Visualize the quality of frozen fish using fluorescence imaging aided with excitation-emission matrix.

The quality monitoring of frozen marine products has become essential in the fishery industry, where efficient and effective quality assurance is becoming increasingly important. In this study, we proposed a novel method of evaluating fish quality by combining the fluorescence excitation-emission matrix (EEM) with imaging techniques to visualize the spatial-temporal changes of freshness indices such as K-value and taste component IMP content. The result showed that the distribution of K-value and IMP content could be visualized with accuracy of R2 = 0.78 and R2 = 0.83, respectively. Furthermore, this innovative approach was applied to differentiate burnt meat, which is a type of abnormal meat found in many types of fish, and it was found that burnt meat could be detected even when in a frozen condition.

Joint lavage followed by viscosupplementation and triamcinolone in patients with severe haemophilic arthropathy: objective functional results.

INTRODUCTION: Viscosupplementation can improve function in haemophilia patients. Viscosupplementation results can be improved by prior joint lavage and triamcinolone administration. AIM: To objectively assess whether viscosupplementation and associated triamcinolone use in patients with severe haemophilic arthropathy following joint lavage improves force and balance and reduces bleeding events. METHODS: Fourteen patients with haemophilic knee arthritis with and without the involvement of other joints underwent joint lavage and subsequent injections of Hylan G-F20 and triamcinolone into all affected joints. Patients were evaluated with NeuroCom® force and balance platforms using the step-up-and-over task (STP), sit-to-stand test (STS), one-leg stance (UNI) and weight-bearing squat (WBS) at baseline and at 1, 3, 6 and 12 months after the procedure. Bleeding events in the year prior to and the year after the procedure were analysed. RESULTS: Sixteen knees, 15 ankles, eight elbows and one shoulder were treated. The STP results indicated improvements in the lift-up indices (right leg) at all time points evaluated (P = 0.03). The STS results revealed coupled improvement in weight transfer and the rising index for up to a year (P = 0.02). Balance (UNI) with eyes open or closed improved in all evaluations. The WBS results revealed improvements at all degrees of flexion (0, P = 0.003; 30°, P = 0.001; 60°, P < 0.001 and 90°, P < 0.001). The numbers of total and traumatic bleeding events were reduced (P = 0.04). CONCLUSION: Joint lavage followed by injections of triamcinolone and Hylan G-F20 improved balance, function and bleeding events in severe haemophilic arthropathy patients.

Toxic shock-like syndrome caused by non-group A beta-hemolytic streptococci.

Two patients with rapidly developing shock, multisystem organ failure, and destructive soft-tissue infection caused by groups G and C streptococci are described. Both patients died rapidly despite aggressive treatment. The clinical characteristics cannot be distinguished from those of toxic shock-like syndrome, but Streptococcus pyogenes was not recovered. These strains did not produce any previously identified type of streptococcal pyrogenic exotoxins. These findings suggest that toxic shock-like syndrome can be caused not only by group A but also groups G and C streptococci. The causative strains of toxic shock-like syndrome may have something in common with unknown virulent factors for this syndrome.

Sertoli-stromal cell tumor of the ovary: immunohistochemical, ultrastructural, and genetic studies.

The Sertoli-stromal cell tumor (SSCT) of the ovary shows a histologic resemblance to developing or adult testes and is often associated with virilization caused by tumor-produced androgenic hormone. In spite of the unique manifestation of SSCT, detailed characteristics of this tumor are still obscure. The mechanism by which SSCT occurs has not yet been determined. Six SSCTs were studied immunohistochemically, ultrastructurally, and by polymerase chain reaction (PCR) for the presence of sex-determining region Y (SRY) gene and the X chromosome activation state. Immunohistochemically, Sertoli-like cells of SSCT were positive not only for alpha-inhibin but also low-molecular-weight cytokeratin. In control testes, the expression of alpha-inhibin and cytokeratin was limited to a Sertoli cell component and rete testis, respectively. Ultrastructurally, tumor cells composing hollow tubules had an elongated nucleus with deep indentation and annulate lamellae, which are characteristic structures of mature Sertoli cells. In addition, they had studded microvilli on the apical surface and frequent desmosomes, which are structures noted in the cells of rete testis. Histologically, tumor cells of hollow tubules sometimes pouted into the lumen, as did the cells of tubulae rete, entrance into rete testis from seminiferous tubules. All of these findings indicate that some tumor cells of a SSCT show simultaneous differentiation into both Sertoli cells and cells of rete testis. SRY gene was not detected in any cases, and the X chromosome activation pattern was the same as that of the female control.

Chronic lymphocytic leukemia associated with von Recklinghausen neurofibromatosis.

We report a 62-year-old female, with von Recklinghausen neurofibromatosis and chronic lymphocytic leukemia, whose mother and son both had neurofibromatosis and died of digestive tract cancers. The patient died of pneumonia 3 years after the initiation of therapy. Leukemia reported in association with neurofibromatosis are predominantly nonlymphocytic and limited to childhood. The type of association found in our patient has not been reported previously.

Flow-cytometric analysis on kainate-induced decrease in the cellular content of non-protein thiols in dissociated rat brain neurons.

In order to study the kainate-induced oxidative stress on brain neurons, the effect of kainate on cellular content of glutathione in rat cerebellar neurons were examined using a flow cytometer and 5-chloromethylfluorescein, a fluorescent dye for cellular non-protein thiols (mainly glutathione). Kainate at concentrations ranging from 30 microM to 1 mM produced a dose-dependent decrease in cellular content of glutathione. Exposure of neurons to kainate at concentrations of 300 microM or greater seemed to deplete cellular glutathione. Potency of kainate in reducing cellular content of glutathione was greater than those of glutamate and N-methyl-D-aspartate (NMDA). Kainate-induced decrease in cellular content of glutathione was partly attenuated by 6-nitro-7-cyano-quinoxaline-2,3-dione, a blocker of non-NMDA receptors and removal of external Ca2+. Results indicate that kainate causes Ca2(+)-dependent oxidative stress that decreases the cellular content of glutathione via activation of non-NMDA type of glutamate receptors.

Flow-cytometric estimation on glutamate- and kainate-induced increases in intracellular Ca2+ of brain neurons: a technical aspect.

Effects of glutamate and kainate on the intracellular Ca2+ concentration ([Ca2+]i) in a large population (several thousand) of dissociated cerebellar granule cell neurons were evaluated using a flow-cytometer and a combination of two fluorescent dyes, fluo-3-AM for estimating [Ca2+]i and ethidium bromide for removing neurons that had compromised membranes from the cell population examined. The number of neurons responding to glutamate or kainate in augmenting the fluo-3 fluorescence increased in a dose-dependent manner. The number of neurons responding to kainate was much greater than that to glutamate. CNQX, a blocker of non-NMDA receptors, completely blocked the response elicited by kainate while the complete blockade of this glutamate-induced response was made by a combination of MK-801, a NMDA receptor blocker, and CNQX. Nicardipine, a calcium antagonist, decreased the number of neurons responding to glutamate and kainate, suggesting involvement of voltage-dependent calcium channels. These results indicate that the flow-cytometric measurement of glutamate and kainate responses has the potential to provide answers to such questions as what percentage of the population of neurons respond to these amino acids and what is the resulting distribution of [Ca2+]i.

Toxic shock-like syndrome with flu-like prodrome: a possible role of 'enhancing tissue focus' for streptococcal toxic shock.

We describe three patients with invasive group A streptococcal infection, admitted during the 3 months between November 1996 and February 1997. All patients were previously healthy Japanese women who developed a profound shock, with a rapidly fatal outcome, after experiencing flu-like symptoms. All cases conformed to the case definition of toxic shock-like syndrome (TSLS).Currently, the pathogenic mechanism of TSLS remains unclear. Known microbial virulence factors can not sufficiently explain the occurrence of TSLS, and it has been generally considered that host factors may be contributory. On pathological examination, each patient had one organ or tissue that was most severely involved: Case 1 a non-penetrating trauma; Case 2 a pregnant uterus; and Case 3 a pulmonary lesion reminiscent of lymphocytic interstitial pneumonia. On the basis of clinicopathological features of these cases, we propose that the coexistence of 'enhancing tissue focus' may be one of host factors for the progression of TSLS in patients infected with non-invasive GAS.

Immunohistochemical properties of malignant mesothelioma cells in histologic and cytologic specimens.

Although there is general agreement that immunohistochemical methods can aid in the pathologic diagnosis of malignant mesothelioma, some studies have produced conflicting results. To obtain comparable and reproducible results, unequivocal malignant mesotheliomas were studied with the biotin-streptavidin-peroxidase complex method in 14 formalin-fixed, paraffin-embedded tissue specimens, 5 ethanol-fixed smear slides and 3 cold acetone-fixed smear slides. The expression of CA-125, epithelial membrane antigen (EMA) and vimentin by malignant epithelial mesothelioma cells was hindered by their poor preservation in formalin fixative. Cytologic specimens fixed in cold acetone were the best type for immunohistochemistry. The majority of malignant epithelial mesothelioma cells in the smear slides fixed in cold acetone were positive for CA-125, EMA, low-molecular-weight cytokeratin and vimentin, but none of them were positive for carcinoembryonic antigen, CA-19-9, epithelial antigen, high-molecular-weight cytokeratin or Leu-M1.

Effects of triphenyltin on growth and viability of K562 leukemia cells.

The effects of triphenyltin on growth and viability of K562 human leukemia cells were examined using a flow cytometer with fluorescent dyes, ethidium bromide, fluo-3-AM, and propidium iodide. Triphenyltin at concentrations ranging from 30 nM to 1 µM inhibited the growth of K562 cells in a dose-dependent manner when the cells were incubated with triphenyltin at respective concentrations for 72 h. Triphenyltin at 100 nM slowed the rate of growth without affecting the viability. Triphenyltin at 300 nM or higher greatly decreased the viability of K562 cells. Triphenyltin at 300 nM increased the concentration of intracellular Ca(2+) and induced cell cycle arrest in G2/M phase and apoptosis in K562 cells. The concentration of triphenyltin inducing 50% inhibition of growth of K562 cells was lower than those of cisplatin, diphenyltin, monophenyltin, triethyltin and trimethyltin. However, tributyltin was equally toxic. Results suggest that there are several types of mechanisms for the inhibitory action of triphenyltin on the growth of K562 cells, being dependent on its concentration.

Fluorescent estimation on cytotoxicity of methylmercury in dissociated rat cerebellar neurons: its comparison with ionomycin.

To study the cellular basis of the neurotoxicity of methylmercury, the effects of methylmercury on dissociated rat cerebellar neurons were examined using a flow cytometer, a confocal laser microscope and three fluorescent dyes, fluo-3 for monitoring the changes in intracellular Ca(2+) concentration ([Ca(2+)](i)) and for detecting live neurons, ethidium for assessing the neurons that are dead or have compromised membranes, and 5-chloromethylfluorescein (CMF) for estimating the cellular content of nonprotein thiols. Methylmercury at concentrations of 1 µM or greater increased the [Ca(2+)](i) of almost all neurons. Prolonged exposure to methylmercury (3 and 10 µM) produced a further increase in [Ca(2+)](i), in association with compromising membranes in some neurons. Thereafter, methylmercury induced blebs on membranes of some neurons with increased [Ca(2+)](i). Methylmercury at concentrations of 0.3 µM or greater dose-dependently decreased the cellular content of nonprotein thiols. Results suggest that methylmercury may induce the loss of membrane integrity through destabilized Ca(2+) homeostasis and oxidative stress in mammalian brain neurons.

[An autopsied case of pachymeningitis associated with a ruptured, cerebral aneurysm due to Aspergillus infection].

We reported a 64-years-old woman with pachymeningitis associated with a ruptured mycotic cerebral aneurysm due to Aspergillus infection. She had suffered from diabetes mellitus and been treated since she was 49 years old. She complained of headache at the age of 62 and loss of her left visual acuity three months later. She was treated by the pulse therapy of methylprednisolone as neuritis retrobulbaris and her visual acuity recovered. But her headache continued. Three months later, her right visual acuity was lost, and the pulse therapy was not effective this time. Six months later, she died of subarachnoid hemorrhage following acute meningitis. The autopsy was granted, but limited to the cranial cavity. Macroscopically, it disclosed brownish thickened dura around sella turucica involving trigeminal ganglion and optic nerve, and fresh subarachnoid hemorrhage in the basal cisterns and a ruptured aneurysm (3 mm in diameter) between internal carotid and posterior cerebral artery on the left side. Histologically, the brownish thickened dura was infiltrated by lymphocytes, plasma cells, and multinucleated giant cells. The wall around the aneurysm was infiltrated by lymphocytes and plasma cells as well as many fungi. Immunohistochemistry demonstrated the presence of Aspergillus in the thickened dura and the arterial wall around the aneurysm. There were lymphocytes and plasma cell infiltration in the basal subarachnoid space and scattered microabcesses in the brain. Although the first entry of Aspergillus to the dura was unclear, we assume that the final intravascular dissemination of Aspergillus from the dura caused meningitis and mycotic aneurysm.

AKR leukemia-specific surface antigens acquired by malignant transformation: their common and individual specificities.

Antisera against five newly established AKR spontaneous leukemias (KSL) prepared by immunization of KSL cells in (C57B1/6 X C3Hf/He)F1 mice were preabsorbed with non-leukemic AKR lymphoid cells to remove antibodies against virus-associated surface antigens and alloantigens. In absorption tests the antisera showed no cross-reaction by immunofluorescence microscopy with thymocyte, fetal, male-specific H-Y, E and X.1 antigens; with known Gross murine leukemia virus-associated antigens; or with cell surface antigens on Friend, Moloney and Rauscher virus-induced tumors. It was thus shown that the antigens detected were leukemia-specific, were acquired by malignant transformation, and consisted of two types: (1) a common antigen in all the KSL, and (2) individual antigens found in four of the KSL which showed distinct patterns of partial cross-reactivity with the other KSL.

Flow cytometric analysis of the H2O2-induced increase in intracellular Ca2+ concentration of rat thymocytes.

The effect of hydrogen peroxide (H2O2) on the intracellular Ca2+ concentration ([Ca2+]i) of rat thymocytes was examined by a flow cytometer and two fluorescent dyes, fluo-3-AM and ethidium bromide, a dye impermeant to intact membranes, to characterize the H2O2-induced increase in [Ca2+]i. H2O2 at concentrations greater than 30 microM dose-dependently increased the [Ca2+]i of thymocytes which were not stained with ethidium. Removal of external Ca2+ greatly reduced the degree of H2O2-induced increase in [Ca2+]i. However, H2O2 still increased the [Ca2+]i under the external Ca(2+)-free condition. Diethylmaleate, which is known to produce a chemical depletion of cellular nonprotein thiol, significantly increased the [Ca2+]i. Dithiothreitol, which is used to protect cellular nonprotein thiol, slightly decreased the [Ca2+]i, but greatly reduced the H2O2-induced increase in [Ca2+]i. Therefore, it is considered that H2O2 may increase the [Ca2+]i through a mechanism related to the effects of H2O2 on the cellular nonprotein thiol.

Fluorescent estimation of H2O2-induced changes in cell viability and cellular nonprotein thiol level of dissociated rat thymocytes.

We have developed a procedure to simultaneously estimate cell viability and the cellular level of nonprotein thiol (presumably glutathione) using two fluorescent dyes, 5-chloromethylfluorescein (5CMF) and ethidium, and rat thymocytes. Diethylmaleate and N-ethylmaleimide reduced, respectively, the intensity of 5CMF fluorescence to 0.23 and 0.1, relative to the control. Incubation with buthionine sulfoximine, an inhibitor of glutathione synthesis, decreased the intensity of 5CMF fluorescence to 0.61. Results indicate that 5CMF fluorescence can be attenuated by agents that decrease the level of cellular nonprotein thiols, suggesting that 5CMF fluorescence is utilized for estimating the level of cellular glutathione. Hydrogen peroxide (10 microM to 3 mM) reduced the intensity of 5CMF fluorescence in a dose-dependent manner and increased the number of thymocytes stained with ethidium (presumably dead cells or cells with compromised membranes) at concentrations of 300 microM or greater. Reduction of cellular glutathione level seems to precede cell death in which oxidative stress is involved.

New curcuminoids isolated from Zingiber cassumunar protect cells suffering from oxidative stress: a flow-cytometric study using rat thymocytes and H2O2.

Effects of new complex curcuminoids (cassumunin A and cassumunin B) isolated from tropical ginger, Zingiber cassumunar, were examined in dissociated rat thymocytes suffering from oxidative stress induced by 3 mM hydrogen peroxide by using a flow cytometer and ethidium bromide. The effects were compared with those of curcumin, a natural antioxidant, whose chemical structure is included in those of cassumunins A and B. Pretreatment of rat thymocytes with the respective cassumunins at concentrations ranging from 100 nM to 3 microM dose-dependently prevented the hydrogen peroxide (H2O2)-induced decrease in cell viability. It had the same action, although less effective, against the treatment with cassumunin A or B (3 microM) immediately after or 60 min after start of the oxidative stress. Respective potencies of cassumunins A and B in protecting the cells suffering from H2O2-induced oxidative stress were greater than that of curcumin. It is suggested that cassumunins A and B may possess a potent protective action on living cells suffering from oxidative stress.

Oxidative stress-induced increase in intracellular Ca2+ and Ca(2+)-induced increase in oxidative stress: an experimental model using dissociated rat brain neurons.

In order to study the oxidative stress-induced change in intracellular concentration of Ca2+ ([Ca2+]i) and Ca(2+)-induced oxidative stress, effects of hydrogen peroxide and ionomycin, a calcium ionophore, on rat cerebellar neurons were examined using a flow cytometer and fluorescent dyes: fluo-3 for monitoring [Ca2+]i; 2',7'-dichlorofluorescin, for reactive oxygen species; and 5-chloromethylfluorescein, for cellular nonprotein thiols. Oxidative stress induced by hydrogen peroxide dose-dependently increased [Ca2+]i and decreased the content of nonprotein thiols. Ionomycin increased oxidative metabolism and decreased the content of nonprotein thiols. Results suggest that oxidative stress induces an increase in [Ca2+]i while an increase in [Ca2+]i increases oxidative stress in neurons.

An autopsied case of acute myocarditis with myocardial calcification.

A 47-year-old woman was admitted with fever, hypotension, an elevated serum creatinine kinase level, and electrocardiographic abnormalities, which led to the diagnosis of acute myocarditis. She was placed on percutaneous cardiopulmonary support because of hemodynamic collapse on the third hospital day. Serial echocardiography showed gradual recovery of profound hypokinesis and edematous thickening of the left ventricle, but she died of sepsis on the 17th day without overt renal insufficiency or electrolytic abnormalities. Autopsy revealed myocardial necrosis with lymphocytic infiltrates and extensive myocardial calcification. Calcification was dense in the area of severe myocardial necrosis, and the distribution of calcium deposits suggested that the calcification was a consequence of significant inflammation of the myocardium. Recovery of regional wall motion was prominent in the area of severe inflammatory change. Dissociation between the pathologic and echocardiographic findings suggested the possibility of functional reversibility of severely damaged myocardium and possible mechanisms of abnormal contractile function other than inflammatory change.