Heparin and LPS-induced COX-2 expression in airway cells: a link between its anti-inflammatory effects and GAG sulfation.
OBJECTIVE
Previous studies have indicated that the sulfated polysaccharide heparin has anti-inflammatory effects. However, the mechanistic basis for these effects has not been fully elucidated.


METHODS
NCI-H292 (mucoepidermoid) and HBE-1 (normal) human bronchial epithelial cells were treated with LPS alone or in the presence of high-molecular-weight (HMW) fully sulfated heparin or desulfated HMW heparin. Cells were harvested to examine the phosphorylation levels of ERK1/2, p38, and NF-kB p65 and COX-2 protein expression by Western blot and gene expression of both COX-2 and CXCL-8 by TaqMan qRT-PCR.


RESULTS
Heparin is known to exert an influence on receptor-mediated signaling through its ability to both potentiate and inhibit the receptor-ligand interaction, depending upon its concentration. In H292 cells, fully-sulfated HMW heparin significantly reduced LPS-induced gene expression of both COX-2 and CXCL-8 for up to 48 hours, while desulfated heparin had little to no significant suppressive effect on signaling or on COX-2 gene or protein expression. Desulfated heparin, initially ineffective at preventing LPS-induced CXCL8 up-regulation, reduced CXCL8 transcription at 24 hours. In contrast, in normal HBE-1 cells, fully sulfated heparin significantly suppressed only ERK signaling, COX-2 gene expression at 12 hours, and CXCL-8 gene expression at 6 and 12 hours, while desulfated heparin had no significant effects on LPS-stimulated signaling or on gene or protein expression. Sulfation determines heparin's influence and may reflect the moderating role of GAG sulfation in lung injury and health.


CONCLUSIONS
Heparin's anti-inflammatory effects result from its nonspecific suppression of signaling and gene expression and are determined by its sulfation.
a Department of Molecular Biomedical Sciences, College of Veterinary Medicine, North Carolina State University , Raleigh , North Carolina , USA.
Na Young
NY
Yi
a Department of Molecular Biomedical Sciences, College of Veterinary Medicine, North Carolina State University , Raleigh , North Carolina , USA.
Donna R
DR
Newman
a Department of Molecular Biomedical Sciences, College of Veterinary Medicine, North Carolina State University , Raleigh , North Carolina , USA.
Huiying
H
Zhang
a Department of Molecular Biomedical Sciences, College of Veterinary Medicine, North Carolina State University , Raleigh , North Carolina , USA.
Helena
H
Morales Johansson
a Department of Molecular Biomedical Sciences, College of Veterinary Medicine, North Carolina State University , Raleigh , North Carolina , USA.
Philip L
PL
Sannes
D000894:Anti-Inflammatory Agents, Non-Steroidal / Q000737:chemistry / Q000494:pharmacology; D002460:Cell Line; D051546:Cyclooxygenase 2 / Q000235:genetics* / Q000378:metabolism*; D015870:Gene Expression / Q000187:drug effects; D006025:Glycosaminoglycans / Q000737:chemistry / Q000378:metabolism; D006493:Heparin / Q000737:chemistry / Q000494:pharmacology*; D006801:Humans; D016209:Interleukin-8 / Q000235:genetics / Q000378:metabolism; D008070:Lipopolysaccharides / Q000633:toxicity*; D020935:MAP Kinase Signaling System / Q000187:drug effects; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D020545:Respiratory Mucosa / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism*
D000894:Anti-Inflammatory Agents, Non-Steroidal; C508606:CXCL8 protein, human; D006025:Glycosaminoglycans; D016209:Interleukin-8; D008070:Lipopolysaccharides; D012333:RNA, Messenger; C013786:A73025; D006493:Heparin; D051546:Cyclooxygenase 2; C496540:PTGS2 protein, human
COX-2; LPS; heparin; lung; sulfation