Correlation between human health and reactive oxygen species produced in blood: a long-term chemiluminescence and fluorescence analysis.

The previous slide-glass type system could simultaneously detect reactive and highly reactive oxygen species, i.e., superoxide radicals (O2-·) and hypochlorite ions (OCl-) elicited from leucocytes in sample blood, but had some drawbacks, i.e., signal noise from air-flow stirring, potential biohazard risks, etc. because of open samples placed on a slide glass. We overcame these drawbacks by adopting a fluidic-chip container in a new system, which resulted in higher sensitivity and more stable measurements. Using the new system, we conducted a pilot study on nominally healthy volunteers to find whether or not the monitored activities of leukocytes can distinguish more or less unhealthy conditions from healthy ones. At first, healthy volunteers of both genders and of various ages showed that the fluctuation magnitudes (%) of O2-· and OCl- were nearly similar to each other and to that of the neutrophil count fluctuation. These parameters sometimes exceeded the healthy fluctuation range. By comparing these large fluctuations with the data of an inflammation marker C-reactive protein (CRP), the neutrophil count fluctuation and the timings/symptoms of abnormalities found in questionnaire, we could gain information suggesting the factors causing the large fluctuations. The new system could detect bodily abnormalities earlier than CRP or self-aware symptoms.

Rapid on-site dual optical system to measure specific reactive oxygen species (O2-• and OCl-) in a tiny droplet of whole blood.

Oxidative stress has been implicated in various disorders and controlling it would be important for healthy life. We have developed a new optical system for easily and accurately measuring oxidative stress in whole blood. It is optimized for simultaneously detecting reactive oxygen species (ROS) and highly reactive ROS (hROS), elicited mostly by white blood cells in a few microliters of blood. Results obtained by using this system show at least four important findings. 1) chemiluminescence of MCLA was confirmed to be attributable to O2-•. 2) PMA-stimulated cells released O2-• longer and more slowly than fMLP-stimulated ones. 3) fluorescence produced by APF oxidation was confirmed to be attributable to hROS, mostly OCl-, produced by myeloperoxidase. 4) the generation of OCl- was found to be a slower process than the O2-• generation. We also conducted pilot studies of oxidative stress in healthy volunteers.

Evaluation of a Hypertensive Rat Model Using Peripheral Blood Neutrophil Activity, Phagocytic Activity and Oxidized LDL Evaluation.

BACKGROUND/AIM: A system is being developed that can be used to easily evaluate the health condition of an individual with the help of trace amounts of a blood sample by focusing on xenobiotics. The system is called "Multimodal homeostasis evaluation system" (measurement of neutrophil activity, phagocytic activity of phagocytes and quantification of oxidized LDL (OxLDL)). To elucidate the possibility of using this system as an evaluation system for the health condition of an individual, clearly explaining the changes in various diseases is essential. In this study, evaluations were carried out using hypertensive model animals. MATERIALS AND METHODS: Spontaneously hypertensive model rats SHR/NCrlCrlj and control rats WKY/NCrlCrlj were raised for 10 weeks from 6 to 16 weeks of age and their blood pressure was measured over time. Blood neutrophil activity (superoxide anion (O2•-) generation and myeloperoxidase (MPO) activity) and phagocytic activity of phagocytes was measured by our developed apparatus (a simple prototype device under development). OxLDL was measured by an ELISA kit, and biochemical markers were measured using the blood sample. RESULTS: Compared to WKY rats of the control group, systolic blood pressure, diastolic blood pressure, and mean blood pressure of SHR rats increased significantly with age. In SHR rats, there was a significant elevation in O2•- generation and MPO activity of neutrophils, alanine aminotransferase and triglycerides of blood, while phagocytic activity, OxLDL, low-density lipoprotein cholesterol, high-density lipoprotein cholesterol, total cholesterol and total-bilirubin decreased. CONCLUSION: In the hypertensive model, biochemical markers were found to have a relationship with O2•- generation, MPO activity, phagocytic activity of phagocytes, and OxLDL. This system is expected to be useful for clinical monitoring of hypertension diseases.

Oral administration of Pantoea agglomerans-derived lipopolysaccharide prevents metabolic dysfunction and Alzheimer's disease-related memory loss in senescence-accelerated prone 8 (SAMP8) mice fed a high-fat diet.

The pathogenesis of Alzheimer's disease (AD) remains unclear, but an imbalance between the production and clearance of amyloid-ß (Aß) peptides is known to play a critical role in AD progression. A promising preventative approach is to enhance the normal Aß clearance activity of brain phagocytes such as microglia. In mice, the intraperitoneal injection of Toll-like receptor 4 agonist was shown to enhance Aß clearance and exhibit a preventative effect on AD-related pathology. Our previous clinical study demonstrated that orally administered Pantoea agglomerans-derived lipopolysaccharide (LPSp) exhibited an LDL (low-density lipoprotein)-lowering effect in human volunteers with hyperlipidemia, a known risk factor for AD. In vitro studies have shown that LPSp treatment increases Aß phagocytosis by microglial cells; however it is still unclear whether orally administered LPSp exhibits a preventive effect on AD progression. We show here that in senescence-accelerated prone 8 (SAMP8) mice fed a high-fat diet, oral administration of LPSp at 0.3 or 1 mg/kg body weight·day for 18 weeks significantly improved glucose metabolism and lipid profiles. The LPSp treatment also reduced pro-inflammatory cytokine expression and oxidative-burst activity in the peripheral blood. Moreover, LPSp significantly reduced brain Aß burden and memory impairment as seen in the water maze test, although we could not confirm a significant enhancement of Aß phagocytosis in microglia isolated from the brains after treatment. Taken together, our results show that LPSp holds promise as a preventative therapy for AD or AD-related diseases induced by impairment of metabolic functions.

Oral administration of Pantoea agglomerans-derived lipopolysaccharide prevents development of atherosclerosis in high-fat diet-fed apoE-deficient mice via ameliorating hyperlipidemia, pro-inflammatory mediators and oxidative responses.

Pantoea agglomerans (P. agglomerans) is a Gram-negative bacterium that grows symbiotically with various edible plants, and the oral or sublingual administration of lipopolysaccharide derived from P. agglomerans (LPSp) have been suggested to contribute to prevention of immune-related diseases. Our previous study indicated that orally administered LPSp was shown to exhibit an LDL-lowering effect in hyperlipidemic volunteers; however, a preventive effect of LPSp on atherosclerosis is unclear. The present study attempted to evaluate the anti-atherosclerotic effect by LPSp in a mouse model of high-fat diet (HFD)-induced atherosclerosis. For 16 weeks, apoE-deficient mice were fed an HFD and received drinking water containing LPSp (0.3 or 1 mg/kg body weight/day). The results showed that the orally administered LPSp decreased body weight. A significant reduction in atherosclerotic plaque deposition was observed even with the lower dose of LPSp. The biochemical analyses showed that LPSp markedly improved glucose tolerance and reduced plasma LDL and oxidized LDL levels. In addition, LPSp significantly reduced the production of pro-inflammatory mediators including MCP-1 (in the plasma), TNF-α and IL-6 (in the colon), and decreased the oxidative burst activities in the peripheral blood sample. Taken together, these results suggest the possibility that oral administration of LPSp can effectively ameliorate HFD-induced hyperlipidemia and inflammatory/oxidative responses to prevent atherosclerosis and related metabolic disorders.