W. Speidl

Polymorphic Membrane Protein (PMP) 20 and PMP 21 of Chlamydia pneumoniae Induce Proinflammatory Mediators in Human Endothelial Cells In Vitro by Activation of the Nuclear Factor-κB Pathway

We tested whether polymorphic membrane proteins (PMPs) of Chlamydia pneumoniae might play a role in triggering an inflammatory response in human endothelial cells. Of 15 purified, recombinant chlamydial PMPs tested, 2 (PMP 20 and PMP 21) dose-dependently increased the production of the inflammatory mediators interleukin (IL)-6 and monocyte chemoattractant protein-1 (MCP-1), in cultured human endothelial cells; production of IL-8 was also increased. When endothelial cells were infected by live C. pneumoniae, an increase in the production of IL-6, IL-8, and MCP-1 was seen. We used adenovirus-induced overexpression of IkappaBalpha-an inhibitor of nuclear factor (NF)-kappaB-to demonstrate that PMP 20 and PMP 21 increase the production of IL-6 and MCP-1 in human endothelial cells by activation of the NF-kappaB pathway, because, in cells overexpressing IkappaBalpha, treatment with the respective PMP did not result in increased production of IL-6 and MCP-1. Thus, C. pneumoniae could, by interactions of its PMPs with the endothelium, contribute to the process of vascular injury during the development and progression of atherosclerotic lesions.

Chlamydia pneumoniae in Carotid Artery Atherosclerosis A Comparison of Its Presence in Atherosclerotic Plaque, Healthy Vessels, and Circulating Leukocytes From the Same Individuals

Background and Purpose— There is growing clinical and experimental evidence that infections with Chlamydia pneumoniae might contribute to the development and progression of atherosclerosis. However, studies detecting the pathogen in atherosclerotic lesions examined either only atherosclerotic vessels or control vessels without atherosclerosis obtained from a different group of individuals. We analyzed atherosclerotic plaques of the carotid artery, samples of apparently healthy greater saphenous veins, and circulating leukocytes from the same individual patients for the presence of C pneumoniae. Methods— From each of 46 patients undergoing carotid endarterectomy for symptomatic carotid artery stenosis, these samples were analyzed by nested polymerase chain reaction for C pneumoniae–specific DNA. Furthermore, we determined IgA and IgG titers specific for the pathogen and plasma levels of C-reactive protein in these patients. Results— C pneumoniae DNA was detected in 86.9% of the leukocytes and in 82.6% of the atherosclerotic plaques but in only 6.5% of the saphenous veins. In 85% of patients who also had leukocytes positive for C pneumoniae, the atherosclerotic plaques were positive and the saphenous veins were negative. The presence of C pneumoniae–specific DNA in leukocytes significantly coincided with the presence of the respective DNA in the plaques of the carotid arteries (P =0.0002). No association between the presence of C pneumoniae and specific IgA or IgG levels was seen. C-reactive protein levels were significantly higher in patients with chlamydia-positive atherosclerotic plaques and with positive leukocytes than in patients with negative plaques of the carotid arteries or negative leukocytes, respectively (P <0.01, P <0.05). Conclusions— Our observation of >80% incidence of C pneumoniae in atherosclerotic plaques of the carotid artery does not prove causality between an infection with the pathogen and the development of atherosclerosis. It must be emphasized, however, that >90% of apparently healthy saphenous veins were negative for C pneumoniae. Given the structural and functional differences between veins and arteries, careful interpretation of our results regarding a possible causative role of C pneumoniae seems warranted.

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. CONCLUSION 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.