Long-term potentiation in the interpositus and vestibular nuclei in the rat

Summary Previous unpublished experiments from this laboratory had revealed only post-activation depression effects in the cerebellar cortex when its inputs were activated by high frequency trains. In the experiments reported in this paper, we found reliable long-term potentiation (LTP) effects in the deep nuclei (interpositus and vestibular) when stimulation trains were applied to the white matter at the point where inferior peduncle fibers enter the cerebellum. LTP effects were found in both acute and chronic preparations. In the chronic preparations, LTP lasted for at least 8 days in all but one animal.     

Complete nucleotide sequence of the M RNA segment of Rift Valley fever virus

The entire M RNA segment of the phlebovirus Rift Valley fever virus (RVFV) has been molecularly cloned and the complete nucleotide sequence determined. The RNA is 3884 nucleotides in length, corresponding to a molecular weight of 1.38 X 10(6), having a base composition of 27.3% A, 25.4% G, 27.2% U, and 20.1% C. Sequences present at the 3' and 5' termini of the molecule are largely complementary for some 51 residues and can form a stable duplex structure when the potential secondary structure of the entire molecule is considered. A single major open reading frame, capable of encoding 1206 amino acids (131,845 Da), was found in the viral-complementary sequence ("positive" polarity). Amino-terminal amino acid sequencing of the purified viral glycoproteins G1 and G2 allowed for the positioning of the coding sequences for these polypeptides within this major open reading frame in the following orientation with respect to the genomic M RNA: 3'-G2-G1-5'. From the predicted amino acid composition of the two mature viral glycoproteins, both were found to have a high cysteine content (G2, 6%; G1, 5%). Sequences within the open reading frame capable of encoding up to 23,000 Da of polypeptide were found in addition to those required for the viral glycoproteins. The potential contribution of these sequences to the coding capacity of the M RNA, viral protein processing, and intracellular protein distribution is discussed.     

Double nondisjunction during karyotypic progression of chemically induced Syrian hamster cell lines

The karyotypic evolution of three chemically induced cell lines of Syrian hamster embryo in culture are described. The only karyotypic alteration of one clone was a trisomy of chromosome #11, which presumably arose by nondisjunction after carcinogen treatment. A pure population of cells with the trisomy was observed repeatedly upon karyotyping of cells at the first three passages after cloning. However, at a late passage, apparently normal diploid cells appeared in the culture, which we propose resulted from a second nondisjunction of one chromosome #11, reverting the cells from trisomy 11 to disomy 11. The karyotypic evolution of two other cell lines also involved double nondisjunction, which resulted in duplication of a translocated chromosome and concurrent loss of the normal nonrearranged chromosome. Taken together with the reported findings of others, the results indicate that double nondisjunction is a mechanism in karyotypic progression during neoplastic development.     

Methylation of human T-cell leukemia virus proviral DNA and viral RNA expression in short- and long-term cultures of infected cells

Leukemic peripheral blood lymphocytes from individuals infected with the human T-cell leukemia/lymphoma virus (HTLV) were found to express little or no viral RNA before being put into tissue culture. Within 24-48 hr, viral RNA expression increased at least four- to eightfold. Established HTLV-infected cell lines constitutively express viral RNA. Southern blots of DNA from HTLV-infected cells digested with the methylation-sensitive restriction enzyme HpaII showed that the proviral DNA was methylated in all of the uncultured peripheral blood cells tested. In contrast, no proviral methylation was detected in any of the cell lines examined, suggesting a functional correlation between methylation and viral RNA expression. However, DNA from HTLV-infected lymphocytes cultured for 48 hr (by which time increases in viral RNA expression are evident) did not differ detectably with respect to proviral DNA methylation from uncultured cells, suggesting that the increase in viral RNA expression after short-term culture is mediated by mechanisms independent of changes in DNA methylation.     

Familial renal carcinoma

Renal carcinoma developed in two or more members of nine families. Multiple generations were affected in five kindreds, and siblings in four. The median age at cancer diagnosis was a decade earlier than usual, and individual patients had bilateral or multifocal lesions; these are features of hereditary forms of diverse cancers. No patient had von Hippel-Lindau disease or other predisposing genetic syndromes. Karyotypes of the peripheral blood of nine persons showed no instance of a 3;8 chromosome translocation as recently reported in association with renal carcinoma in ten members of one family. The findings show that familial renal cancer is more common than previously reported in the literature.     

Polyorna virus adsorbs to specific sialyloligosaccharide receptors on erythrocytes

Hemagglutination by polyoma virus has been shown to require sialic acid residues on cell surface oligosaccharides. This report presents evidence which suggests that adsorbtion of polyoma virus to erythrocytes is not due simply to a nonspecific electrostatic interaction with negatively charged sialic acids but rather requires the presence of specific sialyloligosaccharide structures. Newcastle disease virus (NDV) neuraminidase hydrolyzes sialic acids in the sequences NeuAcα2,3Gal and NeuAcα2,8NeuAc. Erythrocytes treated with NDV lost 40% of their surface sialic acid, retained full hemagglutination by certain influenza viruses which also bind sialyloligosaccharides, and yet were not agglutinated by polyoma virus. Erythrocytes treated with Vibrio cholerae neuraminidase, which removes virtually all sialic acid, were no longer agglutinated by influenza virus or polyoma virus. Selective replacement of sialic acid on V. cholerae neuraminidase-treated cells with purified β-galactoside α-2,3-sialytransferase in the sequence NeuAcα2, 3Gal completely restored hemagglutination by polyoma virus. In contrast, replacement of sialic acid by other sialyitransferases in the sequences NeuAcα2,6Gal or NeuAcα2,6GalNAc does not restore hemagglutination by polyoma virus.     

Identification and characterization of fast-sedimenting SV40 nucleoprotein complexes.

Simian virus 40 (SV40) nucleoprotein complexes were extracted from nuclei of infected monkey cells and fractionated on neutral sucrose density gradients. Four forms of SV40 chromatin were distinguished by their labeling kinetics and apparent sedimentation coefficients: Replicating viral DNA in 95 S SV40 chromatin was converted to 75 S chromatin when viral DNA replication was completed. A portion of the 75 S chromatin was rapidly assembled into 200 S nucleoprotein complexes and subsequently into 240 S complexes. The 75 S and the fast-sedimenting complexes were further characterized and compared with mature SV40 virus by equilibrium density centrifugation, plaque assay, and electron microscopy. These experiments suggest that the fast-sedimenting forms may represent virion precursors or a subclass of salt-labile virions, but they contain little or no mature virus.     

The COX-2 inhibitor, rofecoxib, ameliorates dextran sulphate sodium induced colitis in mice

Objective: We have evaluated the efficacy of the selective cyclo-oxygenase (COX)-2 inhibitor, rofecoxib, for the prevention of experimental colitis.Material and methods: To induce colitis BALB/c mice received 5% dextran sulphate sodium (DSS) in their drinking water continuously for 7 days. Rofecoxib (2.5–10 mg/kg body weight, p.o.) was administered throughout the treatment period with DSS. Colitis was quantified by a clinical damage score, colon length, weight loss, stool consistency and rectal bleeding. Inflammatory response was assessed by neutrophil infiltration, determined by histology and myeloperoxidase (MPO) activity. Interleukin (IL)-1, prostaglandin (PG)E₂ and PGD₂ levels in colon mucosa and the immunohistochemical expression of COX-1 and –2 were also studied.Results: The COX-2 inhibitor ameliorated severe colitis, reduced the degree of inflammation through reduction of neutrophil infiltration and IL-1 levels. PGE₂, and PGD₂ synthesis were significantly reduced in DSS-treated groups. Indeed, treatment with rofecoxib diminished the lost of COX-1 caused by DSS in the crypt epithelium whereas expression of COX-2 remained unaffected.Conclusions: Rofecoxib is protective in acute DSS – induced colitis, probably by reducing neutrophil infiltration, inhibiting up-regulation of IL-1 and returning to normal COX-1 expression in the inflamed colonic mucosa.Received 19 April 2004; returned for revision 17 June 2004; accepted by I. Ahnfelt-Rønne 23 November 2004     

NADPH oxidase priming and p47phox phosphorylation in neutrophils from synovial fluid of patients with rheumatoid arthritis and spondylarthropathy

Superoxide anion (O2°-)production by neutrophil NADPH oxidase participates in arthritic joint lesion formation. Proinflammatory cytokines such as tumor necrosis factor (TNF), interleukin 8 (IL-8) and granulocyte/macrophage-colony stimulating factor (GM-CSF) have a priming effect on neutrophil NADPH oxidase activity. NADPH oxidase activation is dependent on phosphorylation of p47phox, a cytosolic component of the enzyme. We studied O2°-production and p47phox phosphorylation in synovial fluid (SF) from patients with rheumatoid arthritis (RA) and spondylarthropathy (SpA) according to TNF, IL-8 and GM-CSF levels. O2°-production by neutrophils isolated from SF of all the arthritis patients (RA and SpA) was higher than that of circulating resting neutrophils and when stimulated with fMLP or PMA. In addition, p47phox was partially phosphorylated in SF neutrophils compared to circulating neutrophils. High levels of TNF and IL-8 (but not GM-CSF) are detected in patient's SF (compared to circulating blood levels). TNF levels were significantly higher in RA than in SpA SF. These results suggest that increased NADPH oxidase activity could be involved in arthritic joint inflammation through increased p47phox phosphorylation. This could be the result of the presence of high levels of priming agents such as TNF and IL-8 but not GM-CSF.