The Fanconi anemia proteins FAA and FAC function in different cellular compartments to protect against cross-linking agent cytotoxicity

Fanconi anemia (FA) is an autosomal recessive disease characterized by chromosomal instability, bone marrow failure, and a high risk of developing malignancies. Although the disorder is genetically heterogeneous, all FA cells are defined by their sensitivity to the apoptosis-inducing effect of cross-linking agents, such as mitomycin C (MMC). The cloned FA disease genes, FAC and FAA, encode proteins with no homology to each other or to any known protein. We generated a highly specific antibody against FAA and found the protein in both the cytoplasm and nucleus of mammalian cells. By subcellular fractionation, FAA is also associated with intracellular membranes. To identify the subcellular compartment that is relevant for FAA activity, we appended nuclear export and nuclear localization signals to the carboxy terminus of FAA and enriched its localization in either the cytoplasm or the nucleus. Nuclear localization of FAA was both necessary and sufficient to correct MMC sensitivity in FA-A cells. In addition, we found no evidence for an interaction between FAA and FAC either in vivo or in vitro. Together with a previous finding that FAC is active in the cytoplasm but not in the nucleus, our results indicate that FAA and FAC function in separate subcellular compartments. Thus, FAA and FAC, if functionally linked, are more likely to be in a linear pathway rather than form a macromolecular complex to protect against cross-linker cytotoxicity.     

Orthodontic correction of lingually displaced canine teeth in a young dog using light-cured acrylic resin

PURPOSE: We investigated whether the high resolution ultrasound (13 MHz-scanner) shows smaller lesions and better differentiation than the 7.5 MHz-scanner. METHOD: Prospectively, sonography was performed on forty-seven patients with a 7.5 MHz-scanner as well as with a 13 MHz-scanner in identical slices. RESULTS: Obviously we could obtain more exact diagnoses by using the high resolution scanner. In two patients additional satellite of the primary tumor could be found. In four patients, unclear sonographic findings could be identified as cysts. A disadvantage in the usage of the 13 MHz-scanner is that mastopathy and benign lesions are more difficult to diagnose. With the high resolution more details could be seen although the inhomogeneity as well as the irregularity of the margins are seen more clearly and, therefore, the physician has to reestimate his point of view. To optimize the quality of the pictures made by high resolution ultrasound, it is necessary to regulate the system, which sometimes is quite difficult. CONCLUSION: The recognition of smallest lesions and the reliable presentation of cysts indicates that the 13 MHz-scanner is a good additive diagnostic parameter to the 7.5 MHz-scanner. Therefore, this method may become important for diagnosing multicentrity within carcinomas.     

Quantitative assessment of the operative results after extended myectomy and surgical reconstruction of the subvalvular mitral apparatus in hypertrophic obstructive cardiomyopathy using dynamic three-dimensional transesophageal echocardiography

OBJECTIVES: The aim of this study was to examine the value of dynamic three-dimensional (3D) transesophageal echocardiography (TEE) for the postoperative evaluation after extended myectomy and surgical reconstruction of the subvalvular mitral valve apparatus in patients with hypertrophic obstructive cardiomyopathy (HOCM). BACKGROUND: Two-dimensional imaging techniques such as echocardiography, computed tomography and magnetic resonance imaging have not been able to precisely quantify the effects of surgical therapy on the morphology of the left ventricular outflow tract (LVOT). METHODS: Multiplane TEE with 3D reconstruction was performed in 11 patients before and after the operation and in 16 normal control subjects for comparison. The preoperative maximal systolic pressure gradient in the LVOT was 69 +/- 59 mm Hg. The following variables were measured within the dynamic 3D data set: depth, width, length and cross-sectional area (CSA) gain caused by the myectomy trough, minimal CSA of the LVOT at each time point and its cyclic changes and maximal mitral leaflet deviation during systole. RESULTS: Functional class improved from 3.0 +/- 0.2 before the operation to 1.5 +/- 0.6 after it. The maximal systolic pressure gradient in the outflow tract decreased to 26 +/- 21 mm Hg postoperatively (p < 0.001). Minimal CSA of the outflow tract increased from 1.1 +/- 1.2 to 3.8 +/- 1.9 cm2 postoperatively (p < 0.001), similar to the value of the control group (4.2 +/- 1.5 cm2, p = NS). The area gain due to the myectomy trough was 1.3 +/- 1.0 cm2, corresponding to 48 +/- 12% of the total operative area difference. Maximal systolic depth of the myectomy was 7 +/- 2 mm, maximal width was 20 +/- 8 mm and length was 28 +/- 7 mm. Maximal deviation of the mitral leaflets fell from 15 +/- 7 to 6 +/- 7 mm postoperatively (p < 0.01). In five patients mass measurements of the intracavitary portion of the papillary muscle (PM) revealed an increase from 7.3 +/- 1.0 to 12.1 +/- 2.5 g due to surgical mobilization of PMs (p < 0.01). CONCLUSIONS: 3D TEE quantifies the differences in outflow tract morphology before and after surgery for HOCM. This technique may have an impact on the planning of operative interventions and allow for the evaluation of its results.     

Glutathione S-transferases: two-dimensional electrophoretic protein markers of lead exposure

Glutathione S-transferases (GST) are a family of detoxification isoenzymes that catalyze the conjugation of xenobiotics and their metabolites with reduced glutathione. Lead exposure in rats is known to induce GST isoenzymes in the liver and kidney. These changes in expression have potential use as biomarkers of lead exposure. Because two-dimensional electrophoresis (2-DE) enables one to analyze both protein abundance changes and chemical changes in protein structure, 2-DE was used to determine the effect of in vivo lead exposure on GST isoform expression in rat kidney cytosols. Male Sprague-Dawley rats were exposed to inorganic lead, and proteins were separated by conventional ISO-DALT and NEPHGE-DALT techniques and blotted for immunological identification. Lead exposure caused detectable inductions in both GSTP1 and GSTM1 and quantifiable charge modification in GSTP1. These preliminary data confirm the utility of 2-D electrophoretic GST analysis as indicative of lead exposure and toxicity and support its use for further elaboration of lead's effects on renal protein expression.     

Arthroscopic or open Bankart procedures: what are the costs?

The cost implications and resource utilization of arthroscopic and open Bankart procedures were evaluated to determine if differences exist between these procedures when performed in a community setting. Billing and hospital records of consecutive patients who underwent either open or arthroscopic Bankart procedures at the three facilities in our city during an 18-month period were analyzed. Procedure type (open or arthroscopic), location (hospital or surgicenter), operation time, operating room time, postanesthesia care unit time, step-down area time, charges for each of these, and anesthesiologist charges were analyzed for 11 open and 13 arthroscopic Bankart procedures. Open procedures took longer and required more operating room time than arthroscopic procedures regardless of location (P < .01). Open procedures required longer postanesthesia care unit time than arthroscopic procedures (P < .01). Facility made no difference. Anesthesia fees were less for arthroscopic ($882) than open Bankarts ($1,075) (P = .002). Total facility and anesthesia fees were less for arthroscopic ($4,747) than for open Bankarts ($6,062) (P = .05). The arthroscopic Bankart repair was performed more quickly than the open Bankart procedure, regardless of facility choice, and resulted in lower total charges. A surgicenter is less expensive whether these procedures are performed arthroscopically or open.     

Neutrophil chemiluminescence during phagocytosis is inhibited by abnormally elevated levels of acetoacetate: implications for diabetic susceptibility to infections

Human neutrophils by a chemiluminescence assay exhibit diminished phagocytic activity in the presence of abnormally high levels of the serum metabolite acetoacetate. These findings, along with our previous evidence demonstrating myeloperoxidase inhibition by acetoacetate, implicate metabolic ketosis in the inhibition of neutrophil microbicidal activity and thus in increased susceptibility to infections.     

Immunization of cattle with a BHV1 vector vaccine or a DNA vaccine both coding for the G protein of BRSV

A gE-negative bovine herpesvirus 1 (BHV1) vector vaccine carrying a gene coding for the G protein of bovine respiratory syncytial virus (BRSV) (BHV1/BRSV-G) induced the same high degree of protection in calves against BRSV infection and BHV1 infection as a multivalent commercial vaccine. A DNA plasmid vaccine, carrying the same gene as the BHV1/BRSV-G vaccine, significantly reduced BRSV shedding after BRSV infection compared with that in control calves, but less well than the BHV1/BRSV-G vaccine. Flow cytometric analysis showed a significant relative increase of gamma/delta+ T cells in peripheral blood after BRSV challenge-infection of the calves of the control group but not in the vaccinated groups. These results indicate that the G protein of BRSV can induce significant protection against BRSV infection in cattle, and that the BHV1/BRSV-G vaccine protects effectively against a subsequent BRSV and BHV1 infection.     

Modulation of basal ganglia neurotransmission by the classical antipsychotic fluphenazine is due in part to the blockade of dopamine D1-receptors

Classical antipsychotics, such as fluphenazine, influence neurotransmission by blocking both dopamine D1- and D2-receptors which in turn results in widespread adaptive changes in the neurochemistry of the basal ganglia. The purpose of the present study was to determine the role of D1-receptors in mediating some of these neurochemical events, including changes in D1- and D2-receptor binding, and the expression of preproenkephalin and glutamic acid decarboxylase mRNAs. For these experiments, rats were given a depot injection of fluphenazine decanoate or injected twice daily for 21 days with the D1-receptor antagonist SCH-23390. An additional group received both fluphenazine and SCH-23390 and controls were given saline. Fluphenazine administration decreased D2-receptor binding throughout the basal ganglia while SCH-23390 was without effect. In contrast to the uniform reduction in D2-receptor binding, fluphenazine altered D1-receptor binding in a region-dependent manner. Region-dependent changes were also observed in animals given SCH-23390 which increased binding in the entopeduncular nucleus and posterior caudate-putamen without affecting other brain regions. Both fluphenazine and SCH-23390 significantly enhanced preproenkephalin and glutamic acid decarboxylase (GAD) mRNA expression in the anterior striatum. Fluphenazine also increased GAD mRNA levels in the entopeduncular nucleus. Together, these results indicate that the attenuation of D1-receptor-mediated neurotransmission modulates a number of clinically relevant neurochemical processes in the basal ganglia.     

SopB, a protein required for virulence of Salmonella dublin, is an inositol phosphate phosphatase

Several proteins secreted by enteric bacteria are thought to contribute to virulence by disturbing the signal transduction of infected cells. Here, we report that SopB, a protein secreted by Salmonella dublin, has sequence homology to mammalian inositol polyphosphate 4-phosphatases and that recombinant SopB has inositol phosphate phosphatase activity in vitro. SopB hydrolyzes phosphatidylinositol 3,4,5-trisphosphate, an inhibitor of Ca2+-dependent chloride secretion. In addition, SopB hydrolyzes inositol 1,3,4,5,6 pentakisphosphate to yield inositol 1,4,5, 6-tetrakisphosphate, a signaling molecule that increases chloride secretion indirectly by antagonizing the inhibition of chloride secretion by phosphatidylinositol 3,4,5-trisphosphate [Eckmann, L., Rudolf, M. T., Ptasznik, A., Schultz, C., Jiang, T., Wolfson, N., Tsien, R., Fierer, J., Shears, S. B., Kagnoff, M. F., et al. (1997) Proc. Natl. Acad. Sci. USA 94, 14456-14460]. Mutation of a conserved cysteine that abolishes phosphatase activity of SopB results in a mutant strain, S. dublin SB c/s, with decreased ability to induce fluid secretion in infected calf intestine loops. Moreover, HeLa cells infected with S. dublin SB c/s do not accumulate high levels of inositol 1,4,5,6-tetrakisphosphate that are characteristic of wild-type S. dublin-infected cells. Therefore, SopB mediates virulence by interdicting inositol phosphate signaling pathways.