Activation of protein kinase C pathway contributes to hydrogen peroxide-induced increase in endothelial permeability.

BACKGROUND: We examined the effects of hydrogen peroxide (H2O2) on endothelial permeability and the possible role of protein kinase C (PKC) activation in mediating the response. EXPERIMENTAL DESIGN: Pulmonary microvessel endothelial cell monolayers were grown to confluency on gelatin- and fibronectin-coated microporous filters. Endothelial permeability was measured by determining the transendothelial clearance rate of [125I]albumin. The monolayers in all cases were challenged for 1 hour with H2O2. In some experiments, the monolayers were preincubated with PKC inhibitors H7 (an isoquinolinylsulphonamide derivative) (0.05 mM) or calphostin C (5 x 10(-6) mM) or with the inactive isoquinolinylsulphonamide analog, HA1004 (0.05 mM), before the H2O2 challenge. RESULTS: Addition of H2O2 (0 to 0.5 mM) to endothelial monolayers in the absence of PKC inhibitors resulted in a concentration-dependent increases in endothelial permeability and the response occurred without LDH release and morphologic evidence of cytolysis. The increase in permeability was significantly reduced by H7 and calphostin C, but not by HA1007. Immunocytochemical localization of PKC indicated that PKC isotype II was abundant in these cells and that it was distributed uniformly in the cytosol. H2O2 induced translocation of PKC to the cell membrane indicating enzyme activation. H7 and calphostin C prevented the H2O2-induced PKC translocation, whereas HA1004 had no effect. Both PKC inhibitors also prevented cell "rounding" and formation of interendothelial gaps, whereas HA1004 was ineffective. CONCLUSIONS: The results indicate that PKC activation is an important determinant of the H2O2-induced increase in endothelial permeability.     

Misidentification of toxigenic Corynebacterium diphtheriae as a Corynebacterium species with low virulence in a child with endocarditis.

A 6-year-old boy presented to a university hospital in Malaysia with infective endocarditis complicating cyanotic congenital heart disease. Blood cultures showed a gram-positive, aerobic, coryneform-like bacillus identified by the hospital laboratory as Corynebacterium xerosis, but a reference laboratory identified the organism as a toxigenic strain of Corynebacterium diphtheriae. The two laboratories concurred on all biochemical test results except for sucrose fermentation.     

Primary malignant melanoma of cervix: a case report

Primary malignant melanoma of cervix is a rare entity. A case of primary malignant melanoma of cervix presented with vaginal discharge and black colored growth over cervix. Histology showed malignant pleomorphic cells with melanin pigment in the cytoplasm.     

Bulk and Solution Copolymerization of Butyl Acrylate/Methyl Methacrylate at Elevated Temperatures

A series of bulk and solution (in toluene) copolymerizations of butyl acrylate/methyl methacrylate were performed independently at two laboratories. The runs were at elevated temperatures ranging from 90 to 140 °C conducted to high conversion levels, and samples were characterized for conversion, cumulative copolymer composition and number‐ and weight‐average molecular weights and distribution. Variation of the comonomer feed composition, temperature, and the solvent, initiator and chain transfer agent concentrations was studied. Using a mechanistic model, conversion data were predicted to high conversions using terminal model kinetics at 90 and 115 °C. The copolymer composition data conformed to the terminal kinetic model over the entire temperature range. Solvent effects were reflected by changes in the butyl acrylate rate constants.

Composition vs. conversion. Effect of feed composition for runs at 140 °C.     

First external quality assurance of antibody diagnostic for SARS-new coronavirus.

To confirm an infection with the new coronavirus (SARS-CoV) causing the severe acute respiratory syndrome (SARS) diagnostic assays for detection of SARS-CoV specific antibody are necessary. To evaluate the diagnostic performance of laboratories an external quality assurance (EQA) study was performed in 2004. Participating laboratories (9/20) correctly detected anti-SARS antibodies in serum samples without false positive results in an immunofluorescence assay. In contrast, only 4/13 laboratories detected most of the anti-SARS antibody positive samples without false positive results using enzyme immunoassays (EIA) and/or immunoblot. The overall results clearly demonstrate that serological diagnosis of SARS-CoV remains at an early stage of development, with further technical improvements required, particularly with respect to the use of SARS specific EIAs.     

Presence of two Ca2+ influx components in internal Ca2+-pool-depleted rat parotid acinar cells

The molecular mechanism(s) involved in mediating Ca²⁺ entry into rat parotid acinar and other non-excitable cells is not known. In this study we have examined the kinetics of Ca²⁺ entry in fura-2-loaded parotid acinar cells, which were treated with thapsigargin to deplete internal Ca²⁺ pools (Ca²⁺-pool-depleted cells). The rate of Ca²⁺ entry was determined by measuring the initial increase in free cytosolic [Ca²⁺] ([Ca²⁺]_i) in Ca²⁺-pool-depleted, and control (untreated), cells upon addition of various [Ca²⁺] to the medium. In untreated cells, a low-affinity component was detected with K _Ca = 3.4 ± 0.7 mM (where K _Ca denotes affinity for Ca²⁺) and V _max = 9.8 ± 0.4 nM [Ca²⁺]_i/s. In thapsigargin-treated cells, two Ca²⁺ influx components were detected with K _Ca values of 152 ±  79 μM (V _max = 5.1 ± 1.9 nM [Ca²⁺]_i/s) and 2.4 ±  0.9 mM (V _max = 37.6 ± 13.6 nM [Ca²⁺]_i/s), respectively. We have also examined the effect of Ca²⁺ and depolarization on these two putative Ca²⁺ influx components. When cells were treated with thapsigargin in a Ca²⁺-free medium, Ca²⁺ influx was higher than into cells treated in a Ca²⁺-containing medium and, while there was a 46% increase in the V _max of the low-affinity component (no change in K _Ca), the high-affinity component was not clearly detected. In depolarized Ca²⁺-pool-depleted cells (with 50 mM KCl in the medium) the high-affinity component was considerably decreased while there was an apparent increase in the K _Ca of the low-affinity component, without any change in the V _max. These results demonstrate that Ca²⁺ influx into parotid acinar cells (1) is increased (four- to five-fold) upon internal Ca²⁺ pool depletion, and (2) is mediated via at least two components, with low and high affinities for Ca²⁺. Received: 30 October 1995/Received after revisionand accepted: 13 December 1995     

A pharmacological study of group I muscle afferent terminals and synaptic excitation in the intermediate nucleus and Clarke's column of the cat spinal cord

Summary When administered microelectrophoretically GABA and piperidine-4-sulphonic acid depolarized the central terminations of muscle group Ia and Ib afferent fibres in the lumbar intermediate nucleus and Clarke's column of cats anaesthetised with pentobarbitone sodium. Both this depolarization, and primary afferent depolarization, generated by impulses in other primary afferent fibres which produce prolonged bicuculline-sensitive inhibition of the firing of group I afferent fibre-excited interneurones in the intermediate nucleus and cells in Clarke's column, are reduced by microelectrophoretic bicuculline methochloride. Systemically administered (±)-baclofen hydrochloride (maximum dose 8 mg kg^–1) depressed the monosynaptic excitation of Clarke's column neurones by impulses in muscle and cutaneous afferent fibres. Microelectrophoretically administered (–)-baclofen reduced the bicuculline-sensitive primary afferent depolarization of group I terminations without, however, reducing the depolarizing action of GABA or piperidine-4-sulphonic acid. The depression by (–)-baclofen of the group I monosynaptic excitation of intermediate nucleus neurones is not reduced by concentrations of bicuculline methochloride adequate to suppress prolonged inhibition of these neurones     

Differential effects of complement activation induced by cobra venom factor on pulmonary transvascular fluid and protein exchange

Sheep were prepared with lung lymph fistulas for assessment of the effects of complement activation on pulmonary transvascular fluid and protein exchange in the intact animal. Cobra venom factor (CVF 200 +/- 46 U/kg) was injected intravenously for activation of the complement system. In some animals (n = 6), pulmonary lymph flow (Qlym) and pulmonary arterial pressure (Ppa) increased without a change in the lymph-to-plasma protein concentration ratio (L/P) or in pulmonary blood flow (QL), indicating an increase in pulmonary vascular permeability to proteins. In another group (n = 6), Qlym and the L/P did not change, and there were also no changes in Ppa and QL following a similar injection of CVF. Morphologic evidence showed that leukocytes were trapped in pulmonary vessels and interstitium of both groups. Pulmonary edema was also present in both groups. Complement activation does not uniformly increase pulmonary lymph flow despite pathologic evidence of leukocyte sequestration and pulmonary edema. The lack of change in lymph flow in some animals may be due to lymphatic insufficiency, or lack of generation of humoral mediators, and/or a decrease in pulmonary capillary pressure.