Visiting a forest, but not a city, increases human natural killer activity and expression of anti-cancer proteins.

We previously reported that a forest bathing trip enhanced human NK activity, number of NK cells, and intracellular anti-cancer proteins in lymphocytes. In the present study, we investigated how long the increased NK activity lasts and compared the effect of a forest bathing trip on NK activity with a trip to places in a city without forests. Twelve healthy male subjects, age 35-56 years, were selected with informed consent. The subjects experienced a three-day/two-night trip to forest fields and to a city, in which activity levels during both trips were matched. On day 1, subjects walked for two hours in the afternoon in a forest field; and on day 2, they walked for two hours in the morning and afternoon, respectively, in two different forest fields; and on day 3, the subjects finished the trip and returned to Tokyo after drawing blood samples and completing the questionnaire. Blood and urine were sampled on the second and third days during the trips, and on days 7 and 30 after the trip, and NK activity, numbers of NK and T cells, and granulysin, perforin, and granzymes A/B-expressing lymphocytes in the blood samples, and the concentration of adrenaline in urine were measured. Similar measurements were made before the trips on a normal working day as the control. Phytoncide concentrations in forest and city air were measured. The forest bathing trip significantly increased NK activity and the numbers of NK, perforin, granulysin, and granzyme A/B-expressing cells and significantly decreased the concentration of adrenaline in urine. The increased NK activity lasted for more than 7 days after the trip. In contrast, a city tourist visit did not increase NK activity, numbers of NK cells, nor the expression of selected intracellular anti-cancer proteins, and did not decrease the concentration of adrenaline in urine. Phytoncides, such as alpha-pinene and beta-pinene were detected in forest air, but almost not in city air. These findings indicate that a forest bathing trip increased NK activity, number of NK cells, and levels of intracellular anti-cancer proteins, and that this effect lasted at least 7 days after the trip. Phytoncides released from trees and decreased stress hormone may partially contribute to the increased NK activity.

Forest bathing enhances human natural killer activity and expression of anti-cancer proteins.

In order to explore the effect of forest bathing on human immune function, we investigated natural killer (NK) activity; the number of NK cells, and perforin, granzymes and granulysin-expression in peripheral blood lymphocytes (PBL) during a visit to forest fields. Twelve healthy male subjects, age 37-55 years, were selected with informed consent from three large companies in Tokyo, Japan. The subjects experienced a three-day/two-night trip in three different forest fields. On the first day, subjects walked for two hours in the afternoon in a forest field; and on the second day, they walked for two hours in the morning and afternoon, respectively, in two different forest fields. Blood was sampled on the second and third days, and NK activity; proportions of NK, T cells, granulysin, perforin, and granzymes A/B-expressing cells in PBL were measured. Similar measurements were made before the trip on a normal working day as the control. Almost all of the subjects (11/12) showed higher NK activity after the trip (about 50 percent increased) compared with before. There are significant differences both before and after the trip and between days 1 and 2 in NK activity. The forest bathing trip also significantly increased the numbers of NK, perforin, granulysin, and granzymes A/B-expressing cells. Taken together, these findings indicate that a forest bathing trip can increase NK activity, and that this effect at least partially mediated by increasing the number of NK cells and by the induction of intracellular anti-cancer proteins.

Potential immunosuppressive and antiinflammatory activities of Malaysian medicinal plants characterized by reduced cell surface expression of cell adhesion molecules.

In the search for agents effective against immune-mediated disorders and inflammation, we have screened Malaysian medicinal plants for the ability to inhibit the expression of intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) on the surface of murine endothelial cells (F-2), and mouse myeloid leukaemia cells (M1), respectively. Of 41 kinds (29 species, 24 genera, 16 families) of Malaysian plants tested, 10 and 19 plant samples significantly downregulated the expression of ICAM-1 and VCAM-1, respectively. Bioassay-directed fractionation of an extract prepared from the bark of Goniothalamus andersonii showed that its ingredients, goniothalamin (1) and goniodiol (2) inhibited the cell surface expression of both ICAM-1 and VCAM-1. The present results suggest that Malaysian medicinal plants may be abundant natural resources for immunosuppressive and antiinflammatory substances.

Influence of natural and synthetic compounds on cell surface expression of cell adhesion molecules, ICAM-1 and VCAM-1.

Various natural and synthetic compounds including alkaloids, terpenoids and phenolics were tested for inhibition of the cell surface expression of intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1), both of which are crucial in the regulation of immune response and inflammation. Of 40 compounds tested, two compounds significantly downregulated the expression of VCAM-1 on murine endothelial cells (F-2) and ten compounds that of ICAM-1 on mouse myeloid leukemia cells (M1). Sanguinarine chloride (5) and isoliquiritigenin (13) were capable of lowering the levels of both ICAM-1 and VCAM-1. The structure-activity relationships study on chalcone and flavone derivatives related to 13 suggested that the inhibitory activity of the chalcone derivatives is attributable to the 4-hydroxy group as well as the possible coplanarity between the phenyl ring and the adjacent conjugated ketone.

P300 habituation from auditory single-stimulus and oddball paradigms.

The habituation of auditory P300 event-related brain potential (ERP) from single-stimulus paradigm was evaluated and compared to that from oddball paradigm. Three task conditions were: oddball with a button press (oddball/press) response, single-stimulus with a button-press (single-stimulus/press) response and a silent count (single-stimulus/count) response. The oddball/press condition demonstrated larger P300 amplitude and longer latency overall than either single-stimulus condition, but P300 amplitude decreased and peak latency increased similarly over successive trial blocks for all three tasks. Thus, the oddball and single-stimulus ERP tasks produce analogous changes under repeated measurements and indicate that the single-stimulus task can serve as an alternative method for eliciting the P300 in applied and clinical settings.

Oral taurine supplementation prevents the development of ethanol-induced hypertension in rats.

Taurine is known to lower blood pressure in essential hypertension and some experimental hypertensive models. Taurine has also been reported to activate aldehyde dehydrogenase and to inhibit the elevation of plasma acetaldehyde concentration after ethanol intake. Because acetaldehyde, the first metabolite of ethanol, is suspected to be responsible for many adverse effects of alcohol consumption, we examined the effect of taurine supplementation on ethanol-induced hypertension and abnormalities in the intracellular cation metabolism in Witar-Kyoto rats. In Study 1, systolic blood pressure and intraplatelet free calcium were significantly higher in rats who received 15% ethanol in drinking water than in control rats. Oral taurine supplementation (1% taurine and 15% ethanol in drinking water) completely prevented the development of ethanol-induced hypertension. Intraerythrocyte sodium and intraplatelet free calcium were significantly decreased in taurine-supplemented rats as compared with rats who received 15% ethanol only. In Study 2, hemoglobin-associated acetaldehyde (HbAA) was measured as a marker of protein-bound acetaldehyde. HbAA was significantly elevated in rats who received 5% ethanol in drinking water as compared with control rats. Taurine supplementation (1% taurine and 5% ethanol in drinking water) significantly decreased HbAA. Our findings suggest that the oral supplementation of taurine prevents ethanol-induced hypertension by decreasing protein bound acetaldehyde and altering the cation handling by the membrane.

P300 as a measure of cognitive dysfunction from occupational and environmental insults.

The P300 component of the event-related brain potential (ERP) is a sensitive, non-invasive, and convenient measure of cognitive dysfunction resulting from a variety of etiological agents. Application-orientated research on using the P300 measure as a cognitive probe for a wide range of neurological and psychiatric situations has been expanding rapidly in the last decade.The aim of this paper is to preview issues of application-oriented P300 research in occupational and environment medicine. Firstly, the neurophysiological background of the P300 is outlined. Secondly, the recent findings of P300 abnormalities following various occupational and environmental exposures are overviewed. Thirdly, the empirical issues for application-oriented research such as the potential causes of variability, limitation and difficulty are summarized, with suggestion for controlling them and for future standardization. Finally, it is concluded that P300 assessments demonstrate promising possibility as a sensitive marker for general cognitive dysfunction in occupational and environmental medicine.

[A study of preventive medicine in relation to mental health among middle-management employees (Part 2)--effects of long working hours on lifestyles, perceived stress and working-life satisfaction among white-collar middle-management employees].

Recently, we have introduced the holistic method into health care of workers and aimed at improvement of Quality of Life (QOL). It has been made clear that primary prevention of diseases needs lifestyle appraisal. Therefore, we studied the daily working hours and the effects of long working hours on lifestyles, perceived stress and working-life satisfaction, which we used as a subjective index of Quality of Life (QOL), based on data obtained from a survey of 1,026 department chiefs and 2,902 section chiefs in 110 large companies in Japan in 1990. The results are summarized as follows. (1) The percentage of those working 10 hours per day was 41.6% for department chiefs and 40.4% for section chiefs, and for 11 or more working hours it was 24.1% in department chiefs and 30.9% in section chiefs. The younger both department and section chiefs, the longer their working hours. (2) Both department and section chiefs had a significant relationship between long working hours and poor sleeping habits, poor physical exercise, feeling busy, irregularity of daily life and irregularity of daily meals. The department chiefs had a significant relationship between long working hours and unbalanced nutrition or no hobbies. The section chiefs had a significant relationship between long working hours and drinking many cups of tea or coffee, taking a lot of salt or ill physical condition during the past six months.

[Treatment with arbekacin of surgical infections by resistant strains of Staphylococcus aureus. Arbekacin Study Group].

The frequency of infection by methicillin-resistant Staphylococcus aureus (MRSA) is high in Japan and control of such strains is urgently needed. Arbekacin (ABK), a semisynthetic aminoglycoside, has potent activity against S. aureus, including resistant strains, and against Gram-negative bacteria as well. For this reason, in surgical infections (which are often caused by more than one bacterium), this drug might be particularly effective. We calculated the MIC and the decrease in the MIC when cultures of 59 resistant strains of S. aureus isolated in our wards at Osaka City University Hospital, contained arbekacin in the medium. We also used the drug to treat 12 infections caused by resistant strains of S. aureus. The MICs of vancomycin had a single peak at 0.5 microgram/ml, and those for ABK had double peaks at 0.5 and 4.0 micrograms/ml. The effect of arbekacin in lowering the MIC of minocycline (MINO) was slight because of the low MIC of MINO. Effects on fosfomycin (FOM), ampicillin, clavulanic acid/ticarcillin, cefotiam, cefuzonam, flomoxef, and imipenem/cilastatin were strong; the peaks were lowered by 1/2(7)-1/2(11). When 1.0 micrograms/ml ABK was present in the medium, the efficacy of FOM was increased enough that, by prediction from the pharmacokinetics of FOM (blood level when given at the usual dose), all but one (2%) of the 47 resistant strains would be eradicated clinically. If 2.0 micrograms/ml ABK were in the medium, all strain would be eradicated, by our calculations. We treated 11 infections and one colonization by resistant strains of S. aureus with ABK and evaluated the response in these cases of infection. Four infections were treated with FOM as well. The clinical efficacy was good in four infections (three patients), fair in four, and poor in three, for an efficacy rate of 36%. All presumed causative bacteria were eradicated in two (18%) of the 11 infections and S. aureus strains were eradicated in three (27%) of the 11 infections. No symptoms of side effects were reported, but blood urea nitrogen and creatinine rose in a 72-year-old woman with duodenal perforation and peritonitis. The MIC levels of ABK were satisfactory, but clinical efficacy for staphylococcal infections caused by resistant strains was unsatisfactory.

Effects of bed rest on bone metabolism in patients with femoral neck fracture.

The effect of continuous 1 week bed rest during traction on bone metabolism were studied in patients with femoral neck fracture. Bone demineralization was assessed by measuring serum calcium (Ca), phosphorus (P), and alkaline phosphatase (AlkP) before, during, and after 1 week-bed rest with skeletal traction. In addition, 24-hour urinary excretion of calcium and phosphorus were measured. Serum Ca did not change significantly before, during, and after bed rest, whereas serum P increased after 1 week bed rest and postoperatively remained significantly elevated. Serum AlkP did not increase with bed rest, but became elevated postoperatively. Urinary Ca excretion increased from 105.8 +/- 15.2 to 138.0 +/- 15.4 mg/24h after 1 week of bed rest, and remained elevated postoperatively. In contrast, urinary P excretion initially increased 495.1 +/- 65.0 to 610.8 +/- 87.7 mg/24h, and subsequently decreased to a level significantly less than the admission level. These results suggest that 1 week bed rest for the patients with femoral neck fracture resulted in bone atrophy and demineralization.

Tissue reaction to alumina implants inserted into the tibiae of rats.

We examined the tissue reaction to three alumina implants--single-crystal alumina (SA), dense polycrystal alumina (DPA), and porous polycrystal alumina (PPA)--inserted transcortically, extending into the medullary canal of rat tibiae, and assessed the quantitative differences in bone reaction using an image processing system. There was no difference in the degree of maturation of newly formed bone around the three kinds of alumina. SA and DPA were encapsulated with a continuous bone layer, but some bone tissue was attached focally around PPA. Bone trabeculae in the control site diminished in size and number chronologically. Multinucleated giant cells were observed on the surface of DPA and PPA, but not on SA. Tabulation of the quantitative evaluation indicated that SA showed the highest, DPA a lower, and PPA the lowest in bone contact rate, bone contact thickness, and bone contact area. These data suggest that SA is superior to the other two as an implant material.

Improved method using a bubble-free adhesion technique for the preparation of semi-serial undecalcified histologic sections containing dental implants.

The preparation technique, with the minimum of artifacts for the semi-serial undecalcified histologic sections containing dental implants, is presented in this study. The sections enabled finer light-microscopic observations to be made. A formalin-fixed tissue block containing a dental implant was dehydrated in ethanol and acetone, and then embedded in polyester resin under 76 cm Hg reduced pressure. The embedded block was trimmed by a cutter and ground by abrasive paper. In a 1.5 Kg f/cm2 pressurized chamber, its polished surface was bonded to a methacrylate slide by means of ethylcyanoacrylate used in an adhesion loading device. This meant that no bubbles could arise in the interface between the slide and the block. The slide-block was then attached to an adsorptive specimen-holder of a hard-tissue cutting machine and cut to a thickness of approximately 50 microns, with use of a diamond blade. The slide-section was ground to 15-40 microns with wet-type abrasive paper and film on a polishing table. Etching with weak acid and surface staining with toluidine blue and methylene blue/basic fuchsin/light green were performed on the section.

Differences between the bony interfaces of titanium and hydroxyapatite-alumina plasma-sprayed titanium blade implants.

Histopathologic study was done to reveal the difference of bony interface in the mandibular bone between titanium blade implants and titanium implants coated with the mixture of hydroxyapatite-alumina by the plasma-spray technique, using the improved undecalcified histologic technique and the image analyzing system. In 4 adult shepherd dogs, the implantation was performed in the molar region of the mandible 2 months after tooth extraction. The coated implants and the non-coated ones were inserted respectively in the mandibles. They were sacrificed on the 5th, 7th, 14th and 28th day postoperatively. In 5-day implant after insertion, granulation tissues or fibrous connective tissues were observed around both implants in bone tissues. In 7-day implant, osteoid tissues regenerated around the apex of both implants, but attached a little to them. In 14-day implant, osteoid tissues or woven bone regenerated around both implants and attached more to the coated implant (approximately 13.7%) than the non-coated one (approximately 6.0%). In 28-day implant, regenerative woven or lamellar bone attached much more to the coated implant (approximately 63.4%) than the non-coated one (approximately 8.8%). Bony interface increased remarkably in the coated implants, chronologically. The results showed that the plasma-sprayed hydroxyapatite-alumina coating was effective for the initial fixation of endosseous dental implants.

Immunohistochemical investigation of cerebral ischemia during recirculation.

Immunohistochemical methods for the determination of tubulin, creatine kinase BB-isoenzyme, and astroprotein-glial fibrillary acidic protein were used to investigate recovery of the ischemic lesion after temporary occlusion of a common carotid artery in the gerbil and the evolution of the postischemic lesion following reperfusion. One group of gerbils was followed from 15 minutes to one month after an ischemic period of 30 minutes, and another group was examined after 7 days following an ischemic period of 5 to 30 minutes. It was found that the postischemic lesion, visualized as loss of the immunohistochemical reaction for tubulin and creatine kinase BB-isoenzyme, evolved within 60 minutes after reperfusion in the hippocampus and cerebral cortex and within 3 hours in the caudoputamen and thalamus. Resolution of the preexisting ischemic lesion was possible only after an ischemic period of less than 10 minutes in the cerebral cortex and caudoputamen and less than 15 minutes in the thalamus. In the CA1-CA2 region of the hippocampus, the ischemic lesion already existed after an ischemic period of 5 minutes and was mostly irreversible. The immunohistochemical method of testing for different cellular and subcellular components was very useful for investigation of cerebral ischemia and may also be advantageous for investigation of other pathophysiological conditions of the nervous system.

Immunohistochemical investigation of cerebral ischemia in gerbils.

Experimental cerebral ischemia was produced in gerbils by occlusion of the right common carotid artery in the neck. The evolution of the ischemic lesions was followed from five minutes to six hours by using the immunohistochemical techniques for tubulin and creatine kinase BB-isoenzyme. The earliest lesion was found in the subiculum-CA1 and CA2 regions of the hippocampus in five minutes. There was loss of staining in the apical dendrites and perikarya of the pyramidal cells. The earliest lesion in the cerebral cortex, visible in ten minutes, was a laminar loss of staining for tubulin. Evolution of the ischemic lesions in the thalamus and caudoputamen was delayed. However, in two hours widespread ischemic lesions were seen there. Evolution of the ischemic lesions was slightly slower with the reaction for creatine kinase BB-isoenzyme as compared to the reaction for tubulin, but was far more sensitive than hematoxylin-eosin staining. The distribution of ischemic lesions detected by the immunohistochemical method compared to ischemic areas detected by an India ink perfusion study suggested that both the extent of regional ischemia and regional difference in tissue vulnerability were contributing factors for the emergence of early ischemic lesions. The mechanism for prompt disappearance of the immunohistochemical reaction for tubulin is not clear, but the present investigation demonstrates the usefulness of the immunohistochemical technique for detecting early ischemic lesions and provides a possible biochemical mechanism for cellular damage after ischemic insults.

Absorption and bioavailability of diclofenac after rectal administration of diclofenac-na gel preparation in rat and man.

In order to evaluate diclofenac-Na (DC-Na) micro-enema, DC-Na gel preparations were administered to rats and man. When DC-Na gel preparations were rectally administered at various pH (pH 5- 8) to rats, their bioavailability increased at higher pH. The bioavailability of DC-Na gel preparations (pH 8.0) in rats was significantly higher than that with conventional suppository bases, Witepsol H-15 and polyethylene glycol 1000 (PEG 1000). In man, the DC-Na gel preparation showed higher Cmax and higher bioavailability than commercial suppository made with an oily base. DC-Na gel preparations containing 10% v/v oleic acid showed a prolonged action. The irritative effect of DC-Na gel preparation on rectal mucosa in rats was weaker than that of PEG 1000, but similar to that of Witepsol H-15. Therefore, the present results suggest that gel preparation is a favorable form for rectal administration of diclofenac-Na.

Immunohistochemical localization of creatine kinase BB-isoenzyme and tubulin in gerbil brain.

Cellular distribution of creatine kinase BB-isoenzyme and tubulin was investigated immunohistochemically with tissue sections from various areas of the gerbil brain. Both the reaction for creatine kinase BB-isoenzyme and tubulin visualized nerve cell bodies, dendrites and axons. While the reaction for tubulin was similar between nerve cell bodies and their dendrites, the reaction for creatine kinase BB-isoenzyme tended to be more intense in dendrites. While the reaction for tubulin could be seen in nerve cell bodies of different sizes, the reaction for creatine kinase BB-isoenzyme was more intense in larger neurons. Both the reaction for creatine kinase BB-isoenzyme and tubulin visualized astrocytic cytoplasm but only in the corpus callosum and in the white matter of limited areas. Visualization of dendrites to their peripheries with the reaction for creatine kinase BB-isoenzyme may indicate participation of this isoenzyme in maintainance of high energy-phosphates locally within dendrites. A combination of the immunohistochemical procedure for creatine kinase BB-isoenzyme and tubulin may be very useful for demonstration of various pathophysiological states of the central nervous system which affect dendrites.

Selenite prevents the induction of sister-chromatid exchanges by methyl mercury and mercuric chloride in human whole-blood cultures.

The protective effect of sodium selenite (Na2SeO3) against the cytogenetic toxicity of methyl mercury (CH3HgCl) and mercuric chloride (HgCl2) were investigated on human whole-blood cultures in relation to induction of sister-chromatid exchange (SCE). Both mercurials caused a dose-dependent increase in SCEs, methyl mercury being about 5 times more potent than mercuric chloride. Sodium selenite also induced SCEs. However, the simultaneous addition of selenite (1 x 10(-7) -3 x 10(-5) M) to cell cultures containing either methyl mercury (3 x 10(-6) M) or mercuric chloride (1 x 10(-5) M) prevented the induction of SCEs by the mercurial in a clear dose-related manner. When selenite and mercurial were simultaneously added at a molar ratio of 1:2 Na2SeO3:CH3HgCl, or 1:1 Na2SeO3:HgCl2, cells from treated cultures showed no increase in the SCE frequency. These results indicate that selenite and mercury mutually antagonize their ability to cause DNA damage leading to the formation of SCEs. The formation of bis(methylmercuric)selenide, (CH3Hg)2Se, from Na2SeO3 and CH3HgCl, or a high molecular complex consisting of glutathione-Se-Hg from Na2SeO3 and HgCl2 involving the participation of glutathione in RBCs might play a key role in this antagonism between mercury and selenium.