Atherogenic effects of Chlamydia pneumoniae:: Refuting the innocent bystander hypothesis

Objective: Serologic evidence of Chlamydia pneumoniae infection and atherosclerosis was first demonstrated in patients with ischemic heart disease in 1988. Subsequently, the organism has been detected in several cardiovascular lesions. Outside of observational reports, few studies mechanistically link vascular infection with C pneumoniae and atherogenesis. To better define its pathophysiologic role, we examined the influence of C pneumoniae infection of human vascular smooth muscle cells on vascular smooth muscle cell proliferation, cell-cycle protein expression, and inflammatory cytokine release. Methods: Human aortic vascular smooth muscle cells were inoculated with C. pneumoniae in culture.. Proliferation was assessed by mitochondrial activity, direct cell counting, and immunohistochemical staining for proliferating cell nuclear antigen. Electromobility gel shift assays probed for the antiproliferative cell-cycle protein p53. Supernatants were assayed for the mitogens interleukin-6 and interleukin-8 by enzyme-linked immunosorbent assay. Results: After C pneumoniae inoculation, vascular smooth muscle cell proliferation increased 2-fold by mitochondrial activity and more than 3-fold by cell numbers. C pneumoniae infection promoted a 3-fold increase in proliferating cell nuclear antigen expression, which was associated with decreased nuclear binding of p53. Compared with control, C. pneumoniae inoculation resulted in a 2.5-fold increase in released interleukin-6 and interleukin-8. In each experiment, the influence of C. pneumoniae was abrogated by concomitant treatment with the macrolide antibiotic azithromycin. Conclusions: C. pneumoniae induced human vascular smooth muscle cell proliferation and proliferating cell nuclear antigen expression, down-regulated p53, and promoted release of prototypical atherogenic cytokines. These in vitro findings indicate that C. pneumoniae is more than an innocent bystander, rather it is a pathophysiologic participant in atherogenesis warranting elimination.