Adult Still's disease reflects a Th2 rather than a Th1 cytokine profile

Adult Still's disease (ASD) is a chronic multisystemic disease. Extraordinarily high serum levels of IL-18 in ASD patients have been described, whereas the mechanism remains to be clarified. This study aimed to evaluate proinflammatory cytokines and to consider their pathological roles. In patients with rheumatic diseases (n = 151), blood samples were taken at the active phase and the serum levels of IL-18 and other proinflammatory cytokines were measured by ELISA. The extra-high levels of IL-18 were confirmed selectively in ASD patients (n = 10). In the active phase of ASD patients, the levels of IL-6 were elevated accordingly, but IL-1beta and TNF-alpha were undetectable. As to Th1-Th2 cytokines, the levels of IL-4 and IL-13, but not INF-gamma, IL-12, or IL-2, were elevated in all ASD patients examined. Moreover, the serum levels of IL-18 showed a good correlation with those of IL-4, suggesting that ASD reflects a Th2 rather than a Th1 cytokine profile.     

Retrograde dopaminergic neuron degeneration following intrastriatal proteasome inhibition

Recent studies have suggested that defects in the ubiquitin-proteasome system (UPS) contribute to the etiopathogenetic mechanisms underlying dopaminergic neuronal degeneration in Parkinson's disease. The present study aims to study the effects of proteasome inhibition in the nerve terminals of nigrostriatal dopaminergic neurons in the substantia nigra pars compacta (SNpc). Following a unilaterally intrastriatal injection of lactacystin, a selective proteasome inhibitor, dopaminergic neurons in the ipsilateral SNpc progressively degenerated with alpha-synuclein-immunopositive intracytoplasmic inclusions. When lactacystin was administered at a high concentration, the striatum was simultaneously involved, and alpha-synuclein-immunopositive extracytoplasmic granules appeared extensively within the SN pars reticulata (SNpr). In addition, during the retrograde neuron degeneration in SN, the level of heme oxygenase-1 immunopositivity, an oxidative stress marker, was markedly increased in SNpc neurons. These results reveal that intrastriatal proteasome inhibition sufficiently induces retrograde dopaminergic neuronal degeneration with abundant accumulation of alpha-synuclein in the SN.     

Over-expression of TSC-22 (TGF-beta stimulated clone-22) markedly enhances 5-fluorouracil-induced apoptosis in a human salivary gland cancer cell line

We have recently isolated TSC-22 (transforming growth factor-beta-stimulated clone-22) cDNA as an anticancer, drug-inducible (with vesnarinone) gene in a human salivary gland cancer cell line, TYS. We have also reported that TSC-22 negatively regulates the growth of TYS cells and that down-regulation of TSC-22 in TYS cells plays a major role in salivary gland tumorigenesis (Nakashiro et al, 1998). In this study, we transfected TYS cells with an expression vector encoding the TSC-22-GFP (green fluorescent protein) fusion protein, and we established TSC-22-GFP-expressing TYS cell clones. Next, we examined (a) the subcellular localization of the fusion protein, (b) the sensitivity of the transfectants to several anticancer drugs (5-fluorouracil, cis-diaminedichloroplatinum, peplomycin), and (c) induction of apoptotic cell death in the transfectants by 5-fluorouracil treatment. The TSC-22-GFP fusion protein was clearly localized to the cytoplasm, but not to the nucleus. Over-expression of the TSC-22-GFP fusion protein did not affect cell growth, but significantly increased the sensitivity of the cells to the anticancer drugs (p < 0.01; one-way ANOVA). Furthermore, over-expression of the TSC-22-GFP fusion protein markedly enhanced 5-fluorouracil-induced apoptosis. These findings suggest that over-expression of TSC-22-GFP protein in TYS cells enhances the chemosensitivity of the cells via induction of apoptosis.     

Prefixation treatment with ethidium bromide for high resolution banding analysis of chromosomes from cultured human bone marrow cells

Prefixation treatment of cultured human bone marrow cells with a DNA intercalating agent, ethidium bromide (EBr), induced a dose- and time-related elongation of chromosomes. When compared with EBr-free cultures, a 2.9-fold increase in the yield of early mitotic cells with more than 400 bands per haploid set of chromosomes was achieved by simply adding 10 micrograms/ml of EBr during the last 2 hours of culture. The proportion of early mitotic cells was equal to that obtained in methotrexate synchronized cultures. Fluorescence banding methods using base composition specific agents actinomycin D/DAPI for AT base pairs and chromomycin A3/distamycin A for GC suggested that EBr does not have base specificity, because EBr did not alter the banding patterns of chromosomes obtained with these staining procedures.     

Flow-dependent water permeability of the rabbit descending limb of Henle's loop

Because completely opposite results have been reported on the water permeability of the rabbit descending limbs of Henle's loop (DLH), we rigorously examined water permeability of the upper portion of the descending limb of the rabbit long-looped nephron. Even when the double-cannulation method was used in an attempt to reduce the resistance of tubular outflow, the collected fluid-to-perfusate inulin ratio was equal to or very close to the bathing fluid-to-perfusate osmolality ratio, indicating that osmotic equilibration occurred along the tubule by absorption of water. When perfusion rates were controlled by varying the height of the fluid reservoir connected to the perfusion pipette, osmotic (Pf) as well as diffusional (Pdw) water permeability was shown to be correlated with perfusion rate and/or perfusion pressure. Pf and Pdw at zero perfusion rate as determined from the values of the intercept of regression lines were 253 X 10(-3) and 4.54 X 10(-3) cm X s-1, respectively. The maximal values for Pf and Pdw were 737-1,098 X 10(-3) and 18.3 X 10(-3) cm X s-1, respectively. By changing the resistance to perfusion at the tubular outflow, it was shown that changes in Pf paralleled changes in perfusion rate rather than changes in perfusion pressure. Under stop-flow conditions the luminal fluid volume rapidly decreased after the osmolality of the bathing fluid was increased, suggesting that the segment is highly permeable to water even at zero flow rate. Reflection coefficients for urea and NaCl were 1.01 and 0.82, respectively. These data support the view that this segment is highly permeable to water and that increases in osmolality along the DLH in vivo may be accounted for mainly by abstraction of water rather than addition of solutes.     

Effect of a spider toxin (JSTX) on excitatory postsynaptic current at neuromuscular synapse of spiny lobster

1. We studied the blocking properties of a spider (Nephila clavata) toxin (JSTX) purified from venom on the spiny lobster neuromuscular junction. 2. When a small amount of JSTX was applied to the neuromuscular junction, the excitatory postsynaptic potential (EPSP) was partially suppressed. The amplitude of EPSPs remained at a steady level for several hours during the washing of the preparation, showing that the action of JSTX is irreversible. 3. We recorded the excitatory postsynaptic current (EPSC) from synaptic site using a macro-patch electrode. The amplitude of EPSC increased linearly with hyperpolarization of the membrane potential in the presence and absence of JSTX. 4. The decay phase time constant of EPSC and spontaneous EPSC was decreased by hyperpolarizing the membrane potential both in the absence and in the presence of JSTX. The relationship between the decay time constant and the membrane potential was not modified by JSTX. 5. It is suggested that JSTX irreversibly blocks EPSC by acting on the site that is apart from the ionic channel of the glutamate receptor molecule.     

Age- and sex-related changes in toluene metabolism by rat hepatic microsomes in vitro

Age- and sex-related changes in toluene metabolism by hepatic microsomes of male and female Sprague-Dawley (SD) rats (1 to 20 weeks) were investigated. A major metabolite of toluene, benzyl alcohol (BA), was measured by high-performance liquid chromatography (HPLC). At low substrate toluene concentrations (0.4 mM), in male rats, BA increased dramatically with development, reaching a peak at 5 weeks of age, rapidly decreasing thereafter. In female rats, BA increased dramatically with development at 3 to 5 weeks of age, and then declined gradually to a low level. Gender differences were obtained at 5 and 20 weeks of age, with BA products being higher in males than in females. At high substrate toluene concentrations (5.0 mM), in male rats, the BA formation pattern was similar to that at the low substrate concentration, although the rate of increase with age was slower. In female rats, a peak was obtained at 3 weeks of age, and then declined gradually to a low level. Gender differences were obtained at 5, 15 and 20 weeks of age, with BA products being higher in males than in females. These results indicate that toluene metabolism exhibits age and gender differences.