Secretoneurin increases monolayer permeability in human coronary artery endothelial cells.
BACKGROUND
Secretoneurin (SN), a novel neuropeptide, may play a role in inflammation in the vascular system. However, the interaction between SN and endothelial cells is largely unknown. This study's objective is to investigate the effects of SN on endothelial permeability and its associated molecular mechanisms in human coronary artery endothelial cells (HCAECs).


METHODS
HCAECs were treated with SN. Monolayer permeability was assayed with a transwell system and a Texas Red-labeled dextran tracer. The mRNA and protein levels of endothelial junctional molecules were determined by real-time reverse transcriptase polymerase chain reaction (RT-PCR) and Western blot analysis, respectively. Superoxide anion was determined by fluorescent dye dihydroethidium-based flow cytometry detection. Mitogen-activated protein kinase (MAPK) activation was determined by Bio-Plex immunoassay.


RESULTS
HCAECs were treated with SN (15, 30, and 60 ng/ml) for 24 hours and showed a significant increase of monolayer permeability by 12%, 33%, and 47%, respectively, compared with controls (P < 0.05). SN-treated cells showed a significant reduction of zonula occludens-1 (ZO-1) and occludin at both mRNA and protein levels (P < 0.05). In addition, SN significantly increased superoxide anion in HCAECs (P < 0.05). Furthermore, SN activated MAPKs (JNK and ERK1/2) but not p38. Both antioxidant seleno-L-methionine (SeMet) and specific inhibitors of JNK and ERK1/2 effectively blocked SN-induced monolayer permeability increase in HCAECs.


CONCLUSIONS
SN significantly increases monolayer permeability and reduces the expression of ZO-1 and occludin through oxidative stress and the activation of JNK and ERK1/2 in HCAECs. This study provides direct evidence that SN impairs endothelial barrier function.
Yan
Shaoyu
S
Molecular Surgeon Research Center, Division of Vascular Surgery and Endovascular Therapy, Michael E. DeBakey Department of Surgery, Baylor College of Medicine, Houston, TX 77030, USA.
Wang
Xinwen
X
Chai
Hong
H
Wang
Hao
H
Yao
Qizhi
Q
Chen
Changyi
C
D002199:Capillary Permeability / Q000187:drug effects*; D018929:Cell Culture Techniques; D003331:Coronary Vessels / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D003911:Dextrans / Q000493:pharmacokinetics; D042783:Endothelial Cells / Q000187:drug effects* / Q000378:metabolism*; D006801:Humans; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D009479:Neuropeptides / Q000494:pharmacology*; D010952:Plasma Substitutes / Q000493:pharmacokinetics; D012333:RNA, Messenger / Q000378:metabolism; D053381:Secretogranin II; D013481:Superoxides / Q000378:metabolism
D003911:Dextrans; D008565:Membrane Proteins; D009479:Neuropeptides; D010952:Plasma Substitutes; D012333:RNA, Messenger; D053381:Secretogranin II; D013481:Superoxides; C081159:secretoneurin; D020928:Mitogen-Activated Protein Kinases