A 2,7-diamino-1,4,8-triazanaphthalene derivative selectively binds to cytosine bulge DNA only at a weakly acidic pH.

The synthesis and properties of 2,7-diamino-1,4,8-triazanaphthalene (azaDANP) are described. AzaDANP is protonated only at a weakly acidic pH to bind to the cytosine bulge DNA duplex selectively. Upon binding of azaDANP to the cytosine bulge DNA, a new absorption band at 407 nm appears, and the absorption change of azaDANP on binding to the target is very sensitive to environmental pH with a bell-shaped pH-absorption profile.

Follistatin is an early player in rainbow trout ovarian differentiation and is both colocalized with aromatase and regulated by the wnt pathway.

In mammals, follistatin (FST) plays an important role in early ovarian differentiation, acting downstream of the Wnt pathway. In teleost fish, fst is implicated in folliculogenesis and oocyte maturation, and an early and specific expression during ovarian differentiation has been described in rainbow trout, Oncorhynchus mykiss. By in situ hybridization, we demonstrated that during rainbow trout gonadal differentiation, fst shares a similar expression pattern with cyp19a1a, the gene encoding ovarian aromatase, a key steroidogenic enzyme needed for ovarian differentiation in fish. Expression of fst and cyp19a1a was first detected in a few scattered cells in the embryonic ovary several days before hatching. Then, after histological differentiation, fst and cyp19a1a expression was localized in clusters of cells lining the future ovarian lamellae. As FST expression is known to be induced by the Wnt/ß-catenin pathway in mammals, the Wnt pathway was inhibited in vivo with the IWR-1 molecule, and we analyzed by qPCR the effects of this treatment on fst expression. We found that IWR-1 decreased fst expression in female gonads, consistent with a regulation of fst expression by the Wnt pathway in rainbow trout. Furthermore, expression of cyp19a1a was also downregulated, suggesting an implication of the Wnt pathway in ovarian differentiation.

Alteration of gap junction proteins (connexins) following lateral fluid percussion injury in rats.

Gap junctions are intercellular channels that mediate the cytoplasmic exchange of small hydrophilic molecules and are formed by a family of integral membrane proteins called connexins (Cxs). Cx43 is expressed predominantly in astrocytes, while Cx36 is expressed in neurons. In this study, we show alteration of Cx43 and Cx36 in the hippocampus after traumatic brain injury in rats. Adult male Sprague-Dawley rats were subjected to lateral fluid percussion injury of moderate severity. Brain coronal sections were used for immunohistochemistry with Cx43 and Cx36 antibodies. Cx43 immunoreactivity was increased in reactive astrocytes in the damaged hippocampus 24 hours after injury, and persisted for 72 hours. On the other hand, Cx36 immunoreactivity increased in CA3 neurons 1 hour after injury, and decreased later. These results indicate that gap junctions might participate in the pathophysiological process after traumatic brain injury.

Prediction of flood abnormalities for improved public safety using a modified adaptive neuro-fuzzy inference system.

It is widely accepted that an efficient flood alarm system may significantly improve public safety and mitigate economical damages caused by inundations. In this paper, a modified adaptive neuro-fuzzy system is proposed to modify the traditional neuro-fuzzy model. This new method employs a rule-correction based algorithm to replace the error back propagation algorithm that is employed by the traditional neuro-fuzzy method in backward pass calculation. The final value obtained during the backward pass calculation using the rule-correction algorithm is then considered as a mapping function of the learning mechanism of the modified neuro-fuzzy system. Effectiveness of the proposed identification technique is demonstrated through a simulation study on the flood series of the Citarum River in Indonesia. The first four-year data (1987 to 1990) was used for model training/calibration, while the other remaining data (1991 to 2002) was used for testing the model. The number of antecedent flows that should be included in the input variables was determined by two statistical methods, i.e. autocorrelation and partial autocorrelation between the variables. Performance accuracy of the model was evaluated in terms of two statistical indices, i.e. mean average percentage error and root mean square error. The algorithm was developed in a decision support system environment in order to enable users to process the data. The decision support system is found to be useful due to its interactive nature, flexibility in approach, and evolving graphical features, and can be adopted for any similar situation to predict the streamflow. The main data processing includes gauging station selection, input generation, lead-time selection/generation, and length of prediction. This program enables users to process the flood data, to train/test the model using various input options, and to visualize results. The program code consists of a set of files, which can be modified as well to match other purposes. This program may also serve as a tool for real-time flood monitoring and process control. The results indicate that the modified neuro-fuzzy model applied to the flood prediction seems to have reached encouraging results for the river basin under examination. The comparison of the modified neuro-fuzzy predictions with the observed data was satisfactory, where the error resulted from the testing period was varied between 2.632% and 5.560%. Thus, this program may also serve as a tool for real-time flood monitoring and process control.

Predominant presence of Streptococcus anginosus in the saliva of alcoholics.

Chronic alcohol consumption is known to be a major risk factor for cancers of the upper aerodigestive tract. The incidence of esophageal cancer (4.4%) in alcoholics is reported to be much higher than that in the Japanese population as a whole (0.0001%). This suggests the presence of specific factors in chronic alcohol consumption-related carcinogenesis. Recently, data showing a significant correlation between Streptococcus anginosus and carcinogenesis in the upper aerodigestive tract have been reported. In this study, the ratio of S. anginosus to oral bacteria in the saliva of 38 alcoholic patients was investigated to determine if there is an association between alcoholic patients and S. anginosus infection. The level of S. anginosus in the saliva from 22 healthy people, 41 esophageal cancer patients, 32 gastritis patients, and 24 periodontitis patients was also investigated and compared to the level in alcoholic patients. In the saliva from esophageal cancer patients, the level of S. anginosus was not significantly different from that of healthy people. The levels of S. anginosus in periodontitis and gastritis patients were also similar. In alcoholics, however, there was an extremely high level of S. anginosus, suggesting that they, rather than healthy people and general esophageal cancer patients, have a high risk for S. anginosus infection.

Heat stress-induced modulation of host defense against Toxoplasma gondii infection in mice.

This study examined the effects of burn injury on murine immune response against Toxoplasma gondii infection. Male C57BL/6 mice were divided into 3 groups: T. gondii infection (group T), burn injury (group B), and burn injury followed by T. gondii infection (group BT). The survival of group BT was significantly lower than those of group B and group T. Parasite abundance in the tissues was determined by quantitative competitive-polymerase chain reaction. Group BT exhibited significantly higher numbers of T. gondii than group T. Antibody production against T.g.HSP30 in group BT was significantly lower than that in group T, whereas no significant difference was observed in SAG1-specific antibody production. Delayed-type hypersensitivity (DTH) specific for 2,4-dinitrofluorobenzene (DNFB) of both group B and group BT was significantly lower than that of group T. One week after infection, serum interferon-gamma (IFN-gamma) and interleukin (IL)-10 levels in group BT were significantly lower, whereas serum IL-6 levels were significantly higher than in group T Serum TNF-alpha levels in both group T and group BT were elevated at 1 wk after infection, although there was no significant difference between them. Serum IFN-gamma, IL-10, and TNF-alpha levels in group B were not elevated during the experimental term. In conclusion, the impaired antigen-specific antibody production and DTH response, together with the modulated patterns of cytokine responses, seemed to be strongly involved in the development of burn-induced immunosuppression and the consequent increased susceptibility to T. gondii infection in mice.

Molecular analysis of age-related changes of Streptococcus anginosus group and Streptococcus mitis in saliva.

The purpose of this study was to survey the prevalence of streptococcal species, especially Streptococcus anginosus (which has been reported to be associated with cancer in the upper digestive tract), Streptococcus constellatus, and Streptococcus intermedius in the saliva of different age groups. A sequence analysis of 16S rDNA was performed and DNA quantified using real-time polymerase chain reaction. The S. anginosus level increased with age, whereas the levels of S. constellatus and S. intermedius did not change. Streptococcus mitis was the predominant species in the saliva of all the age groups but, unlike the S. anginosus, the proportion of S. mitis in the salivary bacteria decreased with age. The increase in S. anginosus with age should be carefully monitored because of its association with diseases, including cancer.

Vascular endothelial growth factor expression in pituitary adenomas.

Vascular endothelial growth factor (VEGF) is known to be a mediator of angiogenesis and vascular permeability. A cystic component and hemorrhage are often found in pituitary adenomas. In the present study we assess the VEGF expression based on immunohistochemical examinations in 48 pituitary adenomas. All the adenomas showed some VEGF immunoreactivity mainly in the cytoplasm of tumor cells. Of the 48 adenoma-cases, 16 cases had a strong VEGF immunoreactivity, 26 cases had a moderate one, and 6 cases had a weak one. On the MR images, a cystic component was found in 16 cases (33.3%), and a hemorrhage was found in 18 cases (37.5%). The VEGF immunoreactivity had a significant relationship with the cystic component but neither the hemorrhage, size, recurrence, or HE classification. These findings suggest that VEGF might play a potential role in the pathogenesis of cystic formation in pituitary adenomas.

Characteristic phosphorylation of the extracellular signal-regulated kinase pathway after kainate-induced seizures in the rat hippocampus.

Extracellular signal-regulated kinase (ERK) pathways play a crucial role in cell growth and long-lasting neuronal plasticity. Several studies have shown that phosphorylated-ERK (p-ERK) significantly increases after kainic acid (KA) administration. However, little or no information is available about the spatial distribution of p-ERK after KA-induced seizures. We herein show that KA-induced seizures significantly increase p-ERK in both neurons and astrocytes in rat brain using Western blots and immunohistochemistry. A strong immunoreactivity for p-ERK was induced in the dentate hilar neurons and CA3 neurons 30 mins and 6 hrs after KA injection. In addition, immunoreactivity for p-ERK was seen in astrocytes 6 hrs after KA injection. 72 hrs after KA injection, all pyramidal neurons had died. These findings suggest that the ERK pathway participates in the KA-induced neurotoxicity in the rat hippocampus.

Involvement of MyD88 in host defense and the down-regulation of anti-heat shock protein 70 autoantibody formation by MyD88 in Toxoplasma gondii-infected mice.

This study investigated the influence of TLR (toll-like receptor)4, TLR2, and MyD88 in Toxoplasma gondii-infected wild-type (WT) mice and TLR4-, TLR2-, and MyD88-deficient mice. Ninety-five percent of MyD88-deficient mice died 10-16 days after intraperitoneal infection with 100 cysts of T. gondii Fukaya strain, whereas 95-100% of TLR4- and TLR2-deficient mice and WT C57BL/6 (B6) mice survived for more than 7 wk after T. gondii infection. The distribution of T. gondii in various organs of TLR4-, TLR2-, and MyD88-deficient mice and WT B6 mice was assessed 2 wk after T. gondii intraperitoneal infection using quantitative competitive polymerase chain reaction. In MyD88-deficient mice, high levels of T. gondii load were observed in the brain, tongue, heart, lungs, spleen, liver, mesenteric lymph node, and kidneys after infection. The T. gondii load was significantly increased in the lungs in both TLR4- and TLR2-deficient mice compared with WT B6 mice. High levels of anti-mouse heat shock protein (mHSP)70 autoantibody and anti-T. gondii HSP70 antibody production were detected in the sera from MyD88-deficient mice.

[Dihydropyrimidine dehydrogenase activity in urothelial cancer--influence of UFT administration on DPD activity].

We measured the dihydropyrimidine dehydrogenase (DPD) activity considered to regulate the sensitivity of pyrimidine fluoride anti-cancer drugs in 50 cases of urothelial cancer (preoperative therapy group 10 cases, non-therapy group 40 cases). The association of DPD activity with pathological factors and TS activity as well as the influence of UFT oral administration on the DPD activity in tumors and blood were studied. The DPD activity in cancer tissue tended to be higher than that in the mucous membranes of the non-tumor region, but the difference was not significant. The DPD activity was high in infiltrative cancer (p < 0.05) but was not associated with atypia. The TS activity and DPD activity were manifested independently. The DPD activity in cancer tissue was significantly higher after UFT administration, with levels of 76.7 +/- 71.0 pmol/mg/min before administration and 220.6 +/- 129.1 pmol/mg/min after (p < 0.05). The DPD activity in peripheral blood was elevated after administration, but no significant difference was found. Since DPD activity is found in urothelial cancer tissue, 5-FU stable to DPD should preferably be used. It is also suggested that using a more potent DPD inhibitor in combination will produce higher anti-tumor effects.

Identification and characterization of a novel Golgi protein, GCP60, that interacts with the integral membrane protein giantin.

We demonstrated previously that the integral membrane protein giantin has the Golgi localization signal at the COOH-terminal cytoplasmic domain (Misumi, Y., Sohda, M., Tashiro, A., Sato, H., and Ikehara, Y. (2001) J. Biol. Chem. 276, 6867-6873). In the present study, using this domain as bait in the yeast two-hybrid screening system, we identified a novel protein interacting with giantin. The 3.6-kilobase mRNA encoding a 528-amino acid protein of 60 kDa designated GCP60 was ubiquitously expressed and was especially abundant in the testis and ovary. Immunofluorescence and immunoelectron microscopy confirmed that GCP60 was co-localized with giantin in the Golgi complex. GCP60 was found to be a peripheral protein associated with the Golgi membrane, where a COOH-terminal domain of GCP60 interacts with the COOH-terminal cytoplasmic domain of giantin. Overexpression of the COOH-terminal domain of GCP60 caused disassembly of the Golgi structure and blocked protein transport from the endoplasmic reticulum to the Golgi. Taken together, these results suggest that GCP60 is involved in the maintenance of the Golgi structure by interacting with giantin, affecting protein transport between the endoplasmic reticulum and the Golgi.

[Laparoscopy-assisted total nephroureterectomy for renal pelvic and/or lower ureteral cancer].

PURPOSE: The usefulness of laparoscopy-assisted total nephroureterectomy for patients with renal pelvic and lower ureteral cancer is evaluated. MATERIAL: Seven patients with renal pelvic cancer and four with lower ureteral cancer performed laparoscopy-assisted total nephroureterectomy from May 1997 to December 2000 (Ten males and one female, mean age 68.5 year-old). METHOD: Of the 11 patients, the initial one received preoperative embolization of the renal artery. Under general anesthesia laparoscopy-assisted total nephroureterectomy underwent via transperitoneal approach in three patients and retroperitoneal approach in eight. After the kidney was completely dissected under laparoscopic procedure, it was delivered en bloc with ureter from the skin incision in the lower abdomen. RESULT: Two patients needed conversion to open surgery. The mean operating time of nine patients except for conversion cases was 272 minutes and the mean blood loss was 313 ml. There was no major complication associated with laparoscopic procedure. There was no significant difference in both complication and recurrence rate between laparoscopy-assisted total nephroureterectomy and open surgery. CONCLUSION: Laparoscopy-assisted total nephroureterectomy is an useful procedure for the treatment of patients with renal pelvic and lower ureteral cancer because it enables us to remove out the kidney and ureter from one small lower abdominal incision.

Isolation, structure determination, and synthesis of neodysiherbaine A, a new excitatory amino acid from a marine sponge.

[structure: see text] A new excitatory amino acid, neodysiherbaine A (2), was isolated as a minor constituent of the aqueous extract from the marine sponge Dysidea herbacea. The structure was deduced by spectroscopic methods and established unambiguously by the total synthesis. The present synthesis, including as a key step cross-coupling of the 6/5-bicyclic core with an amino acid residue, is useful in constructing its structural analogues.

Organ infectivity of Toxoplasma gondii in interferon-gamma knockout mice.

To determine the influence of interferon (IFN)-gamma on the organ infectivity and on the genetic susceptibility of susceptible (C57BL/6) and resistant (BALB/c) strains after peroral infection with cysts of Toxoplasma gondii. IFN-gamma knockout (KO) mice in C57BL/6 and BALB/c backgrounds were utilized. The kinetics of the changes in T. gondii abundance were evaluated with a quantitative competitive polymerase chain reaction assay in various organs at different times after peroral infection. In IFN-gamma KO mice, a T. gondii-specific gene, SAG1, was detected in all organs examined, and the protozoan proliferated much more actively than in wild-type mice. The abundance of T. gondii was much higher in mesenteric lymph nodes and the heart than in other organs. In contrast, in the nervous system organs and kidneys, only a weakly detectable reaction was observed. Toxoplasma gondii grew at a more rapid rate in the organs of IFN-gamma KO C57BL/6 mice than in the organs of IFN-gamma KO BALB/c mice during the course of infection. Destruction of the IFN-gamma gene showed remarkable effects on the infectivity in both susceptible and resistant mice.

Pituitary chromophobe carcinoma with a low level of serum gonadotropin and an aspermatogenesis in a dog.

A 5-year-old male Shiba dog with progressive neurologic signs was examined by computed tomography (CT). A CT image of the brain disclosed a large, spherical high-density lesion in the thalamus and diencephalon. Serum LH, FSH and testosterone levels were all low. Macroscopically the large mass was connected with the sella turcia, and it was histopathologically diagnosed as a pituitary chromophobe carcinoma. An aspermatogenesis was observed in the testes. Therefore, it was suggested that the low levels of gonadotropin secretion from the pituitary gland due to the pituitary tumor resulted in the failure of maturation of spermatozoa and spermatids.

Induction of primary root curvature in radish seedlings in a static magnetic field.

Primary roots of radish (Raphanus sativus L.) seedlings were exposed to an inhomogeneous static magnetic field generated by a permanent magnet, during continuous rotation on a 0.06 rpm clinostat, thereby reducing the unilateral influence of gravity. The roots responded tropically to the static magnetic field with the tropism appearing to be negative. These roots responded significantly (P < 0.05) to the south pole of the magnet. The significant tropic response was found for a magnetic flux density of 13-68 mT, for a field gradient of 1.8-14.7 T/m, and for the product of magnetic field and field gradient of 0.023-1.0 T(2)/m. A small, but insignificant, response of the roots to the north pole has also been found.

Establishment of gene-vaccinated skin grafting against Toxoplasma gondii infection in mice.

Vaccine effects of in vivo gene-vaccinated skin graft were evaluated against Toxoplasma gondii (T. gondii) infection. By using a gene gun, cDNA coding T. gondii SAG1 molecule was intracutaneously vaccinated into C57BL/6 (B6; a susceptible strain), BALB/c (a resistant strain) and (C57BL/6 x BALB/c) F1 (CBF1) mice, and the gene-vaccinated skin of these strains was transplanted to CBF1 mice. Regarding the antibody production against SAG1, CBF1-recipient mice transplanted with the SAG1 gene-vaccinated B6 skin were high responders, whereas CBF1 mice skin grafted with vaccinated skin of both BALB/c and CBF1 mice were low responders. The donor-derived LC/DC migrated to the draining lymph nodes of the recipients from the skin graft within 3 days. The vaccine effect against T. gondii challenge infection was obtained in CBF1 mice which received the skin graft of the SAG1 gene-vaccinated BALB/c mice.