Altered Nrf2 signaling mediates hypoglycemia-induced blood-brain barrier endothelial dysfunction in vitro.
Hypoglycemia impairs blood-brain barrier (BBB) endothelial function; a major hallmark in the pathogenesis of various CNS disorders. Previously, we have demonstrated that prolonged hypoglycemic exposure down-regulated BBB endothelial NF-E2 related factor-2 (Nrf2) expression; a redox-sensitive transcr...
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| Format: | Article |
| Language: | English |
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Public Library of Science (PLoS)
2015-01-01
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| Series: | PLoS ONE |
| Online Access: | http://europepmc.org/articles/PMC4373930?pdf=render |
| _version_ | 1818290593870118912 |
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| author | Ravi K Sajja Kayla N Green Luca Cucullo |
| author_facet | Ravi K Sajja Kayla N Green Luca Cucullo |
| author_sort | Ravi K Sajja |
| collection | DOAJ |
| description | Hypoglycemia impairs blood-brain barrier (BBB) endothelial function; a major hallmark in the pathogenesis of various CNS disorders. Previously, we have demonstrated that prolonged hypoglycemic exposure down-regulated BBB endothelial NF-E2 related factor-2 (Nrf2) expression; a redox-sensitive transcriptional factor that regulates endothelial function. Here, we sought to determine the functional role of Nrf2 in preserving BBB integrity and molecular mechanisms underlying hypoglycemia-induced Nrf2 down-regulation in vitro using human cerebral microvascular endothelial cell line (hCMEC/D3). Cell monolayers were exposed to normal or hypoglycemic (5.5 or 2.2mM D-glucose) media for 3-24h. Pharmacological or gene manipulation (by silencing RNA) approaches were used to investigate specific molecular pathways implicated in hypoglycemia-induced Nrf2 degradation. BBB integrity was assessed by paracellular permeability to labeled dextrans of increasing molecular sizes (4-70kDa). Silencing Nrf2 expression in hCMEC/D3 cells abrogated the expression of claudin-5 and VE-cadherin, while ZO-1 was up-regulated. These effects were paralleled by a decrease in electrical resistance of hCMEC/D3 monolayers and potential increase in permeability to all labeled dextrans. Hypoglycemic exposure (3-24h) led to progressive and sustained down-regulation of Nrf2 (without affecting mRNA) and its target, NQO-1, with a concomitant increase in the cytosolic pool of E3 ubiquitin ligase, Siah2 (but not Keap1). Pretreatment with protease inhibitor MG132, or selective knock-down of Siah2 (but not Keap1) significantly attenuated hypoglycemia-induced Nrf2 destabilization. While hypoglycemic exposure triggered a significant increase in BBB permeability to dextrans, silencing Siah2 gene abrogated the effects of hypoglycemia and restored BBB integrity. In summary, our data indicate a potential role for Nrf2 signaling in regulating tight junction integrity and maintaining BBB function. Nrf2 suppression by increased Siah2-driven proteasomal degradation mediates hypoglycemia-evoked endothelial dysfunction and loss of BBB integrity. Overall, this study suggests that sustained activation of endothelial Nrf2 signaling could have therapeutic potential to prevent hypoglycemia-induced cerebrovascular dysfunction. |
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| institution | Directory Open Access Journal |
| issn | 1932-6203 |
| language | English |
| last_indexed | 2024-12-13T02:30:39Z |
| publishDate | 2015-01-01 |
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| spelling | doaj.art-bd6c1c2f82ad402487cbdced0808ff2b2022-12-22T00:02:31ZengPublic Library of Science (PLoS)PLoS ONE1932-62032015-01-01103e012235810.1371/journal.pone.0122358Altered Nrf2 signaling mediates hypoglycemia-induced blood-brain barrier endothelial dysfunction in vitro.Ravi K SajjaKayla N GreenLuca CuculloHypoglycemia impairs blood-brain barrier (BBB) endothelial function; a major hallmark in the pathogenesis of various CNS disorders. Previously, we have demonstrated that prolonged hypoglycemic exposure down-regulated BBB endothelial NF-E2 related factor-2 (Nrf2) expression; a redox-sensitive transcriptional factor that regulates endothelial function. Here, we sought to determine the functional role of Nrf2 in preserving BBB integrity and molecular mechanisms underlying hypoglycemia-induced Nrf2 down-regulation in vitro using human cerebral microvascular endothelial cell line (hCMEC/D3). Cell monolayers were exposed to normal or hypoglycemic (5.5 or 2.2mM D-glucose) media for 3-24h. Pharmacological or gene manipulation (by silencing RNA) approaches were used to investigate specific molecular pathways implicated in hypoglycemia-induced Nrf2 degradation. BBB integrity was assessed by paracellular permeability to labeled dextrans of increasing molecular sizes (4-70kDa). Silencing Nrf2 expression in hCMEC/D3 cells abrogated the expression of claudin-5 and VE-cadherin, while ZO-1 was up-regulated. These effects were paralleled by a decrease in electrical resistance of hCMEC/D3 monolayers and potential increase in permeability to all labeled dextrans. Hypoglycemic exposure (3-24h) led to progressive and sustained down-regulation of Nrf2 (without affecting mRNA) and its target, NQO-1, with a concomitant increase in the cytosolic pool of E3 ubiquitin ligase, Siah2 (but not Keap1). Pretreatment with protease inhibitor MG132, or selective knock-down of Siah2 (but not Keap1) significantly attenuated hypoglycemia-induced Nrf2 destabilization. While hypoglycemic exposure triggered a significant increase in BBB permeability to dextrans, silencing Siah2 gene abrogated the effects of hypoglycemia and restored BBB integrity. In summary, our data indicate a potential role for Nrf2 signaling in regulating tight junction integrity and maintaining BBB function. Nrf2 suppression by increased Siah2-driven proteasomal degradation mediates hypoglycemia-evoked endothelial dysfunction and loss of BBB integrity. Overall, this study suggests that sustained activation of endothelial Nrf2 signaling could have therapeutic potential to prevent hypoglycemia-induced cerebrovascular dysfunction.http://europepmc.org/articles/PMC4373930?pdf=render |
| spellingShingle | Ravi K Sajja Kayla N Green Luca Cucullo Altered Nrf2 signaling mediates hypoglycemia-induced blood-brain barrier endothelial dysfunction in vitro. PLoS ONE |
| title | Altered Nrf2 signaling mediates hypoglycemia-induced blood-brain barrier endothelial dysfunction in vitro. |
| title_full | Altered Nrf2 signaling mediates hypoglycemia-induced blood-brain barrier endothelial dysfunction in vitro. |
| title_fullStr | Altered Nrf2 signaling mediates hypoglycemia-induced blood-brain barrier endothelial dysfunction in vitro. |
| title_full_unstemmed | Altered Nrf2 signaling mediates hypoglycemia-induced blood-brain barrier endothelial dysfunction in vitro. |
| title_short | Altered Nrf2 signaling mediates hypoglycemia-induced blood-brain barrier endothelial dysfunction in vitro. |
| title_sort | altered nrf2 signaling mediates hypoglycemia induced blood brain barrier endothelial dysfunction in vitro |
| url | http://europepmc.org/articles/PMC4373930?pdf=render |
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