Metastasis-associated alterations in phospholipids and fatty acids of human prostatic adenocarcinoma cell lines.

Metastatic variants of human prostatic adenocarcinoma cell lines (DU-145, LNCaP, and ND-1) were studied by using soft agar colony forming efficiency, nude mice tumorigenicity, in vitro invasion assay, and type IV collagenase assay. The DU-145 and ND-1 cell line showed higher metastatic potential than LNCaP. Lipids from DU-145, ND-1, and LNCaP cells were extracted and analyzed by thin-layer chromatography and gas-liquid chromatography. The major lipids were phosphatidylcholine, phosphatidylethanolamine, sphingomyelin, fatty acids, and cholesterol. The sphingomyelin level was significantly higher in highly metastatic cells (DU-145 and ND-1) compared with the lower metastatic variant (LNCaP). The increase in the synthetic pathway and decrease in degradation pathway of sphingomyelin in microsomal fractions was sufficient to account for the measured increase in sphingomyelin in DU-145 cells compared with LNCaP cells. The major fatty acids of these lipids were palmitic (16:0), stearic (18:0), oelic (18:1), and arachidonic acid (20:4). The arachidonic acid level was significantly decreased in DU-145 and ND-1 compared with LNCaP cells. Electron microscopic studies showed no significant changes in the morphology of DU-145, ND-1, and LNCaP cells. The results of these investigations demonstrate for the first time that sphingomyelin and arachidonic acid contents are different in high and low metastatic variants of human prostatic adenocarcinoma cell lines.     

A single-amino-acid substitution in polyomavirus VP1 correlates with plaque size and hemagglutination behavior.

The plaque size and hemagglutination characteristics of five cloned wild-type strains of polyomavirus were determined. The strains fell into two groups, those with large or small plaques, each with distinctive hemagglutination behavior at different temperatures and pHs. The nucleotide sequence of VP1, the major capsid protein of the virus, was determined for each of the viral strains. The PTA (large-plaque) and RA (small-plaque) strains differed only at residue 92 of VP1, where there is a glutamic acid or glycine, respectively (R. Freund, A. Calderone, C. J. Dawe, and T. L. Benjamin, J. Virol. 65:335-341, 1991). The same amino acid difference in VP1 correlated with plaque size and hemagglutination properties of the other sequenced viruses. Mutagenesis converting amino acid 92 from glutamic acid to glycine converted the plaque size and hemagglutination behavior of the large-plaque PTA strain to that of a small-plaque strain. Furthermore, PTA and RA VP1 proteins produced in Escherichia coli behaved as their parental viruses did in hemagglutination assays. These results demonstrate that amino acid residue 92 of VP1 is involved in determining the plaque size and hemagglutination behavior of polyomavirus and strongly suggest that this region of the VP1 polypeptide interacts directly with cell receptors.     

Integumental lesions caused by ectoparasites in a wild population of the side-blotched lizard (Uta stansburiana)

Histopathological effects of ectoparasites on integument were examined for a wild population of the side-blotched lizard Uta stansburiana. These included the trombiculid Neotrombicula californica, the pterygosomatid mite Geckobiella texana; the macronyssid mite Ophionyssus natricis (Macronyssidae) and the ixodid tick Ixodes pacificus. A diffuse inflammatory response occurred at the site of chigger and tick attachment which consisted of histiocyte, heterophil, fibroblast and lymphocyte infiltration that often extended into the dermis. Granuloma formation also was noted. The most prevalent parasite was N. californica which frequently occurred in large aggregations above the eyelids. Ectoparasites were most abundant from February through April.     

Thermodynamics of the disproportionation of adenosine 5'-diphosphate to adenosine 5'-triphosphate and adenosine 5'-monophosphate. I. Equilibrium model

The thermodynamic treatment of the disproportionation reaction of adenosine 5′-diphosphate to adenosine 5′-triphosphate and adenosine 5′-monophosphate is discussed in terms of an equilibrium model which includes the effects of the multiplicity of ionic and metal bound species and the presence of long range electrostatic and short range repulsive interactions. Calculated quantities include equilibrium constants, enthalpies, heat capacities, entropies, and the stoichiometry of the overall reaction. The matter of how these calculations can be made self-consistent with respect to both calculated values of the ionic strength and the molality of the free magnesium ion is discussed. The thermodynamic data involving proton and magnesium-ion binding data for the nucleotides involved in this reaction have been evaluated.     

Co-distribution of annexin VI and actin in secretory ameloblasts and odontoblasts of rat incisor

Summary Annexin VI and actin were detected by immunoblot analysis in the enamel- and dentin-related portions of dental tissues. Annexin VI was found mainly in the particulate fraction whereas actin was detected in both the soluble and particulate fractions. By immunoelectron microscopy, annexin VI antibodies conjugated with colloidal gold were seen to label the mitochondria, the cytosol and the nucleus of secretory ameloblasts and odontoblasts of rat incisor. In the processes of these cell, the plasmalemmal undercoat was labeled. Antiactin antibodies labeled the desmosome-like junctions, the cytosol, and the mitochondria of the cell bodies. Extensive labeling was seen at the periphery of the Tomes' processes and odontoblast processes. These results suggest that annexin VI may play a role in Ca²⁺-regulation in the cell bodies, especially as a calcium receptor protein in the mitochondria. Moreover, annexin VI and actin seem to be co-distributed in secretory processes. Thus, these proteins might be both involved in exocytotic and endocytotic events.     

A high melting structure in DNA distinguishes phases of the cell cycle

Differential scanning microcalorimetry of the nuclei of dividing CHO cells revealed DNA structures that showed structural transitions at 60, 76, 88, and 105 degrees C (transitions I to IV, respectively). In cultures synchronized by isoleucine deprivation the enthalpies of transitions I and II were rather constant throughout the cell cycle. While the sum of the enthalpies of III and IV was nearly constant, the ratio of IV to III varied substantially from one phase of the cycle to another. A high IV:III ratio of 6 characterized G1 while S phase gave a IV:III ratio of about 2. Cells containing metaphase chromosomes also showed a IV:III ratio near 2. The IV:III ratio for CHO cells showed a progressive decrease as the cells were maintained in isoleucine-free medium from 0 to 6 days.     

Mineral-binding proteoglycans of fetal porcine calvarial bone

To provide a more definitive characterization of the hydroxylapatite-associated proteoglycans (HAPG) of bone, proteins were extracted from the mineralized matrix of fetal porcine calvaria with 0.5 M EDTA in the absence of guanidine HCl. The small proteoglycans obtained in the extract were fractionated by gel filtration on Sepharose CL-6B, purified by ion-exchange chromatography on Polyanion matrix (fast protein liquid chromatography), and then separated into three major populations of chondroitin sulfate proteoglycans by chromatography on hydroxylapatite, all in the presence of 7 M urea. Based on immunological and chemical properties, two classes of bone proteoglycan were resolved. In one class (HAPG1), the proteoglycan and specific CNBr-derived peptides cross-reacted with three monoclonal antibodies that recognize different epitopes of the protein core of bovine skin proteodermatan sulfate. The other class of proteoglycan included two species (HAPG2, HAPG3) which were not recognized by these antibodies. In addition, these proteoglycans did not stain with Coomassie Blue R-250 nor with silver stain nor did they bind to nitrocellulose membranes used in Western blots. However, the cationic dye Stains-all stained both HAPG2 and HAPG3; the protein cores of these proteoglycans were stained a characteristic turquoise blue, whereas the protein core of HAPG1 was stained pink. The average Mr values of the bone proteoglycans, from gradient sodium dodecyl sulfate-polyacrylamide gel electrophoresis were: HAPG1, 120,000, with a protein core (chondroitinase AC-digested) of 45,000; HAPG2 and HAPG3, 110,000, with protein cores of 37,000-38,000. On 15% polyacrylamide gel electrophoresis, the protein cores of HAPG2 and HAPG3 migrated with an Mr 30,000, while HAPG1 protein core was unchanged (Mr 45,000). Based on amino acid analysis, the protein chains of HAPG2 and HAPG3 appear to be identical, although minor differences in the relative amount of glucosamine were evident. In contrast, the composition of HAPG1 was quite different, with higher relative amounts of hydrophobic and aromatic residues and lower amounts of Asx and Glx. The presence of 360 residues/1,000 of Asx and Glx in HAPG2 and HAPG3 may in part explain the characteristic staining and immunotransfer properties of these proteoglycans. The unique amino-terminal sequence of HAPG2 (Asn-Pro-Val-Ala-Arg-Tyr-Gln), together with the immunological and chemical properties, would indicate that HAPG2 and HAPG3 are novel proteoglycans and, unlike HAPG1, could be unique to mineralized tissues.