Gemcitabine Induces Radiosensitization Through Inhibition of RAD51-dependent Repair for DNA Double-strand Breaks.

BACKGROUND/AIM: Gemcitabine (GEM) is used in clinical chemo-radiotherapy; however, the mechanism that contributes to enhanced radiosensitivity by GEM is not fully-understood. We evaluated the effect of GEM on radiosensitization in pancreatic cancer cell lines. MATERIALS AND METHODS: Pancreatic cell lines PK-59 and PK-45p were used. A total of 5 µM GEM for 4 h were administered pre- or post-gamma irradiation. RESULTS: Enhanced cell killing effects by GEM in radiotherapy were observed for pre-treatment but not post-treatment GEM. We focused on the dynamics of RAD51 and phospho-H2AX foci after irradiation. Significantly higher numbers of phospho-H2AX foci were observed in GEM pre-treated cells than in untreated cells after irradiation. We also found inhibition of the formation and degradation of RAD51 foci by GEM pre-treatment. The radiosensitizing effect of GEM was suppressed by knockdown of RAD51. CONCLUSION: RAD51-dependent homologous recombination is one of the key targets in the GEM-induced radiosensitizing effect.

Expiratory computed tomographic techniques: a cause of a poor rate of change in lung volume.

Ninety-nine patients (29 males and 70 females; mean age, 57.1 years; range, 22-81 years) were included in this study to evaluate the factors affecting smaller lung volume changes in expiratory high-resolution computed tomography performed to depict air trapping. All patients underwent inspiratory and expiratory chest thin-section CT examinations and pulmonary function tests. Air trapping on CT images was graded subjectively. All variables (age, sex, diagnosis, pulmonary function index, and air trapping score) were compared with the degree of change in lung volume between the inspiratory and expiratory CT examinations. The variables affecting a lower degree of volume change were vital capacity, forced vital capacity (FVC), forced expiratory volume in 1 s (FEV1.0), and the FEV1.0/FVC ratio. Bronchiolitis obliterans was the dominant diagnosis in patients with insufficient degrees of breath holding and in patients with negative air trapping scores despite an abnormal air trapping index. An insufficient degree of lung changes between inspiration and expiration on CT examinations represented bronchiolitis obliterans, which resulted in low FEV1.0 and FEV1.0/FVC values. Changes in the time gap from the announcement of exhalation and breath holding to the start of scanning most effectively indicated air trapping in patients with bronchiolar disorders.

[Effective treatment with tranexamic acid for chronic disseminated intravascular coagulation associated with aortic dissection].

An 82-year-old female with a history of aortic dissection was admitted to our hospital for evaluation of thrombocytopenia and a bleeding tendency. Blood coagulation test data demonstrated that the patient had disseminated intravascular coagulation (DIC) secondary to aortic dissection. We initiated anti-fibrinolytic therapy with tranexamic acid. After this anti-fibrinolytic therapy, thrombocytopenia and the levels of fibrinogen and fibrinogen degradation products improved. In addition, we reviewed 7 other patients who were treated with tranexamic acid for DIC associated with aortic dissection. Anti-fibrinolytic therapy with tranexamic acid is effective for treating DIC secondary to aortic dissection.

Central nervous system lymphoma newly developed 12 years after remission of an ocular adnexal lymphoma.

Recurrence of non-Hodgkin's lymphoma more than 5 years after the initial diagnosis is rare. When late relapse occurs, it is difficult to determine whether it is a true recurrence or a new lesion. We experienced a case of an 81-year-old woman who developed central nervous system (CNS) lymphoma 12 years after remission of ocular adnexal lymphoma. Both showed the histology of diffuse large B-cell lymphoma. To elucidate whether the CNS lymphoma was clonally related to the first lymphoma, rearrangement of the immunoglobulin heavy chain genes of each lymphoma was studied using a polymerase chain reaction-based method. The results revealed that the sizes of the amplified products of the rearranged regions from the two lymphomas were different. This suggested different clonal origins of the lymphomas. It is clinically important to determine the origin of a second neoplasm because patients with a clonally related second lymphoma are usually treated with more intensive regimens, while those with a clonally unrelated lymphoma receive standard first-line therapy. The present case shows that, in the case of recurrent non-Hodgkin's lymphoma, not only histological confirmation but also genetic assessment is important to clarify the origin of the second lymphoma.

[Acute upper airway obstruction after extubation due to excessive anteflexion of the neck with occipitocervical fusion].

A patient developed upper airway obstruction immediately after tracheal extubation due to excessive anteflexion of the neck with occipitocervical fusion. A 59-year-old woman who had undergone mastectomy 17 years previously was scheduled for occipitocervical fusion for C2 vertebral metastasis. Retroflexion of her neck was restricted. Nasal intubation under sedation was performed using bronchial fiberscopy under fentanyl and propofol anesthesia. Emergence from anesthesia was smooth, and extubation was performed. Immediately after extubation, the patient could not breathe, and manual mask ventilation was impossible. Re-intubation was performed 30 minutes after the extubation. Oral fiberscopy revealed pharyngeal obstruction, and laryngeal edema was not observed. Fixation of her neck in excessive anteflexion was suspected to have caused her dyspnea. Therefore, re-operation was performed, and she was transferred to the intensive care unit under anesthesia. One day postoperatively, extubation was performed successfully with no dyspnea. Fixation of the neck in excessive anteflexion is one of the causes of upper airway obstruction after occipitocervical fusion. We must carefully observe cervical X-ray films to locate the upper airway obstruction, and careful extubation using a tube exchanger is strongly recommended in this operation.

All-trans retinoic acid displays multiple effects on the growth, lipogenesis and adipokine gene expression of AML-I preadipocyte cell line.

Adipose tissue is a potential site of retinoic acid (RA) action, but its physiological significance remains to be clarified. We have examined the effect of all-trans retinoic acid (ATRA) on growth and differentiation of preadipocytes, and on adipokine gene expression in mature adipocytes using human preadipocyte cell model, AML-I. Both ATRA and 9-cis RA induced growth arrest in AML-I preadipocyte at between 50 and 100 µM, which was accompanied by apoptosis. Western blotting showed a loss of NF-κB, Bcl-2 and p-Akt, and the accumulation of Bad and Akt in cytoplasm of ATRA-treated AML-I preadipocytes. Exposure of AML-I to ATRA or 9-cis RA increased intracellular lipid accumulation in a time-dependent manner compared to vehicle-treated cells. Expression of fatty acid synthase (FAS) and peroxisome proliferator-activated receptor-γ (PPAR-γ) proteins was increased in ATRA-treated cells. Thus, both ATRA and 9-cis RA promoted differentiation, inhibited proliferation and induced apoptosis in AML-I preadipocytes. ATRA also modulated adipokine expression by increasing the mRNA level of adipocytokines (adiponectin, leptin and LPL), and by inhibiting PAI-1 mRNA expression in mature AML-I adipocytes. The data suggest that ATRA exerts a wide range of effects--growth arrest, apoptosis, lipogenesis and modulation of adipokine gene expression--during the maturation of preadipocytes into adipocytes.

[A case of femoral neuropathy after radical ovariectomy].

We experienced a 55-year-old female patient who was diagnosed as femoral neuropathy after radical ovariectomy. An epidural catheter was introduced at T11-12 interspace without any problems and general anesthesia was induced and maintained. The operation ended uneventfully. On the first postoperative day, she noticed hypesthesia of the inner surface of her left thigh and could not raise the left leg. The symptom remained after the removal of epidural catheter on the second postoperative day, and the influence of insertion of the epidural catheter on the symptom was suspected. We performed neurological examinations and found weakness of the left quadriceps femoris muscle, weakness of the left patellar reflex, and weakness of touch sensation and cold sensation and hypalgesia on the anterior surface of the left thigh and the inner surface of the left lower leg. Those findings led us to diagnose with femoral neuropathy probably due to abdominal retractors or the operation itself, and insertion of epidural anesthesia could not be the cause of neuropathy. Her symptom was ameliorated with a conservative therapy after four months. We should perform fine neurological examinations when neurological complications occur, especially when we use epidural catheters, and also should have the knowledge about those complications.

IGF-1 induction by acylated steryl ß-glucosides found in a pre-germinated brown rice diet reduces oxidative stress in streptozotocin-induced diabetes.

BACKGROUND: The pathology of diabetic neuropathy involves oxidative stress on pancreatic ß-cells, and is related to decreased levels of Insulin-like growth factor 1 (IGF-1). Acylated steryl ß-glucoside (PR-ASG) found in pre-germiated brown rice is a bioactive substance exhibiting properties that enhance activity of homocysteine-thiolactonase (HTase), reducing oxidative stress in diabetic neuropathy. The biological importance of PR-ASG in pancreatic ß-cells remains unknown. Here we examined the effects of PR-ASG on IGF-1 and glucose metabolism in ß-cells exposed to oxidative stress. METHODOLOGY/PRINCIPAL FINDINGS: In the present study, a pre-germinated brown rice (PR)-diet was tested in streptozotocin (STZ)-induced diabetic rats. Compared with diabetic rats fed control diets, the PR-diet fed rats showed an improvement of serum metabolic and neurophysiological parameters. In addition, IGF-1 levels were found to be increased in the serum, liver, and pancreas of diabetic rats fed the PR-diet. The increased IGF-1 level in the pancreas led us to hypothesize that PR-ASG is protective for islet ß-cells against the extensive injury of advanced or severe diabetes. Thus we examined PR-ASG to determine whether it showed anti-apoptotic, pro-proliferative effects on the insulin-secreting ß-cells line, INS-1; and additionally, whether PR-ASG stimulated IGF-1 autocrine secretion/IGF-1-dependent glucose metabolism. We have demonstrated for the first time that PR-ASG increases IGF-1 production and secretion from pancreatic ß-cells. CONCLUSION/SIGNIFICANCE: These findings suggest that PR-ASG may affect pancreatic ß-cells through the activation of an IGF-1-dependent mechanism in the diabetic condition. Thus, intake of pre-germinated brown rice may have a beneficial effect in the treatment of diabetes, in particular diabetic neuropathy.

Induction of apoptosis and lipogenesis in human preadipocyte cell line by n-3 PUFAs.

We examined the effect of n-3 PUFAs (polyunsaturated fatty acids) on the growth and maturation of human preadipocyte cell line AML-I. On day 3 of the culture, n-3 fatty acids such as DHA (docosahexaenoic acid) and EPA (eicosapentaenoic acid), but not n-6 fatty acid LA (linoleic acid), induced growth arrest accompanied by the appearance of characteristics of apoptosis in AML-I cells at concentrations between 250 and 500 µM by Annexin V-FITC staining. In Western blotting analysis, the loss of NF-κB, Bcl-2 and p-Akt and the accumulation of Bad and Akt were observed in the cytoplasmic protein from the EPA-treated cells. Exposure of AML-I to EPA or DHA increased the cytoplasmic lipid accumulation compared with the vehicle-treated cells in a time-dependent manner during 4 and 6 days culture period by Oil Red O staining. The expression of FAS (fatty acid synthase) and PPAR-γ (peroxisome proliferator-activated receptor-γ) were increased in EPA-treated cells. These results suggest that EPA and DHA promote differentiation, inhibit proliferation and induce apoptosis in preadipocyte cell line AML-I.

The HPB-AML-I cell line possesses the properties of mesenchymal stem cells.

BACKGROUND: In spite of its establishment from the peripheral blood of a case with acute myeloid leukemia (AML)-M1, HPB-AML-I shows plastic adherence with spindle-like morphology. In addition, lipid droplets can be induced in HPB-AML-I cells by methylisobutylxanthine, hydrocortisone, and indomethacin. These findings suggest that HPB-AML-I is similar to mesenchymal stem cells (MSCs) or mesenchymal stromal cells rather than to hematopoietic cells. METHODS: To examine this possibility, we characterized HPB-AML-I by performing cytochemical, cytogenetic, and phenotypic analyses, induction of differentiation toward mesenchymal lineage cells, and mixed lymphocyte culture analysis. RESULTS: HPB-AML-I proved to be negative for myeloperoxidase, while surface antigen analysis disclosed that it was positive for MSC-related antigens, such as CD29, CD44, CD55, CD59, and CD73, but not for CD14, CD19, CD34, CD45, CD90, CD105, CD117, and HLA-DR. Karyotypic analysis showed the presence of complicated abnormalities, but no reciprocal translocations typically detected in AML cases. Following the induction of differentiation toward adipocytes, chondrocytes, and osteocytes, HPB-AML-I cells showed, in conjunction with extracellular matrix formation, lipid accumulation, proteoglycan synthesis, and alkaline phosphatase expression. Mixed lymphocyte culture demonstrated that CD3+ T-cell proliferation was suppressed in the presence of HPB-AML-I cells. CONCLUSIONS: We conclude that HPB-AML-I cells appear to be unique neoplastic cells, which may be derived from MSCs, but are not hematopoietic progenitor cells.

Effect of genistein and daidzein on the proliferation and differentiation of human preadipocyte cell line.

Isoflavones are known to have several biological activities, including a hypolipidemic effect. However, the mechanism of the lipid lowering effect of genistein remains to be elucidated. There is conflicting evidence on the effect of genistein for the deposition of adipocyte tissues. We examined the effect of the isoflavones on the growth and differentiation of human preadipocyte cells, AML-I. Growth arrest accompanied by the appearance of characteristics of apoptosis was observed by genistein or daidzein treatment under the adipogenic stimulation. The expressions of apoptosis-related proteins, Bad, Akt, and p-Akt, were modulated in the genistein-treated cells by Western blot analysis. On the other hand, exposure of AML-I to the isoflavones increased accumulation of cytoplasmic lipid droplets. Actually, the cytoplasmic expressions of fatty acid synthase (FAS) and peroxisome proliferator-activated receptor (PPAR)-gamma were increased in the genistein-treated cells. Glycosylated forms of the isoflavones genistein and puerarin did not have such activities. These results suggested that only aglycon forms of isoflavones induced not only apoptosis but also lipogenesis in the preadipocyte cell line AML-I. The possible mechanism of these phenomena has been discussed in the text.

Anesthetic management of a patient with aortocaval fistula.

Aortocaval fistula is a rare complication of ruptured abdominal aortic aneurysm (AAA), and patients with an aortocaval fistula show multiple symptoms. We report an 87-year-old man who was diagnosed as having an AAA with aortocaval fistula and who developed refractory hypotension after induction of anesthesia. Following a phenylephrine injection for slight hypotension induced by anesthetic induction, he developed severe hypotension and bradycardia, and his skin became cyanotic. Vasopressor agents had no immediate effect on the hypotension, but blood pressure gradually increased in about 30 min with continuous infusion of dopamine and noradrenaline. Transesophageal echocardiography (TEE) showed right ventricle (RV) hypokinesis and massive tricuspid regurgitation (TR). Central venous pressure (CVP) showed a remarkably high value. After the repair of the aortocaval fistula, the hemodynamics became stable, RV motion was improved, TR was reduced, and CVP became normal. Anesthetic management of the repair of an aortocaval fistula is very difficult. The hemodynamics changed dramatically throughout anesthesia in our patient with this disorder, even though low-dose anesthetics were used. For the successful treatment of this disorder, preparation for the operation is required before the induction of anesthesia, and urgent closure of the fistula is necessary after the induction of anesthesia. TEE is a useful tool for monitoring hemodynamics in such patients.

The mechanisms of the direct action of etomidate on vascular reactivity in rat mesenteric resistance arteries.

BACKGROUND: Etomidate minimally influences hemodynamics at a standard induction dose in young healthy patients, but can cause significant systemic hypotension at higher doses for induction or electroencephalographic burst suppression (i.e., cerebral protection) in patients with advanced age or heart disease, and during cardiopulmonary bypass. However, less is known about its action on systemic resistance arteries. METHODS: Using an isometric force recording method and fura-2-fluorometry, we investigated the action of etomidate on vascular reactivity in small mesenteric arteries from young (7-8 wk old, n = 179) and aged (96-98 wk old, n = 10) rats. RESULTS: In the endothelium-intact strips from young rats, etomidate enhanced the contractile response to norepinephrine or KCl (40 mM) at 3 microM but inhibited it at higher concentrations (>or=10 microM). The enhancement was still observed after treatment with N(G)-nitro l-arginine, tetraethylammonium, diclofenac, nordihydroguaiaretic acid, losartan, ketanserin, BQ-123, or BQ-788, but was not observed in aged rats. In the endothelium-denuded strips from young rats, etomidate (>or=10 microM) consistently inhibited the contractile response to norepinephrine or KCl without enhancement at 3 microM. In the fura-2-loaded, endothelium-denuded strips from young rats, etomidate inhibited norepinephrine- or KCl-induced increases in both intracellular Ca(2+) concentration ([Ca(2+)]i) and force. Etomidate still inhibited the norepinephrine-induced increase in [Ca(2+)]i after depletion of the intracellular Ca(2+) stores by ryanodine, which was sensitive to nifedipine. Etomidate had little effect on norepinephrine- or caffeine-induced Ca(2+) release from the intracellular stores or Ca(2+) uptake into the intracellular stores. During stimulation with norepinephrine or KCl, etomidate had little effect on the [Ca(2+)]i-force relation at low concentrations (

Roles of endothelial oxidases in endothelium-derived hyperpolarizing factor responses in mice.

The endothelium synthesizes and releases several vasodilator substances, including prostacyclin, nitric oxide (NO), and endothelium-derived hyperpolarizing factor (EDHF). We have demonstrated that endothelium-derived hydrogen peroxide (H2O2) is an EDHF in animals and humans and that superoxide anions derived from endothelial nitric oxide synthases (NOSs) system are an important precursor for EDHF/H2O2 in mice. There are several intracellular sources of superoxide anions other than NOSs, including NAD(P)H oxidase, xanthine oxidase, lipoxygenase, and mitochondrial electron transport chain. In this study, we examined the possible role of endothelial oxidases other than NOSs in the EDHF-mediated responses. In angiotensin II-infused mice, both EDHF-mediated relaxations and hyperpolarizations to acetylcholine were significantly reduced, nitric oxide-mediated relaxations were rather enhanced, and vascular smooth muscle responses were preserved. Antihypertensive treatment normalized blood pressure but failed to improve EDHF-mediated responses in those mice. Acute inhibition of endothelial oxidases other than NOSs, including NAD(P)H oxidase, xanthine oxidase, lipoxygenase, or mitochondrial electron transport chain, had no inhibitory effects on EDHF-mediated responses. Furthermore, in p47phox-knockout mice, EDHF-mediated responses were unaltered. These results suggest that endothelial oxidases other than NOSs are not involved in EDHF/H2O2 responses in mice, suggesting a specific link between endothelial NOSs system and EDHF responses under physiological conditions.

Naringenin and hesperetin induce growth arrest, apoptosis, and cytoplasmic fat deposit in human preadipocytes.

Citrus flavonoids are reported to be promising bioactive compounds against hyperlipidemia and lipid biosynthesis. However, the mechanism of the lipid lowering effect by flavonoids remains unknown. The present study examines the effect of some flavanones on the adipocytic conversion of the human preadipocyte cell line, AML-I. Among four structure-related flavanones including naringenin, naringenin-7-rhamnoglucoside (naringin), hesperetin, and hesperetin-7-rhamnoglucoside (hesperidin), the aglycones such as naringenin and hesperetin exhibited the growth arrest of AML-I cells. When the cells were examined by Annexin V-FITC staining method, it was noticed that growth arrest was induced by apoptotic cell death. In the study of apoptosis-related protein in the naringenin-treated cells, anti-apoptotic proteins such as p-Akt, NF-kappaB, and Bcl-2 were decreased, and pro-apoptotic protein Bad was accumulated by Western blot analysis. Interestingly, exposure of AML-I cells to naringenin or hesperetin during short-term cultures increased cytoplasmic lipid droplets by Sudan Black B staining. Furthermore, expression of fatty acid synthase (FAS) and peroxisome proliferator activated receptor (PPAR)-gamma was enhanced in naringenin-treated cells. These data suggest that apoptosis by flavanones does not inhibit the adipocytic conversion of AML-I preadipocytes. The result also indicates that adipocyte may not be a direct target for the lipid-lowering activity of the flavanones.

Effect of body mass index on perioperative complications in thoracic surgery.

Obesity is perceived as a risk factor in general thoracic surgery. We conducted a single-center retrospective evaluation of perioperative complications in 822 patients who underwent thoracic surgery between 2000 and 2005. According to body mass index, 82 were underweight (< 18.5 kg m(-2)), 568 were normal (18.5-24.9 kg m(-2)), 155 were overweight (25.0-29.9 kg m(-2)), and 17 were obese (>or=30 kg m(-2)). A significant increase in preoperative comorbidity (hypertension and ischemic heart disease) was observed with increasing body mass index. There was no significant difference in operation time or length of stay in the operating room, but extubation time was significantly different among the 4 groups. Of the intraoperative complications, alveolar-arterial oxygen difference increased significantly with increasing obesity, and hypoxia was least common in the normal group. Postoperatively, there was more pulmonary leakage in the underweight group and less pneumonia in the normal group. Both the underweight and the obese are at increased risk of perioperative complications and need to be carefully observed and managed intraoperatively and postoperatively.

Crucial role of nitric oxide synthases system in endothelium-dependent hyperpolarization in mice.

The endothelium plays an important role in maintaining vascular homeostasis by synthesizing and releasing several relaxing factors, such as prostacyclin, nitric oxide (NO), and endothelium-derived hyperpolarizing factor (EDHF). We have previously demonstrated in animals and humans that endothelium-derived hydrogen peroxide (H(2)O(2)) is an EDHF that is produced in part by endothelial NO synthase (eNOS). In this study, we show that genetic disruption of all three NOS isoforms (neuronal [nNOS], inducible [iNOS], and endothelial [eNOS]) abolishes EDHF responses in mice. The contribution of the NOS system to EDHF-mediated responses was examined in eNOS(-/-), n/eNOS(-/-), and n/i/eNOS(-/-) mice. EDHF-mediated relaxation and hyperpolarization in response to acetylcholine of mesenteric arteries were progressively reduced as the number of disrupted NOS genes increased, whereas vascular smooth muscle function was preserved. Loss of eNOS expression alone was compensated for by other NOS genes, and endothelial cell production of H(2)O(2) and EDHF-mediated responses were completely absent in n/i/eNOS(-/-) mice, even after antihypertensive treatment with hydralazine. NOS uncoupling was not involved, as modulation of tetrahydrobiopterin (BH(4)) synthesis had no effect on EDHF-mediated relaxation, and the BH(4)/dihydrobiopterin (BH(2)) ratio was comparable in mesenteric arteries and the aorta. These results provide the first evidence that EDHF-mediated responses are dependent on the NOSs system in mouse mesenteric arteries.

Structural analysis of novel bioactive acylated steryl glucosides in pre-germinated brown rice bran.

Previous studies from our laboratory indicated that pre-germinated brown rice (PR) contained certain unknown bioactive lipids that activated two enzymes related to diabetes: Na+/K+ATPase and homocysteine-thiolactonase. In this paper, we report on the isolation and structural characterization of the activator lipids from PR bran as acylated steryl glucosides (ASGs). The activator lipid was isolated by silica gel column chromatography, and its chemical structure was determined by NMR, GC-MS, and tandem mass spectrometry. We demonstrated that the bioactive component consists of a mixture of acylated steryl beta-glucosides. Delta8-cholesterol and 2-hydroxyl stearic acid were identified as constituents of ASGs. The steryl glucosides (SGs) subsequent to alkaline hydrolysis lost this enzyme activator activity. Soybean-derived ASGs were not active. This activity may be quite peculiar to PR-derived ASGs. Our findings suggest that the molecular species of ASG may play an important contributing role in the anti-diabetic properties of a PR diet.