Differential genetic responses to ionizing irradiation in individual families of Japanese medaka, Oryzias latipes.

Although no statistically significant hereditary effects have yet been detected in the children of survivors from the atomic bombings in Hiroshima and Nagasaki, recent animal studies have found that exposure to ionizing radiation can cause genomic and epigenomic instability in the exposed individuals, as well as their offspring, and therefore, may have much larger genetic effects than predicted by earlier studies. When individuals are exposed to various environmental insults, including radiation, individual sensitivity to the insults often varies. Variance in germ-line response to radiation among individuals has been widely recognized, but it is difficult to address due to the use of inbred strains and the limited number of offspring that can be produced by a pair of mice, the common model used to study genetic effects of radiation. Herein is the first study to examine individual family responses to ionizing radiation using a parent-pedigree approach in an outbred strain of a vertebrate model, the Japanese medaka fish. Changes in frequencies of radiation-induced germline mutations at nine microsatellite loci were examined in the same families before and after exposure to one of four acute doses of ionizing radiation (0.1, 0.5, 2.5, 5Gy, plus sham-exposed controls). Families varied significantly in pre-exposure mutation frequencies and responses to irradiation, but germline mutations were elevated in at least one family after 0.1, 0.5, and 5Gy exposures. Variance among individuals in sensitivity to radiation is well documented for many endpoints, and our work now extends these endpoints to include germ-line mutations. Further studies are needed to elucidate dose response, effects at varying stages of spermatogenesis, and the mechanisms underlying the variance in these individual responses to radiation.

DHEA improves impaired activation of Akt and PKC zeta/lambda-GLUT4 pathway in skeletal muscle and improves hyperglycaemia in streptozotocin-induced diabetes rats.

AIM: Addition of dehydroepiandrosterone (DHEA) to a cultured skeletal muscle locally synthesizes 5alpha-dihydrotestosterone (DHT). It induced activation of glucose metabolism-related signalling pathway via protein kinase B (Akt) and protein kinase C zeta/lambda (PKC zeta/lambda)-glucose transporter-4 (GLUT4) proteins. However, such an effect of DHEA in vivo remains unclear. METHODS: Using streptozotocin (STZ)-induced rats with type 1 diabetes mellitus, we tested the hypothesis that a single bout of DHEA injection in the rats improves hyperglycaemia and muscle GLUT4-regulated signalling pathway. After 1 week of STZ injection (55 mg kg(-1)) with male Wistar rats, fasting glucose concentrations were determined in a blood sample taken from the tail vein. Blood glucose levels were then monitored for 180 min after DHEA or sesame oil (control) was injected (n = 10 for each group). RESULTS: Blood glucose levels decreased significantly for 30-150 min after 2 mg DHEA injection in the STZ rats. In the skeletal muscle, expression and translocation of GLUT4 protein, phosphorylation of Akt and PKC zeta/lambda, and phosphofructokinase and hexokinase enzyme activities increased significantly by DHEA injection. However, DHEA-induced improvements in Akt and PKC zeta/lambda-GLUT4 pathways were blocked by a DHT inhibitor. CONCLUSION: These results suggest that a single bout of DHEA injection can improve hyperglycaemia and activate the glucose metabolism-related signalling pathway via Akt and PKC zeta/lambda-GLUT4 proteins of skeletal muscles in rats. Moreover, these results show that a DHEA-induced increase in muscle glucose uptake and utilization might contribute to improvement in hyperglycaemia in type 1 diabetes mellitus.

Communication of radiation-induced signals in vivo between DNA repair deficient and proficient medaka (Oryzias latipes).

Radiation-induced bystander effects are established consequences of exposure to ionizing radiation. The operation of this mechanism has been seen in vitro and also between fish, mammals, and plants in vive where stress signals from treated organisms induce responses in neighbors. In vitro research shows that DNA repair deficient cells produce more toxic bystander responses. To test this in vivo two strains of Japanese medaka were tested. One is a mutant, repair deficient strain (ric2) and the other, the wildtype repair proficient strain (CAB). Irradiated fish swam with unirradiated partners in a strain mix and match protocol. The data suggest that medaka produce signals, when exposed to radiation, that induce unirradiated fish ofthe same strain swimming with them to produce an altered response to that seen in bystanders to sham irradiated fish. More apoptosis was seen in bystanders to repair deficient fish. When the strains are mixed, the bystanders of either strain respond like the donor strain. Measurements of Bcl-2 and cmyc proteins in the explants confirmed these observations. A possible role for p53 was also identified in that the use of reporters with mutant p53 demonstrated that CAB signals killed all the reporter cells by apoptosis. Use of a similar but p53 wildtype cell line had no such effect. The data add to the body of knowledge showing that bystander signals operate at hierarchical levels of organization greater than the individual and may therefore have relevance in radioecology and (eco)systems biology.

[Repair of paravalvular leak after a third mitral valve replacement].

This study reports on a 57-year-old woman who underwent a 3rd mitral valve replacement and presented with complaints of fatigue. Laboratory examination revealed severe hemolytic anemia, and trans-esophageal echocardiography revealed a paravalvular leak (PVL) around the prosthetic valve at the posterior trigone in the mitral position. PVL was regarded as the cause of hemolytic anemia. At surgery, a small tissue defect was detected around the calcified posterior trigone of the mitral annulus with no evidence of infective endocarditis. The mitral PVL was successfully repaired with suture closure of the annular defect. The postoperative course was uneventful: postoperative echocardiography revealed no evidence of PVL, and the hemolytic anemia subsided.

[Ascending aortic aneurysm following aortic valve replacement due to aortitis syndrome; report of a case].

A 60-year-old woman had previously undergone aortic valve replacement for aortic regurgitation. As the aortic wall was elastic hard, inflammatory change was suspected; therefore, we undertook a partial biopsy of the ascending aortic wall and the intraoperative pathological specimens were compatible with aortitis syndrome. As there was no active inflammatory change, she was diagnosed as inactive aortitis syndrome and steroid therapy was not applied. Seven years later, a follow-up computed tomography (CT) showed an ascending aortic aneurysm of 65 mm in diameter. Aortic root replacement was planned based on a clinical diagnosis of an aneurysm of the ascending aorta. The patient was discharged without complication 21 days after surgery. It is possible that an inactive stage of aortitis may lead to late dilatation of the ascending aorta; therefore, careful postoperative follow-up is necessary in such cases.

[Cardiac papillary fibroelastomas: report of 2 cases].

We report 2 cases of cardiac papillary fibroelastomas in adults. Case 1: A 61-year-old man was admitted because echocardiography showed a 1 cm pedunculated papillary tumor in the left atrium. In an operation, it was located in the left atrium near the mitral valve and was resected along with a 5 mm margin of endocardium. Case 2: A 60-year-old woman had a 1 cm mobile tumor in the right ventricle near the tricuspid valve located by echocardiography in a preoperative examination of a ventricular septal defect. In an operation, a pedunculated tumor located in the right ventricle was resected. In these 2 cases, histopathology showed the tumor to be a papillary fibroelastoma. Almost all cardiac papillary fibroelastoma are closely related to the cardiac valve, but in these cases, the tumors were located in the left atrium, and the right ventricle, respectively, which is quite rare.

Significance of the transcription factor KLF5 in cardiovascular remodeling.

Structural remodeling of the heart and blood vessels is an important pathologic process in the development of many cardiovascular diseases. However, transcriptional regulation of altered gene expression during cardiovascular remodeling is not well understood. We previously isolated KLF5/basic transcription element-binding (BTEB)2, a Krüppel-like factor, as a transcription factor that binds the promoter of the embryonic smooth muscle myosin heavy chain gene (SMemb). KLF5 activates many genes inducible during cardiovascular remodeling, such as platelet-derived growth factor (PDGF)-A/B, Egr-1, plasminogen activator inhibitor-1 (PAI-1), inducible nitric oxide synthase (iNOS), and vascular endothelial growth factor (VEGF) receptors. KLF5 is abundantly expressed in embryonic smooth muscles and is down-regulated with vascular development, but reinduced in proliferative neointimal smooth muscles in response to vascular injury. In KLF5 gene-targeted mice, homozygotes die at an early embryonic stage whereas heterozygotes are apparently normal. However, in response to external stress, arteries of heterozygotes exhibit diminished levels of smooth muscle and adventitial cell activation. Furthermore, angiotensin II-induced cardiac hypertrophy and fibrosis are attenuated in heterozygotes. KLF5 activities are regulated by many transcriptional regulators and nuclear receptors, such as retinoic acid receptor-alpha (RAR alpha), NF-kappaB, PPAR gamma, p300, and SET. Interestingly, RAR alpha agonist suppresses KLF5 and cardiovascular remodeling, whereas RAR alpha antagonist activates KLF5 and induces angiogenesis. These results indicate that KLF5 is an essential transcription factor in cardiovascular remodeling and a potential therapeutic target for cardiovascular disease.

Morphologic evaluation of the antitumor activity of photodynamic therapy (PDT) using mono-L-aspartyl chlorin e6 (NPe6) against uterine cervical carcinoma cell lines.

In order to elucidate the antitumor effect and mechanism of action of photodynamic therapy (PDT) using the photosensitizing agent mono-L-aspartyl chlorin e6 (NPe6) and a semiconductor laser, we conducted a morphologic study on uterine cervical cancer cell lines. First, tumor shrinkage was confirmed in a tumor growth inhibition test. Next, morphologic changes after PDT were examined, and since the major change appeared to be tumor necrosis secondary to obstruction of the blood vessels around the tumor, an NPe6 cell uptake experiment was performed. The results confirmed that a significantly greater amount of NPe6 was incorporated by human umbilical vein endothelial cells (HUV-EC1) and the cervical cancer cell lines than by human umbilical cord-derived fibroblasts. Based on these findings it was concluded that NPe6 possesses tumor affinity, and necrosis secondary to vascular obstruction was postulated to be the principal mechanism of the antitumor effect of PDT using NPe6.

Bilirubin calculi crushing by laser irradiation at a molecular oscillating region wavelength based on infrared absorption spectrum analysis using a free-electron laser: an experimental study.

We investigated a new laser technique of crushing bilirubin calculi, our aim being to crush calculi in isolation using a minimally invasive procedure. Infrared absorption spectrum analysis of the bilirubin calculi was conducted, revealing maximum absorption spectrum at a wavelength of the C=O stretching vibration of ester binding that exists within the molecular structure of bilirubin calcium. As an experiment to crush calculi using the free-electron laser, we set the laser at the effective irradiation wavelength of ester binding, and conducted noncontact irradiation of the bilirubin calculi. The calculi began to slowly ablate until the irradiated site had been completely obliterated after 20s of irradiation. Moreover, absorption spectrum analysis of the irradiated site, from a comparison of absorption peak ratios, revealed that absorption peak intensities decreased over time at the absorption wavelength of ester binding. These findings suggest that irradiation of molecular oscillating region wavelengths peculiar to calculi based on infrared absorption spectrum analysis results in the gradual crushing of calculi in isolation by breaking down their molecular structure.

Lipid analysis of peripheral blood monocytes in psoriatic patients using Fourier-transform infrared microspectroscopy.

In psoriasis vulgaris, there are immunological abnormalities of T cells and monocytes. We previously demonstrated that monocytes in the peripheral blood of patients with psoriasis vulgaris are activated and produce an excess of inflammatory cytokines. It has long been suggested that fat metabolism is impaired in patients with this illness. In addition, it has been reported that macrophages activated by engulfing low density lipoprotein (LDL) immune complexes release large quantities of tumor necrosis factor (TNF)-alpha and interleukin (IL)-1beta. Hence we hypothesized that the monocytes of psoriatic patients are activated by engulfing lipids and overproduce inflammatory cytokines. Therefore we measured both the serum and monocyte levels of lipids in the peripheral blood of psoriatic patients. At the same time, we calculated the psoriasis area and severity index (PASI) scores and analyzed their correlation with the lipid kinetics. The results showed that the serum cholesterol ester level and the cholesterol ester level in monocytes of psoriatic patients were significantly higher than those in healthy individuals. However, the cholesterol ester level in monocytes of patients with hyperlipidemia was also high, and there was no correlation between cholesterol ester level in monocytes of psoriatics and PASI scores. The cholesterol ester level in the monocytes of psoriatic patients was high, but this does not seem to play an important role in the pathogenesis of psoriasis.

Endovascular photodynamic therapy using mono-L-aspartyl-chlorin e6 to inhibit Intimal hyperplasia in balloon-injured rabbit arteries.

BACKGROUND AND OBJECTIVE: Intimal hyperplasia (IH) leading to restenosis is a major complication of arterial revascularization. The purpose of this study was to investigate the effect of photodynamic therapy (PDT) using mono-L-aspartyl chlorin e6 (NPe6) as a photosensitizer and intraluminal radial irradiation for inhibition of IH experimentally. STUDY DESIGN/MATERIALS AND METHODS: Study of laser transmission through the blood indicated that exclusion of blood is a prerequisite for intraluminal PDT. For homogeneous radial laser irradiation to the vessel wall, we used a newly developed cylindrical diffusing balloon laser fiber. Injuries were induced by pulling a balloon catheter through the right iliac artery of rabbits. One and 6 hours after the NPe6 injection (5mg/kg i.v.), drug distribution was examined by fluorescence microscopy. Nineteen rabbits received NPe6 at the time of injuries and PDT was performed with 664-nm laser at 30 and 10 J/cm(2) (20, 30, 40 mW/cm(2)) 1 hour after the injuries. The arteries were harvested at 2 days. In a second group of rabbits, PDT was given at 30 mW/cm(2) (30 J/cm(2)). Two weeks after treatment, the arteries were removed and examined histologically. RESULTS: NPe6 was found to be distributed selectively in the injured media. Endovascular NPe6-PDT showed complete depletion of smooth muscle cells even with 10 J/cm(2) at 2 days. IH was significantly inhibited at 14 days after PDT. CONCLUSIONS: Endovascular PDT of injured artery using NPe6 can prevent IH in this model of arterial wall injury and may become clinically useful for the prophylaxis of IH.

Oozing from the pericardium as an etiology of cardiac tamponade associated with screw-in atrial leads.

Screw-in atrial pacing leads are widely used. Cardiac tamponade is a complication. An 81-year-old woman with advanced atrioventricular block underwent permanent pacemaker implantation and subsequently developed cardiac tamponade. At surgery, the lead-tip screw was found penetrated through the right atrium but not through the pericardium. The source of bleeding was confirmed to scratching the inner pericardial membrane by the screw tip. Although cardiac tamponade due to perforation and leakage is known, tamponade caused by the trauma of an atrial screw on the pericardium with resultant ooze is less well described.

[Photodynamic diagnosis and photodynamic therapy for experimental hepatoma with ME 2906].

We studied the effects of photodynamic diagnosis (PDD) and photodynamic therapy (PDT) in confirming the existence of hepatoma, using the new photosensitizer mono-L-aspartyl chlorine 6. Japanese white rabbits were selected for abdominal incision under intravenous anesthesia, and VX 2 tumor cells were transplanted into the left liver lobe to create a hepatoma model. In the experiment, hepatoma of 1 cm in diameter (at one week after transplantation) was radiated with a semiconducter laser (664 nm, 200 J/cm2) for treatment.

Effects of photodynamic therapy using mono-L-aspartyl chlorin e6 on vessels and its contribution to the antitumor effect.

The effect of photodynamic therapy (PDT) on the vascular system has a significant role in tumor tissue destruction. We investigated the contribution of vascular damage to the antitumor effects of PDT and analyzed the quantitative vascular changes after PDT. Fibrosarcoma-bearing BALB / c male mice were injected with mono-L-aspartyl chlorin e6 (NPe6) at a dose of 0.25, 5 or 15 mg / kg, and photoradiation was performed with a diode laser 10 min, 2 h or 24 h after injection, respectively. Ten minutes after injection of 0. 25 mg / kg, NPe6 was found to be present only in plasma, while at 2 h after injection of 5 mg / kg it was present in both plasma and tumor, and 24 h after injection of 15 mg / kg it was present only in the tumor. The antitumor effects observed in the 5 mg / kg-2 h and 0. 25 mg / kg-10 min groups were virtually the same, whereas the effect in the 15 mg / kg-24 h group was weaker. The damage to the tumor vasculature and tumor cells in the 15 mg / kg-24 h group occurred later than under the other conditions, and vascular damage in the tumor-surrounding tissue was also less marked even 24 h after PDT. These results suggested that the plasma NPe6 concentration during laser irradiation contributed more than the tumor NPe6 concentration to the antitumor effect, and that the minimal damage to blood vessels around the tumor at the low plasma NPe6 concentration may be one reason for the failure to obtain a marked antitumor effect.

Effects on Rb(+)(K+) uptake of HeLa cells in a high K(+) medium of exposure to a switched 1.7 Tesla magnetic field.

Effects of a switched, time-varying 1.7 T magnetic field on Rb(+)(K+) uptake by HeLa S3 cells incubated in an isosmotic high K(+) medium were examined. The magnetic flux density was varied intermittently from 0.07-1.7 T at an interval of 3 s. K(+) uptake was activated by replacement of normal medium by high K(+) medium. A membrane-permeable Ca(2+) chelating agent (BAPTA-AM) and Ca(2+)-dependent K(+) channel inhibitors (quinine, charibdotoxin, and iberiotoxin) were found to reduce the Rb(+)(K+) uptake by about 30-40%. Uptake of K(+) that is sensitive to these drugs is possibly mediated by Ca(2+)-dependent K(+) channels. The intermittent magnetic field partly suppress ed the drug-sensitive K(+) uptake by about 30-40% (P < 0.05). To test the mechanism of inhibition by the magnetic fields, intracellular Ca(2+) concentration ([Ca(2+)]c) was measured using Fura 2-AM. When cells were placed in the high K(+) medium, [Ca(2+)]c increased to about 1.4 times the original level, but exposure to the magnetic fields completely suppressed the increase (P < 0.01). Addition of a Ca(2+) ionophore (ionomycin) to the high K(+) medium increased [Ca(2+)]c to the level of control cells, regardless of exposure to the magnetic field. But the inhibition of K(+) uptake by the magnetic fields was not restored by addition of ionomycin. Based on our previous results on magnetic field-induced changes in properties of the cell membrane, these results indicate that exposure to the magnetic fields partly suppresses K(+) influx, which may be mediated by Ca(2+)-dependent K(+) channels. The suppress ion of K(+) fluxes could relate to a change in electric properties of cell surface and an inhibition of Ca(2+) influx mediated by Ca(2+) channels of either the cell plasma membrane or the inner vesicular membrane of intracellular Ca(2+) stores.

Renal artery stenosis associated with epidermal nevus syndrome.

Epidermal nevus syndrome is an unusual neurocutaneous disorder in which epidermal nevi are associated with abnormalities of the skeleton and central nervous system, including the eyes and somtimes the cardiovascular system. We treated a patient in whom the latter included renal artery stenosis. An 18-year-old man with epidermal nevi was diagnosed as having the syndrome based on the additional presence of scoliosis, an arachnoid cyst in the middle cranial fossa, and microphthalmos. Hypertension was diagnosed when the patient was 15 years old. The plasma renin activity (9.7 ng/ml/h) was elevated. Right renal artery stenosis was demonstrated by angiography, and the abdominal aorta was narrowed distal to the ostium of the superior mesenteric artery. The plasma renin activity in the right renal vein (16 ng/ml/h) was higher than contralaterally (10 ng/ml/h). Several cardiovascular manifestations have been reported as a complication of epidermal nevus syndrome. Hypertension in an individual with epidermal nevi and congenital anomalies should prompt a search for a vascular anomaly.

Localization of experimental submucosal esophageal tumor in rabbits by using mono-L-aspartyl chlorin e6 and long-wavelength photodynamic excitation.

BACKGROUND AND OBJECTIVE: To increase the applicability of photodynamic diagnosis with regard to deep-seated tumor, we illuminated tumors with a long-wavelength laser beam after photosensitization with mono-L-aspartyl chlorin e6 (NPe6). STUDY DESIGN/MATERIALS AND METHODS: Rabbits with VX2 esophageal tumors were divided into four groups. The control group was not treated, and the other three groups were injected with 1, 2.5, and 5 mg/kg mono-L-aspartyl chlorin e6 (NPe6), respectively. After excitation with a 664-nm laser beam (10 mW, 10 seconds), the fluorescence image and the relative fluorescence intensity (tumor/normal tissue) were recorded every 2 hours up to 8 hours by a newly developed diode laser endoscopic fluorescence imaging system. The tissue concentration of NPe6 was examined by high performance liquid chromatography at 2, 4, and 6 hours after injection with 1 and 5 mg/kg NPe6. RESULTS: The diode laser endoscopic fluorescence imaging system was able to selectively detect fluorescence from submucosal tumor by comparison with the surrounding normal mucosa after NPe6 injection. The fluorescence intensity correlated with NPe6 dose, selectively accumulated in the tumor tissue and relative intensity peaked at 6 hours after injection. No fluorescent images were detected in controls. CONCLUSION: Given intravenously, NPe6 at a dose of 5 mg/kg and excited with a 664-nm wavelength laser beam 6 hours later can define experimentally induced deep-seated esophageal carcinoma in rabbits, by using an endoscopic fluorescence imaging system.

Direct interaction between a quinoline derivative, MS-209, and multidrug resistance protein (MRP) in human gastric cancer cells.

MS-209 is a novel quinoline derivative reversing P-glycoprotein-mediated multidrug resistance (MDR). We investigated the interaction between MS-209 and multidrug resistance protein (MRP) in MRP-overexpressing human gastric cancer cells. We measured [3H]leukotriene C4 uptake into the membrane vesicles of the cells and intracellular calcein and [3H]vincristine accumulation with or without MS-209. In multi-drug-resistant MKN45R0.8 cells selected by doxorubicin, MS-209 dose dependently reduced MRP-mediated [3H]leukotriene C4 uptake and increased calcein accumulation. In both resistant and unselected cell lines expressing the MRP gene, MS-209 increased [3H]vincristine accumulation in proportion with the level of MRP mRNA expression and enhanced the cytotoxicity of etoposide, doxorubicin, and vincristine. The reversal effects correlated with the level of MRP mRNA expression in these cells. Our results indicate that MS-209 effectively reverses intrinsic and acquired MRP-mediated MDR of gastric cancer cells by interacting directly with MRP.

Apoptosis and Bbcl-2 expression in gastric carcinomas: correlation withclinicopathological variables, p53 expression, cell proliferation and prognosis.

We investigated apoptosis and Bcl-2 expression in 221 advanced gastric carcinomas in correlation with clinicopathological variables, p53 expression, cell proliferation and prognosis, using the in situ DNA nick end labeling method and immunohistochemistry. Apoptosis was associated with high immunoreactivity of proliferating cell nuclear antigen. Bcl-2 expression correlated with a low apoptotic index and less malignant behavior of tumors. Prognostically, Bcl-2 expression was associated with a better prognosis, whereas p53 expression was the most important prognostic risk factor. Thus, apoptosis in gastric carcinomas is associated with cell proliferation, and Bcl-2 expression may have a prognostic importance as well as p53 expression.