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Heme oxygenase-1 protects corexit 9500A-induced respiratory epithelial injury across species.

Li FJ, Duggal RN, Oliva OM, Karki S, Surolia R, Wang Z, Watson RD, Thannickal VJ, Powell M, Watts S, Kulkarni T, Batra H, Bolisetty S, Agarwal A, Antony VB - PLoS ONE (2015)

Bottom Line: CE induced the expression of HO-1 as well as C-reactive protein (CRP) and NADPH oxidase 4 (NOX4), which are associated with ROS production.Treatment with carbon monoxide releasing molecule-2 (CORM-2) significantly inhibited CE-induced ROS production, while the addition of HO-1 inhibitor, significantly increased CE-induced ROS production and apoptosis, suggesting a protective role of HO-1 or its reaction product, CO, in CE-induced apoptosis.Using HO-1 knockout mice, we further demonstrated that HO-1 protected against CE-induced inflammation and cellular apoptosis and corrected CE-mediated inhibition of E-cadherin and FAK.

View Article: PubMed Central - PubMed

Affiliation: Division of Pulmonary, Allergy, and Critical Care Medicine, Department of Medicine, University of Alabama at Birmingham, Birmingham, AL, United States of America.

ABSTRACT
The effects of Corexit 9500A (CE) on respiratory epithelial surfaces of terrestrial mammals and marine animals are largely unknown. This study investigated the role of CE-induced heme oxygenase-1 (HO-1), a cytoprotective enzyme with anti-apoptotic and antioxidant activity, in human bronchial airway epithelium and the gills of exposed aquatic animals. We evaluated CE-mediated alterations in human airway epithelial cells, mice lungs and gills from zebrafish and blue crabs. Our results demonstrated that CE induced an increase in gill epithelial edema and human epithelial monolayer permeability, suggesting an acute injury caused by CE exposure. CE induced the expression of HO-1 as well as C-reactive protein (CRP) and NADPH oxidase 4 (NOX4), which are associated with ROS production. Importantly, CE induced caspase-3 activation and subsequent apoptosis of epithelial cells. The expression of the intercellular junctional proteins, such as tight junction proteins occludin, zonula occludens (ZO-1), ZO-2 and adherens junctional proteins E-cadherin and Focal Adhesion Kinase (FAK), were remarkably inhibited by CE, suggesting that these proteins are involved in CE-induced increased permeability and subsequent apoptosis. The cytoskeletal protein F-actin was also disrupted by CE. Treatment with carbon monoxide releasing molecule-2 (CORM-2) significantly inhibited CE-induced ROS production, while the addition of HO-1 inhibitor, significantly increased CE-induced ROS production and apoptosis, suggesting a protective role of HO-1 or its reaction product, CO, in CE-induced apoptosis. Using HO-1 knockout mice, we further demonstrated that HO-1 protected against CE-induced inflammation and cellular apoptosis and corrected CE-mediated inhibition of E-cadherin and FAK. These observations suggest that CE activates CRP and NOX4-mediated ROS production, alters permeability by inhibition of junctional proteins, and leads to caspase-3 dependent apoptosis of epithelial cells, while HO-1 and its reaction products protect against oxidative stress and apoptosis.

No MeSH data available.


Related in: MedlinePlus

HO-1 stabilizes the adhesion proteins E-cadherin and FAK.(A) HO-1 WT and HO-KO mice were exposed to either 20 μl of CE or held as controls for 24 hours. Cell lysates were prepared from lung epithelial cells and analyzed by western blotting with anti-E-cadherin and FAK antibodies. (B) The densities of protein bands were determined by densitometry and the data represent a one-fold increase from the control density. Data are shown as a mean ± SD from three different experiments. * p < 0.05, ** p < 0.01 vs HO-1 WT (-) CE; ## p < 0.01 vs HO-1 KO (-) CE; && p < 0.01 vs HO-1 WT (+) CE by a one-way ANOVA with HSD test.
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pone.0122275.g008: HO-1 stabilizes the adhesion proteins E-cadherin and FAK.(A) HO-1 WT and HO-KO mice were exposed to either 20 μl of CE or held as controls for 24 hours. Cell lysates were prepared from lung epithelial cells and analyzed by western blotting with anti-E-cadherin and FAK antibodies. (B) The densities of protein bands were determined by densitometry and the data represent a one-fold increase from the control density. Data are shown as a mean ± SD from three different experiments. * p < 0.05, ** p < 0.01 vs HO-1 WT (-) CE; ## p < 0.01 vs HO-1 KO (-) CE; && p < 0.01 vs HO-1 WT (+) CE by a one-way ANOVA with HSD test.

Mentions: To further investigate the role of HO-1 in cellular protection, we evaluated the effect of CE exposure in vivo in HO-1 KO mice. CE-induced acute lung injury was associated with increased inflammation and alveolar-capillary permeability, as measured by BAL cell counts and protein content, respectively. Compared to HO-1 WT mice, HO-1 KO mice had significantly more alterations in permeability, and higher number of lymphocytes, neutrophils and eosinophils as well as more protein in their BAL fluid in the presence of CE, which is consistent with an increase in lung inflammation and permeability (Fig. 7 A, B and C). More accumulation of TUNEL-positive cells was found in HO-1 KO mice when compared with WT mice on exposure to CE (Fig. 7D). We also found that epithelial cells isolated from HO-1 KO mice were more susceptible to CE-induced destruction of E-cadherin and FAK (Fig. 8). These data suggest that HO-1 has functional significance and that HO-1 may contribute to protection against the increased membrane permeability, disruption of intercellular junctions and the induction of ROS and apoptosis.


Heme oxygenase-1 protects corexit 9500A-induced respiratory epithelial injury across species.

Li FJ, Duggal RN, Oliva OM, Karki S, Surolia R, Wang Z, Watson RD, Thannickal VJ, Powell M, Watts S, Kulkarni T, Batra H, Bolisetty S, Agarwal A, Antony VB - PLoS ONE (2015)

HO-1 stabilizes the adhesion proteins E-cadherin and FAK.(A) HO-1 WT and HO-KO mice were exposed to either 20 μl of CE or held as controls for 24 hours. Cell lysates were prepared from lung epithelial cells and analyzed by western blotting with anti-E-cadherin and FAK antibodies. (B) The densities of protein bands were determined by densitometry and the data represent a one-fold increase from the control density. Data are shown as a mean ± SD from three different experiments. * p < 0.05, ** p < 0.01 vs HO-1 WT (-) CE; ## p < 0.01 vs HO-1 KO (-) CE; && p < 0.01 vs HO-1 WT (+) CE by a one-way ANOVA with HSD test.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4383564&req=5

pone.0122275.g008: HO-1 stabilizes the adhesion proteins E-cadherin and FAK.(A) HO-1 WT and HO-KO mice were exposed to either 20 μl of CE or held as controls for 24 hours. Cell lysates were prepared from lung epithelial cells and analyzed by western blotting with anti-E-cadherin and FAK antibodies. (B) The densities of protein bands were determined by densitometry and the data represent a one-fold increase from the control density. Data are shown as a mean ± SD from three different experiments. * p < 0.05, ** p < 0.01 vs HO-1 WT (-) CE; ## p < 0.01 vs HO-1 KO (-) CE; && p < 0.01 vs HO-1 WT (+) CE by a one-way ANOVA with HSD test.
Mentions: To further investigate the role of HO-1 in cellular protection, we evaluated the effect of CE exposure in vivo in HO-1 KO mice. CE-induced acute lung injury was associated with increased inflammation and alveolar-capillary permeability, as measured by BAL cell counts and protein content, respectively. Compared to HO-1 WT mice, HO-1 KO mice had significantly more alterations in permeability, and higher number of lymphocytes, neutrophils and eosinophils as well as more protein in their BAL fluid in the presence of CE, which is consistent with an increase in lung inflammation and permeability (Fig. 7 A, B and C). More accumulation of TUNEL-positive cells was found in HO-1 KO mice when compared with WT mice on exposure to CE (Fig. 7D). We also found that epithelial cells isolated from HO-1 KO mice were more susceptible to CE-induced destruction of E-cadherin and FAK (Fig. 8). These data suggest that HO-1 has functional significance and that HO-1 may contribute to protection against the increased membrane permeability, disruption of intercellular junctions and the induction of ROS and apoptosis.

Bottom Line: CE induced the expression of HO-1 as well as C-reactive protein (CRP) and NADPH oxidase 4 (NOX4), which are associated with ROS production.Treatment with carbon monoxide releasing molecule-2 (CORM-2) significantly inhibited CE-induced ROS production, while the addition of HO-1 inhibitor, significantly increased CE-induced ROS production and apoptosis, suggesting a protective role of HO-1 or its reaction product, CO, in CE-induced apoptosis.Using HO-1 knockout mice, we further demonstrated that HO-1 protected against CE-induced inflammation and cellular apoptosis and corrected CE-mediated inhibition of E-cadherin and FAK.

View Article: PubMed Central - PubMed

Affiliation: Division of Pulmonary, Allergy, and Critical Care Medicine, Department of Medicine, University of Alabama at Birmingham, Birmingham, AL, United States of America.

ABSTRACT
The effects of Corexit 9500A (CE) on respiratory epithelial surfaces of terrestrial mammals and marine animals are largely unknown. This study investigated the role of CE-induced heme oxygenase-1 (HO-1), a cytoprotective enzyme with anti-apoptotic and antioxidant activity, in human bronchial airway epithelium and the gills of exposed aquatic animals. We evaluated CE-mediated alterations in human airway epithelial cells, mice lungs and gills from zebrafish and blue crabs. Our results demonstrated that CE induced an increase in gill epithelial edema and human epithelial monolayer permeability, suggesting an acute injury caused by CE exposure. CE induced the expression of HO-1 as well as C-reactive protein (CRP) and NADPH oxidase 4 (NOX4), which are associated with ROS production. Importantly, CE induced caspase-3 activation and subsequent apoptosis of epithelial cells. The expression of the intercellular junctional proteins, such as tight junction proteins occludin, zonula occludens (ZO-1), ZO-2 and adherens junctional proteins E-cadherin and Focal Adhesion Kinase (FAK), were remarkably inhibited by CE, suggesting that these proteins are involved in CE-induced increased permeability and subsequent apoptosis. The cytoskeletal protein F-actin was also disrupted by CE. Treatment with carbon monoxide releasing molecule-2 (CORM-2) significantly inhibited CE-induced ROS production, while the addition of HO-1 inhibitor, significantly increased CE-induced ROS production and apoptosis, suggesting a protective role of HO-1 or its reaction product, CO, in CE-induced apoptosis. Using HO-1 knockout mice, we further demonstrated that HO-1 protected against CE-induced inflammation and cellular apoptosis and corrected CE-mediated inhibition of E-cadherin and FAK. These observations suggest that CE activates CRP and NOX4-mediated ROS production, alters permeability by inhibition of junctional proteins, and leads to caspase-3 dependent apoptosis of epithelial cells, while HO-1 and its reaction products protect against oxidative stress and apoptosis.

No MeSH data available.


Related in: MedlinePlus