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.

[Cardiovascular surgery for patient with pulmonary dysfunction].

Pulmonary dysfunction is one of the major factors for postoperative pulmonary complication. Preoperative pulmonary function test reveals possible operative risk. Particularly, in the patient with stage Ⅲ and Ⅳ chronic obstructive pulmonary disease (COPD), open heart surgery with cardiopulmonary bypass is at higher risk of complication. In these patients, safe and reliable surgical procedures are required. Preoperatively, quitting smoking and incentive spirometry can reduce the risk of pulmonary complications. Furthermore, noninvasive positive pressure ventilation (NPPV) on the appropriate indication can help to improve the outcome in the patients with pulmonary dysfunction.

Compensatory cortical activation during performance of an attention task by patients with diffuse axonal injury: a functional magnetic resonance imaging study.

OBJECTIVE: To determine how cortical compensation occurs in higher cognitive systems during the recovery phase of diffuse axonal injury (DAI). DESIGN: 12 right-handed patients with a magnetic resonance imaging (MRI) lesion pattern compatible with pure DAI were identified. Pure DAI was defined as finding of traumatic microbleeds on T2*-weighted gradient-echo images in the absence of otherwise traumatic or non-traumatic MRI abnormalities. 12 matched healthy controls were also enrolled. Functional magnetic resonance imaging (fMRI) was used to assess brain activation during a working memory test (Paced Visual Serial Attention Test (PVSAT)). RESULTS: No significant group differences were observed in reaction times for the PVSAT. Although patients with pure DAI committed a few errors during the PVSAT, controls respond correctly to each probe. Controls showed activations in the left frontal gyrus, left parietal gyrus and right inferior parietal gyrus. Patients with pure DAI showed activations in the left inferior frontal gyrus, right inferior frontal gyrus and right middle frontal gyrus. Between-group analysis of the PVSAT task showed significantly greater activation of the right inferior frontal gyrus (BA 45) and right middle frontal gyrus (BA 9) in patient with pure DAI versus controls. CONCLUSIONS: Patients with pure DAI require compensatory activation of the contralateral (right) prefrontal region to carry out activities similar to healthy controls. These findings provide further evidence for the adaptive capacity of neuronal systems and brain plasticity during the recovery stages of DAI.

Rehabilitation technique facilitates association cortices in hemiparetic patients: functional MRI study.

We used fMRI to study brain activation with facilitative rehabilitation techniques (passive hand movements and visual feedback) in two patients with subcortical lesions. Two tasks were given in a sequence. The first task (trial 1) was repetitive hand grasping by the paretic hand at a rate of 0.5 Hz with the eyes closed. The second task (trial 2), the facilitative rehabilitation technique, included task 1 plus support by a trainer to move the paretic hand with the eyes open to get visual feedback of the movement. The data were analyzed by a subtractive method. When task 1 was subtracted from task 2, it was found that the bilateral visual cortex, contralateral premotor cortex and posterior parietal cortex were involved with the passive hand movement and visual feedback. These facilitative rehabilitation techniques may integrate networks between sensory information and motor commands, and lead to functional reorganization.

Prognostic factors of nosocomial pneumonia in general wards: a prospective multivariate analysis in Japan.

To determine prognostic factors of nosocomial pneumonia in general wards, we performed prospective clinical study using multivariate statistical analysis. Eighty patients with nosocomial pneumonia in our units were enrolled in the study between December, 1996 and January 1998. Clinical setting and severity of pneumonia were evaluated, and laboratory data were collected at the occurrence of nosocomial pneumonia. Death due to nosocomial pneumonia occurred in 29 of 80 patients (mortality rate 36%). Univariate analysis showed the following factors associated with mortality: the presence of an ultimately or rapidly fatal underlying condition, prior antibiotics use, use of antacids, presence of 'high-risk' micro-organisms, sepsis, respiratory failure, multiple organ failure, bilateral chest X-ray infiltrates, a Simplified Acute Physiology Score (SAPS) index > or = 11, albumin < 3.0 g dl(-1), and lactate dehydrogenase (LDH) > or = 796 IUI(-1). Furthermore, multivariate analysis identified three factors significantly associated with mortality: the presence of an ultimately or rapidly fatal underlying condition [odds ratio (OR)=7.0; 95% confidence interval (CI)=1.2-41.1; P=0.03]; SAPS index > or = 11 (OR=7.6; 95% CI=1.1-51.9, P=0.04); LDH > or = 796 IUI(-1) (OR=28.2; 95% CI=2.0-406, P=0.01). Our study indicates that host factors and disease severity factors are important prognostic factors of nosocomial pneumonia in general wards.

Beta(3)-adrenoceptor agonists for the treatment of frequent urination and urinary incontinence: 2-[4-(2-[[(1S,2R)-2-hydroxy-2-(4-hydroxyphenyl)-1-methylethyl]amino]ethyl)phenoxy]-2-methylpropionic acid.

In a search for novel analogues of beta(3)-adrenoceptor (AR) agonists relaxing the bladder for treatment of urinary dysfunction, 2-[4-(2-[[(1S,2R)-2-hydroxy-2-(4-hydroxyphenyl)-1-methylethyl]amino]ethyl)phenoxy]-2-methylpropionic acids (1a-e), into which a fibrate-like structure had been incorporated, were synthesised. Compound 1a was found to be a selective beta(3)-AR agonist in functional assays using the ferret detrusor (beta(3)-AR), rat uterus (beta(2)-AR), and rat atrium (beta(1)-AR); beta(3): EC(50)=7.8 nM, beta(2): IC(50)=7,300 nM, beta(1): EC(20)=23,000 nM. The introduction of a chlorine atom or methyl substituent at the ortho-position on the phenyl ring of 1a further improved beta(3)-AR selectivity. In an in vivo study, 1a lowered intrabladder pressure (ED(50)=31 microg/kg) in rats, without increasing heart rate, in keeping with the in vitro results. Consequently, it is proposed that 1a and its analogues (1b-e), possess beta(3)-AR agonistic activity in the absence of undesirable beta(1)- or beta(2)-AR mediated actions, and may be useful for clinical treatment and pharmacological studies.

Effect of cryopreservation period on rat tracheal allografts.

BACKGROUND: The effect of cryopreservation on tracheal allogenicity is still unclear. Therefore, in this study, we assessed the effect of cryopreservation period on tracheal allografts in 62 rats. METHODS: Each transplant consisted of a 3-ring segment of the trachea harvested from 8 Lewis rats, immersed in preservation solution, and cryopreserved and stored in a Bicell biofreezing vessel in a deep freezer at -80 degrees C. Six tracheal grafts without cryopreservation that underwent a heterotopically syngeneic transplantation into the omentum served as controls. Forty-eight tracheal segments were randomly assigned to 8 groups according to period of cryopreservation, which ranged from 0 to 12 months (0, 0.5 [2 weeks], 1, 2, 3, 6, 9 and 12 months). The cryopreserved grafts were then thawed and heterotopically implanted into the omentum of recipient Brown-Norway rats. After 28 days, the tracheal segments were then evaluated histologically. RESULTS: All isografts in Group 1 were intact, whereas the allografts undergoing a particularly shorter period of cryopreservation showed a more stenotic lumen. Prolonged periods of cryopreservation tended to show decreasing tendencies of viability of chondrocytes, mononuclear cell infiltration and sub-epithelial thickness, whereas all allografts showed a uniformly denuded epithelium, irrespective of the length of cryopreservation. CONCLUSIONS: A longer period of cryopreservation may help to maintain a better patency of tracheal allografts by preventing an allogeneic response. Reduced tracheal allogenicity may be associated with a decreased viability of chondrocytes by cryopreservation.

Discovery of novel N-phenylglycine derivatives as potent and selective beta(3)-adrenoceptor agonists for the treatment of frequent urination and urinary incontinence.

With a novel assay using isolated ferret detrusor to estimate beta(3)-adrenoceptor agonistic activity, we found that a series of glycine derivatives of ritodrine, a beta(2)-adrenoceptor agonist, are potent beta(3)-adrenoceptor agonists, with excellent selectivity versus beta(1) and beta(2) subtypes. Substitution of halogens in the phenyl ring increased potency and selectivity for the beta(3)-adrenoceptor, and this was dependent upon the position of the halogens. The chlorine-substituted derivatives 3f-i exhibited potent beta(3)-adrenoceptor-mediated relaxation of ferret detrusor (EC(50) = 0.93, 11, 14, and 160 nM) and higher potency at beta(3)-adrenoceptors than at beta(1) or beta(2). The intravenous administration of 3h significantly reduced the urinary bladder pressure in anesthetized male rats (ED(50) = 48 microg/kg) without cardiovascular side effects. This article is the first report of structure-activity relationships (SAR) concerning beta(3)-adrenoceptor agonists as agents for the treatment of urinary frequency and incontinence.

Feasibility of cryopreserved tracheal xenotransplants with the use of short-course immunosuppression.

OBJECTIVE: We evaluated the feasibility of discordant xenotransplantation of the cryopreserved trachea with intermittent immunosuppression to help solve the shortage of donor tracheas. METHODS: Two experiments were performed with heterotopic transplantation models in 14 guinea pigs and 85 rats. So that the minimal dose of FK506 for viable fresh xenografts could be determined, FK506 was given in escalating doses (0, 1.5, 2.5, and 3.5 mg/kg) for recipient animals after xenogeneic transplantation. With the goal of obtaining a long-term survival of the xenografts, the effect of cryopreservation on xenografts was assessed and thereafter different cycles of immunosuppression every third week were evaluated in fresh or cryopreserved xenografts in the second experiment. RESULTS: An FK506 dosage of more than 2.5 mg/kg per day was much more effective than smaller dosages, as demonstrated by morphologic assessment. A higher dosage of FK506 potentially delayed the rejection of xenografts and can thus maintain tracheal xenograft viability for less than 4 weeks in rat recipients. In experiment 2, the cryopreserved xenografts showed less histologic viability than fresh xenografts but greater patency of the lumen. The patency of cryopreserved xenografts was favorably maintained for a longer period than that of fresh xenografts with either the same number or more cycles of immunosuppression. CONCLUSIONS: We conclude that the synergistic effect of cryopreservation and adequate intermittent immunosuppression may enable tracheal xenografts to remain viable over longer periods.

Visual symptoms in epilepsy and migraine: localization and patterns.

PURPOSE: To clarify differences in the localization of visual symptom patterns in epilepsy and migraine, we analyzed patient-generated illustrations of visual symptoms. METHODS: Patients were asked to draw their visual symptoms from memory using marker pens of 12 colors. All patients illustrated their symptoms on a template sheet representing the binocular visual field. We analyzed a total of 67 illustrations from 54 patients aged 6-40 years: 28 with epilepsy, 23 with migraine, and 3 with migraine-epilepsy syndrome. RESULTS: With respect to positive visual manifestations, those of epileptic patients were predominantly centrally localized (20 of 24, 83%), whereas those of migraine patients were predominantly peripherally localized (10 of 13, 77%) (p < 0.0001). With respect to negative visual symptoms, those in epilepsy were commonly diffuse (10 of 14, 71%) compared with those in migraine, which were peripheral (9 of 12, 75%) (p < 0.05). CONCLUSIONS: Results of this study suggest that the localization of visual symptoms differs between epilepsy and migraine.

Short-course immunosuppression using FK506 for rat tracheal allografts.

BACKGROUND: A minimizing immunosuppression after a tracheal allotransplantation is desirable. METHODS: We examined the usefulness of a short-course of immunosuppression after tracheal allotransplantation in rat. Each transplant consisting of a 5-ring segment was heterotopically implanted into the omentum. Four animals underwent a syngeneic transplantation and thus served as controls (Group A). Thirty animals underwent an allogeneic transplantation and were randomly classified into 4 groups as follows: No immunosuppression (Group B, n=6), treatment with 0.5 mg/kg of Tacrolims (FK506) (Group C, n=8), 1.0 mg/kg of FK506 (Group D, n=8), and 1.5 mg/kg of FK506 (Group E, n=8). Different doses of FK506 were administered intramuscularly for only three consecutive days after heterotopic tracheal allotransplantation. The serum levels of FK506 were then investigated 3, 7, 14, 21, and 28 days after transplantation in groups C, D, and E. All rats were killed 28 days after transplantation and then the implanted tracheae were harvested, and evaluated histologically. RESULTS: All animals survived for the protocol period. The graft morphology of Group E was significantly better than that of groups B, C, and D regarding both macro- and microscopy, and also showed the same findings as that of Group A, except for low-grade mononuclear cell infiltration. Only in Group E, the FK506 blood level was maintained at over 0.5 ng/ml, which is the lowest detectable limit in this assay, until 21 days after transplantation. CONCLUSIONS: We thus conclude that 1.5 mg/kg of FK506 which was administered for only three consecutive days after surgery may be used to maintain the morphology of tracheal allografts in rats for 28 days after transplantation.

[Recurrent tongue biting due to hypnic myoclonia in infancy].

We report a 15-month-old female infant who had recurrent tongue biting due to hypnic myoclonia. She began to bite her tongue and bleed when she was 9 months old. The tongue biting was accompanied by generalized myoclonus and was seen only during drowsiness. On admission at 15 months of age, she was observed by video-EEG monitoring. There were hypnic jerks with powerful closure of the mouth and severe screaming, but no accompanying evidence of epileptic discharges. The tongue biting with hypnic myoclonia disappeared spontaneously at 22 months of age, and was considered to be an age-dependent phenomenon related to the maturation of the brain and to the development of oral functions.

Chronic expanding hematoma in the chest.

We report the successful surgical treatment of chronic expanding hematoma in the chest. Four patients who had previously undergone artificial pneumothorax, thoracoplasty or tumor extirpation more than 30 years earlier recently became aware of a slowly growing mass. Chronic expanding hematoma which developed into very large masses over a long period of time were thus successfully resected. These patients are now all in good health with no recurrence after the operation. It is important to monitor such patients' laboratory data for hemostasis including the platelet cell counts, the % prothrombin time and the D-dimer, both before and immediately after operation, and the intraoperative bleeding volume.

Combined effects of inhaled nitric oxide and positive end-expiratory pressure during mechanical ventilation in acute respiratory distress syndrome.

We studied the combined effects of inhaled nitric oxide (INO) and positive end expiratory pressure (PEEP) during mechanical ventilation in patients with acute respiratory distress syndrome (ARDS). Eleven patients received 0 and 4 parts per million of INO in random order for 30 min at PEEP levels of 0, 5, and 10 cm H2O. Respiratory and cardiovascular parameters were measured. The addition of INO and PEEP significantly improved arterial oxygenation (p < 0.005 and p < 0.0001, respectively). The combined effect of INO and PEEP on arterial oxygenation was remarkable during 10 cm H2O PEEP. There was synergistic effect on arterial oxygenation by combining INO and 10 cm H2O PEEP. The present study showed that the combination of INO and 10 cm H2O PEEP enhanced arterial oxygenation in patients with ARDS.

A novel mutation of KCNQ3 (c.925T-->C) in a Japanese family with benign familial neonatal convulsions.

At present, only one mutation of KCNQ3, a KCNQ potassium channel gene, has been identified as a cause of benign familial neonatal convulsions type 2 (BFNC2). We found a T to C substitution (c.925T-C) on one allele of affected individuals in a Japanese family with BFNC but not on 200 alleles from healthy subjects. c.925T-->C replaced Trp309, a conserved residue within the P-loop of the KCNQ potassium channel family that holds the channel pore open, with an Arg (W309R). We report c.925T-->C as the second mutation of KCNQ3 responsible for BFNC2.

Limit of warm ischemia time before cryopreservation in rat tracheal isografts.

BACKGROUND: The viability of cadaveric tracheal grafts undergoing cryopreservation is still unclear. We evaluated the limit of warm ischemia time before cryopreservation in rat tracheal isografts. METHODS: Each isograft was harvested from donor rats 0 to 48 hours (0, 6, 12, 18, 24, and 48 hours) after circulatory arrest, immersed in the preservative solution, and stored in a deep freezer until reaching -80 degrees C and then was kept in liquid nitrogen for 3 months. Heterotopic transplantation into the omentum was performed after the isografts were thawed. Graft morphology 3 months after transplantation was assessed. RESULTS: The stepwise increase of warm ischemia time significantly reduced graft survival. A prolonged period of warm ischemia had a degenerative effect on both the epithelium and cartilage. The morphology of the epithelium and cartilage in isografts undergoing warm ischemia for less than 18 hours was better preserved, whereas it deteriorated in isografts undergoing warm ischemia for more than 24 hours. CONCLUSIONS: We thus conclude that the permissible period of warm ischemia before 3-month cryopreservation to maintain tracheal isograft viability is 18 hours in rats.

[Case report of muscle cramp versus focal epilepsy].

We report here a boy suffering from muscle cramps in the right upper extremity. At 32 days of age, he developed purulent meningitis followed by paresis of the right upper extremity. From infancy he had intermittent episodes myoclonus-like involving the right hand. Since he also had true epileptic seizures with loss of consciousness, ocular deviation, and vomiting at 6 and 8 years of age, he was treated with anti-epileptic drugs as therapy for focal motor seizures. At 6 years of age, these episodes increased in frequency. The cramps spread from the right hand to involve the entire upper extremity with pain. At the age of 10, he was referred to Hirosaki University Hospital and was admitted. Using closed circuit television with continuous EEG and EMG monitoring we observed during his episodes repeated EMG abnormalities consisting of continuous discharges of polyphasic motor unit potentials, but no epileptic EEG discharges. We diagnosed these episodes as muscle cramp. His muscle cramps were controlled by medication with muscle relaxants and Chinese medicines. This case illustrates that the differential diagnosis between muscle cramps and epileptic seizures is important for proper treatment.

Maximal period of cryopreservation with the Bicell biofreezing vessel for rat tracheal isografts.

OBJECTIVE: The maximal period of cryopreservation for the trachea is still unsolved. We assessed the maximal period of cryopreservation using the Bicell biofreezing vessel as an easy and cheap slow-freezing instrument for viable tracheal grafts in 95 rats. METHODS: Each isograft was harvested from 17 donor rats, immersed in the preservative solution, and stored in a Bicell device in a deep freezer at -80 degrees C. The tracheal isografts were then randomly assigned to 9 groups according to cryopreservation periods ranging from 0 to 12 months. Included in the 9 groups were 2 subgroups (n = 6 per subgroup) that were observed immediately after being thawed and 1 month after heterotopic transplantation into the omentum after being thawed. Four subgroups (n = 6 per subgroup) were added according to the cryopreservation period for 1, 3, 6, and 12 months to evaluate the graft morphology 3 months after being thawed and transplanted heterotopically. RESULTS: A prolonged period of cryopreservation had a degenerative effect on both the epithelium and cartilage. One month after transplantation, degeneration was more pronounced in the cartilage than in the epithelium, as characterized by the viable chondrocyte ratio and the epithelial score of isografts undergoing cryopreservation for more than 9 months. Three months after transplantation, the morphology of the epithelium and cartilage in isografts undergoing cryopreservation for less than 3 months was better preserved, whereas the morphology of both deteriorated in isografts undergoing cryopreservation for more than 6 months. CONCLUSIONS: We conclude that the permissible period of cryopreservation to maintain tracheal isograft viability in this simple system using a Bicell biofreezing vessel is 3 months.