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The extracellular matrix modulates H2O2 degradation and redox signaling in endothelial cells.

Bagulho A, Vilas-Boas F, Pena A, Peneda C, Santos FC, Jerónimo A, de Almeida RF, Real C - Redox Biol (2015)

Bottom Line: Instead, we found that the ECM regulated GPx activity, a known H2O2 scavenger.Thus, our results unraveled a new mechanism by which the ECM regulates endothelial cell function by altering redox balance.These results pinpoint the ECM as an important component of redox-signaling.

View Article: PubMed Central - PubMed

Affiliation: Centro de Química e Bioquímica, Faculdade de Ciências, Universidade de Lisboa, 1749-016 Lisboa, Portugal. Electronic address: anabagulho@gmail.com.

No MeSH data available.


Catalase and GPx activity in HUVEC plated on different ECMs. In-gel assays to assess the activity of catalase and GPx (band intensity) in HUVEC cultured in gelatin or laminin showed that (A) catalase activity was similar between cells cultured in the two ECMs, whereas (B) GPx activity in cells cultured in the presence of laminin was lower than when cultured in the presence of gelatin (a.u., arbitrary units; values show the mean and SEM; N=5; two-tailed Student's t test, **p=0.005).
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f0015: Catalase and GPx activity in HUVEC plated on different ECMs. In-gel assays to assess the activity of catalase and GPx (band intensity) in HUVEC cultured in gelatin or laminin showed that (A) catalase activity was similar between cells cultured in the two ECMs, whereas (B) GPx activity in cells cultured in the presence of laminin was lower than when cultured in the presence of gelatin (a.u., arbitrary units; values show the mean and SEM; N=5; two-tailed Student's t test, **p=0.005).

Mentions: Our fluorescence spectroscopy results showed that endothelial cells presented similar membrane biophysical properties in the presence of different ECMs, with the observed different hydrogen peroxide consumptions probably not caused by differences in the rate of passive permeation. An alternative explanation for the variations observed in the H2O2 consumption rates of HUVEC in the different ECMs is the alteration of activity of enzymes responsible for H2O2 consumption. To test this hypothesis, we analyzed the expression of catalase, GPx-1 and -2, and Prdx-I, -II and -IV by Western blotting. Our results showed no differences in expression of these proteins (Supplementary Fig. 4). Differences in H2O2 consumption rates of HUVEC in the different ECMs could still be due to alterations in expression of other H2O2 scavengers or alterations in enzyme activity due to post-translational modifications. We, therefore, used in-gel activity assays to assess total catalase and GPx activity in HUVEC cultured in gelatin or laminin. Catalase activity was similar between the cells cultured in the two substrates (Fig. 3A). However, GPx activity of endothelial cells cultured in the presence of laminin was 56% lower than when cultured in the presence of gelatin (Fig. 3B). This is in the same proportion as the variation of the corresponding consumption rates (44%, Fig. 1), strongly suggesting that the alteration in GPx activity was the main responsible for the differences observed in the consumption rates of endothelial cells when cultured in gelatin or laminin.


The extracellular matrix modulates H2O2 degradation and redox signaling in endothelial cells.

Bagulho A, Vilas-Boas F, Pena A, Peneda C, Santos FC, Jerónimo A, de Almeida RF, Real C - Redox Biol (2015)

Catalase and GPx activity in HUVEC plated on different ECMs. In-gel assays to assess the activity of catalase and GPx (band intensity) in HUVEC cultured in gelatin or laminin showed that (A) catalase activity was similar between cells cultured in the two ECMs, whereas (B) GPx activity in cells cultured in the presence of laminin was lower than when cultured in the presence of gelatin (a.u., arbitrary units; values show the mean and SEM; N=5; two-tailed Student's t test, **p=0.005).
© Copyright Policy - CC BY-NC-ND
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4588420&req=5

f0015: Catalase and GPx activity in HUVEC plated on different ECMs. In-gel assays to assess the activity of catalase and GPx (band intensity) in HUVEC cultured in gelatin or laminin showed that (A) catalase activity was similar between cells cultured in the two ECMs, whereas (B) GPx activity in cells cultured in the presence of laminin was lower than when cultured in the presence of gelatin (a.u., arbitrary units; values show the mean and SEM; N=5; two-tailed Student's t test, **p=0.005).
Mentions: Our fluorescence spectroscopy results showed that endothelial cells presented similar membrane biophysical properties in the presence of different ECMs, with the observed different hydrogen peroxide consumptions probably not caused by differences in the rate of passive permeation. An alternative explanation for the variations observed in the H2O2 consumption rates of HUVEC in the different ECMs is the alteration of activity of enzymes responsible for H2O2 consumption. To test this hypothesis, we analyzed the expression of catalase, GPx-1 and -2, and Prdx-I, -II and -IV by Western blotting. Our results showed no differences in expression of these proteins (Supplementary Fig. 4). Differences in H2O2 consumption rates of HUVEC in the different ECMs could still be due to alterations in expression of other H2O2 scavengers or alterations in enzyme activity due to post-translational modifications. We, therefore, used in-gel activity assays to assess total catalase and GPx activity in HUVEC cultured in gelatin or laminin. Catalase activity was similar between the cells cultured in the two substrates (Fig. 3A). However, GPx activity of endothelial cells cultured in the presence of laminin was 56% lower than when cultured in the presence of gelatin (Fig. 3B). This is in the same proportion as the variation of the corresponding consumption rates (44%, Fig. 1), strongly suggesting that the alteration in GPx activity was the main responsible for the differences observed in the consumption rates of endothelial cells when cultured in gelatin or laminin.

Bottom Line: Instead, we found that the ECM regulated GPx activity, a known H2O2 scavenger.Thus, our results unraveled a new mechanism by which the ECM regulates endothelial cell function by altering redox balance.These results pinpoint the ECM as an important component of redox-signaling.

View Article: PubMed Central - PubMed

Affiliation: Centro de Química e Bioquímica, Faculdade de Ciências, Universidade de Lisboa, 1749-016 Lisboa, Portugal. Electronic address: anabagulho@gmail.com.

No MeSH data available.