ABSTRACT
The mitogen activated protein kinases-extracellular signal regulated kinases (MAPK-ERK) pathway is involved in regulation of multiple cellular processes including the cell cycle.In the present study using a Huh7 cell line Con1 with an HCV replicon,we have shown that the MAPK-ERK pathway plays a significant role in the modulation of HCV replication and protein expression and might influence IFN-α signalling.Epithelial growth factor (EGF) was able to stimulate ERK activation and decreased HCV RNA load while a MAPK-ERK pathway inhibitor U0126 led to an elevated HCV RNA load and higher NS5A protein amounts in Con1 cells.It could be further demonstrated that the inhibition of the MAPK-ERK pathway facilitated the translation directed by the HCV internal ribosome entry site.Consistently,a U0126 treatment enhanced activity of the HCV reporter replicon in transient transfection assays.Thus,the MAPK-ERK pathway plays an important role in the regulation of HCV gene expression and replication.In addition,cyclin-dependent kinases (CDKs) downstream of ERK may also be involved in the modulation of HCV replication since roscovitine,an inhibitor of CDKs had a similar effect to that of U0126.Modulation of the cell cycle progression by cell cycle inhibitor or RNAi resulted consistently in changes of HCV RNA levels.Further,the replication of HCV replicon in Conl cells was inhibited by IFN-α.The inhibitory effect of IFN-α could be partly reversed by pre-incubation of Con-1 cells with inhibitors of the MAPK-ERK pathway and CDKs.It could be shown that the MAPK-ERK inhibitors are able to partially modulate the expression of interferon-stimulated genes.
ABSTRACT
Adenoviruses are recognized as common human pathogens that are responsible for a wide variety of infectious syndromes. Bone marrow transplant patients are prone to life threatening opportunistic infections like adenoviruses. The nested polymerase chain reaction has provided an alternative, sensitive diagnostic method for detection of Adenoviruses. In this study we developed PCR from hexon genes as rapid diagnostic method of Advs infections on different clinical samples. Adenovirus infections was defined as the presence of DNA in the blood, urine, stool, or respiratory lavage from bone marrow transplant patients. Two sets of primers [Group specific primers and internal primers] were required to optimize the PCR protocol. This highly sensitive method could detect different types of Advs in two separate sets of PCR. Therefore, DNA amplification in BMT patients would be valuable screening way to evaluate bone marrow transplant recipients. Early detection of Advs by PCR assay is important to asymptomatic infections or preventing aggressive antiviral therapy
ABSTRACT
The woodchuck model is an excellent animal model to study hepadnaviral infection. The new progresses in this model made possible to examine the T-cell mediated immune responses in acute and chronic hepadnaviral infection. Recently, a new assay for cytotoxic T-cells based on detection of CD107 was established for the woodchuck model. In addition, new immunotherapeutic approaches based on combination of potent antiviral treatment and DNA-protein vaccines were proven to be useful for treatment of chronic hepatitis B.
ABSTRACT
<p><b>OBJECTIVE</b>To investigate the expression profile of genes which are involved in IFN antiviral activity and IFN signal transduction pathway in Hep G2 and HepG2.2.15 cells.</p><p><b>METHODS</b>Genes of interest were selected from the UniGene database (http://www.ncbi.nlm.gov/UniGene/Hs.Home.html). The 5'IMAGE clones with 0.5-0.8 kb length were chosen and ordered from RZPD company. The cDNA inserts were amplified by PCR and then were spotted onto the Hybond-N+ membranes. The membranes were denatured and neutralized for Macroarray analysis. HepG2.2.15 and Hep G2 cells were treated without or with IFN-alpha for 6 h, and the total cellular RNA was isolated using Trizol Reagent. Radio-labelled cDNA was generated from 20 microgram of RNA by reverse transcription using 360 units of reverse transcriptase in the presence of 30 microCi of alpha-32P dCTP. Hybridization was performed between 32P-labelled cDNA and membrane arrays. The membranes were then scanned, and the intensity of autoradiographic spots was quantitated by Cyclone Storage Phosphor System. The images were subsequently analysed by the OptiQuant Imager Analysis Software and converted into digital data.</p><p><b>RESULTS</b>The authors found that just partially IFN-inducible genes were expressed in Hep G2 and HepG2.2.15 cells, and the majority of IFN-inducible genes was lowly responsive or non-responsive to IFN-a treatment. Some interferon-stimulated genes (ISGs) were inhibited or blocked, especially in HepG2.2.15 cells. Interestingly, the authors found that the IFN signal transduction pathway (Jak-STAT) was intact and unimpaired in HepG2.2.15 cells.</p><p><b>CONCLUSION</b>Differential gene expression profiles in response to IFN were found between Hep G2 and HepG2.2.15 cells.</p>