Presence of nanobacteria in psammoma bodies of ovarian cancer: evidence for pathogenetic role in intratumoral biomineralization

Abstract

We found a 100% concordance between the expression of nanobacteria and the presence of psammoma bodies in malignant ovarian tumours. Several lines of evidence suggest the involvement of these organisms in the process of biomineralization. We therefore conclude that nanobacterial infection of malignant ovarian tissue contributes to mechanisms leading to the formation of calcified deposits known as psammoma bodies.

The role of calcifying nanoparticles in biology and medicine

“…Bacterioscopic (DNA-specific dyes Hoechst 33258, propidium iodide, PicoGreen, staining after a demineralization is optimal) using scanning and transmission electron microscopy, [16][17][18][19][20][21][22][23][24] von Kossa staining, which is specific to calcium compounds, 16 staining by 2% uranyl acetate (possibly with lead citrate) to detect specific mucus on the hydroxyapatite shell, 1 staining by alizarin red S in the mineralized state, 26 staining by phosphotungstic acid, 16 after the long-term cultivation light microscopy with von Kossa staining is possible to be used for detection. 1,16 Bacteriological (cultivation in DMEM or RPMI-1640 without serum under 37°C for 4-6 weeks after filtration through 0.10-0.22 μm pores), 1,16 replication can be assessed by spectrophotometry (650 nm wavelength).…”

“…Sedivy and Battistutti 46 found CNP antigens in 100% (7 out of 7) of ovarian malignant tumors in which psammoma bodies (which are microcalcifications) were detected, whilst in samples without such microcalcifications these antigens were not observed. 46 Their results were completely proved by Hudelist et al, 19 who identified CNP antigens in eight out of eight ovarian neoplasms containing psammoma bodies, and did not detect them in ten out of ten tumors without these microcalcifications. Interestingly, in this investigation, the extraction of mRNA of CNPs from all tumor samples with psammoma bodies was successful, whilst all attempts to do this in the cases with tumors without psammoma bodies failed.…”

“…[16][17][18][19][20][21][22][23][24] Furthermore, Hudelist et al 19 and Kumar et al 22 were successful in extracting DNA and RNA from CNPs, indicating the presence of nucleic acids. The suggestion that nucleic acids are only bound to highly charged molecules of mineralo-protein complexes outside and are not synthesized in CNPs 12 can be disproved by the results of Ciftçioglu et al, 17 Khullar et al, 20 Miller et al, 21 and Kumar et al, 22 who observed DNA-specific staining of already demineralized CNPs, which could not have been possible in the case of a simple binding of nucleic acids outside by a mineralo-protein shell.…”

“…Interestingly, in this investigation, the extraction of mRNA of CNPs from all tumor samples with psammoma bodies was successful, whilst all attempts to do this in the cases with tumors without psammoma bodies failed. 19 Wen et al 47 have shown that CNP antigens are significantly more often detected in the serum of patients with cholecystolithiasis (31.6%) compared with control group (8%). Moreover, it was observed that the gallbladder mucosa is infected by CNPs during cholecystolithiasis in 35% of cases.…”

“…Many already performed basic investigations (for instance, those where the presence of DNA in CNPs was proved by positive DNA-specific staining, but DNA was not extracted) 17,18,20,21,23,24 are close to this group, and they can be represented as "first generation" studies. Investigations by Hudelist et al 19 and Kumar et al 22 may be represented as prototypes for further investigation (in another words, they can be represented as "second generation" studies), since CNP nucleic acids were successfully extracted, although accurate sequencing was not conducted. Consequently, investigations in which fairly accurate sequencing of the CNP genome will be carried out will represent "third generation" studies.…”