VEGF is upregulated by hypoxia-induced mitogenic factor via the PI-3K/Akt-NF-kappaB signaling pathway.
BACKGROUND
Hypoxia-induced mitogenic factor (HIMF) is developmentally regulated and plays an important role in lung pathogenesis. We initially found that HIMF promotes vascular tubule formation in a matrigel plug model. In this study, we investigated the mechanisms which HIMF enhances expression of vascular endothelial growth factor (VEGF) in lung tissues and epithelial cells.


METHODS
Recombinant HIMF protein was intratracheally instilled into adult mouse lungs, VEGF expression was examined by immunohistochemical staining and Western blot. The promoter-luciferase reporter assay, RT-PCR, and Western blot were performed to examine the effects of HIMF on VEGF expression in mouse lung epithelial cell line MLE-12. The activation of NF-kappa B (NF-kappaB) and phosphorylation of Akt, IKK and IkappaBalpha were examined by luciferase assay and Western blot, respectively.


RESULTS
Intratracheal instillation of HIMF protein resulted in significant increase of VEGF, mainly localized to airway epithelial and alveolar type II cells. Deletion of NF-kappaB binding sites within VEGF promoter abolished HIMF-induced VEGF expression in MLE-12 cells, suggesting that activation of NF-kappaB is essential for VEGF upregulation induced by HIMF. Stimulation of lung epithelial cells by HIMF resulted in phosphorylation of IKK and IkappaBalpha, leading to activation of NF-kappaB. In addition, HIMF strongly induced Akt phosphorylation, and suppression of Akt activation by specific inhibitors and dominant negative mutants for PI-3K, and IKK or IkappaBalpha blocked HIMF-induced NF-kappaB activation and attenuated HIMF-induced VEGF production.


CONCLUSIONS
These results suggest that HIMF enhances VEGF production in mouse lung epithelial cells in a PI-3K/Akt-NF-kappaB signaling pathway-dependent manner, and may play critical roles in pulmonary angiogenesis.
Tong
Qiangsong
Q
Department of Anesthesiology and Critical Care Medicine, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA. qtong@slu.edu
Zheng
Liduan
L
Lin
Li
L
Li
Bo
B
Wang
Danming
D
Huang
Chuanshu
C
Li
Dechun
D
D000818:Animals; D002460:Cell Line; D004305:Dose-Response Relationship, Drug; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism; D005786:Gene Expression Regulation; D036341:Intercellular Signaling Peptides and Proteins; D007442:Intubation, Intratracheal; D008168:Lung / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D016328:NF-kappa B / Q000378:metabolism; D020932:Nerve Growth Factor / Q000008:administration & dosage / Q000494:pharmacology; D019869:Phosphatidylinositol 3-Kinases / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation; D011401:Promoter Regions, Genetic; D011506:Proteins / Q000008:administration & dosage / Q000494:pharmacology*; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D011994:Recombinant Proteins / Q000008:administration & dosage / Q000494:pharmacology; D015398:Signal Transduction; D013997:Time Factors; D014162:Transfection; D015854:Up-Regulation; D042461:Vascular Endothelial Growth Factor A / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*
D036341:Intercellular Signaling Peptides and Proteins; D016328:NF-kappa B; D011506:Proteins; D012333:RNA, Messenger; D011994:Recombinant Proteins; C413576:Retnla protein, mouse; D042461:Vascular Endothelial Growth Factor A; C467480:vascular endothelial growth factor A, mouse; D020932:Nerve Growth Factor; D019869:Phosphatidylinositol 3-Kinases; D051057:Proto-Oncogene Proteins c-akt