Induction of immunologic refractoriness in adults by meningococcal C polysaccharide vaccination

Thirty-four adults were vaccinated with 1/50 of the usual dose of meningococcal polysaccharide vaccine (1 microg of A, C, Y, and W135 polysaccharides, given intramuscularly). This dose was selected as a probe to assess B cell memory. The probe elicited meningococcal C bactericidal antibody responses in all 18 adults who had been vaccinated 4 years earlier with an investigational meningococcal A and C oligosaccharide-protein conjugate vaccine and in the majority of the 11 subjects vaccinated for the first time. In contrast, the responses of the 5 adults given a full dose of licensed polysaccharide vaccine 4 years earlier were <1/10 of those of the other 2 groups. Thus, adults previously given a full dose of meningococcal polysaccharide vaccine show evidence of immunologic refractoriness to group C polysaccharide, whereas refractoriness is not observed after conjugate vaccination. These findings have implications for the use of meningococcal polysaccharide vaccine when the risk of disease is low.     

Primary binding domain of bovine von Willebrand factor fragment expressed in E. coli

Bovine vWF cDNA has been cloned from a bovine endothelial cell library. A fragment of this cDNA, corresponding to amino acid sequence Leu 469-Ser 723, called primary adhesion domain (PAD-1), and containing the binding sites for platelet glycoprotein Ib (GPIb), heparin and collagen, has been expressed in E. coli. The reduced and alkylated form of fragment PAD-1 inhibited native vWF binding to GPIb. Fragment PAD-1 bound to heparin and botrocetin in a specific and dose dependent manner as did the native vWF. In a solid-phase assay, fragment PAD-1 bound to calf skin collagen in contrast to a human vWF recombinant fragment (Ser 445-Val 733) which was inactive in the same assay. The studies presented in this paper demonstrated that the A1 domain of bovine vWF contained the GPIb, heparin, botrocetin as well as collagen binding sites and that integrity of the disulfide bond (Cys 509-Cys 695), did not seem to be essential for binding of bovine vWF fragment to GPIb.     

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.     

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.     

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.     

Bimodification of polyacrylonitrile (PAN) fibers

A new technique for the modification of PAN fibers has evolved, in which the starting precursor material is impregnated with an aliphatic carboxylic acid, as well as KMnO₄. The bimodification of PAN, attempted by us for the first time, is aimed at achieving dual advantages in plasticization effect, as well as in a catalytic effect in a single treatment. The unmodified, unimodified, and bimodified precursors have been characterized for their mechanical, thermal, and physical behavior during various stages of processing to carbon fibers. These results are discussed in detail. The study further shows an 18% improvement in the mechanical properties of carbon fibers, prepared from bimodified precursor, as compared to that of unmodified precursor. One of the reasons for such improvement is the lowering in diameter of PAN fibers and in the resulting carbon fibers. © 1993 John Wiley & Sons, Inc.     

Effects of ethanol administration on components of the ubiquitin proteolytic pathway in rat liver

Hepatic protein accumulation during ethanol administration may result partly from an ethanol-elicited decline in hepatic protein degradation, which we have previously shown. We conducted the current studies to examine the effects of ethanol administration on the levels of hepatic ubiquitin, an 8.5-kd protein which is an important mediator of extralysosomal protein catabolism. Rats were pair-fed liquid diets containing either ethanol (36% of calories) or isocaloric maltose-dextrin for 1 to 5 weeks. Ubiquitin was immunochemically quantified by competitive enzyme-linked immunosorbent assay (ELISA) in crude cytosol fractions from whole liver and in 12,000g supernatants of hepatocyte lysates. Ubiquitin levels in hepatic cytosol fractions of ethanol-fed rats exceeded those of controls by about 30%. Isolated hepatocytes from ethanol-fed animals also showed a 40% to 75% elevation of ubiquitin above that in cells of pair-fed controls and this difference exceeded the relative rise in hepatocellular protein. In hepatocyte lysates subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblotting, we detected monomeric ubiquitin and higher molecular mass ubiquitin-protein conjugates. However, the immunoblot analyses revealed no quantitative changes in the level of either free or conjugated ubiquitin. The ubiquitin conjugating activity of crude and diethyl aminoethyl-fractionated liver cytosols of ethanol-fed rats had equal capacities to those from controls in catalyzing the formation of ubiquitin-protein conjugates. Our findings indicate that chronic ethanol consumption increased the level of immunoreactive ubiquitin in rat liver. This may have resulted from enhanced ubiquitin production because of an ethanol-elicited stress response and/or decreased catabolism of ubiquitin and its conjugates. Our findings also provide no indication that the ethanol-elicited reduction in hepatic proteolysis is because of a ubiquitin-mediated mechanisms.     

The strength of the fibre-polymer interface in short glass fibre-reinforced polypropylene

A method for the determination of the interfacial bond strength in well-aligned short glass fibre-reinforced polypropylene samples is discussed. The method takes into account the variation of the interfacial shear stress during the deformation process; consequently, it yields very consistent results at all values of the composite strain. The influence of the fibre orientation with respect to the load axis is appropriately considered using macro-mechanical analyses for stiffness and strength of the composite. The method is compared with two other methods reported in the literature.