Glycoprotein 130 ligand oncostatin-M induces expression of vascular endothelial growth factor in human adult cardiac myocytes.
OBJECTIVE
In murine and rat cardiac myocytes the gp130 system transduces survival as well as hypertrophic signals and via induction of the expression of the potent angiogenic factor VEGF in these cells also indirectly contributes to cardiac repair processes through the development of new blood vessels. There are, however, species differences in receptor specificity and receptor crossreactivity in the gp130-gp130 ligand system. We asked whether gp130 signaling is also involved in the regulation of VEGF in human cardiac myocytes and if so which gp130 ligands are critical for such an effect.


METHODS
Human adult cardiac myocytes (HACMs) were isolated from myocardial tissue and characterised by positive staining for myocardial actin, troponin-I and cardiotin. HACMs were treated with the gp130 ligands CT-1, IL-6, LIF or OSM and VEGF-1 was determined by a specific ELISA in the conditioned media of these cells. RT-PCR and Western blot analysis was used in order to detect gp130, IL-6-receptor, LIF-receptor or OSM-receptor specific protein and mRNA in human adult cardiac myocytes and for detection of VEGF-1 specific mRNA in cardiac myocytes after incubation with OSM. Pieces of myocardial tissue were incubated ex vivo in the presence and absence of OSM and VEGF was determined in supernatants of these cultures and immunohistochemistry was performed on the tissue using specific antibodies for VEGF-1. Immunohistochemistry was also employed to detect VEGF in sections from a healthy human heart and in a heart from a patient suffering from acute myocarditis.


RESULTS
OSM, but not CT-1, IL-6 or LIF increased VEGF-1 production in human adult cardiac myocytes dose-dependently derived from five different donors. This selective stimulation of VEGF by gp130 ligands was also reflected by a specific receptor expression on these cells. We detected high levels of mRNA for gp130 and the OSM receptor in freshly isolated human cardiac myocytes but only low amounts of mRNA for the IL-6 receptor whereas mRNA for the LIF receptor was hardly detectable by RT-PCR. OSM receptor and IL-6 receptor were also detectable by Western blotting whereas LIF receptor was only present as a faint band. OSM also increased the expression of VEGF-1 mRNA in cardiac myocytes. When pieces of human myocardial tissue were incubated with the gp130 ligands in an ex vivo model only OSM resulted in an increase in VEGF-1 in the supernatants of these cultures. Furthermore, VEGF increased in tissue samples treated with OSM in cardiac myocytes as evidenced by immunohistochemistry. In addition, we found increased VEGF-1 expression in myocardial tissue from a patient suffering from acute myocarditis.


CONCLUSIONS
The gp130-gp130 ligand system is also involved in VEGF regulation in human cardiac myocytes and OSM is the gp130 ligand responsible for this effect in the human system whereas LIF and CT-1 which had been shown to regulate VEGF expression in mouse and rat cardiac myocytes had no effect. Thus we have added OSM, which is produced by activated T lymphocytes and monocytes, to the list of regulatory molecules of VEGF production in the human heart. Our results lend further support to the notion that besides hypoxia, inflammation via induction of VEGF through autocrine or paracrine pathways plays a key role in (re)vascularisation of the myocardium.
D000328:Adult; D000704:Analysis of Variance; D015153:Blotting, Western / Q000379:methods; D002352:Carrier Proteins / Q000494:pharmacology; D002478:Cells, Cultured; D006023:Glycoproteins / Q000378:metabolism*; D006131:Growth Inhibitors / Q000378:metabolism / Q000494:pharmacology*; D006801:Humans; D007150:Immunohistochemistry / Q000379:methods; D015850:Interleukin-6 / Q000494:pharmacology; D053605:Leukemia Inhibitory Factor; D018832:Molecular Chaperones / Q000494:pharmacology; D009205:Myocarditis / Q000378:metabolism*; D032383:Myocytes, Cardiac / Q000187:drug effects / Q000378:metabolism*; D053683:Oncostatin M; D027701:Organic Cation Transport Proteins; D010455:Peptides / Q000378:metabolism / Q000494:pharmacology*; D011506:Proteins; D012333:RNA, Messenger / Q000032:analysis; D000074058:Solute Carrier Family 22 Member 5; D042461:Vascular Endothelial Growth Factor A / Q000032:analysis / Q000235:genetics*
D002352:Carrier Proteins; D006023:Glycoproteins; D006131:Growth Inhibitors; D015850:Interleukin-6; C507837:LIF protein, human; D053605:Leukemia Inhibitory Factor; C507838:Lif protein, mouse; D018832:Molecular Chaperones; C508351:OSM protein, human; D027701:Organic Cation Transport Proteins; C508352:Osm protein, mouse; D010455:Peptides; D011506:Proteins; D012333:RNA, Messenger; C492020:SLC22A5 protein, human; C492019:Slc22a5 protein, mouse; D000074058:Solute Carrier Family 22 Member 5; D042461:Vascular Endothelial Growth Factor A; C480482:glycoprotein 130, human; D053683:Oncostatin M