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To assess the use of plasma free amino acids (PFAAs) as biomarkers for metabolic disorders, it is essential to identify genetic factors that influence PFAA concentrations. PFAA concentrations were absolutely quantified by liquid chromatography–mass spectrometry using plasma samples from 1338 Japanese individuals, and genome-wide quantitative trait locus (QTL) analysis was performed for the concentrations of 21 PFAAs. We next conducted a conditional QTL analysis using the concentration of each PFAA adjusted by the other 20 PFAAs as covariates to elucidate genetic determinants that influence PFAA concentrations. We identified eight genes that showed a significant association with PFAA concentrations, of which two, SLC7A2 and PKD1L2, were identified. SLC7A2 was associated with the plasma levels of arginine and ornithine, and PKD1L2 with the level of glycine. The significant associations of these two genes were revealed in the conditional QTL analysis, but a significant association between serine and the CPS1 gene disappeared when glycine was used as a covariate. We demonstrated that conditional QTL analysis is useful for determining the metabolic pathways predominantly used for PFAA metabolism. Our findings will help elucidate the physiological roles of genetic components that control the metabolism of amino acids.

Background: Although lung transplantation from donation after cardiac death (DCD), especially uncontrolled DCD, is limited by warm ischemic periods, the molecular mechanism of warm ischemia-reperfusion-injury (IRI) has not been well elucidated. The purpose of this study was to clarify the particular longitudinal mechanisms of molecular factors involved in warm IRI. Methods: Cold ischemic-time (CIT)-group lungs were retrieved and subjected to 3-h of cold preservation, whereas warm ischemic-time (WIT)-group lungs were retrieved after 3-h of warm ischemia. Orthotopic rat lung transplantation was performed and the grafts were reperfused for 1 or 4-h. The graft functions, gene expression, and activation of inflammatory molecules in the grafts were analyzed. Exhaled-carbon-monoxide-concentration (ExCO-C) was measured during reperfusion. Results: Only the WIT-group showed obvious primary graft dysfunction at 1-h reperfusion, but the graft function was recovered during 4-h reperfusion. Most of pro-inflammatory cytokines and stress-induced molecules showed different expression and activation patterns between CIT and WIT groups. In the WIT-group, the expressions of anti-inflammatory molecules, IL-10 and HO-1, were significantly increased at 1-h reperfusion compared to the CIT-group, and these high levels were maintained through 4-h reperfusion. Furthermore, ExCO-C levels in the WIT-group increased immediately after reperfusion compared to the CIT-group. Conclusions: This study indicates that warm IRI may involve a different mechanism than cold IRI and anti-inflammatory pathways may play important roles in the graft recovery after lung transplantation from uncontrolled DCD.

Objectives Accepting organs donated after cardiac death (DCD) is an effective approach to the donor shortage. However, lung transplantations from DCD donors show severe rapid pulmonary graft dysfunction (PGD) followed by warm ischemia–reperfusion injury (IRI). This study sought to clarify the molecular mediators in warm IRI, including activation of mitogen-activated protein kinase (MAPK) and the downstream cascades. Methods We performed single left lung transplantation using organs from male Sprague-Dawley rats after 0 (CIT group), 30 (30WIT group), or 180 (180WIT group) minutes of warm ischemia time. Pulmonary graft functions were estimated by blood gas analysis. At 1 hour after reperfusion, the phosphorylation status of MAPKs (ERK, p38, and JNK) and the gene expression levels of transcription factors (Egr-1 and ATF-3) and immune mediators (MCP-1, MIP-2, PAI-1, ICAM-1, TNF-α, IL-1β, IL-6, and COX-2) in the grafts were examined using Western blotting and real-time polymerase chain reaction assays. Results Severe PGD was observed in the 180WIT group compared with transplanted lungs in the other groups, which exhibited good pulmonary graft function. ERK and JNK activations, as well as mRNA levels of transcription factors (Egr-1 and ATF3) significantly increased with greater warm ischemic times. The pattern of JNK activation correlated with the severity of PGD. MCP-1, ICAM-1, IL-1β, IL-6, and COX-2 were also up-regulated among the 180WIT group, although MIP-2 and PAI-1 showed no significant differences among the groups. Conclusions We suggest that the ERK and JNK pathways may play important roles to induce the injury caused by prolonged warm ischemia followed by reperfusion in the setting of lung transplantation from DCD donors.

The electrostatic rotary atomizer has the highest paint efficiency in all kind of atomizers. The usage of an electrostatic rotary atomizer, therefore, has contributed to reduce the waste of paint and volatile organic compounds (VOCs). Concurrently, water-borne painting processes which use water-soluble paint have been introduced to reduce the amount of VOCs emissions. With the water-borne painting, the atomizer body is easily contaminated by the paint mists. The contamination has been an important problem to be solved in factories. In this report, the causes of water-borne paint contamination was considered and the experimental results of the contamination proof system were presented. The atomizer is surrounded by the repelling film that is charged and repels the incoming paint droplets which otherwise contaminate the atomizer. From the experimental results, following electrical properties are found to be important for the repelling film such as low capacitance and high insulation property to keep high surface potential. These features were also checked by the space charge measurements inside the repelling film. In addition, safety against incendiary sparking was investigated. The ground electrode of sphere, needle or blush-shape was approached to contact to the charged film. Energy of the discharge was calculated from the measured discharge current and the surface potential of the film. The discharge energy of all measured electrodes was below the minimum ignition energy of 0.24 mJ used in the painting industries. At the meantime, the results indicate that shape of the electrode is an important parameter to affect the discharge energy.

BACKGROUND Early growth response-1 (Egr-1) has been shown to be a trigger-switch transcription factor that is involved in lung ischemia-reperfusion injury (IRI). METHODS Mouse lung transplants were performed in wild-type (WT) C57BL/6 and Egr1-knockout (KO) mice in the following donor → recipient combinations: WT → WT, KO → WT, WT → KO, and KO → KO to determine whether the presence of Egr-1 in the donor or recipient is the most critical factor for IRI. Pulmonary grafts were retrieved after 18 hours of ischemia after 4 hours of reperfusion. We analyzed graft function by analyzing arterial blood gas and histology in each combination and assessed the effects of Egr1 depletion on inflammatory cytokines that are regulated by Egr-1 as well on polymorphonuclear neutrophil (PMN) infiltration. RESULTS Deletion of Egr1 improved pulmonary graft function in the following order of donor → recipient combinations: WT → WT < WT → KO < KO → WT < KO → KO. Polymerase chain reaction assays for Il1B, Il6, Mcp1, Mip2, Icam1, and Cox2 showed significantly lower expression levels in the KO → KO group than in the other groups. Immunohistochemistry demonstrated clear Egr-1 expression in the nuclei of pulmonary artery endothelial cells and PMN cytoplasm in the WT grafts. Flow cytometry analysis showed that Egr1 deletion reduced PMN infiltration and that the extent of reduction correlated with graft function. CONCLUSIONS Both graft and recipient Egr-1 played a role in lung IRI, but the graft side contributed more to this phenomenon through regulation of PMN infiltration. Donor Egr-1 expression in pulmonary artery endothelial cells may play an important role in PMN infiltration, which results in IRI after lung transplantation.

The inhibitory potency on urease, urinary excretion in rats and mutagenicity of scores of substituted hippuro- and N-acylglycino-hydroxamic acid newly synthesized, were examined. Some of these compounds were powerful inhibitors against bacterial intracellular urease, but hippuro-derivatives were mutagenic. Among them, mmethoxyhippuro- and 2,2-dimethylpropionylglycino-hydroxamic acids appeared to be promising because of their high urinary excretion after oral administration to rats. They prevented bladder stone formation as well as alkalinization of urine in vivo, when they were orally administered to rats with urinary tract infection caused by Proteus mirabilis.

Background: A study was performed to compare the effects of immunization with ragweed pollen (RW) in two different adjuvants on the characteristics of a previously described model of experimental immune-mediated blepharoconjunctivitis (EC) in rats. Methods: Lewis or Brown Norway (BN) rats were immunized with 100 µg of RW in emulsion with aluminum hydroxide [Al(OH)3] or complete Freund’s adjuvant (CFA). Three weeks later, the animals were challenged with eye drops containing RW in PBS. Twenty-four hours after topical challenge, eyes, blood, and lymph nodes were obtained for histology, measurement of antigen-specific antibodies, and proliferation or cytokine assays, respectively. In addition to active immunization, recipients of RW-primed lymph node cells were challenged and evaluated as above. Results: RW in both adjuvants induced infiltration with predominantly mononuclear cells in Lewis rats and eosinophils in BN rats. As well as active immunization, eosinophils were detected only in BN rats by adoptive transfer of cells. Lymphocyte proliferative responses to RW were high in immunized Lewis rats when CFA was used as an adjuvant. In contrast, proliferative responses in BN rats were higher when Al(OH)3 was used. RW-specific IgE was detected only in BN rats. There were no significant differences in RW-specific IgG1/ IgG2a ratio among the four groups. Lewis rats had higher level of RW-specific interferon-γ in the culture supernatant. Conclusions: The characteristics of EC are different in Lewis and BN rats, dependent on the genetic background of the rat strains. The response to RW was similar to other previously used antigens, such as ovalbumin.

· Background: Fischer rats were less susceptible to experimental autoimmune uveoretinits (EAU) than Lewis rats, although both strains have the same MHC molecules. The purpose of this study was to compare the susceptibility of experimental allergic/immune-mediated blepharoconjunctivitis (EAC) between these two strains. · Methods: Male Lewis and Fischer rats were immunized with either ovalbumin (OVA) or OVA peptide (OVA323–339) in complete Freund’s adjuvant (CFA). Three weeks later, they were challenged with OVA by eye drops. Twenty-four hours later, after clinical evaluation, they were killed and eyes, blood and lymph nodes were harvested for histology, antibody titer and proliferation assay, cytokine production or flow cytometric analysis, respectively. · Results: Fischer rats developed mild EAC compared with Lewis rats. Cellular proliferative responses, IFN-γ production of culture supernatant and serum IgG specific for OVA were basically the same between the two strains. The same OVA peptides were selected as immunodominant. Flow cytometric analysis demonstrated the same cellular profile of lymph node cells in the two strains. · Conclusion: EAC in Fischer rats was milder than that in Lewis rats, although no apparent differences in immunological parameters between these two strains were detected. These data suggest that factors unrelated to immunological parameters may depend on the susceptibility of EAC.

Purpose. We recently reported the essential role of cellular immunity on the induction of experimental immune-mediated blepharoconjunctivitis (EC, formerly EAC) by using ovalbumin (OVA) as a model antigen in Lewis rats. The purpose of this study was to investigate the possible induction of EC by immunization with an OVA peptide (OVA 323–339).Methods. Lewis rats were immunized with various doses of OVA or OVA 323–339 in complete Freund's adjuvant. Three weeks later they were challenged with OVA or OVA 323–339 by eye drops; 24 h after challenge, eyes including lids, lymph nodes and blood were harvested after clinical evaluation. An OVA 323–339-specific cell line (S816) was established by periodical stimulation with this peptide. Pathogenicity of S816 was tested by adoptive transfer of S816 into syngeneic recipient rats after challenge with OVA or OVA 323–339.Results. All rats immunized with OVA 323–339 developed EC after challenge with OVA or OVA 323–339. Rats immunized with OVA 323–339 at doses as low as 0...

We report for the first time the frequency distributions of HLA-DRB1 and -DQB1 genes in 55 patients with testicular germ cell carcinoma (TGC) using the modified PCR-RFLP method and compare the results with those for 1216 healthy Japanese control subjects. The modified PCR-RFLP method produced accurate, reproducible cleavage patterns that are easily discriminated. HLA-DRB1*0410 was the susceptibility allele (RR = 3.26, P = 0.006) and DQB1*0602 appears to be a candidate protective allele (RR = 0.26, P = 0.02) for TGC in the Japanese. None of the HLA-DRB1 and -DQB1 alleles showed a specific tendency for histological type or clinical stage of the tumours. Previous studies based on serotyping methods failed to show these allelic associations. High-resolution genotyping is essential because the peptide-binding domain of MHC class II molecules is determined more precisely by their genotypes than by their serotypes. In addition, inherent technical difficulties and typing errors of up to 25% make serotyping inefficient. Our results suggest that high-resolution genotyping is a useful genetic marker to determine risk for TGC. Images Figure 1

Non-MHC-restricted rejection mechanisms against the murine MHC-negative F9 embryonal carcinoma cells were analyzed. Strains of C57BL/6 (B6) background were resistant to the tumors irrespective of H-2 haplotypes, while others, including BALB/c background, were susceptible. This resistance was suggested to be mediated primarily by the host thymus-dependent alphabeta T cells, since both athymic B6 nude and normal B6 mice depleted with alphabeta T cells showed susceptible phenotype. The difference of the nature of alphabeta T cells infiltrating in H-2-identical B6- and BALB.B-derived tumors was then comparatively analyzed. It was revealed that unique T cells with NK1+ CD4- CD8- (double negative (DN)) alphabeta TCR+ phenotype were accumulated significantly in B6, but few in BALB.B mice. The population freshly isolated from the F9 tumor tissues preferentially expressed potent IL-4 mRNA, and was suggested to be mostly responsible for the endogenous IL-4 production. Indeed, the injection of either anti-NK1.1 or anti-IL-4-neutralizing Ab into the normal B6 rendered them significantly susceptible to the tumor cells. These results strongly suggested that NK1+ DN alphabeta T cells were responsible primarily for the rejection mechanisms against F9 tumors. Histologically, F9 tumors in B6 mice were characterized by abundant macrophage infiltration and massive tumor necrosis, neither of which was observed in those in BALB.B nor B6 mice preinjected with anti-IL-4 Ab, indicating that both histologic features in the resistant strain were dependent on the endogenous IL-4. Present results provide one of the first instances in which a recently emerging minor T cell subpopulation, thymus-dependent NK1+ DN alphabeta T cells, plays an essential role in anti-tumor responses in vivo.

PURPOSE We assessed the impact of different treatment modalities on sexuality and fertility in long-term survivors of testicular cancer. MATERIALS AND METHODS The sample consisted of 85 testicular cancer patients, of whom 19 had undergone chemotherapy with retroperitoneal lymph node dissection (RPLND), 15 had received chemotherapy only, 42 had received infradiaphragmatic radiotherapy, and nine had received surveillance therapy. The questionnaire reported sexual function, marital status, and issues related to fertility and childbearing. RESULTS One fourth to one half reported some type of sexual impairment in each group. The only significant difference was that approximately 70% of men with RPLND reported inability of ejaculation and a greater decline in semen volume, which is expected. The most striking finding is that the rates and nature of sexual dysfunction of surveillance patients were similar to other treatment groups, except for ejaculatory function. The highest rates of infertility distress were observed in chemotherapy patients. CONCLUSION These data suggest that sexual dysfunction and infertility represent the major persisting side effects, even years after diagnosis. The hypothesis that surveillance patients have fewer sexual problems is not upheld in this study.

We examined the effect of staphylococccal enterotoxin B (SEB)-induced anergy on expression of six different cytokine genes in T cells restimulated with SEB in vitro. We found that although IL-2, IL-3, and IL-4 mRNA levels are substantially reduced in anergic T cells, mRNAs for IL-6, IL-10, IFN-gamma, and TNF-alpha are expressed normally. Thus, there appeared both anergy-sensitive and resistant cytokine mRNA expression in restimulated anergic T cells. The same pattern of cytokine mRNA responses was observed in anergic CD4+ T cells, indicating that the preferential induction of anergy in Th1-like cells is not evident in this in vivo model. Employing TCR V beta 8.2 transgenic mice in which almost all T cells become anergic, we found that the TCR/CD3 complex can transduce both anergy-sensitive and resistant signals. Furthermore, a series of experiments using FK506, A23187, and PMA suggests that signals between TCR and activation of calcineurin and protein kinase C may be blocked in anergic T cells. This is supported by our gel mobility shift assays indicating that calcineurin and/or PMA-inducible NF-ATp, OAP40, and AP-1, but not calcineurin-independent Oct-2, are repressed in anergic spleen T cells upon restimulation with SEB. Taken together, these results suggest that, among signals elicited by stimulation of TCR with SEB, a Ca2+/calcineurin-NF-ATp pathway and other signals, including protein kinase C, are repressed in anergic T cells upstream of their activation, which are essential for the cytokine mRNA expression of the anergy-sensitive type but are dispensible for those of the anergy-resistant type.

In an effort to study the pathophysiological events in the development of insulitis in NOD mice, we have developed ILI- and NOD-nu/nu mice. ILI mice are a nondiabetic inbred strain but are derived from the same JcI:ICR mouse as NOD mice and share the same H-2 allotype with NOD mice. Splenocytes and CD4+ cells from diabetic NOD mice appeared to transfer insulitis to ILI-nu/nu mice, suggesting that ILI mice already express autoantigen(s) responsible for insulitis. But reciprocal thymic grafts from NOD mice into ILI-nu/ nu mice and those from ILI mice into NOD-nu/nu mice failed to allow the development of insulitis, implying that ILI mice possess neither precursor T cells nor the thymic environment responsible for the development of insulitis. In addition, splenocytes from ILI mice appeared to contain regulatory cells which suppress the development of diabetes but not that of insulitis in NOD mice. The use of these nude mice should provide more information on the products of insulitis-susceptibility genes of NOD mice.

Protection against infection with intracellular pathogens operates in two stages, early innate resistance and late acquired protective immunity (API), in inbred mouse strains. Although both C57BL/10 (B10) and BALB/c mice bear the susceptible phenotype of innate resistance, Calmette-Guérin bacillus (BCG) vaccination generated efficient API in B10 but not in BALB/c mice. Employing a specific nitric oxide (NO) synthase inhibitor, we revealed that NO production plays a pivotal role in the API of B10 mice. Consistent with this, expressions of the inducible isoform of NO synthase (iNOS) protein and mRNA were significantly higher in the spleen of B10 mice than in that of BALB/c mice. Furthermore, IFN-gamma, a potent inducer of iNOS, and mRNAs for IL-12 (p40); an inducer of IFN-gamma and IL-2 were also vigorously expressed in the spleen of B10 mice compared with that of BALB/c mice. In an attempt to clarify the mechanism by which the different capacities for API are generated, we analyzed the cytokine network between T cells and macrophages in both B10 and BALB/c mice. We found that multiple functions of macrophages, which include capacities to express IL-12 (p40) mRNA in response to BCG and to express mRNAs for iNOS and IL-12 (p40) in response to IFN-gamma, were impaired in BALB/c mice as compared with B10 mice. However, T cells appeared to express comparable level of IFN-gamma mRNA in both strains when stimulated with IL-12. Taken together, these results indicate that the macrophage functions play a pivotal role in both the induction and effector phases of API to determine the susceptibility of mice to BCG infection.

We investigated the cis-acting sequences that function in the B-cell-specific expression of the HLA-DPB1 gene. Class II B major histocompatibility genes contain a conserved upstream sequence that is important in the expression of these genes. This region has been divided into three major elements, the W, X, and Y boxes. In this paper, we identified an additional positive regulatory element upstream from the DPB1 W box. Using 5' deletion mutants and a substitution mutant, we mapped a positive element, called the W' box, between -184 approximately -169 bp. Sequence comparison revealed that the W' box shares homology with the W box. Electrophoretic mobility shift assay confirmed that the W box binds proteins that also recognize the W' box. Furthermore, deletion and substitution mutants indicate that the W and W' boxes effectively enhance CAT activities only when the X and Y boxes exist.

Five patients with unilateral leg lymph edema had intraarterial injections of lymphocytes in the affected leg with consequent improvement in 3. To assess the degree of lymph edema, T2 and intensity ratio between the 2 legs in STIR images were used. Mean and SD of T2 values in the subcutaneous tissue were measured using a triple echo sequence and found larger on the edematous side than in the opposite extremity. The mean T2 shortening obtained after the lymphocyte therapy correlated well with the reduction of limb circumference. Mean T2 reflects the fluid content, and SD of T2 the fluid distribution in the subcutaneous tissue. The degree of lymph edema and the effect of therapy can be evaluated quantitatively by measuring the value of the T2 relaxation time.

Taking advantage of the reverse transcriptase-polymerase chain reaction (RT-PCR), we have analyzed T cell receptor gamma-chain mRNA of synovial fluid gamma/delta T cells from patients with rheumatoid arthritis (RA) in comparison with those of peripheral blood mononuclear cells (PBMC) from RA patients and healthy individuals. The quantitative RT-PCR method in conjunction with nucleotide sequencing revealed the frequent usage of the V gamma 3 gene segment in RA synovial fluid mononuclear cells (SFMC) (p < 0.01) which in PBMC of healthy individuals occurred rarely. PBMC of most healthy individuals expressed the V gamma 9 gene predominantly (p < 0.01) as expected. However, only half of RA patients showed elevated levels of the V gamma 9 gene expression in their PBMC. The gamma-chain mRNA containing the V gamma 3 gene in RA SFMC showed no conserved junctional sequence (complementarity-determining region 3). To investigate the nature of ligands recognized by the V gamma 3-bearing T cells, we analyzed V gamma gene usage of RA SFMC, RA PBMC, and normal PBMC stimulated with Mycobacterium tuberculosis (MT) or MT plus interleukin-2 since there is mounting evidence of high reactivity of RA SFMC to MT and mycobacterial heat-shock protein 65. However, the V gamma usage appeared to be mostly V gamma 9 in RA SFMC, RA PBMC and normal PBMC. Taken together these results suggest that an as yet unknown antigen(s) (other than MT) might select gamma/delta T cells expressing the V gamma 3 gene in RA SFMC.

Interleukin (IL)-12 is a logical candidate for participation in the differentiation of T helper (Th) 1 cells. IL-12 is produced by macrophages and B cells and induces the production of interferon (IFN)-gamma from Th1 cells and NK cells. In this study, we show that the IFN-gamma itself is capable of inducing IL-12 mRNA expression in murine macrophage cell line J774. The mRNA was apparent by 12h after IFN-gamma treatment, although maximal induction required 36 to 48h. Furthermore, we investigated the signal transduction mechanism responsible for the mRNA expression. The IFN-gamma-inducible IL-12 mRNA was blocked by two protein tyrosine kinase (PTK) inhibitors, genistein and herbimycin A, suggesting that PTK is involved in the IFN-gamma-inducible IL-12 mRNA expression.

Ischemia reperfusion injury (IRI) remains a significant cause of morbidity and mortality after lung transplantation. Early growth response-1 (EGR1) drives the expression of inflammatory mediators and has an important role in IRI. We hypothesized that the severe IRI caused by a long preservation induces a specific expression pattern of EGR1 and its target genes which would correlate with the lung graft function.SD rat lungs were preserved at 4 °C for 3 or 18 h, then transplanted and reperfused. Pulmonary grafts were evaluated for the blood gas oxygenation and pathological findings. The intra-graft mRNA levels of EGR1 and its downstream target genes were measured by real-time PCR. A Western blotting analysis of the EGR1 expression was used to validate the changes in the protein level.There was upregulation of EGR1, MIP-2 and PAI-1 when there was prolonged hypothermic preservation. The expression levels of MIP-2 and PAI-1 were observed to increase for up to 4 h in the 18 h preserved lungs. There were no differences in the expression levels of IL-1β and ICAM-1 between the lungs subjected to short and long periods of ischemia.Our data showed that prolonged hypothermic graft preservation deteriorates the pulmonary graft function, which was associated with the induction of EGR1 and its downstream target genes, which may aggravate IRI following lung transplantation.

We previously reported that detection of telomerase activity in urinary exfoliated cells obtained from urothelial cancer patients by telomeric repeat amplification protocol (TRAP) assay is a more sensitive method of diagnosis than conventional urine cytologic examination, particularly in patients with grade 1 tumors. To establish this method as a noninvasive screening test for the diagnosis of urothelial cancers, we performed the semi-quantitative analysis of telomerase activity using Telomerase PCR ELISA™. Spontaneous voided urine samples were obtained from 65 urothelial and 58 non-urothelial cancer patients. When the mean + 2 standard deviation of telomerase activity in urine sediments of non-urothelial cancer patients was arbitrarily determined as a cut-off, the sensitivity of TRAP enzyme-linked immunosorbent assay (ELISA) and the conventional cytology were 57% and 35%, respectively. Detection rate was significantly higher in semi-quantitative TRAP assay than in conventional cytologic examination in grade 1 cancer patients (52% vs. 5%, p = 0.00195). False positives were detected in 5% of non-urothelial cancer patients without pyuria and in 11% of non-urothelial cancer patients with pyuria (p = 0.395). Telomerase activity was enhanced in some cases after phenol extraction or extracting epithelial cells by using Dynabeads of macroscopic hematuria and pyuria, indicating that hematuria and pyuria might contribute to false negatives. In conclusion, the TRAP-ELISA method is superior to the standard TRAP assay in quantitativeness and simplicity of the experimental procedure. Detection of telomerase activity in urine sediments is particularly useful for the diagnosis of low-grade tumors. However, telomerase activity in patients with high grade tumors often might be underestimated due to the excessive amount of exfoliated epithelia with necrotic tissues, hematuria, and pyuria.

Of a 5 km-long steel bridge constituting a vital part of the projected Trans-Tokyo Bay Highway, a 1,630 m-long section of 10-span continuous box girder construction containing a main span 240 m in length and 41 m in elevation above the water was analyzed for aerodynamic stability. Wind tunnel testing with a coil-spring-supported two-dimensional model of the box girder cross section showed the girder undergoing galloping and vortex-induced oscillations in the heaving mode in either instance, as predicted by computer simulation. Then, for quantitative evaluation of these oscillations, a three-dimensional aeroelastic model representing the entire 10-span bridge section was tested in the wind tunnel. Unlike the case with the two-dimensional sectional model, only the heaving mode of vortex-induced oscillation, without the galloping vibration, was found to take place. Evidently the adjoining bridge spans abutting the main span, being more slender in girder cross section profile than the latter, served to restrain the vibration-promoting aerodynamic force of the main span. It therefore was decided that addressing only the vortex-induced oscillation would suffice in designing the bridge structure adequately for aerodynamic stability.

Brown Norway Katholiek rats, which have very low levels of plasma kininogens, excreted a much smaller amount of kinin in the urine than normal rats of the same strain. The systolic blood pressure of 7-week-old kininogen-deficient rats fed low (0.3%) NaCl diets (131 +/- 4 mm Hg, n = 12) was not different from that in normal rats. Two percent NaCl diets given from 7 weeks of age for 4 weeks caused rapid increases in blood pressure (167 +/- 4 mm Hg, n = 12, 9 weeks old) in deficient rats, although the same diets induced no blood pressure increase in normal rats. Urinary excretion of active kallikrein and prokallikrein remained constant in both rat groups throughout NaCl loading. During this period, the deficient rats secreted less urine (9 weeks old, P < .05) and less urinary sodium (11 weeks old, P < .05). Serum levels of sodium in deficient rats were higher (P < .05) than in normal rats at 9 weeks of age. Intracellular concentrations of sodium in the erythrocytes of deficient rats were higher (P < .05) than in normal rats throughout NaCl loading. Subcutaneous infusion of bovine low molecular weight kininogen with an osmotic pump in NaCl-loaded deficient rats induced a reduction (P < .01) in blood pressure and increases (P < .05) in urine volume and urinary sodium and kinin levels. By contrast, subcutaneous infusion of the bradykinin antagonist Hoe 140 or of aprotinin in NaCl-loaded normal rats induced a hypertensive response. This antagonist treatment reduced urine volume and urinary sodium. These results indicate that the lack of kinin generation observed in the kininogen-deficient rats was related through sodium retention to the hypertensive response to NaCl loading.

BACKGROUND: Morphologic grading of malignancy is considered to be of prognostic value in patients with transitional cell carcinomas of the urinary bladder (TCC). This qualitative approach is, however, associated with low reproducibility. Grading of malignancy can be carried out on a reproducible, quantitative scale.METHODS: A retrospective, prognostic study of 110 patients treated for TCC in clinical Stages Ta-T4 (median follow-up time, 6 years) was performed, evaluating various grading techniques. Unbiased estimates of the volume-weighted mean nuclear volume (nuclear vV), nuclear volume fraction, estimates of nuclear mean profile area (aH(nuc)), nuclear profile density index (NI), and mitotic profile density index (MI) were obtained by stereologic and morphometric techniques.RESULTS: The T-stage and morphologic grade of malignancy were closely cross-correlated (+0.63 0.26). Multiple hazards regression analysis (Cox models) revealed that clinical stage of disease was the sole independent prognostic variable. Only estimates of nuclear vV added significant independent prognostic prediction with regard to recurrence-free survival in the 48 patients with Ta tumors (2P = 0.03).CONCLUSIONS: The results suggested that estimates of nuclear vV are prognostically superior to morphologic grading of malignancy in noninvasive TCC, whereas both morphologically and quantitatively based malignancy grading are without prognostic value in invasive TCC.

A rare example of a benign glandular schwannoma is reported. Further attention is drawn to this unusual subtype of schwannoma and the clinical parameters of the reported cases are compared with those of other types of schwannomas. The tumors tend to be small and superficial, and by contrast with malignant glandular schwannomas, the benign variant does not appear to be associated with von Recklinghausen neurofibromatosis.

We have previously demonstrated that HLA-DR molecule expression induced by IFN-gamma is associated with phosphatidylinositide turnover, activation of protein kinase C, and elevation of intracellular calcium. Because phosphorylation of phospholipase C-gamma 1 on tyrosine residues is known to be involved in the activation of phosphatidylinositide turnover, we investigated the role of tyrosine protein kinase (TPK) in the signal transduction for IFN-gamma-inducible DR molecule expression on T98G cells. The effects of three specific TPK inhibitors, genistein, herbimycin A, and tyrphostin, suggest that TPK is involved in the signal transduction. These inhibitors inhibited the IFN-gamma-inducible DR molecule expression in a dose-dependent manner. Being consistent with this, immunoblotting with an anti-phosphotyrosine mAb revealed that IFN-gamma induces a rapid increase in protein tyrosine phosphorylation. Genistein not only abrogated the IFN-gamma-induced enhancement of tyrosine phosphorylation, but also inhibited the IFN-gamma-induced production of inositol-4-5-triphosphate and the elevation of intracellular calcium. However, these three TPK inhibitors failed to inhibit the DR molecule expression induced by PMA and A23187. These findings suggest that the tyrosine phosphorylation is an early and critical event that precedes phosphatidylinositide turnover leading to activation of protein kinase C and elevation of intracellular calcium concentration during IFN-gamma-inducible DR molecule expression.

Laparoscopic removal of the adrenal gland was performed in 13 patients with primary aldosteronism, Cushing's syndrome or a non-functioning adenoma. Six operations were performed on the right side while the other seven were performed on the left side. Four or five trocars were inserted through the abdominal wall and the adrenal gland was exposed transperito-neally. Eleven patients underwent successful removal of the gland laparo-scopically, while two patients required open surgery as a result of bleeding or severe adhesion due to previous abdominal surgery. The time required for the operation was 161 to 415 min with an average of 297 min and blood loss during the operation was less than 60 ml in eight of the 11 successful patients. Eight of the 11 patients could walk within 48 h post-operatively. Laparoscopic procedures are ideal for adrenalectomy and advancement of the surgical technique will result in shorter operation times and lower morbidity rates.

Gastrointestinal toxicity and various infections are serious problems associated with high-dose chemotherapy. Antibacterial chemoprophylaxis reduces the incidence of gram-negative bacterial infection; however, it may affect the normal intestinal flora and induce drug resistance in organisms. We evaluated the chronological changes in fecal bacteria and organic acids in 6 patients undergoing autologous stem cell transplantation with quinolone-based chemoprophylaxis. All patients developed grade 2-3 diarrhea. Four patients developed grade 3 febrile neutropenia. The total count of obligatory anaerobic bacteria was significantly decreased on Day 7, but total facultative anaerobic bacterial count did not change throughout transplantation. However, Enterobacteriaceae and Lactobacillus were decreased on Day 7 and Staphylococcus was increased after transplantation. Total organic acid concentration and short-chain fatty acids were decreased on Day 7. The bacterial flora and organic acids in the gut were significantly altered in patients who underwent autologous stem cell transplantation with quinolonebased chemoprophylaxis. These changes may contribute to gastrointestinal toxicity and infections.

A 64 year-old woman with a diagnosis of lung adenocarcinoma was admitted to our neurosurgical division in February, 2007, suffering from severe headache and dizziness. Systemic chemotherapy had been repeated for multiple metastases to the bone and cerebral cortex since 7 months before. Reexamination with MRI revealed mild hydrocephalus without cortical metastasis. Cytological analysis of the cerebrospinal fluid (CSF) provided the diagnosis of leptomeningeal metastasis. Removal of 8 ml of CSF dramatically alleviated the patient's symptoms. To improve the quality of her remaining life, she underwent lumboperitoneal (L-P) shunt using a Strata adjustable pressure valve. Severe headache disappeared and other symptoms gradually improved after the operation. She survived for 10 months after the shunt placement, perticipating in family life for 6 months. Pressure level of the Strata valve was changed twice according to the degree of hydrocephalus and functioned well while her life lasted. The present case showed that L-P shunt is one of the effective palliative procedures and an adjustable pressure valve is available for this kind of CSF shunt in patients with leptomeningeal metastasis.

Background Lung grafts can be perfused ex vivo for 2 hours without edema formation; however, prolonged ex vivo lung perfusion (EVLP) eventually induces lung injury. This study evaluated the change in proinflammatory cytokines of the perfusate during EVLP and investigated the effect of cytokine removal using an adsorbent membrane. Methods Porcine heart-lung blocks were harvested after electrically induced cardiac arrest and underwent 12-hour EVLP with an adsorbent membrane (membrane group: n=5) and without an adsorbent membrane (control group: n=6). Results In the control group, both tumor necrosis factor-α and interleukin 8 levels were elevated in the perfusate 2 hours after perfusion. Although tumor necrosis factor-α and interleukin 8 levels were significantly lower in the membrane group than in the control group during the EVLP period, there was no significant difference in oxygenation, pulmonary vascular resistance, edema formation, or myeloperoxidase activity between the two groups. Conclusions Tumor necrosis factor-α and interleukin 8 levels of the perfusate were elevated during EVLP. Although adverse effects of these inflammatory cytokines were anticipated, removal of inflammatory cytokines by the adsorbent membrane did not improve lung function during prolonged EVLP. Factors other than the cytokines may play a major role in causing lung injury during EVLP. Further research is needed to investigate the real mechanism of lung graft injury during prolonged EVLP and to establish longer EVLP duration for graft treatment. This strategy could contribute to the salvage of potentially damaged lungs, especially from cardiac death donors, and to expansion of the donor pool.