Exploiting hepatitis C virus activation of NFkappaB to deliver HCV-responsive expression of interferons alpha and gamma.

Chronic infection with hepatitis C virus (HCV) may lead to liver failure and hepatocellular carcinoma. Current treatment for HCV includes high systemic doses of interferonalpha (IFNalpha), which is effective in less than half of patients and may have severe side effects. We designed conditional IFNalpha and IFNgamma expression constructs to be triggered by HCV-induced activation of NFkappaB, and delivered these using highly efficient recombinant Tag-deleted SV40-derived vectors. NFkappaB activates the HIV-1NL4-3 long terminal repeat (HIVLTR) as a promoter, which accounts for the conditional transgene expression. Human hepatocyte lines and primary rat hepatocytes (PRH) were transduced with SV[HIVLTR](IFN) vectors, and transfected with HCV cDNA. Production of human and murine IFNalpha and IFNgamma in cytosol and culture supernatants was measured. HCV activated the HIVLTR to produce and secrete IFNs, and did so largely through the NFkappaB binding sites of the HIVLTR. Levels of IFNs secreted, and the magnitude of induction in response to HCV, were greater in hepatocyte lines than in primary cultured hepatocytes. However, even in the latter, supernatant IFNalpha concentrations achieved by this approach were similar to therapeutic serum concentrations sought in systemic IFNalpha-treated patients. In coculture studies, secreted IFNalpha activated its cognate response elements in untransduced cells, suggesting that its potential inhibitory effects on HCV may not be limited to transduced cells. Although HCV replication in culture is difficult to assess, HCV-induced IFNalpha production demonstrably reduced HCV transcription. Conditional expression of IFNs within the liver may represent an attractive approach to therapy of severe chronic HCV infection that could avoid the side effects of systemic treatment regimens.

[The tissue specificity of the protective action of cytoplasmic factors on the membrane-bound system of Ca2+ transport in the sarcoplasmic reticulum of the heart and the skeletal muscles]. / Tkanespetsifichnost' protektornogo deistviia tsitoplazmaticheskikh faktorov na membranno-sviazannuiu sistemy transporta Ca2+ v sarkoplazmaticheskom retikulume serdtsa i skeletnykh myshts.

Whether soluble cytoplasmic factors (SCF) can protect the membrane-associated Ca2+ transport system in sarcoplasmic reticulum (SR) was studied in controls, stress and adaptation to it. Ca-transport was damaged in stress much more in the skeletal muscles compared to the heart. Initial performance of Ca-pump in the skeletal muscles was decreased by 43%, in the heart--no change. After stress, thermoresistance of the enzyme in the skeletal muscles was 3-5 times less than in the myocardium. The enzyme reaction on adaptation and stress in both tissues is the same--a significant rise in Ca-pump thermoresistance. When preadapted animals are exposed to stress, deceleration of the initial speed and resistance of the enzyme in the myocardium are fully prevented while in the skeletal muscles Ca-pump resistance is higher than in the controls. In the heart, SCF have a protective effect on thermoresistance. In the skeletal muscles, SCF protect initial speed of the enzyme in the control, in stress and adaptation to it. A cross-over protective SCF (from adapted animals) effect was found on the stress-impaired membrane system of Ca-transport. Adapted SCF elevate 2.5-fold thermoresistance of the enzyme. Correlation between cytoplasmic and membrane protective mechanisms in stress and adaptation to it is considered.