Hiroshi Ochi

Lipid A directly inhibits IL-4 production by murine Th2 cells but does not inhibit IFN-γ production by Th1 cells

Lipopolysaccharide (LPS) is known to be an immunopotentiator but its effect on cytokine production by Th1 and Th2 cells is unknown. We found that high amounts of LPS, its lipid A moiety, and a lipid A analog all induced a decrease in IL‐4 production and an increase in IFN‐γ production when given to keyhole limpet hemocyanin (KLH)‐restimulated lymph node cells prepared from KLH‐primed mice. Lipid A was similarly found to inhibit IL‐4 production by purified CD4+ T cells and Th2 clones activated with immobilized anti‐CD3ϵ and anti‐CD28 antibodies, suggesting that the inhibition is not indirectly mediated through effects on antigen‐presenting cells. No inhibitory effect of lipid A was observed on IFN‐γ production by a Th1 clone. Production of both IL‐4 by the Th2 clones and IFN‐γ by the Th1 clone were inhibited by the immunosuppressive agent cyclosporin A. These findings indicate that lipid A can directly inhibit IL‐4 production by CD4+ T cells without inhibiting the production of IFN‐γ. Lipid A may therefore become a useful tool to study the intracellular events that differentiate Th1 and Th2 cells.

Methamphetamine directly accelerates beating rate in cardiomyocytes by increasing Ca2+ entry via L-type Ca2+ channel

Methamphetamine induces several cardiac dysfunctions, which leads to arrhythmia, cardiac failure and sudden cardiac death. Although these cardiac alterations elicited by methamphetamine were thought to be due to an indirect action of methamphetamine, namely, an excessive catecholamine release from synaptic terminals, while it seems likely that methamphetamine directly modulates the functioning of cardiomyocytes independent of neurotransmitters. However, the direct effects of methamphetamine on cardiomyocytes are still not clear. We show that methamphetamine directly accelerates the beating rate and alters Ca(2+) oscillation pattern in cultured neonatal rat cardiomyocytes. Adrenergic receptor antagonists did not block the methamphetamine-induced alterations in cardiomyocytes. Treatment with a ryanodine receptor type 2 inhibitor and a sarcoplasmic reticulum Ca(2+)-ATPase inhibitor did not affect these responses, either. In contrast, the L-type Ca(2+) channel inhibitor nifedipine eradicated these responses. Furthermore, methamphetamine elevated the internal free Ca(2+) concentration in HEK-293T cells stably transfected with the L-type Ca(2+) channel alpha1C subunit. In neonatal rat cardiomyocytes, methamphetamine accelerates beating rate and alters Ca(2+) oscillation pattern by increasing Ca(2+) entry via the L-type Ca(2+) channels independent of any neurotransmitters.

Peripheral blood T lymphocytes and basophils, freshly isolated from house-dust-mite-sensitive patients, produce interleukin-4 in response to allergen-specific stimulation.

We examined the capacity of interleukin-4 (IL) production from lymphocytes and basophils, isolated from the peripheral blood of allergic patients sensitive to house dust mite, after stimulation with mite extract. IL-4 production was measured by a sensitive bioassay based on coculture with CT.h4S (a human IL-4-responsive cell line). Lymphocytes and basophils from patients with elevated serum IgE specific to mite allergen [radioallergosorbent test (RAST) score > 3] could produce detectable levels of IL-4 in response to mite extract, whereas those from patients with a RAST score of less than 2 or normal volunteers could not. The sensitivity of basophils to mite extract was high, so that a lower concentration of mite extract (1-10 ng/ml) could induce maximal IL-4 production. On the other hand, a higher concentration (10 micrograms/ml) was required for maximal IL-4 production from the lymphocytes. These findings demonstrate that allergen-specific IL-4-producing cells, lymphocytes and basophils, are generated in vivo in allergic patients and also that there exist characteristic differences between lymphocytes and basophils related to the in vivo source of IL-4.

Effects of buttermilk powders on emulsification properties and acid tolerance of cream.

UNLABELLED Emulsifying properties and acid tolerance are 2 of the most important characteristics of cream. The effects of the buttermilk component, especially its phospholipids, on the emulsifying properties and acid tolerance of cream were investigated in this study. Two buttermilks with differing phospholipid contents and skimmed milk were used to evaluate the effects of phospholipids on the aforementioned parameters. The mean diameter of fat globules and the cream viscosity were used as indicators of emulsifying properties. Acid tolerance was evaluated by studying the effect of citric acid on the maximum viscosity of cream. This was tested by adding 400 μL of 10% (w/w) citric acid solution to cream every minute and simultaneously measuring pH and viscosity. In 45% and 40% fat cream systems, buttermilk, and especially that with higher phospholipid content, improved the emulsifying properties and acid tolerance of the cream. The components of buttermilk could alter the properties of the surface of fat globules, thereby altering the emulsification properties of the cream. However, neither of the tested buttermilks affected the emulsifying properties and acid tolerance of lower-fat (35% and 30%) cream systems. Emulsifying components exist in proportionately larger amounts in lower-fat creams, which could render the emulsifying properties resistant to change. The number of fat globules may also influence acid-induced changes in viscosity. The addition of phospholipids or lysophospholipids did not improve the acid tolerance of creams, a finding that may be attributable to the formation of complexes of phospholipids and protein. PRACTICAL APPLICATION The findings presented herein demonstrate the ability to improve the acid tolerance of cream using materials derived from milk. Implementing these findings appropriately may result in a high-quality cooking cream.

Reducing effect of calcium in combination with magnesium and lactulose on body fat mass in middle-aged Japanese women

BACKGROUND It has been reported that adequate calcium intake decreases body fat and appropriate intakes of magnesium suppress the development of the metabolic syndrome. Furthermore, lactulose increases the absorption of calcium and magnesium. An optimal combination of calcium, magnesium and lactulose may therefore reduce body fat mass. METHODS An open-label randomized controlled trial was conducted to investigate the body fat-reducing effects of a test food containing 300 mg calcium, 150 mg magnesium, and 4.0 g lactulose. Body composition parameters and blood hormone and urine mineral concentrations were measured at baseline and at 6 and 12 months thereafter. Whole-body fat mass was measured with dual-energy x-ray absorptiometry. RESULTS Seventy-six middle-aged Japanese women (47.5±4.7 years) were randomized to the intake group (n=48) or the non-intake control group (n=28). At 12 months the difference in body fat mass change between the two groups (intake group - control group) was -0.8 kg (95% CI: -1.5 - 0.0 kg, p=0.046), although there were no differences in anthropometric data between the two groups. Body fat percentage at 12 months tended to be lower in the intake group, but the difference was not significant (p=0.09). CONCLUSIONS These findings may suggest that calcium in combination with magnesium and lactulose can reduce body fat mass in middle-aged Japanese women. However, the contribution of magnesium and lactulose are unclear in this study. Further studies are needed to clarify these contributions.

A case of diffuse axonal injury in violent death

A 59-year-old man was carried to the hospital by three men. The deceased was unconscious at admission and his face was severely swollen with many subcutaneous hemorrhages and extensive edema. His death was confirmed 17 min after resuscitation. A judicial autopsy was performed the next day. Findings showed that the victims face and head were reddish and swollen, and that subscalp bleeding was ubiquitous, but no skull fracture, epi- and subdural hematoma or subarachnoidal bleeding was observed. The brain itself was severely edematous but no bleeding was found. Although small hemorrhages were seen in the limbs and back, there were no marked wounds except to the head. To determine the cause of death, we performed a microscopic histochemical examination. Conventional H.E. staining disclosed eosinophilic change, concentration of nuclei, edema, gliosis, and oozing at the corpus callosum. To identify further details of the cause of death, we used Bodian staining, Kluver-Barrera staining, anti-beta amyloid immunostaining, and anti-neurofilament immunostaining. We found sinusoidal swelling of axons and waving axons, which are typical findings of Diffuse Axonal Injury (DAI), but no positive staining of beta amyloid. Focal lesions of the corpus callosum and of the dorsolateral quadrant of the rostral brain stem, and diffuse damage to axons are considered to constitute the DAI triad. We therefore diagnosed the cause of death as DAI. Our experience shows that it is important to use several staining methods for diagnosis of a variety of neuronal degenerative disorders. Several days later, we were informed by the police that several men had hit and kicked the victim in an attempt to lynch him. To compare with this case, we also report two other cases in which DAI was observed.

Misidentification of ethyl chloride in the routine GC-FID analysis for alcohol.

UNLABELLED GC-FID is the method of choice for alcohol screening and quantitative analysis in modern forensic medical practice. Although specific enough for routine use, some results could be misleading. In the current article we present a case of sexual asphyxia with drug and volatile substance abuse. Toxicological analysis revealed the presence of methamphetamine at a concentration of 1.3 microg/mL in blood. An ethanol-like peak was detected during our routine GC-FID test for alcohol (methylethylketone IS). Subsequent GC-MS analysis identified the peak as ethyl chloride. Levels of 0.05 mg/mL in blood and 0.01 mg/mL in urine were measured. Two facts proved misleading in our case. First: very small difference of 0.027 between the ethyl chloride and ethanol peaks in relative retention times at the GC-FID chromatograms. Second: missing evidence for the use of ethyl chloride at the scene-neither cans of the substance were found, nor such information was available otherwise. CONCLUSION there is a substantial risk for mistaking ethyl chloride for ethanol, when ethyl chloride abuse is unanticipated. In the case of slightest uncertainty a GC-MS analysis should be employed to reliably determine the actual substance.

Functional disturbance of naive T lymphocytes in very high IgE producers: depletion of interleukin-4-induced interleukin-4-producing cells.

We examined the capacity of T cells from normal individuals and allergic patients with very high IgE to differentiate into interleukin-4(IL-4)-producing cells in vitro. T cells incubated with anti-CD3 monoclonal antibody plus IL-4 or plus anti-IL-4 antibody in the presence of antigen-presenting cells for 7 days were restimulated and their capacity to express IL-4 mRNA was examined by RT-PCR. In T cells from normal individuals, there was a marked increase in the expression of IL-4 mRNA following the addition of IL-4. After fractionation of normal T cells into naive T (CD45RA+) and memory T (CD45RO+) cells, induction of the increase of IL-4 mRNA was restricted to the naive T cell population. In contrast, in T cells from allergic patients, the stimulation of whole or naive T cells with anti-CD 3 monoclonal antibody in the presence of IL-4 induced much less IL-4 mRNA. These findings suggest the presence of a functional abnormality in IL-4-dependent development of IL-4-producing T cells in the peripheral-blood naive T cells from allergic patients.