Vasopressin induces dopamine release and cyclic AMP efflux from the brain of water-deprived rats: inhibitory effect of vasopressin V2 receptor-mediated phosphorylation

MR imaging is the best noninvasive method for the evaluation of articular cartilage. Recent studies have clarified the MR appearance of normal articular cartilage and proven that MR imaging can detect chondral lesions with high accuracy. Quantitative imaging holds promise for the accurate determination of cartilage volumes and thickness measurements, as well as the ability to detect early chondral degeneration and biochemical changes before gross morphologic changes occur.     

Recombinant expression and characterization of the major beta-lactamase of Mycobacterium tuberculosis

New antibiotic regimens are needed for the treatment of multidrug-resistant tuberculosis. Mycobacterium tuberculosis has a thick peptidoglycan layer, and the penicillin-binding proteins involved in its biosynthesis are inhibited by clinically relevant concentrations of beta-lactam antibiotics. beta-Lactamase production appears to be the major mechanism by which M. tuberculosis expresses beta-lactam resistance. beta-Lactamases from the broth supernatant of 3- to 4-week-old cultures of M. tuberculosis H37Ra were partially purified by sequential gel filtration chromatography and chromatofocusing. Three peaks of beta-lactamase activity with pI values of 5.1, 4.9, and 4.5, respectively, and which accounted for 10, 78, and 12% of the total postchromatofocusing beta-lactamase activity, respectively, were identified. The beta-lactamases with pI values of 5.1 and 4.9 were kinetically indistinguishable and exhibited predominant penicillinase activity. In contrast, the beta-lactamase with a pI value of 4.5 showed relatively greater cephalosporinase activity. An open reading frame in cosmid Y49 of the DNA library of M. tuberculosis H37Rv with homology to known class A beta-lactamases was amplified from chromosomal DNA of M. tuberculosis H37Ra by PCR and was overexpressed in Escherichia coli. The recombinant enzyme was kinetically similar to the pI 5.1 and 4.9 enzymes purified directly from M. tuberculosis. It exhibited predominant penicillinase activity and was especially active against azlocillin. It was inhibited by clavulanic acid and m-aminophenylboronic acid but not by EDTA. We conclude that the major beta-lactamase of M. tuberculosis is a class A beta-lactamase with predominant penicillinase activity. A second, minor beta-lactamase with relatively greater cephalosporinase activity is also present.     

Effects of gonadectomy in infancy and adulthood on handedness in male and female Mongolian gerbils

When assuming a species-typical tripodal posture, female Mongolian gerbils most often rest on their left forepaws and hold their right forelimbs aloft; male gerbils most often do the reverse. This experiment examined effects of gonadectomy, both in infancy and in adulthood, on the sexually dimorphic asymmetry in forelimb use by Mongolian gerbils when maintaining a tripodal stance. In adulthood, both male and female gerbils that had been gonadectomized at birth reversed their forelimb use while in a tripodal stance: Gonadectomized males used their forelimbs as did sham-operated females, and gonadectomized females used their forelimbs as did sham-operated males. Gonadectomy in adulthood abolished the sexual dimorphism in forelimb use seen in sham-operated subjects. It was concluded that gonadal hormones have organizational as well as possible activational effects on adult patterns of forelimb use by gerbils.     

Oxidative inactivation of gastric peroxidase by site-specific generation of hydroxyl radical and its role in stress-induced gastric ulceration

We have shown earlier that restraint-cold stress-induced gastric ulceration in rats is caused by metal ion-dependent generation of hydroxyl radical (OH.) and oxidative inactivation of the gastric peroxidase (GPO), an important H2O2 scavenging enzyme. To study the mechanism of the oxidative damage of GPO, the purified enzyme was exposed to an OH. generating system containing Cu2+, ascorbate, and H2O2. Kinetic studies indicate that the enzyme is inactivated in a time-dependent process showing saturation with respect to Cu2+ concentration. The enzyme specifically requires Cu2+ and is not inactivated by the same concentration of Fe2+, Mn2+, or Zn2+. Sensitivity to catalase indicates the critical role of H2O2 in the inactivation. Inactivation is insensitive to superoxide dismutase, suggesting no role of superoxide. The rate of inactivation is not increased in D2O excluding the involvement of singlet oxygen in the process. However, OH. scavengers such as benzoate or mannitol cannot prevent inactivation. The results indicate a plausible generation of OH. within the enzyme molecule as the cause of inactivation. Fragmentation of peptide linkage or intramolecular crosslinking, gross change of tertiary structure, or change in intrinsic tryptophan fluorescence which occurs in "global" oxidation are not evident. Inactivation is dependent on pH and from a plot of K(obs) of inactivation against pH, the controlling role of an ionizable group of the enzyme having a pka of 7.8 could be suggested, deprotonation of which favors inactivation. Amino acid analysis shows a specific loss of two lysine residues in the inactivated enzyme. Competitive kinetic studies indicate that pyridoxal phosphate, a specific modifier of the lysine residue, prevents inactivation by competing with Cu2+ for binding at the GPO. A Cu2+ binding motif consisting at least of two lysine residues exists in GPO, which specifically binds Cu2+ and generates OH.. The radical oxidizes the lysine residues and perturbs the heme environment to cause inactivation. We suggest that oxidative damage of GPO is mediated by site-specific generation of OH. and not by the OH. generated in the bulk phase.     

Intravascular large B-cell lymphoma: the CD5 antigen is expressed by a subset of cases

The CD5 antigen is a T-cell associated marker that is also usually expressed by two B-cell neoplasms, chronic lymphocytic leukemia/small lymphocytic lymphoma and mantle cell lymphoma. We observed CD5 antigen expression in a subset of cases of intravascular large B-cell lymphoma (IVLBL), and we report here five cases. The patients, two men and three women, ranged in age from 59 to 81 years. Biopsy specimens were obtained from kidney, lung, bone marrow, abdominal wall, and neck, the latter involving a lymphangioma. All of the cases had histologic features typical of IVLBL, with large and atypical lymphoid cells located predominantly within blood vessels. Immunohistochemical studies performed using routinely fixed, paraffin-embedded tissue sections showed that the neoplastic cells were B cells, positive for the CD20 antigen and negative for the CD3 or CD43 antigens. All cases were also positive for the CD5 antigen. One case had an immunoglobulin heavy chain gene rearrangement shown by using a polymerase chain reaction method. The finding of CD5 antigen expression in a subset of IVLBL cases adds to other evidence in the literature suggesting that IVLBL is a heterogeneous entity. We considered the possibility that these cases were related to or represented unusual histologic forms of transformation from either chronic lymphocytic leukemia/small lymphocytic lymphoma or mantle cell lymphoma. All of the cases, however, were negative for the CD23 antigen and cyclin D1 (bcl-1) protein, which is evidence against this interpretation. The biologic significance of CD5 antigen expression in cases of IVLBL is uncertain. These neoplasms might arise from a separate lineage of CD5-positive B cells or from a specific, early stage of B-cell differentiation. Alternatively, some investigators have suggested that CD5 antigen expression by B cells is a marker of activation.     

A randomized controlled trial of filgrastim during remission induction and consolidation chemotherapy for adults with acute lymphoblastic leukemia: CALGB study 9111

Recombinant human granulocyte colony-stimulating factor (G-CSF; filgrastim) shortens the time to neutrophil recovery after intensive chemotherapy, but its role in the treatment of adults with acute lymphoblastic leukemia (ALL) is uncertain. We randomly assigned 198 adults with untreated ALL (median age, 35 years; range, 16 to 83) to receive either placebo or G-CSF (5 microgram/kg/d) subcutaneously, beginning 4 days after starting intensive remission induction chemotherapy and continuing until the neutrophil count was >/=1, 000/microL for 2 days. The study assignment was unblinded as individual patients achieved a complete remission (CR). Patients initially assigned to G-CSF then continued to receive G-CSF through 2 monthly courses of consolidation therapy. Patients assigned to placebo received no further study drug. The median time to recover neutrophils >/=1,000/microL during the remission induction course was 16 days (interquartile range [IQR], 15 to 18 days) for the patients assigned to receive G-CSF and 22 days (IQR, 19 to 29 days) for the patients assigned to placebo (P < .001). Patients in the G-CSF group had significantly shorter durations of neutropenia (<1, 000/microL) and thrombocytopenia (<50,000/microL) and fewer days in the hospital (median, 22 days v 28 days; P = .02) compared with patients receiving placebo. The patients assigned to receive G-CSF had a higher CR rate and fewer deaths during remission induction than did those receiving placebo (P = .04 by the chi-square test for trend). During Courses IIA and IIB of consolidation treatment, patients in the G-CSF group had significantly more rapid recovery of neutrophils >/=1,000/microL than did the control group by approximately 6 to 9 days. However, the patients in the G-CSF group did not complete the planned first 3 months of chemotherapy any more rapidly than did the patients in the placebo group. Overall toxicity was not lessened by the use of G-CSF. After a median follow-up of 4. 7 years, there were no significant differences in either the disease-free survival (P = .53) or the overall survival (P = .25) for the patients assigned to G-CSF (medians, 2.3 years and 2.4 years, respectively) compared with those assigned to placebo (medians, 1.7 and 1.8 years, respectively). Adults who received intensive chemotherapy for ALL benefited from G-CSF treatment, but its use did not markedly affect the ultimate outcome.     

Cytochrome P450 and dependent activities in unexposed and PAH-exposed terrestrial annelids

The cytochrome P450 system of the oligochaetes Eisenia f. fetida (tiger worm) and Enchytraeus crypticus (pot worm) was analysed using ethoxy-, pentoxy- and benzoxyresorufin as substrates for monooxygenase activity. Whole body microsomes of the earthworm E.f. fetida displayed PentROD activity in the range from 0.26 to 1.05 pmol mg protein-1 min-1 and BenzROD activity in the range from 0.14 to 0.30 pmol mg protein-1 min-1. Exposure of the animals for up to four weeks to 100 mg fluoranthene or benzo[a]pyrene kg-1 soil (dry weight) did not induce significant changes in the activity of these monooxygenases. In E. crypticus EROD activity was in the range from 2.10 to 6.18 pmol mg protein-1 min-1 and PentROD activity in the range from 1.75 to 4.78 pmol mg protein-1 min-1. Short-term exposure to BaP by feeding reduced the EROD activity significantly by 45%, but did not effect PentROD activity. After long-term (8 weeks) exposure to BaP in the agar-agar medium EROD activity was not changed but PentROD had decreased to zero. In both species cytochrome P420 and NADPH-cytochrome C reductase activity were present. In E.f. fetida microsomes are associated with the giant haemoglobin. Both can be separated by gel filtration on a Sepharose B2 column or by hydrophobic interaction chromatography after solubilisation with cholate. NADPH-cytochrome C reductase elutes together with haemoglobin. Cytochrome P420 is eluted with Emulgen 911 and can be further purified by ion exchange chromatography using HA-Ultrogel. By SDS-PAGE of the purified microsomal proteins three protein bands are visualised in the range of cytochrome P450 displaying an apparent molecular mass of 54, 56 and 58 kDa. Only the 54-kDa protein interacts weakly with perch (Perca fluviatilis) CYP1A antibodies, while two proteins with an apparent molecular mass of 65 and 71 kDa give a strong antibody signal.     

Substitutions of aspartate 378 in the phosphorylation domain of the yeast PMA1 H+-ATPase disrupt protein folding and biogenesis

There is strong evidence that Asp-378 of the yeast PMA1 ATPase plays an essential role in ATP hydrolysis by forming a covalent beta-aspartyl phosphate reaction intermediate. In this study, Asp-378 was replaced by Asn, Ser, and Glu, and the mutant ATPases were expressed in a temperature-sensitive secretion-deficient strain (sec6-4) that allowed their properties to be examined. Although all three mutant proteins were produced at nearly normal levels and remained stable for at least 2 h at 37 degrees C, they failed to travel to the vesicles that serve as immediate precursors of the plasma membrane; instead, they became arrested at an earlier step of the secretory pathway. A closer look at the mutant proteins revealed that they were firmly inserted into the bilayer and were not released by washing with high salt, urea, or sodium carbonate (pH 11), treatments commonly used to strip nonintegral proteins from membranes. However, all three mutant ATPases were extremely sensitive to digestion by trypsin, pointing to a marked abnormality in protein folding. Furthermore, in contrast to the wild-type enzyme, the mutant ATPases could not be protected against trypsinolysis by ligands such as MgATP, MgADP, or inorganic orthovanadate. Thus, Asp-378 functions in an unexpectedly complex way during the acquisition of a mature structure by the yeast PMA1 ATPase.     

Pathologic seminal vesicle invasion after radical prostatectomy for patients with prostate carcinoma: effect of early adjuvant radiation therapy on biochemical control

BACKGROUND: The authors evaluated the effect of postoperative radiation therapy on freedom from biochemical failure (bNED) in men with prostate carcinoma who had pathologic seminal vesicle invasion after radical prostatectomy and negative pelvic lymph node dissection (pT3cN0). METHODS: Between 1989 and 1995, 375 men underwent radical prostatectomy at Thomas Jefferson University Hospital. Fifty-three men (13%) had pT3cN0 prostate carcinoma and were the subject of this analysis. Men in whom prostate specific antigen (PSA) could not be detected were deemed free of biochemical failure. RESULTS: Of the 53 men with pT3cN0 prostate carcinoma, 18 had an elevated PSA immediately after surgery and received salvage radiation therapy (RT). The 3-year bNED rate for this group was only 38%. At 3 months, PSA could not be detected in the other 35 men. Fifteen of those 35 men underwent early adjuvant RT, and the other 20 were observed for biochemical failure. The 3-year bNED rate for the 15 patients treated with immediate adjuvant RT was 86%, compared with 48% for the 20 men who were observed (P = 0.01). CONCLUSIONS: These data suggest that early adjuvant RT for men with pT3cN0 prostate carcinoma and no detectable PSA postoperatively reduces the likelihood of future biochemical failure. Men with pT3cN0 prostate carcinoma and a persistently elevated postoperative PSA level are less likely to benefit from RT and should be considered for systemic therapy.     

Ultrasonically activated shears in thyroid surgery

BACKGROUND: Ultrasonically activated shears (UAS) have been documented to be both safe and fast devices in laparoscopic surgery. We studied whether the use of UAS would have some advantage in thyroid surgery. METHODS: Thyroidectomies, performed by one senior endocrine surgeon between December 1996 and February 1997, were retrospectively matched, with patients operated on by the same surgeon using the conventional method. RESULTS: Six pairs of total thyroidectomies and one pair of lobectomies could be matched. Mean operating time was 100 minutes for the patients operated on with the UAS and 154 minutes for the patients operated on with the conventional method. The mean operating time with the UAS was thus on average 64.6% of the operation time with the conventional method, with a 95% confidence interval from 50.1% to 83.5% (t = 4.00, 6 df, P = 0.007). CONCLUSIONS: In this material the use of UAS reduced significantly operating time in thyroidectomies.