Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.2.1.23 (beta-galactosidase)
 14,648 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A solid phase assay for the colorimetric detection of competitively amplified HCV-cDNA has been established and used to investigate clinical samples from patients with chronic hepatitis. The assay is based on the reduction in the amplification of an hepatitis C virus-related competitor molecule by wild-type hepatitis C virus during polymerase chain reaction. The internal standard contains a lac operator sequence, allowing the amount of amplified competitor to be determined using a lac I-repressor/beta-galactosidase fusion protein. The reduction in the amplification of competitor is dependent upon the concentration of HCV-RNA in the original sample. External hepatitis C virus wild-type standards are used to calibrate each concurrently tested set of patients. We present and discuss the potential benefit, but also the limitations of this new approach for quantifying hepatitis C virus viremia. In 47 serum samples from 28 patients with chronic hepatitis C virus infection, including five repeatedly tested alpha HCV positive patients under interferon therapy, viral titer was determined. Sera from nine healthy blood donors served as controls. The sensitivity and specificity of this procedure are identical to those of conventional nested polymerase chain reaction. As both internal and external standards are used in every assay and final detection of amplicons can be carried out in microtiter plates, this reliable and time-saving test system may be routinely applied for monitoring antiviral treatment or for studying the relation of plus- and minus-stranded HCV-RNA in infected tissues.
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PMID:Quantitation of HCV-replication using one-step competitive reverse transcription-polymerase chain reaction and a solid phase, colorimetric detection method. 781 15

Cationic lipid-DNA complexes (lipoplexes) have been widely used as gene transfer vectors which avoid the adverse immunogenicity and potential for viraemia of viral vectors. With the long-term aim of gene transfer into skeletal muscle in vivo, we describe a direct in vitro comparison of two commercially available cationic lipid formulations, Lipofectamine and DOSPER. Optimisation of transfection was performed in the C2C12 mouse muscle cell line, before further studies in primary mouse myoblasts and C2C12 myotubes. Reporter gene constructs expressing either E. coli beta-galactosidase or green fluorescent protein (GFP) were used in order to evaluate transfection efficiency by histochemical staining or FACS analysis, respectively. Both lipid formulations were able to promote efficient, reproducible gene transfer in C2C12 cells, and to transfect primary mouse myoblast cultures successfully. However, DOSPER exhibited the important advantage of being able to transfect cells in the presence of serum of both bovine and murine origin. This feature allowed increased cell survival during in vitro transfections, and may be advantageous for direct in vivo gene transfer efficacy.
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PMID:Lipofection of cultured mouse muscle cells: a direct comparison of Lipofectamine and DOSPER. 961 80

Hepatitis C virus (HCV) is a major human pathogen causing mild to severe liver disease worldwide. This positive strand RNA virus is remarkably efficient at establishing chronic infections. Although a high rate of genetic variability may facilitate viral escape and persistence in the face of Ag-specific immune responses, HCV may also encode proteins that facilitate evasion of immunological surveillance. To address the latter possibility, we examined the influence of specific HCV gene products on the host immune response to vaccinia virus in a murine model. Various vaccinia/HCV recombinants expressing different regions of the HCV polyprotein were used for i.p. inoculation of BALB/c mice. Surprisingly, a recombinant expressing the N-terminal half of the polyprotein (including the structural proteins, p7, NS2, and a portion of NS3; vHCV-S) led to a dose-dependent increase in mortality. Increased mortality was not observed for a recombinant expressing the majority of the nonstructural region or for a negative control virus expressing the beta-galactosidase protein. Examination of T cell responses in these mice revealed a marked suppression of vaccinia-specific CTL responses and a depressed production of IFN-gamma and IL-2. By using a series of vaccinia/HCV recombinants, we found that the HCV core protein was sufficient for immunosuppression, prolonged viremia, and increased mortality. These results suggest that the HCV core protein plays an important role in the establishment and maintenance of HCV infection by suppressing host immune responses, in particular the generation of virus-specific CTLs.
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PMID:Suppression of host immune response by the core protein of hepatitis C virus: possible implications for hepatitis C virus persistence. 991 17