Parametric and Nonparametric Analyses of Repeated Ordinal Categorical Data

We compare two models for the analysis of repeated ordinal categorical data: the classical parametric model for means of scores assigned to the categories of the response variable and a nonparametric model based on relative effects derived from the marginal distribution functions of the response. An example in the field of Dentistry is used to illustrate and to compare the models. We also consider a simulation study to evaluate the type‐I error rates and the power of tests under both models in a balanced design setup. The simulation results suggest that both approaches behave similarly for equally spaced scores but may perform differently otherwise. (© 2004 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)     

Effects of PGE1 or PGE2 and/or acetazolamide on expulsion of Nippostrongylus brasiliensis from rats

PGE1 and PGE2 have been reported to enhance natural expulsion of Nippostrongylus brasiliensis, a nematode parasite, from the intestine of the rat. Mucus production may also be a key element of worm rejection. Our study attempts to determine if 1) PGE1 or PGE2 alter the normal course of infection with N. brasiliensis in rats, 2) a known mucous enhancing drug, acetazolamide, can augment the rate of worm expulsion, and 3) combinations of prostaglandins and acetazolamide affect N. brasiliensis in the rat. Rats were inoculated with approximately 1,000 infective larvae of N. brasiliensis. Animals were administered, intraduodenally, one of the following: 0.2 ml 0.9% NaCl; 0.2 ml 100% ethanol; 250 micrograms PGE1/0.2 ml 100% ethanol; 250 micrograms PGE2/0.2 ml 100% ethanol; 250 micrograms acetazolamide/0.2 ml 100% ethanol; 250 micrograms PGE1 or PGE2 + 250 micrograms acetazolamide/0.2 ml 100% ethanol. These solutions were given in a single bolus on day 6 postinoculation (PI) or twice daily on days 6-9 PI. Following these treatments the number of parasite ova per gram feces per day for days 6-10 PI and numbers of worms present at necropsy on day 10 PI were determined. Treatment with prostaglandins or acetazolamide or both failed to adversely affect egg deposition by adult female worms or the number of worms in the small intestine. These results do not support the involvement of prostaglandins in the expulsion of N. brasiliensis from the host intestine.     

Different Characteristics of the Two Glutamate Synthases in the Green Leaves of Lycopersicon esculentum

The two glutamate synthases, NAD(P)H- and ferredoxin-dependent, from the green leaves of tomato plants (Lycopersicon esculentum L. cv Hellfrucht frühstamm) differed in their chemical properties and catalytic behavior. Gel filtration of NAD(P)H enzyme gave an apparent molecular size of 158 kilodalton, whereas the ferredoxin enzyme molecular size was 141 kilodalton. Arrhenius plots of the activities of the two enzymes showed that the NAD(P)H enzyme had two activation energies; 109.6 and 70.5 kilojoule per mole; the transition temperature was 22°C. The ferredoxin enzyme however, had only one activation energy; 56.1 kilojoule per mole. The respective catalytic activity pH optima for the NAD(P)H- dependent and the ferredoxin dependent enzymes were around 7.3 and 7.8. In experiments to evaluate the effects of modulators aspartate enhanced the NAD(P)H-linked activity, with a K_a value of 0.25 millimolar, but strongly inhibited that of the ferredoxin-dependent glutamate synthase with a K_i of 0.1 millimolar. 3-Phosphoserine was another inhibitor of the ferredoxin dependent enzyme with a K_i value of 4.9 millimolar. 3-Phosphoglyceric acid was a potent inhibitor of the ferredoxin-dependent form, but hardly affected the NAD(P)H-dependent enzyme. The results are discussed and interpreted to propose different specific functions that these activities may have within the leaf tissue cell.     

Production and characterization of human renin antibodies with region-oriented synthetic peptides

Polyclonal and monoclonal antibodies were raised against pure human renin, but nothing was known about the regions against which they were directed. Using a three-dimensional model of mouse submandibular renin, we selected seven peptide sequences as belonging to potential epitopes. The main criteria for their choice were the location of the peptide sequences near the catalytic region and on the surface of the renin molecule and their hydrophilicity. After transposition of the regions to the 340-amino acid sequence of human renin, the seven peptides (corresponding to amino acids 50-60, 63-71, 81-90, 118-126, 162-169, 247-255, and 287-295) were synthesized, coupled to bovine serum albumin, and injected into rabbits. Five of these peptides elicited antibodies, and 50-68% binding of the corresponding iodinated peptide was obtained with a 1:25 dilution of antiserum. The antisera titers ranged from 1:5,000 to 1:100,000 when tested by enzyme-linked immunosorbent assay. The same antisera bound 15-65% of labeled pure human renin at a final dilution of 1:2.5, the highest percentage being obtained with peptide 81-90 antiserum. At a 1:5 dilution, the five antisera inhibited renin activity by 23-68% in human plasma with a high renin activity (40 ng of angiotensin I/h/ml). At a final dilution of 1:50, peptide 81-90 antiserum was still capable of producing 25% inhibition. Purified IgG (0.6 mg) from this antiserum inhibited pure human renin activity by up to about 40%, as measured by its reaction with pure synthetic human tetradecapeptide substrate. Antigenic peptides that mimic a part of the human renin sequence, especially peptide 81-90 representing the "flap" covering the cleft between the two renin lobes, constitute promising tools for the development of a synthetic antirenin vaccine.     

Structural analysis of an HLA-B27 population variant, B27f. Multiple patterns of amino acid changes within a single polypeptide segment generate polymorphism in HLA-B27

The structure of a new HLA-B27 variant, B27f, distinguishable from other HLA-B27 subtypes by isoelectric focusing and serologic criteria, has been established by comparative peptide mapping and radiochemical sequence analysis. HLA-B27f differs from the major B27.1 subtype in three clustered amino acid replacements: Asp74, Asp77, and Leu81 in B27.1 are changed to Tyr74, Asn77, and Ala81, respectively in B27f. This pattern of differences is analogous to that of HLA-B27.2 in that this subtype also differs from B27.1 in multiple clustered substitutions within the same segment. Thus, polymorphism within the HLA-B27 system is being achieved by introducing different sets of amino acid changes within a particular short segment of the alpha 1 domain. The most likely mechanism for the introduction of multiple changes within this segment is a nonreciprocal recombination event, such as gene conversion. The structural analogies and ethnic distribution of B27f and B27.2 as compared with those of B27.3, and B27.4 support a dynamic model of HLA-B27 evolution in which polymorphism has been created after the separation of the major ethnic groups. In this model, a Caucasian branch would be characterized by subtypes differing from B27.1 in a few changes within the alpha 1 domain, which were probably generated by single genetic steps. An Oriental branch would include those subtypes which differ from B27.1 by changes in both alpha 1 and alpha 2, involving multiple genetic steps for their generation.     

Nitrogen metabolism in tumor bearing mice

In experiments with whole animals infested with a highly malignant strain of Ehrlich ascites tumor cells, serial concentrations of amino acids were determined for host plasma, ascitic fluid, and tumor cells, throughout tumor development. Concentration gradients of glutamine, asparagine, valine, leucine, isoleucine, phenylalanine, tyrosine, histidine, tryptophan, arginine, serine, methionine, and taurine from the host plasma toward the ascitic liquid were established; while on the other hand, concentration gradients from the ascitic liquid toward the plasma were established for glutamate, aspartate, glycine, alanine, proline, and threonine. With the exception of aspartate the concentrations of these amino acids were highest inside the cells. Arginine was the only amino acid not detected in tumor cells. In vitro incubations of tumor cells in the presence of glutamine and/or glucose, as the energy and nitrogen sources, confirmed the amino acid fluxes previously deduced from the observed relative concentrations of amino acids in plasma, ascitic liquid, and tumor cells, suggesting that glutamate, alanine, aspartate, glycine, and serine can be produced by tumors. These findings support that changes in amino acid patterns occurring in the host system are related to tumor development.     

Coated nuts as an enrichment device to elicit tool use in tufted capuchins (Cebus apella)

The present paper describes a simple technique that hardens the shell of nuts and makes the use of a tool to crack them open more compelling. Walnuts were coated with a dough of sawdust and nontoxic white glue in different combinations; they were tested for hardness by using machines normally used to test different kinds of wood. Data on relative hardness for uncoated walnuts and walnuts coated with dough of two different combinations are presented. The coated walnuts were significantly harder to break than the uncoated ones, whereas no significant difference was found when comparing the hardness of two types of coated walnuts. Furthermore, observations on a captive group of tufted capuchins (Cebus apella) are described. The monkeys needed significantly more time to break open the coated walnuts. Early results show that coated walnuts may favor acquistion of tool use skills in a juvenile capuchin.     

Purification of phosphate-dependent glutaminase from isolated mitochondria of Ehrlich ascites-tumour cells.

Phosphate-dependent glutaminase was purified to homogeneity from isolated mitochondria of Ehrlich ascites-tumour cells. The enzyme had an Mr of 135,000 as judged by chromatography on Sephacryl S-300. SDS/polyacrylamide-gel electrophoresis displayed two protein bands, with Mr values of 64,000 and 56,000. Two major immunoreactive peptides of Mr values of 65,000 and 57,000 were found by immunoblot analysis using anti-(rat kidney glutaminase) antibodies. The concentration-dependences for both glutamine and phosphate were sigmoidal, with S0.5 values of 7.6 mM and 48 mM, and Hill coefficients of 1.5 and 1.6, respectively. The glutaminase pH optimum was 9. The activation energy of the enzymic reaction was 58 kJ/mol. The enzyme showed a high specificity towards glutamine. A possible explanation for the different kinetic behaviour found for purified enzyme and for isolated mitochondria [Kovacević (1974) Cancer Res. 34, 3403-3407] should be that a conformational change occurs when the enzyme is extracted from the mitochondrial inner membrane.