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Am J Physiol Heart Circ Physiol 287: H1115-H1124, 2004. First published May 13, 2004; doi:10.1152/ajpheart.00075.2004
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Evidence of nanobacterial-like structures in calcified human arteries and cardiac valves

Virginia M. Miller,1,2 George Rodgers,6 Jon A. Charlesworth,3 Brenda Kirkland,7 Sandra R. Severson,1 Todd E. Rasmussen,1 Marineh Yagubyan,1 Jeri C. Rodgers,7 Franklin R. Cockerill, III,4 Robert L. Folk,7 Ewa Rzewuska-Lech,1 Vivek Kumar,5 Gerard Farell-Baril,5 and John C. Lieske5

Departments of 1Surgery, 2Physiology and Bioengineering, 3Biochemistry and Molecular Biology, 4Clinical Microbiology, and 5Internal Medicine, Division of Nephrology, Mayo Clinic, Rochester, Minnesota 55905; and 6Heart Hospital of Austin and 7Department of Geological Sciences, The University Texas at Austin, Austin, Texas 78712

Submitted 28 January 2004 ; accepted in final form 11 May 2004

Mechanisms mediating vascular calcification remain incompletely understood. Nanometer scale objects hypothesized to be a type of bacteria (nanobacteria) are associated with calcified geological specimens, human kidney stones, and psammona bodies in ovarian cancer. Experiments were designed to evaluate human vascular tissue for the presence of similar nanometer-scale objects. Calcified human aneurysms (n = 8), carotid plaques (n = 2), femoral arterial plaques (n = 2), and cardiac valves (n = 2) and noncalcified aneurysms from patients with bicuspid aortic valve disease (n = 2) were collected as surgical waste from the Heart Hospital of Austin, Austin, Texas, and Mayo Clinic, Rochester, Minnesota. Whole mounts or adjacent sections from each specimen were examined by electron microscopy, stained for calcium phosphate, or stained with a commercially available antibody (8D10). Filtered (0.2 µm) homogenates of aneurysms were cultured and costained with 8D10 antibody followed by PicoGreen to detect DNA or incubated with [3H]uridine. Staining for calcium phosphate was heterogeneously distributed within all calcified tissues. Immunological staining with 8D10 was also heterogeneously distributed in areas with and without calcium phosphate. Analysis of areas with positive immunostaining identified spheres ranging in size from 30 to 100 nm with a spectral pattern of calcium and phosphorus (high-energy dispersive spectroscopy). Nanosized particles cultured from calcified but not from noncalcified aneurysms were recognized by a DNA-specific dye and incorporated radiolabeled uridine, and, after decalcification, they appeared via electron microscopy to contain cell walls. Therefore, nanometer-scale particles similar to those described as nanobacteria isolated from geological specimens and human kidney stones can be visualized in and cultured from calcified human cardiovascular tissue.

aneurysm; chlamydia; infection; inflammation; nanobacteria


Address for reprint requests and other correspondence: V. M. Miller, Medical Science Bldg. 4-62; Mayo Clinic Rochester; 200 First St. SW, Rochester, MN 55905 (E-mail: miller.virginia{at}mayo.edu).




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