Anxiolytic effect of hepatocyte growth factor infused into rat brain

BACKGROUND: Hepatocyte growth factor (HGF) has the capacity to selectively direct thalamocortical projections into an intermediate target, the pallidum, and eventually to their final cortical destination. HGF may have a role in the mediation of anxiety. Very little is known about other central behavioral effects of HGF. OBJECTIVE: Our aim was to determine what effect HGF has on anxiety in rats. METHODS: HGF was infused at a constant rate into cerebral lateral ventricles and its effect on anxiety in rats was monitored. RESULTS: In the elevated plus maze test and the black and white box test, HGF administration caused all indicators of anxiety to increase. No significant effect on general locomotor activity was seen. CONCLUSION: HGF infusion into the brain produces an anxiolytic effect.     

Development of projection-specific interneurons and projection neurons in the embryonic mouse and rat spinal cord

Interneurons and projection neurons in the lumbar spinal cord of mouse and rat embryos were labeled retrogradely with fluorescent dextran amines from a distance of one segment from the segment of origin [lumbar segment (L) 2]. Six classes with specific axonal projections (ipsilateral ascending, descending, and bifurcating, and commissural ascending, descending, and bifurcating) were identified by differential labeling in both species and followed from embryonic day (E)12 to birth in the mouse. Neurons with shorter projections (intrasegmental interneurons) were not studied. We show that the four nonbifurcating neuron classes occupy characteristic, partially overlapping domains in the transverse plane, indicating a systematic pattern of migration and settlement related to axon trajectories. The number of neurons in each of the nonbifurcating classes increased steadily during development. Bifurcating neurons represented a minor fraction of the total throughout development and had relatively scattered positions within the ipsilateral and commissural neuron domains. Combination of retrograde tracing and immunohistochemistry for the inhibitory neurotransmitter gamma-aminobutyric acid (GABA) showed that none of the spinal neurons in the six projection-specific classes was GABA positive, suggesting that all GABA-positive spinal neurons, including previously described GABA-positive commissural neurons, are unlikely to have projections exceeding one or two segments in either direction.     

A randomized multicenter trial comparing resection and radiochemotherapy for resectable locally invasive pancreatic cancer

BACKGROUND: Though the outcome of resection for locally invasive pancreatic cancer is still poor, it has gradually improved in Japan, and the 5-year survival is now about 10%. However, the advantage of resection over radiochemotherapy has not yet been confirmed by a randomized trial. We conducted this study to compare surgical resection alone versus radiochemotherapy without resection for locally invasive pancreatic cancer using a multicenter randomized design. METHODS: Patients with pancreatic cancer who met our preoperative criteria for inclusion (pancreatic cancer invading the pancreatic capsule without involvement of the superior mesenteric artery or the common hepatic artery, or without distant metastasis) underwent laparotomy. Patients with operative findings consistent with our criteria were randomized into a radical resection group and a radiochemotherapy group (200 mg/m(2)/day of intravenous 5-fluorouracil and 5040 cGy of radiotherapy) without resection. The 2 groups were compared for mean survival, hazard ratio, 1-year survival, quality of life scores, and hematologic and blood chemical data. RESULTS: Twenty patients were assigned to the resection group and 22 to the radiochemotherapy group. There was 1 operative death. The surgical resection group had better results than the radiochemotherapy group as measured by 1-year survival (62% vs 32 %, P=.05), mean survival time (>17 vs 11 months, P < .03), and hazard ratio (0.46, P=.04). There were no differences in the quality of life score or laboratory data apart from increased diarrhea after surgical resection. CONCLUSIONS: Locally invasive pancreatic cancer without distant metastases and major arterial invasion appears to be best treated by surgical resection.     

A new structural class of proteasome inhibitors identified by microbial screening using yeast-based assay

A yeast-based growth interference assay was developed utilizing a yeast strain in which expression of Xenopus cyclin A1 was induced to elevate cell division cycle 28 (Cdc28) kinase activity. Since the hyperactivation of Cdc28 kinase in yeast results in a growth-arrest phenotype, compounds which could rescue the cyclin A1-induced growth arrest might be potential new, antitumor drug candidates acting on the cyclin-dependent, kinase-mediated, cell cycle regulation pathway. In the course of our microbial screening program, the new Streptomyces metabolites, belactosins, were identified. As reported previously, belactosin A induced cell cycle arrest at G2/M phase in human cancer cells. However, the molecular mechanism of action was unknown. We herein demonstrate the proteasome inhibition by belactosin A. Belactosin A did not inhibit yeast Cdc28 kinase and human cyclin-dependent kinase in vitro. On the other hand, it inhibited the chymotrypsin-like activity of the rabbit 20S proteasome. From the initial SAR studies, we identified a hydrophobic belactosin A derivative, KF33955, which exhibited a 100-fold greater growth-inhibitory activity against HeLa S3 cells than belactosin A, presumably due to its higher cell permeability. The biochemical analysis using KF33955 suggested that the proteasome inhibitory activity of KF33955 were irreversible and required the beta-lactone moiety to inhibit the proteasome. KF33955 increased the intracellular levels of protein ubiquitination in NIH3T3 cells. In addition, KF33955 treatment resulted in the accumulation of known proteasome substrates in HeLa S3 cells. These results identify belactosin A as a useful lead compound to target proteasome for the treatment of disease whose etiology is dependent on the unregulated ubiquitin-proteasome pathway.     

Oxidative DNA damage induced by a melatonin metabolite, 6-hydroxymelatonin, via a unique non-o-quinone type of redox cycle

Melatonin, an indolic pineal hormone, is produced primarily at night in mammals and is important in controlling biological rhythms. Although melatonin is known to be effective as a free radical scavenger and has an anti-cancer effect, carcinogenic properties have also been reported. In relation to its carcinogenic potential, we have examined whether 6-hydroxymelatonin, a major melatonin metabolite, can induce DNA damage in the presence of metal ion using [32P]-5'-end-labeled DNA fragments obtained from genes relevant to human cancer. 6-Hydroxymelatonin induced site-specific DNA damage in the presence of Cu(II). Formamidopyrimidine-DNA glycosylase treatment induced cleavage sites mainly at G residues of the 5'-TG-3' sequence, whereas piperidine treatment induced cleavage sites at T mainly of 5'-TG-3'. Interestingly, 6-hydroxymelatonin strongly damaged G and C of the 5'-ACG-3' sequence complementary to codon 273 of the p53 gene. These results suggest that 6-hydroxymelatonin can cause double-base lesions. DNA damage was inhibited by both catalase and bathocuproine, Cu(I)-specific stabilizer, suggesting that reactive species derived from the reaction of H2O2 with Cu(I) participate in DNA damage. Cytochrome P450 reductase efficiently enhanced 6-hydroxymelatonin-induced oxidative DNA damage and oxygen consumption, suggesting the formation of redox cycle. It is noteworthy that 6-hydroxymelatonin can efficiently induce DNA damage via non-o-quinone type of redox cycle. Formation of 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG), a characteristic oxidative DNA lesion, in calf thymus DNA was significantly increased by 6-hydroxymelatonin in the presence of Cu(II). Furthermore, 6-hydroxymelatonin significantly increased the formation of 8-oxodG in human leukemia cell line HL-60 but not in HP100, a hydrogen peroxide (H2O2)-resistant cell line derived from HL-60. The 6-hydroxymelatonin-induced 8-oxodG formation in HL-60 cells significantly decreased by the addition of bathocuproine or o-phenanthroline. Therefore, it is concluded that melatonin may exhibit carcinogenic potential through oxidative DNA damage by its metabolite.     

Comparison of intraperitoneal continuous infusion of floxuridine and bolus administration in a peritoneal gastric cancer xenograft model

Purpose To identify the optimal schedule for intraperitoneal (i.p.) infusion of floxuridine (FUDR) against peritoneal micrometastases from gastric cancer.Methods The efficacy of continuous i.p. infusion of FUDR was compared with that of bolus i.p. administration in peritoneal gastric cancer (MKN45) xenografts. The FUDR continuous delivery system in this study was in the form of injectable poly(lactic-coglycolic) acid (PLGA) microspheres intended for i.p. injection. Animals were treated by continuous i.p. infusion using FUDR-loaded microspheres or bolus i.p. administration of FUDR.Results In vitro testing demonstrated that FUDR was released slowly from the microspheres at a rate of approximately 5% of the total encapsulated drug per day. In in vivo studies, the peritoneal level was found to persist and was approximately 5- to 50-fold higher than that of plasma for more than 2 weeks following a single injection of the microspheres. An in vitro MTT assay showed that exposure time clearly influenced the cytotoxic potency of FUDR. In vivo, continuous infusion was more effective against peritoneal tumor than bolus administration at equivalent doses. However, compared with bolus administration, toxicity was increased, resulting in a reduced maximum tolerated dose (MTD) with continuous infusion. When the treatment was carried out at each MTD (continuous 1 mg/kg, bolus 600 mg/kg), continuous infusion had no advantage in inhibiting tumor growth.Conclusions Owing to the higher toxicity and the equal efficacy of continuous infusion compared with bolus administration, continuous infusion is not recommended in i.p. FUDR treatment.This study was supported in part by a grant from the Japan Society for the Promotion of Science and by a grant from the Setsuro Fujii Memorial Osaka Foundation for Promotion of Fundamental Medical Research.     

Mechanism of carcinogenesis induced by a veterinary antimicrobial drug, nitrofurazone, via oxidative DNA damage and cell proliferation

Nitrofurazone, a veterinary antimicrobial drug, causes mammary and ovarian tumors in animals. We investigated the mechanisms of carcinogenesis by nitrofurazone. Nitrofurazone significantly stimulated the proliferation of estrogen-dependent MCF-7 cells. Nitrofurazone caused Cu(II)-mediated damage to 32P-5'-end-labeled DNA fragments obtained from human genes only when cytochrome P450 reductase was added. DNA damage was inhibited by catalase and bathocuproine. DNA damage was preferably induced at the 5'-ACG-3' sequence, a hotspot of the p53 gene. These findings suggest that nitrofurazone metabolites are involved in tumor initiation through oxidative DNA damage and nitrofurazone itself enhances cell proliferation, leading to promotion and/or progression in carcinogenesis.     

Significance of working environment improvement based on determination of source of harmful substance--example of an endoscope syringe room using glutaraldehyde

Because a worker complained of irritated eyes and throat due to glutaraldehyde (GA) in an endoscope syringe room, the automated endoscope washers and GA liquid stored in reserve were isolated in a separate room. A ventilation system was installed in this room, and the packing of the automated endoscope washers was changed. However, since the obnoxious smell of GA still remained in the endoscope syringe room, we had to determine the source of the smell. A plastic bucket with a cap was found to be filled with GA for disinfection of the endoscope apparatus. GA had evaporated when dispersed around the bucket, resulting in the obnoxious smell. The plastic bucket was replaced with a different type of container. Moreover, GA from the separate room did not affect the concentration of GA in the working area because the separate room for the automated endoscope washers had twice the ventilation volume proposed by the guidelines of the Society of Gastroenterology Nurses and Associates of the USA and the Healthcare Engineering Association of Japan. Consequently, we reconfirmed the significance of working environment improvement after clarifying the source of the harmful substance.     

Diagnostic significance of serum soluble transferrin receptors in various anemic diseases: the first multi-institutional joint study in Japan

Serum soluble transferrin receptor (sTfR) has been reported to be higher in patients with iron deficiency or with elevated erythropoiesis. In the present study, serum sTfR was measured in various anemic diseases and their clinical significance was examined in a multi-institutional joint study. Serum sTfRs in patients with the following anemic diseases were markedly higher than those in normal healthy adults: non-treated iron deficiency anemia (IDA) (9.13 +/- 7.04 mg/l, n = 52, p < 0.0001), anemia of chronic disorders (ACD) (3.45 +/- 1.38 mg/l, n = 20, p < 0.0001), hemolytic anemia (HA) (5.57 +/- 3.26 mg/l, n = 17, p < 0.0001), and myelodysplastic syndrome (MDS) (4.03 +/- 2.83 mg/l, n = 20, p < 0.0001). There were significant differences between IDA and ACD (p < 0.0001), between aplastic anemia (AA) (1.58 +/- 1.26 mg/l, n = 16) and MDS (p < 0.001), and between AA and MDS with refractory anemia (MDS-RA) (4.16 +/- 3.40 mg/l, n = 9) (p < 0.02). In patients with chronic renal failure (CRF), serum sTfR levels and serum sTfR/log serum ferritin ratios (sTfR/F index) were compared in the two classified groups according to Muirhead's criteria, as IDA and non-IDA groups with or without recombinant human erythropoietin (rHuEPO) treatment. Significantly high levels of both serum sTfR (p < 0.0001) and the sTfR/F index (p < 0.0001) were observed in IDA without rHuEPO treatment. Especially in CRF with rHuEPO treatment, the sTfR/F index showed marked elevation in the IDA group (p < 0.0001) compared with serum sTfR (p < 0.001), indicating more diagnostic efficacy of the sTfR/F index for CRF with IDA. In conclusion, the serum sTfR concentration is a useful diagnostic tool for discrimination between IDA and ACD, and between AA and MDS-RA, and for the detection of iron deficiency in CRF patients in the Japanese population.     

Blood flow does not correlate with the size of metastasis in our new intravital observation model of Lewis lung cancer

We previously reported a novel in situ observation model for microcirculation of lung metastasis from subcutaneously implanted Lewis lung cancer into mouse. Using this model, we studied the correlation of blood flow and the size of lung metastasis. It was revealed that metastatic growth and its angiogenesis are suppressed by circulating angiogenesis inhibitors, such as angiostatin or endostatin, released from primary tumor. When we removed the primary tumor, the metastasized lung cancer significantly grew faster and larger. But the blood flow per area did not increase either inside or outside of the metastatic tumor. This suggests that the growth of metastatic tumor is directly regulated not by blood flow increase but by the other effects of the circulating factors.