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Dexamethasone Suppressed LPS-Induced Matrix Metalloproteinase and Its Effect on Endothelial Glycocalyx Shedding.

Cui N, Wang H, Long Y, Su L, Liu D - Mediators Inflamm. (2015)

Bottom Line: The aim of this study is to determine the mechanism of sepsis-induced vascular hyperpermeability and the beneficial effect of glucocorticoid in protecting vascular endothelium.The inhibition of MMPs by dexamethasone was significantly lower than that by doxycycline, while the rescue of syndecan-1 expression from LPS-induced endotoxemic rat thoracic aorta was significantly higher in the dexamethasone-treated compared to the doxycycline-treated (p = 0.03).In conclusion, activation of MMPs plays important role in regulating ZO-1 and syndecan-1 protein levels in LPS mediated endothelial perturbation.

View Article: PubMed Central - PubMed

Affiliation: Department of Critical Care Medicine, Peking Union Medical College Hospital, Beijing 100730, China.

ABSTRACT
The aim of this study is to determine the mechanism of sepsis-induced vascular hyperpermeability and the beneficial effect of glucocorticoid in protecting vascular endothelium. Male Sprague-Dawley rats were given either a bolus intraperitoneal injection of a nonlethal dose of LPS (Escherichia coli 055:B5, 10 mg/kg, Sigma) or vehicle (pyrogen-free water). Animals of treatment groups were also given either dexamethasone (4 mg/kg, 30 min prior to LPS injection) or the matrix metalloproteinases (MMPs) inhibitor doxycycline (4 mg/kg, 30 min after LPS injection). Both activities and protein levels of MMP-2 (p < 0.001) and MMP-9 (p < 0.001) were significantly upregulated in aortic homogenates from LPS-treated rats, associated with decreased ZO-1 (p < 0.001) and syndecan-1 (p = 0.011) protein contents. Both dexamethasone and doxycycline could significantly inhibit MMPs activity and reserve the expressions of ZO-1 and syndecan-1. The inhibition of MMPs by dexamethasone was significantly lower than that by doxycycline, while the rescue of syndecan-1 expression from LPS-induced endotoxemic rat thoracic aorta was significantly higher in the dexamethasone-treated compared to the doxycycline-treated (p = 0.03). In conclusion, activation of MMPs plays important role in regulating ZO-1 and syndecan-1 protein levels in LPS mediated endothelial perturbation. Both dexamethasone and doxycycline inhibit activation of MMPs that may contribute to the rescue of ZO-1 and syndecan-1 expression.

No MeSH data available.


Related in: MedlinePlus

Immunoblotting of MMP-2 and MMP-9. Proteins were extracted from the aortic tissues of indicated treatment groups and immunoblotted for MMP-2 and MMP-9 as described in the method. (a) Immunoblotting image of MMP-2 and MMP-9 among different groups. GAPDH was used as loading control. Latent MMP-2 (72 kDa) and MMP-9 (92-kDa) were detected as indicated. (b and c) Densitometric analysis of 8 biological replicates of MMP-2 and MMP-9, respectively. ∗p < 0.05; ∗∗p < 0.001.
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fig3: Immunoblotting of MMP-2 and MMP-9. Proteins were extracted from the aortic tissues of indicated treatment groups and immunoblotted for MMP-2 and MMP-9 as described in the method. (a) Immunoblotting image of MMP-2 and MMP-9 among different groups. GAPDH was used as loading control. Latent MMP-2 (72 kDa) and MMP-9 (92-kDa) were detected as indicated. (b and c) Densitometric analysis of 8 biological replicates of MMP-2 and MMP-9, respectively. ∗p < 0.05; ∗∗p < 0.001.

Mentions: In order to determine the effect of dexamethasone or doxycycline on MMP expression and activation, a gelatin zymogram was performed. To accomplish this, equal amount of protein from each group aortic tissue homogenate was loaded in a 10% PAGE-SDS gel with gelatin as substrate. As shown in Figure 2, under “Control” condition or in the presence of dexamethasone alone, neither MMP-2 nor MMP-9 was detected by gelatin zymogram. In contrast, both MMP-2 and MMP-9 were significantly induced by LPS. Interestingly, however, only MMP-9 but not MMP-2 was activated by LPS in the current study (Figure 2(a)). In the presence of dexamethasone, LPS-induced MMP-9 (both latent and active) and MMP-2 (latent form) were significantly suppressed (Figures 2(b), 2(c), and 2(d), p < 0.05). Similarly, doxycycline not only almost completely blocked latent MMP-2 and MMP-9 production, but also significantly inhibited MMP-9 conversion from latent to active form (Figures 2(b), 2(c), and 2(d), p < 0.05). Consistent with results of gelatin zymogram, immunoblotting of MMP-2 and MMP-9 confirmed that both dexamethasone and doxycycline could significantly suppress LPS-induced MMP-2 or MMP-9 expression in the aortic tissues (Figures 3(a) and 3(b)).


Dexamethasone Suppressed LPS-Induced Matrix Metalloproteinase and Its Effect on Endothelial Glycocalyx Shedding.

Cui N, Wang H, Long Y, Su L, Liu D - Mediators Inflamm. (2015)

Immunoblotting of MMP-2 and MMP-9. Proteins were extracted from the aortic tissues of indicated treatment groups and immunoblotted for MMP-2 and MMP-9 as described in the method. (a) Immunoblotting image of MMP-2 and MMP-9 among different groups. GAPDH was used as loading control. Latent MMP-2 (72 kDa) and MMP-9 (92-kDa) were detected as indicated. (b and c) Densitometric analysis of 8 biological replicates of MMP-2 and MMP-9, respectively. ∗p < 0.05; ∗∗p < 0.001.
© Copyright Policy - open-access
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4493300&req=5

fig3: Immunoblotting of MMP-2 and MMP-9. Proteins were extracted from the aortic tissues of indicated treatment groups and immunoblotted for MMP-2 and MMP-9 as described in the method. (a) Immunoblotting image of MMP-2 and MMP-9 among different groups. GAPDH was used as loading control. Latent MMP-2 (72 kDa) and MMP-9 (92-kDa) were detected as indicated. (b and c) Densitometric analysis of 8 biological replicates of MMP-2 and MMP-9, respectively. ∗p < 0.05; ∗∗p < 0.001.
Mentions: In order to determine the effect of dexamethasone or doxycycline on MMP expression and activation, a gelatin zymogram was performed. To accomplish this, equal amount of protein from each group aortic tissue homogenate was loaded in a 10% PAGE-SDS gel with gelatin as substrate. As shown in Figure 2, under “Control” condition or in the presence of dexamethasone alone, neither MMP-2 nor MMP-9 was detected by gelatin zymogram. In contrast, both MMP-2 and MMP-9 were significantly induced by LPS. Interestingly, however, only MMP-9 but not MMP-2 was activated by LPS in the current study (Figure 2(a)). In the presence of dexamethasone, LPS-induced MMP-9 (both latent and active) and MMP-2 (latent form) were significantly suppressed (Figures 2(b), 2(c), and 2(d), p < 0.05). Similarly, doxycycline not only almost completely blocked latent MMP-2 and MMP-9 production, but also significantly inhibited MMP-9 conversion from latent to active form (Figures 2(b), 2(c), and 2(d), p < 0.05). Consistent with results of gelatin zymogram, immunoblotting of MMP-2 and MMP-9 confirmed that both dexamethasone and doxycycline could significantly suppress LPS-induced MMP-2 or MMP-9 expression in the aortic tissues (Figures 3(a) and 3(b)).

Bottom Line: The aim of this study is to determine the mechanism of sepsis-induced vascular hyperpermeability and the beneficial effect of glucocorticoid in protecting vascular endothelium.The inhibition of MMPs by dexamethasone was significantly lower than that by doxycycline, while the rescue of syndecan-1 expression from LPS-induced endotoxemic rat thoracic aorta was significantly higher in the dexamethasone-treated compared to the doxycycline-treated (p = 0.03).In conclusion, activation of MMPs plays important role in regulating ZO-1 and syndecan-1 protein levels in LPS mediated endothelial perturbation.

View Article: PubMed Central - PubMed

Affiliation: Department of Critical Care Medicine, Peking Union Medical College Hospital, Beijing 100730, China.

ABSTRACT
The aim of this study is to determine the mechanism of sepsis-induced vascular hyperpermeability and the beneficial effect of glucocorticoid in protecting vascular endothelium. Male Sprague-Dawley rats were given either a bolus intraperitoneal injection of a nonlethal dose of LPS (Escherichia coli 055:B5, 10 mg/kg, Sigma) or vehicle (pyrogen-free water). Animals of treatment groups were also given either dexamethasone (4 mg/kg, 30 min prior to LPS injection) or the matrix metalloproteinases (MMPs) inhibitor doxycycline (4 mg/kg, 30 min after LPS injection). Both activities and protein levels of MMP-2 (p < 0.001) and MMP-9 (p < 0.001) were significantly upregulated in aortic homogenates from LPS-treated rats, associated with decreased ZO-1 (p < 0.001) and syndecan-1 (p = 0.011) protein contents. Both dexamethasone and doxycycline could significantly inhibit MMPs activity and reserve the expressions of ZO-1 and syndecan-1. The inhibition of MMPs by dexamethasone was significantly lower than that by doxycycline, while the rescue of syndecan-1 expression from LPS-induced endotoxemic rat thoracic aorta was significantly higher in the dexamethasone-treated compared to the doxycycline-treated (p = 0.03). In conclusion, activation of MMPs plays important role in regulating ZO-1 and syndecan-1 protein levels in LPS mediated endothelial perturbation. Both dexamethasone and doxycycline inhibit activation of MMPs that may contribute to the rescue of ZO-1 and syndecan-1 expression.

No MeSH data available.


Related in: MedlinePlus