The effects of essential oil, povidone-iodine, and chlorhexidine mouthwash on salivary nitrate/nitrite and nitrate-reducing bacteria.

Dietary nitrate is reduced to nitrite and nitric oxide by microbial flora, and this activity is beneficial to vascular health. It has been reported that this bacterial process is inhibited by chlorhexidine mouthwash, although the effects of other products are largely unknown. This study examined the effects of several treatments on salivary nitrate/nitrite and nitrate-reducing bacteria. Twelve university staff and students performed mouth-washing with water (control), essential oil, 0.35% povidone-iodine, or 0.0025% chlorhexidine and then ate 100 g lettuce (110 mg nitrate content), followed by collection of saliva and tongue bacteria at the baseline, and 1, 5, and 10 h thereafter. The individual treatments were separated by an interval of one week. Salivary nitrate/nitrite was measured by the calorimetric method, and a representative nitrate-reducing bacterial species, Veillonella dispar, was detected and semi-quantified using a polymerase chain reaction (PCR) assay. Significant increases in salivary nitrate/nitrite were observed for all treatments (all P < 0.05). The PCR assay showed that water, essential oil, and povidone-iodine mouthwash had little effect, whereas V. dispar DNA bands were markedly inhibited after washing with chlorhexidine. These results suggest that essential oil and povidone-iodine mouthwash have little effect on oral nitrate-reducing activity. Salivary nitrite production was not reduced by chlorhexidine, but the fainter band of V. dispar DNA suggests that longer daily use might blunt this nitrate-reducing activity.

Serological survey to determine the occurrence of malignant catarrhal fever infection in the Japanese small ruminant population from northern districts.

Ovine herpesvirus 2 (OvHV-2) causes sheep-associated malignant catarrhal fever (SA-MCF), and is responsible for economic losses in cattle and other susceptible species around the world. A survey of 154 serum samples from 14 flocks in 3 Japanese prefectures (Hokkaido, Aomori and Iwate) was undertaken between 2007 and 2008 to test for antibodies to OvHV-2. OvHV-2 was present in 56 sheep and 2 goats, with 37.66% of samples having a positive reaction using a serum neutralization test. The immune reaction reported in goats could result from Caprine herpesvirus-2. These results indicate that sheep are reservoirs for OvHV-2 in the field in Japan, and they might transmit the virus to susceptible cattle and wild fauna.

Acidic environments induce differentiation of Proteus mirabilis into swarmer morphotypes.

Although swarmer morphotypes of Proteus mirabilis have long been considered to result from surfaced-induced differentiation, the present findings show that, in broth medium containing urea, acidic conditions transform some swimmer cells into elongated swarmer cells. This study has also demonstrates that P. mirabilis cells grown in acidic broth medium containing urea enhance virulence factors such as flagella production and cytotoxicity to human bladder carcinoma cell line T24, though no significant difference in urease activity under different pH conditions was found. Since there is little published data on the behavior of P. mirabilis at various hydrogen-ion concentrations, the present study may clarify aspects of cellular differentiation of P. mirabilis in patients at risk of struvite formation due to infection with urease-producing bacteria, as well as in some animals with acidic or alkaline urine.

Nitric oxide causes anoikis through attenuation of E-cadherin and activation of caspase-3 in human gastric carcinoma AZ-521 cells infected with Mycoplasma hyorhinis.

Mycoplasma hyorhinis (M. hyorhinis) infection leads cultured cells to various biological alterations in cell metabolism including apoptosis. Apoptosis induced by M. hyorhinis has mainly been considered to be due to mycoplasmal endonucleases. We previously reported that apoptosis in a human carcinoma cell line AZ-521 infected with M. hyorhinis was enhanced by addition of L-ascorbic acid to cell cultures. Since both L-ascorbic acid addition and M. hyorhinis infection activated cellular iNOS, we examined the hypothesis that nitric oxide (NO) exerts an apoptotic effect on M. hyorhinis-infected cells and down-regulates E-cadherin. In this study, we showed that M. hyorhinis infection activates iNOS mRNA synthesis, NO production, and caspase-3 activity and attenuates E-cadherin mRNA synthesis by quantitative real-time PCR, Griess assay and fluorescence caspase-3 detection. L-NAME decreased the numbers of apoptotic cells through inhibition caspase-3 activity. Our results indicate that NO causes anoikis throughout attenuation of E-cadherin and activation of caspase-3 in human gastric carcinoma cell line AZ-521 cells infected with M. hyorhinis.

Growth inhibition of human melanoma cells by a recombinant arginine deiminase expressed in Escherichia coli.

We have cloned the arginine deiminase (ADI) gene from Mycoplasma hominis PG21 genomic DNA by polymerase chain reaction, and changed four TGA tryptophan codons (stop codon in E. coli) to TGG codons in the coding region by site-directed mutagenesis in order to express in E. coli. The recombinant ADI (rADI) was purified to apparent homogeneity by Ni-affinity chromatography after extraction from inclusion bodies followed by refolding. The rADI expressed in E. coli was estimated to be 50 kDa. Dimeric forms of rADI exerted enzymatic activity. We found that high concentration of potassium dihydrogenphosphate (PDP) and L-arginine addition in refolding reaction increases the enzyme activity. The specific activity of rADl was calculated as 0.618 U/mg. In addition, the enzyme activity of purified rADI remained for at least one month in 100 mM PDP solution (pH 6.5), but diminished within one week in 100 mM PDP solution (pH 7.4). Anti-tumor activity of the purified rADI was estimated to be 0.036 U/ml as 50% growth inhibitory activity against human melanoma cell line G-361.

L-ascorbic acid enhances apoptosis in human gastric carcinoma cell line AZ-521 cells infected with Mycoplasma hyorhinis.

Mycoplasma hyorhinis (M. hyorhinis) exerts multiple effects on cell metabolisms including apoptosis mediated by their endonucleases and nitric oxide production in vitro. Although AsA is preferable to health in general because of its reactive oxygen species scavenging activity, we found that in a human carcinoma cell line AZ-521 infected with M. hyorhinis, apoptosis was enhanced by addition of L-ascorbic acid (AsA) to the cell cultures. No significant differences were evident between the AZ-521 cells with and without AsA (AsA-) after 24 hr of incubation in the mitochondrial fluorescence. M. hyorhinis-infected AZ-521 cells treated with AsA (AsA +) have developed distinct DNA ladders as compared to the control cells AsA- after 24 hr of incubation. Marked cytopathic effects were rather apparent in AsA-treated cells than in control cells AsA- after 24 hr. Our data demonstrate that AsA addition to cell cultures enhances apoptosis induced by M. hyorhinis infection. We suggest that the presence of another external apoptotic pathway by M. hyorhinis infection.

Self-propagating calciferous particles detected in a human cell line Kasumi-6 (JCRB1024).

Tiny particles were found in the medium in the presence of the human leukemia cell line Kasumi-6. The particles were separated from human cells by filtration and incubated in RPMI1640 supplemented with 10% fetal calf serum at 37 C. The particles increased in number very slowly in the liquid medium but did not reveal any biological activity. Transmission electron microscopy of the particles showed a spheroid or ovoid shape in ultrathin section. No specific polypeptides from the purified particles were detected by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), except for bovine fetuin that adsorbed to the surface of the particles. X-ray diffractometry as well as Fourier transform infrared spectrometry suggested the particles consisted of hydroxyapatite. The mechanism of self-propagation of the hydroxyapatite particles in liquid medium is currently unknown. This type of particle has been overlooked for a long period because it is noncultivable. It will be necessary to examine its biological effects to the cultured cells.