Isolation of porcine circovirus-like viruses from pigs with a wasting disease in the USA and Europe

Sinus pericranii is an anomalous extracranial vascular malformation that is in continuity with the intracranial dural venous sinuses. Five case reports, three congenital and two traumatic, are described. Clinical management, including evaluation, diagnosis, and treatment, is discussed. Awareness of this entity by plastic surgeons will allow for earlier diagnosis and appropriate surgical management, resulting in decreased risk of complications.     

Cyclic stretch down-regulates calcium transporter gene expression in neonatal rat ventricular myocytes

Abnormal intracellular Ca2+ handling in hypertrophied and failing hearts is partly due to changes in Ca2+ transporter gene expression, but the mechanisms responsible for these alterations remain largely unknown. We previously showed that intrinsic mechanical load (i.e. spontaneous contractile activity) induced myocyte hypertrophy, and down-regulated SR Ca2+ ATPase (SERCA2) gene expression in cultured neonatal rat ventricular myocytes (NRVM). In the present study, we examined whether extrinsic mechanical load (i.e. cyclic stretch) also induced NRVM hypertrophy, and led to down-regulation of SERCA2 and other Ca2+ transporter genes which have been associated with cardiac hypertrophy and failure in vivo. NRVM were maintained in serum-free culture medium under control conditions, or subjected to cyclic mechanical deformation (1.0 Hz, 20% maximal strain, 48 h). Under these conditions, cyclic stretch induced NRVM hypertrophy, as evidenced by significant increases in total protein/DNA ratio, myosin heavy chain (MHC) content, and atrial natriuretic factor (ANF) secretion. Cyclic stretch also induced the MHC isoenzyme "switch" which is characteristic of hemodynamic overload of the rat heart in vivo. Cyclic stretch significantly down-regulated SERCA2 and ryanodine receptor (RyR) mRNA and protein levels, while simultaneously increasing ANF mRNA. In contrast, Na+-Ca2+ exchanger and phospholamban mRNA levels were unaffected. Load-dependent SERCA2 and RyR down-regulation was independent of Ca2+ influx via voltage-gated, L-type Ca2+ channels, as cyclic stretch down-regulated SERCA2 and RyR mRNA levels in both control and verapamil-treated NRVM. These results indicate that extrinsic mechanical load (in the absence of other exogenous stimuli) induces NRVM hypertrophy and causes down-regulation of Ca2+ transporter gene expression. This in vitro model system should prove useful to dissect the intracellular signaling pathways responsible for transducing this phenotype during cardiac hypertrophy and heart failure in vivo.     

Crystal structure of the lectin from Dioclea grandiflora complexed with core trimannoside of asparagine-linked carbohydrates

The seed lectin from Dioclea grandiflora (DGL) has recently been shown to possess high affinity for 3, 6-di-O-(alpha-D-mannopyranosyl)-alpha-D-mannopyranose, the core trimannoside of asparagine-linked carbohydrates, but lower affinity for biantennary complex carbohydrates. In the previous paper, the thermodynamics of DGL binding to deoxy analogs of the core trimannoside and to a biantennary complex carbohydrate were determined by isothermal titration microcalorimetry. The data suggest that DGL recognizes specific hydroxyl groups of the trimannoside similar to that of the jack bean lectin concanavalin A (ConA) (Gupta, D. Dam, T. K., Oscarson, S., and Brewer, C. F. (1997) J. Biol. Chem. 272, 6388-6392). However, the thermodynamics of DGL binding to certain deoxy analogs and to the complex carbohydrate are different from that of ConA. In the present paper, the x-ray crystal structure of DGL complexed to the core trimannoside was determined to a resolution of 2.6 A. The overall structure of the DGL complex is similar to the structure of the ConA-trimannoside complex (Naismith, J. H., and Field, R. A. (1996) J. Biol. Chem. 271, 972-976). The location and conformation of the bound trimannoside as well as its hydrogen-bonding interactions in both complexes are nearly identical. However, differences exist in the location of two loops outside of the respective binding sites containing residues 114-125 and 222-227. The latter residues affect the location of a network of hydrogen-bonded water molecules that interact with the trisaccharide. Differences in the arrangement of ordered water molecules in the binding site and/or protein conformational differences outside of the binding site may account for the differences in the thermodynamics of binding of the two lectins to deoxy analogs of the trimannoside. Molecular modeling studies suggest how DGL discriminates against binding the biantennary complex carbohydrate relative to ConA.     

Z/I and A-band lattice spacings in frog skeletal muscle: effects of contraction and osmolarity

A-band and Z-line/I-band lattice spacings were measured by small-angle X-ray diffraction from relaxed and isometrically-contracting whole frog sartorius muscles with lattice spacings reduced or swollen by changing the osmolarity of the bathing solution. A-band spacing increased by approximately 3% upon isometric contraction at reduced lattice spacings (245-356 mOsm) and decreased by approximately 1% at swollen spacings (172 mOsm), similarly to the behaviour of skinned muscles upon changing from the relaxed state to rigor. The Z/I lattice underwent a significant lattice expansion (3-8%) upon isometric contraction at all osmolarities, in qualitative agreement (but quantitative disagreement) with results from electron microscopy on mammalian skeletal muscle. Lattice areas calculated for the Z/I and A-band lattices indicate a barrel-shaped sarcomere in the resting state, which may provide a partial explanation for how longitudinal forces produced in the A-band can produce a radial expansive force in the Z-line during contraction. The radial component of cross-bridge stiffness was calculated from the A-band data for contracting muscle, using a lattice stability model incorporating structural, osmotic and electrostatic forces. The calculations gave a radial cross-bridge stiffness during contraction of about 9 x 10(5) N m-2, and outward radial force per thick filament in normal Ringer's solution of 6 x 10(-9) N, corresponding to a radial force per cross-bridge of 10(-11) N.     

Augmentation of facial soft-tissue defects with Alloderm dermal graft

The aim of the study was TPS diagnostic value determination in the serum of women with breast cancer as compared with MCA and CA 15-3. The relationship between the serum concentration of these antigens and patient age, clinical stage, histological grade, presence of metastases to lymph nodes and histological type of neoplasm was evaluated. Studies were conducted on the sera of 139 women before surgical procedure aged 28-81, treated in the Clinic of Oncology, at Karol Marcinkowski University of Medical Sciences in Poznań. The TPS concentration was determined using the "BEKI Diagnostics" immunoenzymatic method, MCA - by the "Roche" test and CA 15-3 concentration was determined by the "Abbott" immunofluorescent test. The study showed significantly higher levels of TPS, CA 15-3 and MCA in women with cancer, compared with values in healthy women and women with mastopathy. The highest median of concentration and frequency of occurrence was obtained for TPS. A correlation between enhancement of TPS and CA 15-3 concentration with clinical stage was observed. A similar connection was noted in women with metastases to the lymph nodes. Serum MCA concentration results did not demonstrate the above effects. The study suggests, that in estimating the clinical condition of women with breast cancer, the simultaneous determination of TPS with CA 15-3 seems to be a more useful prognostic factor than TPS or CA 15-3 with MCA.     

Ribonuclease A variants with potent cytotoxic activity

Select members of the bovine pancreatic ribonuclease A (RNase A) superfamily are potent cytotoxins. These cytotoxic ribonucleases enter the cytosol, where they degrade cellular RNA and cause cell death. Ribonuclease inhibitor (RI), a cytosolic protein, binds to members of the RNase A superfamily with inhibition constants that span 10 orders of magnitude. Here, we show that the affinity of a ribonuclease for RI plays an integral role in defining the potency of a cytotoxic ribonuclease. RNase A is not cytotoxic and binds RI with high affinity. Onconase, a cytotoxic RNase A homolog, binds RI with low affinity. To disrupt the RI-RNase A interaction, three RNase A residues (Asp-38, Gly-88, and Ala-109) that form multiple contacts with RI were replaced with arginine. Replacing Asp-38 and Ala-109 with an arginine residue has no effect on the RI-RNase interaction. In addition, these variants are not cytotoxic. In contrast, replacing Gly-88 with an arginine residue yields a ribonuclease (G88R RNase A) that retains catalytic activity in the presence of RI and is cytotoxic to a transformed cell line. Replacing Gly-88 with aspartate also yields a ribonuclease (G88D RNase A) with a decreased affinity for RI and cytotoxic activity. The cytotoxic potency of onconase, G88R RNase A, and G88D RNase A correlate with RI evasion. We conclude that ribonucleases that retain catalytic activity in the presence of RI are cytotoxins. This finding portends the development of a class of chemotherapeutic agents based on pancreatic ribonucleases.     

Effect of feed withdrawal on Campylobacter in the crops of market-age broiler chickens

The presence of Campylobacter and Salmonella on poultry meat products remains a significant public health concern. Previous research has indicated that feed withdrawal may significantly increase Salmonella contamination of broiler crops and that crop contents may serve as an important source of Salmonella carcass contamination at commercial processing. The present study evaluated the effect of preslaughter feed withdrawal on the incidence of Campylobacter isolation in crops of market-age commercial broiler chickens prior to capture and transport to the processing plant. The incidence of Campylobacter isolation from the crop was determined immediately before and after feed withdrawal in 40 7-wk-old broiler chickens obtained from each of nine separate broiler houses. Ceca were collected from broilers in six of the same flocks for comparison with the crop samples. Feed withdrawal caused a significant (P < 0.025) increase in Campylobacter-positive crop samples in seven of the nine houses sampled. Furthermore, the total number of Campylobacter-positive crops increased significantly (P < 0.001) from 90/360 (25%) before feed removal to 224/359 (62.4%) after the feed withdrawal period. Alternatively, feed withdrawal did not significantly alter the Campylobacter isolation frequency from ceca. Similar to our previous studies with Salmonella, the present results suggest that preharvest feed withdrawal increases the frequency of Campylobacter crop contamination and, thus, provides a source of Campylobacter contamination of carcasses at commercial processing.     

Epitope specificity of CD44 for monoclonal antibody-dependent facilitation of marrow engraftment in a canine model

Primary graft rejection after marrow transplantation occurs more frequently in patients receiving HLA-haploidentical compared with HLA-identical sibling transplants. Both human and experimental animal data suggest that the cells responsible for this phenomenon are either host natural killer (NK) cells, T cells, or both. To investigate the mechanisms of graft rejection, we have developed a canine model of marrow transplantation, which uses DLA-nonidentical unrelated donors in the absence of postgrafting immunosuppression. In this model most animals rejected their marrow grafts after a preparative regimen of 9.2 Gy total body irradiation (TBI). However, engraftment of DLA-nonidentical marrow can be facilitated when the recipients are pretreated with monoclonal antibody (MoAb) S5, which recognizes CD44. In this report, we extended these observations by first cloning the canine CD44 and, next, mapping the epitope recognized by S5, which was located in a region conserved among human and canine CD44 and was distinct from the hyaluronan binding domain. However, in vitro binding of S5 caused a conformational change in CD44, which allowed increased hyaluronan binding. Then, we reexamined the in vivo model of marrow transplantation and compared results with MoAb S5 to those with two other anti-CD44 MoAbs, IM7 and S3. Only MoAb S5 significantly increased the engraftment rate of DLA-nonidentical unrelated marrow, whereas the two other anti-CD44 MoAbs were ineffective. The enhanced in vivo effect was not related to differences in the MoAbs' avidities, since both S5 and IM7 had equivalent binding to CD44, but most likely related to the specific epitope that S5 recognizes. Thus, this study shows that the effect of the anti-CD44 MoAb S5 in facilitating engraftment is epitope specific and if one is to use an anti-CD44 to facilitate engraftment of marrow in humans, one cannot assume that any anti-CD44 would work.