Effect of milk centrifugation and incorporation of high-heat-treated centrifugate on the composition,texture, and ripening characteristics of Maasdam cheese

This study investigated the effect of centrifugation (9,000 × g, 50°C, flow rate = 1,000 L/h), as well as the incorporation of high-heat-treated (HHT) centrifugate into cheese milk on the composition, texture, and ripening characteristics of Maasdam cheese. Neither centrifugation nor incorporation of HHT centrifugate into cheese milk had a pronounced effect on the compositional parameters of any experimental cheeses, except for moisture and moisture in nonfat substance (MNFS) levels. Incorporation of HHT centrifugate at a rate of 6 to 10% of the total milk weight into centrifuged milk increased the level of denatured whey protein in the cheese milk and also increased the level of MNFS in the resultant cheese compared with cheeses made from centrifuged milk and control cheeses; moreover, cheese made from centrifuged milk had ～3% higher moisture content on average than control cheeses. Centrifugation of cheese milk reduced the somatic cell count by ～95% relative to the somatic cell count in raw milk. Neither centrifugation nor incorporation of HHT centrifugate into cheese milk had a significant effect on age-related changes in pH, lactate content, and levels of primary and secondary proteolysis. However, the value for hardness was significantly lower for cheeses made from milk containing HHT centrifugate than for other experimental cheese types. Overall, centrifugation appeared to have little effect on composition, texture, and ripening characteristics of Maasdam cheese. However, care should be taken when incorporating HHT centrifugate into cheese milk, because such practices can influence the level of moisture, MNFS, and texture (particularly hardness) of resultant cheeses. Such differences may have the potential to influence subsequent eye development characteristic, although no definitive trends were observed in the present study and further research on this is recommended.     

Method for three-dimensional data registration from disparate imaging modalities in the NOGA Myocardial Viability Trial

Region-by-region comparison of data concerning left ventricular (LV) status is difficult to perform quantitatively if the data was acquired from disparate imaging modalities. We validated a method for comparing measurements obtained by electromechanical mapping (EMM) catheter with dobutamine stress echocardiography (DSE) via biplane contrast ventriculography, with the assistance of three-dimensional (3-D) echocardiographic data. The ventriculograms were traced and the borders were used to reconstruct the LV in 3-D with the aid of a database of 3-D echocardiographic studies. The 3-D LV was oriented to the EMM data based on the body coordinates and then manually scaled and translated to fit. The EMM data were mapped to the 3-D surface. The 3-D surface was divided into the 16 regions defined for echocardiographic assessment. The mean EMM value for local linear shortening, a parameter of function, was computed in each segment. The EMM and semiquantitative echocardiographic assessments of regional myocardial function were compared by segment, and the volume of the 3-D LV was compared with the volume computed from the ventriculogram. The volume of the 3-D surface correlated closely with that of the ventriculogram (r = 0.97, SEE = 27.4 ml) but with a significant overestimation of 63 +/- 35 ml. There was a highly significant (p < 0.0001) agreement in regional function between EMM and echo. Local linear shortening correlated significantly (p < 0.0001) with echocardiographic severity of wall motion, averaging 9.5 +/- 6.5, 8.1 +/- 5.4, 5.9 +/- 4.8, and 6.2 +/- 3.3 in segments read as normal, hypokinetic, akinetic, and dyskinetic, respectively. The method presented is valid for comparing cardiac parameters derived from disparate image data on a region-by-region basis by employing anatomic landmarks on 3-D reconstructions of the LV endocardial surface.     

Cloning, expression, and characterization of a DNA binding domain of gpNu1, a phage lambda DNA packaging protein

Terminase is an enzyme from bacteriophage lambda that is required for insertion of the viral genome into an empty pro-capsid. This enzyme is composed of the viral proteins gpNu1 (20.4 kDa) and gpA (73.3 kDa) in a holoenzyme complex. Current models for terminase assembly onto DNA suggest that gpNu1 binds to three repeating elements within a region of the lambda genome known as cosB which, in turn, stimulates the assembly of a gpA dimer at the cosN subsite. This prenicking complex is the first of several stable nucleoprotein intermediates required for DNA packaging. We have noted a hydrophobic region within the primary amino acid sequence of the terminase gpNu1 subunit and hypothesized that this region constitutes a protein-protein interaction domain required for cooperative assembly at cosB and that is also responsible for the observed aggregation behavior of the isolated protein. We therefore constructed a mutant of gpNu1 in which this hydrophobic "domain" has been deleted in order to test these hypotheses. The deletion mutant protein, gpNu1DeltaK, is fully soluble and, unlike full-length protein, shows no tendency toward aggregation; However, the protein is a dimer under all experimental conditions examined as determined by gel permeation and sedimentation equilibrium analysis. The truncated protein is folded with evidence of secondary and tertiary structural elements by circular dichroism and NMR spectroscopy. While physical and biological assays demonstrate that gpNu1DeltaK does not interact with the terminase gpA subunit, the deletion mutant binds with specificity to cos-containing DNA. We have thus constructed a deletion mutant of the phage lambda terminase gpNu1 subunit which constitutes a highly soluble DNA binding domain of the protein. We further propose that the hydrophobic amino acids found between Lys100 and Pro141 define a self-association domain that is required for the assembly of stable nucleoprotein packaging complexes and that the C-terminal tail of the protein defines a distinct gpA-binding site that is responsible for terminase holoenzyme formation.     

Characterization of the radical trapping activity of a novel series of cyclic nitrone spin traps

alpha-Phenyl-tert-butyl nitrone (PBN) is a nitrone spin trap, which has shown efficacy in animal models of oxidative stress, including stroke, aging, sepsis, and myocardial ischemia/reperfusion injury. We have prepared a series of novel cyclic variants of PBN and evaluated them for radical trapping activity in vitro. Specifically, their ability to inhibit iron-induced lipid peroxidation in liposomes was assessed, as well as superoxide anion (O2(-.)) and hydroxyl radical ((.)OH) trapping activity as determined biochemically and using electron spin resonance (ESR) spectroscopy. All cyclic nitrones tested were much more potent as inhibitors of lipid peroxidation than was PBN. The unsubstituted cyclic variant MDL 101,002 was approximately 8-fold more potent than PBN. An analysis of the analogs of MDL 101,002 revealed a direct correlation of activity with lipophilicity. However, lipophilicity does not solely account for the difference between MDL 101,002 and PBN, inasmuch as the calculated octanol/water partition coefficient for MDL 101,002 is 1.01 as compared to 1.23 for PBN. This indicated the cyclic nitrones are inherently more effective radical traps than PBN in a membrane system. The most active compound was a dichloro analog in the seven-membered ring series (MDL 104,342), which had an IC50 of 26 mum, which was 550-fold better than that of PBN. The cyclic nitrones were shown to trap (.)OH with MDL 101,002 being 20 25 times more active than PBN as assessed using 2-deoxyribose and p-nitrosodimethylaniline as substrates, respectively. Trapping of (.)OH by MDL 101,002 was also examined by using ESR spectroscopy. When Fenton's reagent was used, the (.)OH adduct of MDL 101,002 yielded a six-line spectrum with hyperfine coupling constants distinct from that of PBN. Importantly, the half-life of the adduct was nearly 5 min, while that of PBN is less than 1 min at physiologic pH. MDL 101,002 also trapped the O2(-.) radical to yield a six-line spectrum with coupling constants very distinct from that of the (.)OH adduct. In mice, the cyclic nitrones ameliorated the damaging effects of oxidative stress induced by ferrous iron injection into brain tissue. Similar protection was not afforded by the lipid peroxidation inhibitor U74006F, thus implicating radical trapping as a unique feature in the prevention of cell injury. Together, the in vivo activity, the stability of the nitroxide adducts, and the ability to distinguish between trapping of (.)OH and O2(-.) suggest the cyclic nitrones to be ideal reagents for the study of oxidative cell injury.     

The alpha attenuation test: assessing excessive daytime sleepiness in narcolepsy-cataplexy

Daytime sleep tendency was assessed in 10 drug-free patients with narcolepsy-cataplexy and 10 normals matched for age and gender. Following nocturnal polysomnography, the alpha attenuation test (AAT) and the multiple sleep latency test (MSLT) were administered during five sessions occurring at 2-hour intervals beginning at 0900 and 1000 hours, respectively. For the AAT, participants were polysomnographically recorded for 8 minutes while seated in an illuminated room with their eyes alternately opened and closed. Power spectral analyses of electroencephalograph (EEG) activity at 02-A1 (10 second epochs) were calculated using fast Fourier transformations (FFT) within the alpha frequency range (8-12 Hz) to obtain ratios of mean eyes-closed to mean eyes-open alpha power (i.e. the alpha attenuation coefficient, AAC). The narcoleptics were sleepier than the normals as indicated by a significantly smaller mean AAC and a significantly shorter mean latency to stage 1 on the MSLT. These findings suggest that the AAT may provide a quick and practical objective assessment of the excessive daytime sleepiness (EDS) associated with narcolepsy.     

Orange fruit is more effective than are dark-green, leafy vegetables in increasing serum concentrations of retinol and beta-carotene in schoolchildren in Indonesia

The objectives of this study were to quantify the effectiveness of dietary retinol sources, orange fruit, and dark-green, leafy vegetables in improving vitamin A status, and to test whether orange fruit is a better source of vitamin A and carotenoids than are leafy vegetables. Anemic schoolchildren aged 7-11 y (n = 238) in West Java, Indonesia, were randomly allocated to 1 of 4 groups to consume 2 complete meals/d, 6 d/wk, for 9 wk: 1) 556 retinol equivalents (RE)/d from retinol-rich food (n = 48); 2) 509 RE/d from fruit (n = 49); 3) 684 RE/d from dark-green, leafy vegetables and carrots (n = 45); and 4) 44 RE/d from low-retinol, low-carotene food (n = 46). Mean changes in serum retinol concentrations of the retinol-rich, fruit, vegetable, and low-retinol, low-carotene groups were 0.23 (95% CI: 0.18, 0.28), 0.12 (0.06, 0.18), 0.07 (0.03,0.11), and 0.00 (-0.06, 0.05) micromol/L, respectively. Mean changes in serum beta-carotene concentrations in the vegetable and fruit groups were 0.14 (0.12, 0.17) and 0.52 (0.43, 0.60) micromol/L, respectively. Until now, it has been assumed that 6 microg dietary beta-carotene is equivalent to 1 RE. On the basis of this study, however, the equivalent of 1 RE would be 12 microg beta-carotene (95% CI: 6 microg, 29 microg) for fruit and 26 microg beta-carotene (95% CI: 13 microg, 76 microg) for leafy vegetables and carrots. Thus, the apparent mean vitamin A activity of carotenoids in fruit and in leafy vegetables and carrots was 50% (95% CI: 21%, 100%) and 23% (95% CI: 8%, 46%) of that assumed, respectively. This has important implications for choosing strategies for controlling vitamin A deficiency. Research should be directed toward ways of improving bioavailability and bioconversion of dietary carotenoids, focusing on factors such as intestinal parasites, absorption inhibitors, and food matrixes.     

Osteopenia in children surviving brain tumours

Mature and post-translational precursor gastrin forms are growth factors for colorectal tumours. The immunogen Gastrimmune is composed of the amino terminus of gastrin-17 linked to diphtheria toxoid and raises antibodies in situ which neutralise amidated and glycine-extended gastrin-17. The aim of the study was to determine the effect of treatment with 5-fluorouracil(5-FU)/leucovorin on the antibody titres induced by Gastrimmune and the effect of combination therapy on the growth of the rat colon tumour DHDK12. Gastrimmune was administered to rats s.c. at 3 weekly intervals. The rat colon tumour line DHDK12 was injected into the abdominal wall of BDIX rats. Combinations of 5-FU/leucovorin were injected i.v. on days 1, 3 and 5, with the cycle repeated every 4 weeks. Antibody titres were measured by an ELISA technique. Antibody titres were followed for 40 weeks after Gastrimmune (500 microg.ml(-1)) immunization, with titres peaking between 10 and 20 weeks after a single immunisation and falling by week 30. At termination, no effect was observed on either the histological appearance of the gastro-intestinal tract or the proliferation of the colonic mucosa. Pre- and post-treatment with 5-FU/leucovorin (30 mg.kg(-1)) had no effect on the kinetics and level of antibody response to Gastrimmune. Gastrimmune (200 microg.ml(-1)) and 5-FU/leucovorin combinations (12.5 and 20 mg.kg(-1)) increased the therapeutic effects on the in vivo growth of DHDK12 tumors when compared to the agents given singly. Gastrimmune immunisation may be a therapeutic option for the treatment of colorectal cancer in combination with 5-FU/leucovorin.     

Long-term T cell memory requires the surface expression of self-peptide/major histocompatibility complex molecules

How memory T cells are maintained in vivo is poorly understood. To address this problem, a male-specific peptide (H-Y) was identified and used to activate female anti-H-Y T cells in vitro. Anti-H-Y T cells survived in vivo for at least 70 days in the absence of antigen. This persistence was not because of the intrinsic ability of memory T cells to survive in vivo. Instead, the survival and function of adoptively transferred memory cells was found to require transporter of antigen protein 1-dependent expression of self-peptide/major histocompatibility complex class I molecules in recipient animals. Therefore, it appears that the level of T cell receptor engagement provided by transporter of antigen protein 1-dependent, self-peptide/major histocompatibility complexes is sufficient to maintain the long-term survival and functional phenotype of memory cells in the absence of persistent antigen. These data suggest that positive selection plays a role not only in T cell development but also in the maintenance of T cell memory.     

Development of AMPA-selective glutamate receptors in the auditory brainstem of the barn owl

AMPA receptor specific antibodies were used to study the distribution and development of glutamate receptor subtypes (GluR1-4) in nucleus magnocellularis, angularis, laminaris, and the superior olive of the barn owl. Each nucleus in the adult barn owl expresses characteristic levels of AMPA receptor subtypes, and all are enriched in the subunits associated with rapid desensitization (GluR2 and 4). In the auditory hindbrain of the barn owl, the levels of expression of all AMPA receptors were very low at the time of hatching. In all nuclei, the level of GluR1 immunoreactivity was low to undetectable at all ages studied. In the cochlear nuclei, angularis and magnocellularis, levels of GluR2/3 and GluR4 immunoreactivity increased over the first 2 weeks after hatching, coinciding with the morphological maturation of auditory nerve terminals in NM. In the nucleus laminaris and in the superior olive, GluR2/3 and GluR4 immunoreactivity reached adult-like patterns by 3 weeks after hatching. Thus, adult-like patterns of immunoreactivity appeared at least 1 month before the end of the sensitive period in all nuclei studied.