A full digital, high definition video system (1080i) for laryngoscopy and stroboscopy.

OBJECTIVE: This study aimed to estimate the effectiveness of a full digital, high definition video system for laryngeal observations. METHODS: A newly available, full digital, high definition video camera and high definition video monitor were used. With an endoscopic adaptor and rigid telescope, laryngoscopy and stroboscopy were performed on patients with various kinds of laryngeal lesions. RESULTS: All laryngeal lesions were observed and recorded by the full digital, high definition video camera without incident. The image quality for laryngoscopy and stroboscopy was far superior to that of a conventional video system, including video-endoscopy. Even tiny structures or lesions could clearly be visualised on the monitor. The still image obtained from the full digital, high definition video camera was 1920 x 1080 pixels and was comparable to that obtained from a still camera. CONCLUSIONS: Full digital, high definition video cameras are now commonplace products and can easily be applied to patients with laryngeal disorders. They provide superior laryngeal images, compared with conventional video systems. Furthermore, high definition video systems are cheaper than proprietary medical video systems. We consider our system to represent an accessible technique of gaining superior laryngeal observation in otolaryngological clinics.

Head torsion technique for detailed observation of larynx and hypopharynx.

We introduce here an easy but effective method for detailed observation of the larynx and hypopharynx. During the endoscopic observation, the patient's head is turned to one side. Anatomical structures on the same side of the endolarynx, such as the laryngeal ventricle and inferior surface of the vocal fold, are easily observed. In addition, observation of the opposite side of the hypopharynx also becomes easier. Such head turning is also useful in patients with an oblique larynx, in whom the epiglottis obstructs insertion of the endoscope. This is a simple but very effective technique for laryngeal and hypopharyngeal observation.

An actin barrier to resealing.

Plasma membrane disruption is a common form of cell injury in many normal biological environments, including many mammalian tissues. Survival depends on the initiation of a rapid resealing response that is mounted only in the presence of physiological levels of extracellular Ca(2+). Vesicle-vesicle and vesicle-plasma membrane fusion events occurring in cortical cytoplasm surrounding the defect are thought to be a crucial element of the resealing mechanism. However, in mammalian cells, the vesicles used in this fusion reaction (endosomes/lysosomes) are not present in a 'pre-docked' configuration and so must be brought into physical contact with one another and with the plasma membrane. We propose that a requisite prelude to fusion is the disassembly in local cell cortex of the physical barrier constituted by filamentous actin. Consistent with this hypothesis, we found that rat gastric epithelial (RGM1) cell cortical staining with phalloidin was apparently reduced at presumptive disruption sites. Moreover, flow cytofluorometric analysis of wounded RGM1 populations revealed a small, but significant, Ca(2+)-dependent reduction in whole cell phalloidin staining. The functional significance of this disruption-induced depolymerization response was confirmed in several independent tests. Introduction into RGM1 cells of the filamentous actin-depolymerizing agent, DNase1, enhanced resealing, although cytochalasin treatment, by itself, had no effect. By contrast, when the filamentous actin cytoskeleton was stabilized experimentally, using phalloidin or jasplakinolide, resealing was strongly inhibited. Cells in wounded cultures displayed an enhanced cortical array of filamentous actin, and resealing by such cells was enhanced strongly by both cytochalasin and DNase 1, demonstrating the specific reversibility of a biologically mediated, polymerization-induced inhibition of resealing. We conclude that localized filamentous actin disassembly removes a cortical barrier standing in the way of membrane-membrane contacts leading to resealing-requisite homotypic and exocytotic fusion events.

Follicular dendritic cells in vitro modulate the expression of Fas and Bcl-2 on germinal center B cells.

Germinal center (GC) B cells are highly susceptible to apoptosis. The cellular mechanism regulating this sensitivity, however, has not yet been fully delineated. To investigate whether follicular dendritic cells (FDC) are capable of regulating the susceptibility to apoptosis of GC B cells, we constructed a GC model in vitro: emperipolesis of tonsillar B cells by FDC. We then analyzed the expressions of apoptosis-related proteins (Bcl-2 and Fas) on the cells by three-color flow cytometry. B cells nonentrapped by FDC decreased rapidly in number owing to early apoptosis in vitro, whereas entrapped B cells were rescued for at least 18 h and showed peculiar regulation of Fas and Bcl-2. GC founder cells (CD38+, IgD+; GCFC) and GC B cells (CD38+, IgD-) showed approximately a twofold increased expression of Fas; in contrast, mantle zone B cells (CD38-, IgD+) and memory B cells (CD38-, IgD-) showed no changes. Bcl-2 expression in mantle zone and memory B cells was reduced by approximately one-half; however, GCFC and GC B cells continued to express little Bcl-2 and this did not change. Our findings strongly suggest that FDC play a part in the modulation of the susceptibility to apoptosis on B cells within GC.

Highly augmented cytopathic effect of a fiber-mutant E1B-defective adenovirus for gene therapy of gliomas.

An E1B 55-kDa gene-defective adenovirus (Adv), ONYX-015, has been reported to be a highly useful replication-competent Adv that shows cytopathic effect for cancers with an abnormal p53 gene, without damaging normal tissues. In this study, we combined this Adv (Adv-E1AdB) with a fiber mutation, F/K20, which has a stretch of 20 lysine residues added at the COOH-terminus of the fiber and shows high transduction efficiency to gliomas. In U-373 MG glioma cells, the transduction efficiency of Adv-F/ K20 for lacZ was nine times higher than that of the Adv with wild-type fiber (Adv-F/wt) for lacZ. At a multiplicity of infection of 30, the replication efficiency of Adv-E1AdB-F/K20 was 11 times higher than that of Adv-E1AdB with wt fiber (Adv-E1AdB-F/wt). The ED50 value of AdvE1AdB-F/K20 to U-373 MG cells, which is a measure of the in vitro cytopathic effect, was 32 times greater than that of Adv-E1AdB-F/wt. injection of Adv-E1AdB-F/K20 suppressed the in vivo growth of tumors. The antitumoral effect of Adv-E1AdB-F/K20 was remarkably stronger than that of Adv-E1AdB-F/wt. A greater quantity of replicated virus protein (hexon) by infection with Adv-E1AdB-F/K20 was demonstrated in vitro and in vivo, compared with that of Adv-E1AdB-F/wt. In conclusion, gene therapy using Adv-E1AdB-F/K20, which drastically augmented the antitumoral effect of Adv-E1AdB, will be a promising therapeutic approach for gliomas.

Remnant lipoprotein levels in fasting serum predict coronary events in patients with coronary artery disease.

BACKGROUND: Remnant lipoproteins are atherogenic, but assays of remnants have not been available in routine clinical laboratories because of the lack of practical and validated methods. A simple and reliable method for such an assay, using an immunochemical approach, has recently been developed. This study prospectively examined whether remnant lipoprotein levels in fasting serum, measured by our method, may have prognostic value in patients with coronary artery disease (CAD). METHODS AND RESULTS: Remnant lipoprotein levels in fasting serum were measured in 135 patients with CAD by an immunoaffinity mixed gel containing anti-apolipoprotein (apo) A-1 and anti-apoB-100 monoclonal antibodies. Patients were followed up for 5.1 mg cholesterol/dL; 75th percentile of distribution of remnant levels) than in those with the lowest tertile of remnant levels (

Vitamin E administration improves impairment of endothelium-dependent vasodilation in patients with coronary spastic angina.

OBJECTIVES: We examined the effects of oral administration of vitamin E, an antioxidant, on endothelium-dependent vasodilation in patients with coronary spastic angina. BACKGROUND: We have recently reported that endothelium-dependent vasodilation is impaired in patients with coronary spastic angina (CSA). Furthermore, it is known that oxidative stress may play an important role in the impairment of endothelium-dependent vasodilation in cardiovascular diseases. METHODS: With the ultrasound technique, flow-dependent vasodilation of the brachial arteries during reactive hyperemia was examined before and after treatment for a month with either oral administration of vitamin E (alpha-tocopherol acetate, 300 mg/day) or placebo, which is randomly assigned, in patients with CSA (n=60). RESULTS: Before treatment, patients with CSA had impaired flow-dependent vasodilation, lower plasma levels of alpha-tocopherol and higher plasma levels of thiobarbituric acid reactive substances (TBARS), as compared with age- and sex-matched control subjects (n=60) (flow-dependent vasodilation: 3.1+/-1.8 vs. 7.1+/-2.5%, p < 0.001; alpha-tocopherol levels: 8.9+/-1.8 vs. 10.8+/-1.8 microg/ml, p < 0.001). In patients with CSA, treatment with vitamin E restored flow-dependent vasodilation (3.1+/-1.7 vs. 8.3+/-2.0%, p < 0.001), and this improvement was associated with the decreases in plasma TBARS levels and anginal attacks. CONCLUSIONS: The results indicate that vitamin E treatment improved endothelium-dependent vasodilation and decreased plasma TBARS levels in patients with CSA. Thus, increased oxidative stress may contribute to endothelial dysfunction and anginal attacks in patients with CSA.

Influence of chronic nitric oxide inhibition of coronary blood flow regulation: a study of the role of endogenous adenosine in anesthetized, open-chested dogs.

The effect of chronic inhibition of endothelium-derived nitric oxide (NO) synthesis on the regulation of coronary blood flow (CBF) is yet to be elucidated. A chronic canine model of inhibited NO synthesis was created and the role of adenosine in the regulation of coronary blood flow in this model was examined. Dogs were fed a diet supplemented with 40 mg/kg per day N(G)-nitro-L-arginine methyl ester (L-NAME group, n=8) or a regular diet without L-NAME supplementation (control group, n=8) for 4 weeks. The experiments were performed in an anesthetized, open-chest state and the results were compared in the L-NAME and control groups. Chronic L-NAME treatment significantly increased arterial pressure. Neither basal CBF in the left anterior descending artery nor heart rate differed between the L-NAME and control groups. In the L-NAME group, the response of CBF to intracoronary acetylcholine and adenosine was blunted, but that to glyceryl trinitrate was not. In addition, myocardial reactive hyperemia following 20 sec coronary occlusion was blunted in the L-NAME group. During atrial pacing at a rate 60 beats/min faster than the sinus rate, CBF increased to a similar degree in the L-NAME and control groups, and systolic wall thickening (SWT) changed similarly in both groups. Intracoronary 8-phenyltheophylline (8-PT), an adenosine receptor blocker, decreased basal CBF in the L-NAME group but not in the control group. In the L-NAME group, pacing-induced increase in CBF was abolished and SWT deteriorated after 8-PT administration. Basal myocardial adenosine release was significantly increased in the L-NAME group compared with the control group. It is concluded that in anesthetized, open-chest dogs with chronic inhibition of NO synthesis, adenosine may play a compensatory role in the regulation of coronary blood flow, as concomitant blockade of adenosine causes deterioration of coronary circulation and cardiac function.

Occult thyroid carcinoma manifested as a large mediastinal tumor.

The authors report a rare case of clinically occult thyroid carcinoma that initially manifested as a metastatic mediastinal tumor with tracheal stenosis. A curative operation was performed by complete removal of the tumor through a median sternotomy approach combined with resection of the tracheal invasion. Histopathological examination revealed transition of a well-differentiated papillary carcinoma into a poorly differentiated component in the lesion invading the trachea, which suggested that this transition might have rendered the tumor more aggressive, leading to tracheal invasion.

Human follicular dendritic cells in vitro and follicular dendritic-cell-like cells.

Human follicular dendritic cell (FDC)-like cells (FLC) have been utilized for the in vitro analysis of germinal center reactions. However, there is no consensus whether FLC represent FDC in vitro. The purpose of the present study has therefore been to determine distinguishing features of FDC and FLC in vitro. The expression of CD40, CD54, CD49d, cytokine (gamma-IFN and IL-4)-dependent MHC-class II, and CD106 was observed to be specific for the determination of FDC in long-term culture. The cytokine-dependent emperipolesis of germinal center B cells was establised as another discriminating property for FDC in vitro. In 2 out of 72 long-term cultures of FDC, we encountered dividing cells among the non-dividing population of FDC. The dividing cells expressed accessory molecules similar to those of FDC but showed emperipolesis only for the initial few days of their growth. FDC did not enhance the CD40-dependent proliferation of germinal center B cells; in contrast, FLC augumented it. Both types of cells produced a significant amount of cytokine-dependent IL-6. Further studies are needed to determine whether FLC originate from FDC in vitro.

Extracranial meningioma presenting as a tumour of the external auditory meatus: a case report.

A rare case of extracranial meningioma presenting as a tumour of the external auditory meatus is reported. Biopsy indicated a diagnosis of meningioma, but the radiological appearance was unusual. For example, computed tomography (CT) scans showed an unenhanced tumour mainly located in the squamous part of the temporal bone which expanded into the external meatus destroying the temporal bone. Magnetic resonance imaging (MRI) revealed that the tumour did not extend into the intradural space. This meningioma, had an obvious tendency for extracranial development. According to the operative findings, the tumour arose from the middle cranial fossa dura and extended through the air cells of the temporal bone into the external meatus, instead of growing intracranially. Secondary extracranial meningiomas of the temporal bone usually have a large intracranial component and cause neurological symptoms. However, this was a very rare case of a small meningioma causing no symptoms except for conductive hearing loss.

Stavudine selectively induces apoptosis in HIV type 1-infected cells.

We evaluated the cytotoxic effects of various human immunodeficiency virus (HIV-1) reverse transcriptase inhibitors (zidovudine, didanosine, zalcitabine, stavudine, and nevirapine) on HIV-1-infected and uninfected T cell lines. Among the compounds, only stavudine (not the others) proved to be more cytotoxic to MOLT-4/IIIB cells (MOLT-4 cells chronically infected with HIV-1) than to uninfected MOLT-4 cells. Its 50% cytotoxic concentrations were 59.8 and 2.2 microM for MOLT-4 and MOLT-4/IIIB cells, respectively. Stavudine was also more cytotoxic to CEM/ROD (CEM cells chronically infected with HIV type 2) than to uninfected CEM cells. Microscopic analysis revealed that stavudine induced apoptosis in MOLT-4/IIIB cells. Apparent chromatin condensation in the nucleus was observed by electron microscopy. Furthermore, a DNA fragmentation ladder was detected by agarose gel electrophoresis. Addition of thymidine to the culture medium could rescue the cells from stavudine-induced apoptosis. The expression of anti-apoptotic protein Bcl-2 was partially downregulated in MOLT-4/IIIB cells after treatment with stavudine. This downregulation was not identified in MOLT-4 cells. These results indicate that stavudine selectively induces apoptosis in HIV-1-infected T cells and may have potential as a novel strategy for effective chemotherapy of the acquired immune deficiency syndrome (AIDS).

[Immunohistochemical study on the histogenesis of malignant fibrous histiocytoma].

Forty cases of malignant fibrous histiocytomas (MFHs) consisting of 33 cases of storiform-pleomorphic subtype MFH and 7 cases of myxoid subtype MFH, were studied immunohistochemically. Immunohistochemical stains were performed by the labelled streptavidin biotin method. The primary antibodies used in this study were classified into 4 groups according to their main specificities: 1) epithelial and non-epithelial (mesenchymal) markers, 2) histiocytic markers, 3) myogenic markers, 4) and that considered as possible lipogenic marker. In addition to these markers, alcian-blue stain, periodic acid-Schiff stain, Masson-trichrome stain, silver impregnation stain, oil red O stain, and Sudan III stain were performed in some cases at the same time. Over 85% of the cases were positive for mesenchymal and histiocytic markers, and no cases were positive for epithelial markers. About 40% of MFH were positive for some myogenic markers, but none of these cases were positive for any skeletal muscle markers. S-100 protein, considered as a possible lipogenic marker through the results of other special stains, was positive in about 10% of the MFHs. The immunohistochemical results of myogenic and lipogenic markers were parallel to the histological subtype; the positive percentages for myogenic markers were about 50% in the cases of storiform-pleomorphic subtype, but were 17% in the cases of myxoid subtype. On the other hand, the positive percentage for lipogenic marker was only 4% in the cases of storiform-pleomorphic subtype, but was 50% in the cases of myxoid subtype. These were supposed to be additional and occult characteristics of MFH. Therefore, the interrelation between the immunohistochemical results and histological subtype of MFH interestingly suggested a new entity of MFH concerning the histogenesis of MFH: MFH was suspected to contain some groups of very poorly differentiated and/or dedifferentiated sarcomas and to be the precondition of some sarcomas.

GM-CSF-induced in vivo expansion of splenic dendritic cells and their strong costimulation activity.

Dendritic cells (DC), with their potent antigen-presenting and accessory activities, are involved in the stimulation of naive T cells. To examine the biological functions of DC, we developed a method for generating large numbers of murine splenic DC. First, DC were propagated in vivo by using a granulocyte-macrophage colony-stimulating factor-secreting tumor as a continuous in vivo source of the cytokine. The DC enriched in the spleen, especially in the white pulps, were purified after an overnight culture. We could reproducibly obtain 6 to 10 X 10(6) splenic DC per mouse. These DC were morphologically similar to interdigitating cells, expressed high levels of MHC class II and costimulatory molecules, and were highly allo-stimulatory in mixed lymphocyte reactions. Further analysis on T cell stimulation activity revealed that the DC had strong costimulatory activity on human T cells. Activated B cells, which express both B7-1 and B7-2, had little T cell costimulatory activity under the same assay conditions. A human histiocytic leukemia cell line, U937, that showed only weak costimulatory activity by itself, worked synergistically with DC and further intensified the T cell stimulation by DC. These findings suggest the presence of a T cell costimulation mechanism in DC, which is activated synergistically by monocytes or macrophages, and deserves further study.

Impairment of coronary blood flow regulation by endothelium-derived nitric oxide in dogs with alloxan-induced diabetes.

Diabetes mellitus is a major cause of ischemic coronary artery disease. Endothelial dysfunction is implicated in the pathogenesis of diabetic vascular disease. To examine coronary blood flow (CBF) regulation with endothelium-derived nitric oxide (EDNO) in the diabetic state, we compared the effects of both acetylcholine (ACh) and adenosine (Ado) on left circumflex coronary artery (LCX) blood flow in 12 vehicle-treated and 21 dogs made diabetic with alloxan anesthetized with pentobarbital. All dogs were pretreated with aspirin to inhibit endogenous prostaglandins. None of the hemodynamic parameters were significantly different in the two groups. The percent change in coronary vascular resistance (CVR) after ACh (100 ng/kg) infusion was significantly attenuated in diabetic dogs (-56.5 +/- 1.4%) as compared with vehicle-treated dogs (-64.5 +/- 1.2%) (p < 0.01), whereas the effect of Ado (1 microgram/kg) was not different between the two groups (-71.1 +/- 1.5% in vehicle, -67.0 +/- 1.3% in diabetes). After infusion of incremental doses of NG-nitro-L-arginine methyl ester (L-NAME) 10(-5)-10(-3)M, the effect of ACh was progressively inhibited in both groups and was different no longer between the two groups after the maximal dose. L-Arginine (L-ARG), but not D-ARG, significantly restored the effect of ACh in diabetic dogs but did not affect vehicle-treated dogs. The effect of Ado did not change after L- and D-ARG administration. Cu, Zn-superoxide dismutase (Cu, Zn-SOD) had no effect on any of the effects of ACh and Ado in diabetic dogs. Regulation of CBF with EDNO is impaired in dogs with alloxan-induced diabetes, and this impairment is partially restored by L-ARG.

Intensified antitumor immunity by a cancer vaccine that produces granulocyte-macrophage colony-stimulating factor plus interleukin 4.

Vaccination with irradiated tumor cells genetically modified to secrete granulocyte-macrophage colony-stimulating factor (GM-CSF tumor vaccine) induces a potent systemic antitumor immunity. To develop a protocol for cancer therapy to further augment the host immune response, we examined the effects of the GM-CSF tumor vaccines simultaneously producing additional cytokines. We prepared cancer vaccines expressing double cytokines by sequential recombinant retrovirus-mediated genetic transductions. We then used a murine intracerebral tumor model in which the GM-CSF tumor vaccine was less effective in immunopotentiation and evaluated tumor vaccines producing various cytokines in conjunction with GM-CSF. The cytokine combination of GM-CSF and interleukin 4 induced more potent antitumor immunity than GM-CSF alone. An in vivo depletion test showed that CD4+, CD8+, and asialoGM1+ cells were required for the optimum function of the GM-CSF plus interleukin 4 tumor vaccine. Histological examinations revealed infiltration of inflammatory cells at the site of tumor cell challenge as well as at the site of vaccination, indicating the induction of a systemic antitumor immune response which reached the central nervous system. Our findings suggest the feasibility of applying the intensified vaccination strategy to treat human cancers including malignant brain tumors.

Enhancement of myocardial reactive hyperemia with manganese-superoxide dismutase: role of endothelium-derived nitric oxide.

OBJECTIVE: To test the hypothesis that superoxide radicals generated during myocardial ischemia and reperfusion influence reactive hyperemia (RH) by reacting with endothelium-derived nitric oxide (EDNO), we examined the effect of manganese (Mn)-superoxide dismutase (SOD) on RH in anesthetized dogs. METHODS: Twelve dogs were pretreated with 8-phenyltheophylline (8PT) to block adenosine's effect. Five dogs were pretreated with 8PT and NG-nitro-L-arginine methyl ester (L-NAME) to block adenosine's and EDNO's effects. Following occlusion of the left circumflex artery (LCX) for 10 and 60 s, RH was observed before and after Mn-SOD. In another group of 6 dogs pretreated with 8PT, RH following 60-s LCX occlusion was observed before and after Mn-SOD and catalase. For comparison with the effect of Mn-SOD, that of copper, zinc (Cu,Zn)-SOD was also examined in another group of 5 dogs. RESULTS: In the dogs pretreated with 8PT, Mn-SOD significantly increased excess flow and repayment of flow debt during RH after 60-s LCX occlusion but did not affect RH after 10-s LCX occlusion. Mn-SOD-induced augmentation of RH following 60-s LCX occlusion was not affected by catalase, while it was completely abolished by L-NAME. In contrast to Mn-SOD, Cu,Zn-SOD showed no effect on RH following 60-s LCX occlusion in the dogs pretreated with 8PT. CONCLUSIONS: Superoxide radicals generated during ischemia for 60 s and reperfusion attenuates myocardial RH through inactivation of EDNO. Mn-SOD shows more beneficial effects on myocardial RH than Cu,Zn-SOD.

Role of adenosine in regulation of coronary flow in dogs with inhibited synthesis of endothelium-derived nitric oxide.

Endothelium-derived nitric oxide (NO) regulates coronary blood flow, but it is unclear how NO synthesis inhibition affects myocardial metabolism. In pentobarbital sodium-anesthetized dogs, myocardial oxygen metabolism, adenosine release, lactate extraction rate (LER), and systolic ventricular wall thickening (SWT) at baseline and during atrial pacing were estimated before and after intracoronary NG-nitro-L-arginine methyl ester (L-NAME) infusion. Coronary blood flow and PO2 in the anterior interventricular vein at baseline were both significantly decreased by L-NAME (3 x 10(-4) M in the coronary blood). Coronary flow was increased during pacing, which was not affected by L-NAME. Myocardial adenosine release remained unchanged during pacing before L-NAME, but it was significantly increased after L-NAME infusion. Neither LER nor SWT changed during pacing performed before and after L-NAME. The experiment was also performed in dogs pretreated with 8-phenyltheophyl-line. After L-NAME, pacing-induced increase in coronary flow was suppressed, and both LER and SWT were significantly decreased during pacing. In conclusion, when NO synthesis is inhibited, adenosine release is increased in response to the increase in myocardial oxygen demand. With this compensatory adenosine release, coronary flow is increased and ventricular function is unaffected.

In vivo antitumor effect of cytotoxic T lymphocytes engineered to produce interferon-gamma by adenovirus-mediated genetic transduction.

Immunotherapy with adoptive transfer of genetically-modified cytotoxic T lymphocytes (CTL) is a promising approach for cancer gene therapy. We developed an adoptive therapy model with murine tumor-specific CTL, to which very efficient (up to 100%) gene introduction was achieved by using recombinant adenoviral vectors. Through a comparative study on the antitumor effects of CTL genetically modified with cytokine genes, transduction with interferon-gamma gene resulted in a prominent increase in therapeutic efficacy of CTL in both metastatic and subcutaneous tumor models. Further additive effect was obtained by the adoptive cellular therapy in combination with vaccination of cytokine gene-modified tumor cells. Our findings provide a hopeful strategy of adoptive immunotherapy for human cancers.