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Gastroenterology. 1999 Mar;116(3):636-42.
Real-time detection system for quantification of hepatitis C virus genome.

Takeuchi T, Katsume A, Tanaka T, Abe A, Inoue K, Tsukiyama-Kohara K, Kawaguchi R, Tanaka S, Kohara M.

Department of Microbiology, The Tokyo Metropolitan Institute of Medical Science, Tokyo, Japan.

BACKGROUND & AIMS: For diagnosis of hepatitis C virus infection and monitoring of viral load in patients, a highly sensitive and accurate hepatitis C virus quantification system is essential. Methods: Hepatitis C virus genome was detected by real/time detection system using an ABI Prism 7700 sequence detector (Perkin Elmer Corp./Applied Biosystems, Foster City, CA). RESULTS: As few as 10 copies of the genome were detected, and the quantification range was between 10(1) and 10(8) copies (r > 0.99). This system was 10-100-fold more sensitive than an Amplicor monitor (Roche Diagnostic Systems, Branchburg, NJ). The coefficient of variation values for both intra-assay precision and interassay reproducibility of identifying the genome quantification ranged from 0.37% to 2.00% and 0.88% to 4.66%, respectively. The system could detect the genome in 98% of patients with chronic hepatitis, 95.8% of patients with liver cirrhosis, and 100% of patients with hepatocellular carcinoma who had the antibody to hepatitis C virus, but could not detect the genome in patients without the antibody. CONCLUSIONS: The establishment of a real-time detection system enables more accurate diagnosis of infection and monitoring of viral load in interferon-treated patients via quantification of viral genome.

online pharmacy ref. source: www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=10029622&dopt=Abstract

Hum Gene Ther. 1999 Jan 20;10(2):223-34.
Contribution of plasmid DNA to inflammation in the lung after administration of cationic lipid:pDNA complexes.

Yew NS, Wang KX, Przybylska M, Bagley RG, Stedman M, Marshall J, Scheule RK, Cheng SH.

Genzyme Corporation, Framingham, MA 01701-9322, USA.

Cationic lipid-mediated gene transfer to the mouse lung induces a dose-dependent inflammatory response that is characterized by an influx of leukocytes and elevated levels of the cytokines interleukin 6 (IL-6), tumor necrosis factor alpha (TNF-alpha), and interferon gamma (IFN-gamma). We have examined the contribution of plasmid DNA (pDNA) to this observed toxicity, specifically the role of unmethylated CpG dinucleotides, which have been previously shown to be immunostimulatory. We report here that complexes of cationic lipid GL-67 and unmethylated pDNA (pCF1-CAT) instilled into the lungs of BALB/c mice induced highly elevated levels of the cytokines TNF-alpha, IFN-gamma, IL-6, and IL-12 in the bronchoalveolar lavage fluids (BALF). In contrast, BALF of animals administered either GL-67 alone or GL-67 complexed with SssI-methylated pDNA contained low levels of these cytokines. Similar results were observed using a plasmid (pCF1-null) that does not express a transgene, demonstrating that expression of chloramphenicol acetyltransferase (CAT) was not responsible for the observed inflammation. The response observed was dose dependent, with animals receiving increasingly higher amounts of unmethylated pDNA exhibiting progressively higher levels of the cytokines. Concomitant with this increase in cytokine levels were also elevated numbers of neutrophils in the BALF, suggesting a possible cause- and-effect relationship between neutrophil influx and generation of cytokines. Consistent with this proposal is the observation that reduction of neutrophils in the lung by administration of antibodies against Mac-1alpha and LFA-1 also diminished cytokine levels. This reduction in cytokine levels in the BALF was accompanied by an increase in transgene expression. In an attempt to abate the inflammatory response, sequences in the pDNA encoding the motif RRCGYY, shown to be most immunostimulatory, were selectively mutagenized. However, instillation of a plasmid in which 14 of the 17 CpG sites were altered into BALF/c mice did not reduce the levels of cytokines in the BALF compared with the unmodified vector. This suggests that other unmethylated motifs, in addition to RRCGYY, may also contribute to the inflammatory response. Together, these findings indicate that unmethylated CpG residues in pDNA are a major contributor to the induction of specific proinflammatory cytokines associated with instillation of cationic lipid:pDNA complexes into the lung. Strategies to abate this response are warranted to improve the efficacy of this nonviral gene delivery vector system for the treatment of chronic diseases.

online pharmacy ref. source: www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=10022547&dopt=Abstract

Exp Hematol. 1999 Feb;27(2):203-9.
Interferon-gamma impairs physiologic downregulation of cyclin-dependent kinase inhibitor, p27Kip1, during G1 phase progression in macrophages.

Matsuoka M, Nishimoto I, Asano S.

Department of Pharmacology, Keio University School of Medicine, Tokyo, Japan.

Cell cycle progression of mouse macrophage cells was impaired by interferon-gamma (IFN-gamma). The blockage of G1/S transition was associated with diminution of cyclin-dependent kinase-2 (CDK2)-associated kinase activities. The expression of p21Cip1 was not upregulated by IFN-gamma. Instead, the physiologic downregulation of p27Kip1 necessary for normal cell cycle progression did not take place sufficiently in the presence of IFN-gamma. During normal cell cycle progression, the levels of p27Kip1 were maximal at early G1 and then decreased gradually. In the presence of IFN-gamma, however, the levels of p27Kip1 discontinued to decrease at a late mid G1 point and were consistently as high as, or higher than, levels observed there. The steady, relatively high-level attachment of p27Kip1 to CDK2 contributed to the insufficient formation of active cyclin/CDK2, possibly deferring cells from entering S phase.

online pharmacy ref. source: www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=10029157&dopt=Abstract

J Am Acad Dermatol. 1999 Feb;40(2 Pt 2):303-7.
Scleromyxedema: treatment with interferon alfa.

Tschen JA, Chang JR.

St. Joseph's Hospital, Baylor College of Medicine, Houston, Texas, USA.

Scleromyxedema is a variant of papular mucinosis characterized by fibroblast proliferation and mucin deposition in the dermis. Historically, it has been very difficult to treat and can cause significant morbidity and mortality with systemic involvement. We describe a case of a woman with scleromyxedema and systemic manifestations treated with interferon alfa. Her skin responded very well to therapy within 3 months; however, her systemic manifestations showed little change. We conclude that interferon alfa may be a useful therapy for patients with scleromyxedema, particularly if the disease process is limited to the skin.

online pharmacy ref. source: www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=10025854&dopt=Abstract

Science. 1999 Feb 19;283(5405):1183-6.
Reciprocal control of T helper cell and dendritic cell differentiation.

Rissoan MC, Soumelis V, Kadowaki N, Grouard G, Briere F, de Waal Malefyt R, Liu YJ.

Schering-Plough, Laboratory for Immunological Research, 27 chemin des Peupliers, Boite Postale 11, 69571, Dardilly, France.

It is not known whether subsets of dendritic cells provide different cytokine microenvironments that determine the differentiation of either type-1 T helper (TH1) or TH2 cells. Human monocyte (pDC1)-derived dendritic cells (DC1) were found to induce TH1 differentiation, whereas dendritic cells (DC2) derived from CD4+CD3-CD11c- plasmacytoid cells (pDC2) induced TH2 differentiation by use of a mechanism unaffected by interleukin-4 (IL-4) or IL-12. The TH2 cytokine IL-4 enhanced DC1 maturation and killed pDC2, an effect potentiated by IL-10 but blocked by CD40 ligand and interferon-gamma. Thus, a negative feedback loop from the mature T helper cells may selectively inhibit prolonged TH1 or TH2 responses by regulating survival of the appropriate dendritic cell subset.

online pharmacy ref. source: www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=10024247&dopt=Abstract

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