A distinct function of the retinoblastoma protein in the control of lipid composition identified by lipidomic profiling.

Here, by combining lipidomics with transcriptome analysis, we demonstrate that Rb depletion in mouse embryonic fibroblastss induces significant alterations in their lipid composition. We discovered that Rb depletion induced increase in lysophosphatidylserine, diacylglycerol (DAG), fatty acid (FA), acylcarnitine, phosphatidylcholine (PC), arachidonoyl ethanolamine, and decrease in phosphatidylglycerol, monoacylglycerol, without change in total lipid per protein levels. Analysis of the acyl chain composition of DAG, PC and phosphatidylserine revealed increase of saturated and mono-unsaturated acyl chains with specific carbon chain length. Consistently, we observed that Rb depletion increased the levels of fatty acids with the corresponding carbon chain length and number of carbon-carbon double bondssuch as myristic acid (14:0), palmitic acid (16:0), stearic acid (18:0) and all forms of FA 18:1. Microarray analysis revealed that Rb depletion induced significant upregulation of enzymes involved in elongation and desaturation of fatty acids. Among these, we found that elongation of long chain fatty acid family member 6 (Elovl6) and stearoyl-CoA desaturase 1 (Scd1) are the most robustly controlled by Rb possibly through E2F and sterol regulatory element-binding protein transcription factors. Depletion of Elovl6 or Scd1 significantly suppressed colony formation, sphere formation and xenograft tumor growth of Rb-deficient tumor cells. Suppression of self-renewal by the SCD1 inhibitor was rescued upon supplementation of the mono-unsaturated fatty acids generated by this enzyme. This study suggests a novel role for Rb in suppressing the malignant progression of tumors by controlling the lipid composition.

The RB-IL-6 axis controls self-renewal and endocrine therapy resistance by fine-tuning mitochondrial activity.

Retinoblastoma (RB) protein inactivation during tumor progression is often associated with acquisition of immature phenotypes and resistance to therapy. Determination of an RB inactivation signature in a context of gaining undifferentiated phenotype in a p53-null sarcoma system revealed a critical role for interleukin (IL)-6. Using a Gene Set Enrichment Analysis (GSEA), we discovered that poorly differentiated breast cancers are enriched for this RB inactivation signature. Accelerated IL-6 secretion following RB inactivation in an RB-intact luminal-type breast cancer cell line MCF-7 promoted a positive feed forward loop between IL-6 and STAT3 driving tumor growth and endocrine therapy resistance. In addition, some of RB-intact basal-like type breast cancer cell lines exhibited a similar phenotype following RB depletion. The mechanism whereby RB inactivation increases IL-6 production in MCF-7 cells appeared to involve fatty acid oxidation (FAO)-dependent mitochondrial metabolism and c-Jun NH(2)-terminal kinase (JNK). In addition, IL-6, via STAT3-mediated feedback to mitochondria, autonomously adjusts mitochondrial superoxide to levels suitable to maintain stem cell-like activity. The gene expression profile of luminal-type breast cancer patients with low RB expression revealed high enrichment of genes involved in mitochondrial respiration and downstream targets of IL-6. These findings unveiled an unexpected strategy whereby RB suppresses malignant features of cancer cells through metabolic reprogramming and cell-autonomous inflammation.

Prevalence of patent foramen ovale with right-to-left shunting in dogs with pulmonic stenosis.

BACKGROUND: Right-to-left (R-L) shunt caused by patent foramen ovale (PFO) concurrent with pulmonic stenosis (PS) is considered common, although there is a lack of published evidence. OBJECTIVES: To investigate the prevalence of R-L shunt caused by a PFO in dogs with PS. ANIMALS: Thirty-one client-owned dogs with PS, without obvious extracardiac disease detected on the clinical examinations. METHODS: Case control study: R-L shunt probably caused by PFO was diagnosed when IV injected microbubbles appeared at the left atrial level with an intact atrial septum on echocardiography (bubble-positive dogs). The severity of PS concurrent tricuspid regurgitation (TR), relative thickness of the right ventricle, and relative right atrial area were compared between bubble-positive and bubble-negative dogs. RESULTS: The prevalence of R-L shunts caused by PFO was 39% (12 of 31 cases). The instantaneous pressure gradient (PG) across the pulmonic valve and relative thickness of the right ventricle were significantly increased in bubble-positive compared with those in bubble-negative dogs. None of the dogs with mild or moderate PS (pressure gradient < 80 mm Hg, n = 2) demonstrated R-L shunt. The prevalence of TR in bubble-positive dogs was significantly higher than that in bubble-negative dogs. DISCUSSION AND CLINICAL RELEVANCE: Patent foramen ovale PFO with R-L shunt was more common in dogs with very severe PS and absent in dogs with mild PS.

Morphology and rheology of an immiscible polymer blend subjected to a step electric field under shear flow.

We have investigated the structural change of an immiscible polymer blend in shear flow when subjected to a step electric field. During the process three-dimensional images were successfully constructed with a confocal scanning laser microscope and at the same time the transient shear stress was also measured. The interface tensor was calculated from the images. Several factors are incorporated into the shear stress, such as the bulk viscosity, the interfacial tension and the Maxwell stress. We performed an experiment to separate the Maxwell stress from the total shear stress. The results are discussed in terms of the interface tensor.

Relationship between real-time monitoring of the graft motility and mucosal histology in swine intestinal transplantation.

We studied the correlation between the motility and the mucosal histology of the small bowel seeking to detect rejection in an early stage by real-time monitoring using a swine model. Intestinal transplantation (ITx) was performed orthotopically using FK506 immunosuppression. The distal about 20 cm segment of the allograft was exteriorized as a Thiry-Vella stoma for biopsies. Strain gauge (SG) force transducers were attached to the graft for real-time monitoring of graft motility. Pigs without ITx were used as controls (group 1). Rejection was classified into four groups by histologic findings: nonrejection (group 2), mild rejection (group 3), moderate rejection (group 4), and severe rejection (group 5). Migrating motor complex (MMC) phase III was analyzed for the following parameters: duration, amplitude, interval, motility index, velocity, and frequency of propagation. In group 2, all parameters were almost the same as those for group 1. In contrast, groups 4 and 5 showed most parameters significantly lower than those in group 1. In group 3, the contractility of the MMC was not significantly altered, but the frequency of the propagation was decreased significantly. In conclusion, graft motility detected by a real-time SG method correlated with the grade of mucosal histology. This method is useful to detect rejection at an early stage by examining the frequency of MMC propagation.

[Quality control (QC) of CT on rail system (FOCAL Unit) with a micro-multi leaf collimator (mMLC) using new GafChromic film for stereotactic radiotherapy].

PURPOSE: Recent years, CT on rail system was reported to be useful as a tool for image-guided radiotherapy (IGRT). This system was clinically developed with the aim of stereotactic irradiation (STI) for brain, lung, liver, prostate and other sites. Quality assurance and quality control (QC) is an important issue in CT on rail system to assure geometric accuracies. The purpose of this study is to estimate the geometric accuracies of our CT on rail system using a detachable micro-multi leaf collimator (mMLC) with new type radiochromic films. Carrying out our original QC program, translational errors, setup reproducibility, beam misalignment and beam characteristics were evaluated. METHODS AND MATERIALS: We have studied with CT on rail system (FOCAL unit, Toshiba Medical systems, Tokyo, Japan) and mMLC unit (Accuknife, Direx Inc., Tokyo, Japan). We have developed original alignment phantom and small steel markers (2 mm phi) were implanted on its surface at certain intervals. Firstly, we have evaluated the accuracy of self-moving CT gantry and CT resolutions for cranio-caudal directions by changing slice thickness. And then using the phantom, we have measured the accuracy and reproducibility of geometric isocenter of the linac side and the CT gantry side by scanning the phantom. We have also measured the geometric changes of the common treatment couch by weight-loaded test (up to 135 kgw). To estimate dosimetric and geometric accuracies with the mMLC unit, the misalignment of the beam axes (gantry, collimator and couch rotation axis), mMLC leaf positions, and dose distributions for the verification plan were measured with new type GafChromic films (GafChromic-RTQA, ISP Inc., USA) and cylindrical phantom. The dose characteristics of the GafChromic film were also evaluated. RESULTS: The reproducibility of the self-moving CT gantry have a good agreement within 1 mm. Weight-load test have shown a good reliability within 2 mm at the common treatment couch. The translational precision of the common treatment couch was 0.0 +/- 0.1 mm at linac side and -0.2 +/- 0.5 mm at CT gantry side. The misalignments of beam axes have been kept within 0.4 mm at maximum. Gap test have shown the accuracies of the mMLC leaf positions, which is needed to keep within 1 mm by a routine calibration. CONCLUSIONS: To practice quality control program for the FOCAL unit and the mMLC unit is essential for a regular interval to reduce systematic errors. New type radiochromic film would be useful for a verification tool as alternative to conventional film.

Endoscopic diverticulotomy for symptomatic pediatric esophageal diverticula.

There have so far been few reports on esophageal diverticulum in children. We experienced two symptomatic pediatric cases with esophageal diverticulum. Our cases manifested high fever and dysphagia with chest pain during swallowing. The patients underwent endoscopic diverticulotomy. The septum between the diverticulum and the esophagus was cut using the argon plasma coagulation (APC 3000) system. We recommend an endoscopic diverticulotomy as an effective treatment modality for such symptomatic cases.

Immunoglobulin improves a model of acute motor axonal neuropathy by preventing axonal degeneration.

BACKGROUND: The action mechanism of IV immunoglobulin (IVIg) for Guillain-Barré syndrome has yet to be clarified. OBJECTIVE: To evaluate clinical, histologic, and immunologic effects in a disease model of acute motor axonal neuropathy (AMAN) treated by IVIg. METHODS: Rabbits were sensitized with gangliosides including GM1 and divided randomly into two groups at disease onset. One group received IV homologous gamma-globulin (400 mg/kg/day) for 5 days (n = 15), and the other received saline (n = 15). Disease severity was scored (0 to 13 points) daily. Sixty days after onset, anti-GM1 antibodies were tested by ELISA, and the number of degenerative axons was counted in spinal anterior roots. RESULTS: Between both groups at onset, there was no difference in any characteristics including clinical score. The IVIg group had faster recovery than the saline group (p = 0.03). The percentage of rabbits that improved by a score of < or =4 was higher in the IVIg (53%) than in the saline (13%) group 60 days after onset (p = 0.03). Anti-GM1 IgG titers 60 days after onset did not differ between the groups. The anterior roots of rabbits surviving 60 days after onset showed lower frequency of axonal degeneration in the IVIg-treated (n = 11; mean 4.5%) than in the saline-treated (n = 8; mean 11.1%) rabbits (p = 0.01). CONCLUSIONS: The therapeutic efficacy of IVIg in an AMAN model was confirmed. IVIg may not affect the production or catabolism of anti-GM1 IgG, but it may prevent axonal degeneration of motor nerves.

Real-time monitoring for detecting rejection using strain gauge force transducers in porcine small bowel transplantation.

PURPOSE: The clinical results of small bowel transplantation (SBT) have not been satisfactory mainly because of the immunological barrier. It is important to detect the presence of and to perform adequate treatment of rejection as early as possible to improve graft survival. Therefore, we have established a pig model to monitor graft motility as a means to detect rejection in real time. METHODS: Orthotropic SBT was performed in 25 pigs using FK-506 (0.05 to 0.1 mg/kg/d) immunosuppression. The interdigestive motor patterns were evaluated using strain gauge force transducers (SG). Seven pigs without SBT were treated as controls (C). Animals that displayed migrating motor complex (MMC) activity as evidenced by duration, amplitude, and interval in the graft were alive more than 10 days with adequate oral feeding: the functional graft (FG) group. In contrast the rejection (R) group did not show these activities on data recorded within 10 days before death due to rejection. RESULTS: The FG group showed MMC propagated throughout the graft with all parameters almost the same as the control group except for the duration. In contrast, all parameters in the group R were significantly lower than those in group FG, suggesting that group R motility was obviously impaired by rejection. CONCLUSIONS: The SG method may afford real-time monitoring of transplanted bowel motility that could be useful to detect rejection after SBT.

Usefulness of anti-GQ1b IgG antibody testing in Fisher syndrome compared with cerebrospinal fluid examination.

Fisher syndrome (FS), a variant of Guillain-Barré syndrome (GBS), is a rare disorder, and there are few reported studies of a large number of patients with FS. Cerebrospinal fluid (CSF) albuminocytological dissociation was found in 59% of 123 FS patients during the first 3 weeks of illness, while serum anti-GQ1b IgG antibody was positive in 85%. Whereas the incidence of CSF albuminocytological dissociation increased from the first to second weeks in FS, anti-GQ1b IgG antibody peaked in the first week, but there was no CSF albuminocytological dissociation. Statistically, anti-GQ1b antibody testing was superior to a CSF examination in supporting a diagnosis of FS during the first 3 weeks of illness, especially in the first week.

Effects of omega-3 fatty acids in rat allogenic small intestinal transplantation.

The aim of this study was to estimate the effect of omega-3 fatty acids on the recipient and graft immune response after rat allogenic small intestinal transplantation. Seven-week-old Lewis rats were randomly assigned to one of three groups according to the diet received: an FO group (fish oil supplemented), an SB group (soy bean oil supplemented) or a control group (normal rat chow). The recipient Lewis rats were each given their respective group diet for 12 days, and then, on the 19th day of gestation, a 2 cm jejunum from the donor fetal Fischer rat was transplanted into the abdominal wall of the recipient rats using a non-vascular anastomotic technique. The recipient rats were killed on day 2 after transplantation, and the recipient plasma IL-2, IFN-gamma and IL-1 beta levels were determined. In addition, the histological findings of the graft were analyzed. The cytokine levels of the FO group were significantly lower than the other two groups. In order to determine the grade of rejection, the morphological findings were blindly graded on a scale of 0-4. The mean grade of the FO group was also significantly lower than the other two groups. Omega-3 fatty acids are therefore considered to have an immunosuppressive effect on rat allogenic small intestinal transplantation based on the recipient plasma IL-1 beta, TNF and IL-2 levels and the histological findings of the grafts.

[Surgical management of acute type A aortic dissection with a complaint of disturbance of consciousness; report of a case].

A 50-year-old female was admitted to our hospital with a chief complaint of disturbance of consciousness (DOC). Left-sided hemiparalysis was noted on examination and cerebral infarction was diagnosed with total occlusion of the right common carotid artery revealed by cerebral angiography. Pharmacological thrombolysis (urokinase 720,000 U) was performed. Dissection of the right common carotid artery was noted after successful thrombolytic therapy. Enhanced chest computed tomography (CT) showed the acute type A aortic dissection involving the cerebral artery. Ascending aortic replacement was performed 4 days after the thrombolytic therapy to avoid brain edema and hemorrhagic infarction during cardiopulmonary bypass. The postoperative course was uneventful. In the case of acute type A aortic dissection with DOC, proper indication and optimal timing of the operation may help to improve patient survival.

Protective effect of sulfobutyl ether beta-cyclodextrin on DY-9760e-induced hemolysis in vitro.

The hemolytic behavior of a novel cytoprotective agent, DY-9760e (3-[2-[4-(3-chloro-2-methylphenyl)-1-piperazinyl]ethyl]-5,6-dimethoxy-1-(4-imidazolylmethyl)-1H-indazole dihydrochloride 3.5 hydrate) was investigated using rabbit erythrocytes. Further, the effects of water-soluble cyclodextrin derivatives, such as 2-hydroxypropyl-beta-cyclodextrin (HP-beta-CyD) and sulfobutyl ether of beta-cyclodextrin (SBE-beta-CyD), on the hemolytic activity of DY-9760e were studied. DY-9760e induced hemolysis at concentrations >0.2-0.3 mM in phosphate buffered saline (PBS) of pH 4.0 and 6.0, where DY-9760e is predominantly in dicationic and monocationic forms, respectively. The hemolytic activity of the monocationic DY-9760e was higher than that of the dicationic species, and the hemolysis at pH 4.0 involved the formation of methemoglobin. DY9760e induced the morphological change of erythrocytes towards membrane invagination at both pH 4.0 and 6.0. SBE7-beta-CyD significantly suppressed the DY-9760e-induced hemolysis and morphological change at both pH 4.0 and 6.0, as well as the formation of methemoglobin at pH 4.0. On the other hand, HP-beta-CyD suppressed only the hemolysis, but neither the morphological change nor the formation of methemoglobin. In addition, the inhibitory effect of SBE7-beta-CyD on the hemolysis was greater than that of HP-beta-CyD. The superior inhibitory effect of SBE7-beta-CyD on the DY-9760-induced hemolysis, the morphological change, and the formation of methemoglobin may be attributable to the formation of a stable inclusion complex with DY-9760e and to the weaker hemolytic activity of SBE7beta-CyD than HP-beta-CyD. These results suggest potential use of SBE7-beta-CyD as a parenteral carrier for DY-9760e.

Preoperative cisplatin for initial treatment of limb osteosarcoma: its local effect and impact on prognosis.

PURPOSE: The significance of preoperative cisplatin (CDDP) as a single agent has not been assessed in terms of its effect on prognosis. The purpose of this multi-institution study was to assess the local effect of preoperative CDDP as a single agent as well as its impact on the prognosis of limb osteosarcoma. PATIENTS AND METHODS: The study group comprised 44 patients with stage IIB limb osteosarcoma who were treated with single-agent CDDP as initial preoperative chemotherapy. Two to five courses of CDDP (mean 2.4 courses) were administered intravenously and/or intraarterially as an initial preoperative treatment. The mean dose of CDDP was 3.0 mg/kg (2.5-3.4 mg/kg). The effect of the treatment was evaluated clinically and histologically. RESULTS: The clinical and histological response rates to preoperative CDDP were 56.8% and 47.6%, respectively. The survival rate was 59.1% among all patients in the study, 64.0% among those with a grade III or IV clinical response, and 52.6% among those with a grade I or II clinical response, with no significant differences between the groups. The survival rate was 70% among patients with a grade III or IV histological response, and 54.5% among those with a grade I or II histological response, with no statistical differences between the groups. CONCLUSIONS: We consider that CDDP is a useful chemotherapeutic agent for preoperative induction therapy for osteosarcoma because of the excellent local effect observed. Good responders to preoperative CDDP showed a better survival rate, but a correlation between the local response to CDDP and the survival rate was not demonstrated statistically. Systemic multidrug chemotherapy should follow preoperative CDDP to diminish the microscopic foci of metastatic disease.

[A successful surgical case of chronic DeBakey IIIb dissecting aortic aneurysm associated with right-sided aortic arch].

A case of chronic DeBakey IIIb dissecting aneurysm associated with right-sided aortic arch and aberrant left subclavian artery is reported. A 51-year-old man was admitted to our institute for surgical treatment of chronic dissecting aortic aneurysm which occurred 3 years ago. A right posterolateral thoracotomy was made through the 4th intercostal space. Closure of entry was performed under temporary hypothermic circulatory arrest and then, this was followed by plication of false lumen under hypothermic CPB. Post-operative clinical course was uneventful and an intra-venous digital subtraction angiography demonstrated that false lumen in the thorax was completely closed. The patient was discharged from the hospital on 21th POD. Dissecting aortic aneurysm associated with right-sided aortic arch is extremely rare. In operative case, pertinent selection of operative methods considering anatomical diversity is required.

Digital subtraction angiogram registration method with local distortion vectors to decrease motion artifact.

We have been investigating registration methods for improving digital subtraction angiography (DSA) images to extract blood vessels by reducing artifacts due to body motion, such as rotation, contraction, and dilation. In this paper, we propose a new and simple DSA registration algorithm with local distortion vectors to reduce artifacts. According to the results, the proposed method works well for vascular system around the nasal cavity and the orbit of the head and neck DSA images, which cannot be observed clearly by conventional methods. Additionally, we have applied the proposed method to abdominal and leg DSA images.

The development of a pollen information system for the improvement of QOL.

The Kochi Prefecture Japanese Cedar and Cypress Pollen Information System (P-Net Kochi) was established in 1991 on the initiative of the Pharmaceuticals and Sanitation Division to improve the quality of life of people in Kochi Prefecture, particularly patients with pollinosis. Kochi Prefecture has the highest forests percentage (84%) in Japan. In addition, 40% of the population of the prefecture is concentrated in Kochi City. The average pollen count at 10 observation points an one year was about 20,000/cm2 during the observation period, but it exceeded 110,000/cm2 in 1995, when it was also high nationwide. Kochi Prefecture organized a system to promote people's understanding of pollinosis in connection with environmental problems, and to enlighten people on appropriate preventive measures. The system has been improved in the rapid and efficient transmission of information over the years with technological advances. The introduction of an automatic monitoring system not dependent on human labor and the support of the users proved to be indispensable for the maintenance of the system.

Inhibitory effects of novel hydrophilic cyclodextrin derivatives on nitric oxide production in macrophages stimulated with lipopolysaccharide.

PURPOSE: The objective of this study is to examine the effects of cyclodextrins (CyDs) on nitric oxide (NO) production in macrophages stimulated with lipopolysaccharide (LPS). METHODS: RAW264.7 cells, a mouse macrophage-like cell, were used. Cytotoxicity of CyDs was evaluated by WST-1 method. Nitrite, iNOS, and iNOS mRNA were determined by Griess method, Western blotting, and reverse transcription-polymerase chain reaction (RT-PCR) analysis, respectively. The interaction of LPS with CyDs was evaluated by utilizing a competitive inclusion phenomenon. The binding of FITC-labeled LPS to the surface of RAW264.7 cells was measured by a flow cytometry. RESULTS: Of 15 CyDs, 2,6-di-O-methyl-alpha-CyD (DM-alpha-CyD), and 2,6-di-O-methyl-3-O-acetyl-beta-cyclodextrin (DMA-beta-CyD) had greater inhibitory activity than did the other CyDs against NO production in RAW264.7 cells stimulated with LPS, without showing any cytotoxicity. DM-alpha-CyD and DMA-beta-CyD specifically inhibited the increase in iNOS and iNOS mRNA levels elicited by stimulation with LPS in RAW264.7 cells. DM-alpha-CyD and DMA-beta-CyD suppressed the binding of FITC-labeled LPS to the surface of cells, probably resulting in inhibitory effects on iNOS expression and NO production. DM-alpha-CyD had a greater interaction with RAW264.7 cells than did DMA-beta-CyD. The pretreatment of RAW264.7 cells with DM-alpha-CyD, not DMA-beta-CyD, decreased the LPS binding to the cell surface. The results suggested that the inhibitory mechanism of the LPS binding to the cell surface is different between DM-alpha-CyD and DMA-beta-CyD. CONCLUSIONS: The present results suggest that DM-alpha-CyD and DMAbeta-CyD attenuates NO production by inhibiting iNOS gene expression in RAW264.7 cells stimulated with LPS, probably due to the suppression of LPS binding to LPS receptors on the cells in the different way.