Characterization of Granulations of Calcium and Apatite in Serum as Pleomorphic Mineralo-Protein Complexes and as Precursors of Putative Nanobacteria

Calcium and apatite granulations are demonstrated here to form in both human and fetal bovine serum in response to the simple addition of either calcium or phosphate, or a combination of both. These granulations are shown to represent precipitating complexes of protein and hydroxyapatite (HAP) that display marked pleomorphism, appearing as round, laminated particles, spindles, and films. These same complexes can be found in normal untreated serum, albeit at much lower amounts, and appear to result from the progressive binding of serum proteins with apatite until reaching saturation, upon which the mineralo-protein complexes precipitate. Chemically and morphologically, these complexes are virtually identical to the so-called nanobacteria (NB) implicated in numerous diseases and considered unusual for their small size, pleomorphism, and the presence of HAP. Like NB, serum granulations can seed particles upon transfer to serum-free medium, and their main protein constituents include albumin, complement components 3 and 4A, fetuin-A, and apolipoproteins A1 and B100, as well as other calcium and apatite binding proteins found in the serum. However, these serum mineralo-protein complexes are formed from the direct chemical binding of inorganic and organic phases, bypassing the need for any biological processes, including the long cultivation in cell culture conditions deemed necessary for the demonstration of NB. Thus, these serum granulations may result from physiologically inherent processes that become amplified with calcium phosphate loading or when subjected to culturing in medium. They may be viewed as simple mineralo-protein complexes formed from the deployment of calcification-inhibitory pathways used by the body to cope with excess calcium phosphate so as to prevent unwarranted calcification. Rather than representing novel pathophysiological mechanisms or exotic lifeforms, these results indicate that the entities described earlier as NB most likely originate from calcium and apatite binding factors in the serum, presumably calcification inhibitors, that upon saturation, form seeds for HAP deposition and growth. These calcium granulations are similar to those found in organisms throughout nature and may represent the products of more general calcium regulation pathways involved in the control of calcium storage, retrieval, tissue deposition, and disposal

Polyphenol-rich cranberry juice has a neutral effect on endothelial function but decreases the fraction of osteocalcin-expressing endothelial progenitor cells

PURPOSE: Cranberry juice (CJ) contains a remarkably high concentration of polyphenols, considered to be beneficial for cardiovascular and bone health. The current double-blind, randomized study was designed to test whether daily consumption of double-strength Ocean Spray light CJ (2 x 230 ml) over 4 months has beneficial effects on vascular function and on endothelial progenitor cells (EPCs) carrying the osteoblastic marker osteocalcin in particular. METHODS: A total of 84 participants (49.5 +/- 16.2 years) with peripheral endothelial dysfunction and cardiovascular risk factors were enrolled in this double-blind, randomized, controlled trial (69 completed the 4-month protocol-32 in the CJ group and 37 in the placebo group, respectively). Vascular responses to reactive hyperemia were measured non-invasively by peripheral arterial tonometry (EndoPAT). Peripheral blood mononuclear cells were stained for EPC markers, as well as osteocalcin, and counted by flow cytometry. RESULTS: Baseline characteristics were similar in both groups. The effect of CJ on peripheral endothelial function and on circulating EPC counts (CD34(+)/CD133(+)/KDR(+)) did not change during the study. A high percentage of EPCs expressed osteocalcin (59.4 +/- 35.7%). CJ, as compared to placebo, induced a decrease in the fraction of EPCs expressing osteocalcin (-8.64 +/- 48.98 and 19.13 +/- 46.11%, respectively, p = 0.019). Systemic levels of the adhesion marker ICAM correlated significantly with the number of EPCs expressing osteocalcin. CONCLUSIONS: The study demonstrated that long-term supplementation of polyphenol-rich CJ did not improve peripheral endothelial function. However, the decrease in the fraction of osteocalcin+ EPCs suggests a potential beneficial effect of polyphenol-rich CJ