the action of TGF-β1. We therefore propose that TGF-β1 promotes angiogenesis at least in part via the autocrine secretion of TGF-α, a cell survival growth factor, activating PI3K/Akt and p42/p44 MAPK.

Selective optogenetic stimulation of fibroblasts enables quantification of hetero-cellular coupling to cardiomyocytes in a three-dimensional model of heart tissue

EP Europace ◽  
2020 ◽  
Vol 22 (10) ◽  
pp. 1590-1599
Author(s):  
Maximilian Funken ◽  
Tobias Bruegmann ◽  
Philipp Sasse
Keyword(s):  
Membrane Potential ◽  
Growth Factor ◽  
Connexin 43 ◽  
Transforming Growth Factor ◽  
Three Dimensional ◽  
Transforming Growth Factor Β1 ◽  
Resting Membrane Potential ◽  
Human Cardiomyocytes ◽  
Tgf Β1

Abstract Aims Besides providing mechanical stability, fibroblasts in the heart could modulate the electrical properties of cardiomyocytes. Here, we aim to develop a three-dimensional hetero-cellular model to analyse the electric interaction between fibroblasts and human cardiomyocytes in vitro using selective optogenetic de- or hyperpolarization of fibroblasts. Methods and results NIH3T3 cell lines expressing the light-sensitive ion channel Channelrhodopsin2 or the light-induced proton pump Archaerhodopsin were generated for optogenetic depolarization or hyperpolarization, respectively, and characterized by patch clamp. Cardiac bodies consisting of 50% fibroblasts and 50% human pluripotent stem cell-derived cardiomyocytes were analysed by video microscopy and membrane potential was measured with sharp electrodes. Myofibroblast activation in cardiac bodies was enhanced by transforming growth factor-β1 (TGF-β1)-stimulation. Connexin-43 expression was analysed by qPCR and fluorescence recovery after photobleaching. Illumination of Channelrhodopsin2 or Archaerhodopsin expressing fibroblasts induced inward currents and depolarization or outward currents and hyperpolarization. Transforming growth factor-β1-stimulation elevated connexin-43 expression and increased cell–cell coupling between fibroblasts as well as increased basal beating frequency and cardiomyocyte resting membrane potential in cardiac bodies. Illumination of cardiac bodies generated with Channelrhodopsin2 fibroblasts accelerated spontaneous beating, especially after TGF-β1-stimulation. Illumination of cardiac bodies prepared with Archaerhodopsin expressing fibroblasts led to hyperpolarization of cardiomyocytes and complete block of spontaneous beating after TGF-β1-stimulation. Effects of light were significantly smaller without TGF-β1-stimulation. Conclusion Transforming growth factor-β1-stimulation leads to increased hetero-cellular coupling and optogenetic hyperpolarization of fibroblasts reduces TGF-β1 induced effects on cardiomyocyte spontaneous activity. Optogenetic membrane potential manipulation selectively in fibroblasts in a new hetero-cellular cardiac body model allows direct quantification of fibroblast–cardiomyocyte coupling in vitro.

scholarly journals Microcarriers Based on Glycosaminoglycan-Like Marine Exopolysaccharide for TGF-β1 Long-Term Protection

Marine Drugs ◽  
2019 ◽  
Vol 17 (1) ◽  
pp. 65 ◽  
Author(s):  
Agata Zykwinska ◽  
Mélanie Marquis ◽  
Mathilde Godin ◽  
Laëtitia Marchand ◽  
Corinne Sinquin ◽  
...  
Keyword(s):  
Tissue Engineering ◽  
Growth Factor ◽  
Transforming Growth Factor ◽  
Target Location ◽  
Cartilage Regeneration ◽  
Transforming Growth Factor Β1 ◽  
Hydrogel Matrix ◽  
Vitro Model ◽  
Tgf Β1

Articular cartilage is an avascular, non-innervated connective tissue with limited ability to regenerate. Articular degenerative processes arising from trauma, inflammation or due to aging are thus irreversible and may induce the loss of the joint function. To repair cartilaginous defects, tissue engineering approaches are under intense development. Association of cells and signalling proteins, such as growth factors, with biocompatible hydrogel matrix may lead to the regeneration of the healthy tissue. One current strategy to enhance both growth factor bioactivity and bioavailability is based on the delivery of these signalling proteins in microcarriers. In this context, the aim of the present study was to develop microcarriers by encapsulating Transforming Growth Factor-β1 (TGF-β1) into microparticles based on marine exopolysaccharide (EPS), namely GY785 EPS, for further applications in cartilage engineering. Using a capillary microfluidic approach, two microcarriers were prepared. The growth factor was either encapsulated directly within the microparticles based on slightly sulphated derivative or complexed firstly with the highly sulphated derivative before being incorporated within the microparticles. TGF-β1 release, studied under in vitro model conditions, revealed that the majority of the growth factor was retained inside the microparticles. Bioactivity of released TGF-β1 was particularly enhanced in the presence of highly sulphated derivative. It comes out from this study that GY785 EPS based microcarriers may constitute TGF-β1 reservoirs spatially retaining the growth factor for a variety of tissue engineering applications and in particular cartilage regeneration, where the growth factor needs to remain in the target location long enough to induce robust regenerative responses.

Fibroblast growth factor 2 and transforming growth factor β1 interactions in human liver myofibroblasts

1995 ◽  
Vol 109 (6) ◽  
pp. 1986-1996 ◽  
Author(s):  
Jean Rosenbaum ◽  
Sylvie Blazejewski ◽  
Anne-Marie Préaux ◽  
Ariane Mallat ◽  
Daniel Dhumeaux ◽  
...  
Keyword(s):  
Growth Factor ◽  
Fibroblast Growth Factor ◽  
Human Liver ◽  
Transforming Growth Factor ◽  
Transforming Growth Factor Β1 ◽  
Fibroblast Growth Factor 2 ◽  
Fibroblast Growth ◽  
Liver Myofibroblasts
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