OBJECTIVETo explore the action mechanism of ginsenoside Rb1 (GRb1) on protecting herpes simplex virus-1 (HSV-1) infected nerves by studying its inhibitory effects on abnormal changes of apoptosis and nerve growth factor (NGF) mRNA expression in HSV-1 infected human glioma cells U251.

METHODSThe inhibitory effects of GRb1 on HSV-1 induced abnormal apoptosis of U251 cells were detected using MTT colorimetry and flow cytometry. The NGF mRNA expressions in different treatment groups were detected using semiquantitative RT-PCR.

RESULTS(1) In 400 microg/mL GRb1 + HSV-1 group, MTT value was higher than HSV-1 group at 24, 36, and 48 h after infection (P < 0.05). (2) Cytopathic effects (CPE) were observed in HSV-1 group at 36 h after infection. In 400 microg/mL GRb1 + HSV-1 group merges increased at 36 h after infection, but most cells were in normal shapes. (3) Results of flow cytometry showed that the cell apoptosis rate was lower in 400 microg/mL GRb1 + HSV-1 group than in the HSV-1 group at24 and 36 h after infection (P < 0.05). (4) Results of RT-PCR showed that in 400 microg/mL GRb1 + HSV-1 group, NGF mRNA expressions decreased at 6-12 h after infection (P < 0.05), but it increased at 24, 36, and 48 h after infection, and was obviously higher than that in the HSV-1 group (P < 0.05).

CONCLUSIONSGRb1 at an appropriate concentration could inhibit abnormal cell apoptosis and changes of NGF mRNA expressions in HSV-1 infection. Therefore, we inferred that GRb1 could protect nerves possibly through up-regulating NGF mRNA expressions and inhibiting apoptosis.

Apoptosis ; drug effects ; Cell Line, Tumor ; Ginsenosides ; pharmacology ; Herpes Simplex ; metabolism ; Herpesvirus 1, Human ; Humans ; Nerve Growth Factor ; metabolism

PURPOSE: To investigate the replication of HSV within cultured cell and axonal transport of HSV within the axon of the ciliary nerve following the injection of HSV into a cultured ciliary nerve. METHODS: The explant of the ciliary nerve was cultured with a medium containing nerve growth factor for 30 days when the suspension of HSV-1 (Kos strain) was introduced into the culture dish to co-culture with the ciliary nerve. The ciliary nerve was examined with transmission electron microscopy 30 days after culture and 6 days after co-culture with HSV. RESULTS: The ultrastructure of the explant of the ciliary nerve co-cultured with HSV showed that the viral capsid acquired a viral envelope and viral core, and a capsid and inclusion body within the nucleus. The enveloped virus was scattered within the vesicles of the cytoplasm. The virus-like particles were identified at the axonal fibers. CONCLUSIONS: The co-culture of the explant of the ciliary nerve and HSV showed the replicative process of the HSV within the cultured cell. The virus-like particles within the axon showed the evidence axonal transport of the virus under culture conditions.
Axonal Transport* ; Axons ; Capsid ; Cells, Cultured ; Coculture Techniques ; Cytoplasm ; Herpes Simplex* ; Herpesvirus 1, Human ; Inclusion Bodies ; Microscopy, Electron, Transmission ; Nerve Growth Factor ; Simplexvirus*

Co-infection of herpes simplex virus type 1 (HSV-1) and human cytomegalovirus (HCMV) is not uncommon in immunocompromised hosts. Importantly, organ transplant recipients concurrently infected with HSV-1 and HCMV have a worse clinical outcome than recipients infected with a single virus. However, factors regulating the pathologic response in HSV-1, HCMV co-infected tissues are unclear. We investigated the potential biologic role of HCMV gene product immediate early 1 (IE1) protein in HSV-1-induced syncytial formation in U373MG cells. We utilized a co-infection model by infecting HSV-1 to U373MG cells constitutively expressing HCMV IE1 protein, UMG1-2. Syncytial formation was assessed by enumerating nuclei number per syncytium and number of syncytia. HSV-1-induced syncytial formation was enhanced after 24 hr in UMG1-2 cells compared with U373MG controls. The amplified phenotype in UMG1-2 cells was effectively suppressed by roscovitine in addition to inhibitors of viral replication. This is the first study to provide histological evidence of the contribution of HCMV IE1 protein to enhanced cytopathogenic responses in active HSV-1 infection.
Cell Line, Tumor ; Giant Cells/*virology ; Herpesvirus 1, Human/*growth & development ; Humans ; Immediate-Early Proteins/biosynthesis/*metabolism ; Protein Kinase Inhibitors/pharmacology ; Purines/pharmacology ; Transfection ; Virus Replication/drug effects

Nerve growth factor (NGF) is mainly secreted by the neuroglia cells, which can exert biological effect through its receptors on the specific target cell surface. NGF is closely related to neurocyte growth, differentiation and apoptosis. As a neurotropic virus, HSV-1 an easily lead to neurocyte, neuroglia cells death or apoptosis. In this study, the U251 human glioma cells were chosen as target cells to study the change of NGF and its receptors in the apoptosis process of HSV-1 infection. Our results showed that U251 cells were permissive to HSV-1 replication. In the apoptosis process of HSV-1 infected U251 cells, the expression of both NGF and P75NTR increased and then decreased, while the expression of TrkA decreased gradually. These result indicated that HSV-1 was able to induce the abnormal expression of NGF and its receptors in U251 cells.
Apoptosis ; Cell Line, Tumor ; Gene Expression ; Glioma ; genetics ; metabolism ; physiopathology ; virology ; Herpes Simplex ; genetics ; metabolism ; physiopathology ; virology ; Herpesvirus 1, Human ; genetics ; physiology ; Humans ; Nerve Growth Factor ; genetics ; metabolism ; Receptor, Nerve Growth Factor ; genetics ; metabolism ; Receptor, trkA ; genetics ; metabolism ; Virus Replication

RT-PCR was used to detect expression level of VP16 mRNA and IFN-gamma mRNA in Herpes simplex virus type-1 infected mice brains at 4th day, 7th day, 10th day, 14th day, 21st day post infection and investigate the effects of the Gardenia extracts-T9 on viral replication and host immunity. The results showed that expression of VP16 mRNA in Gardenia extracts-T9 high dose and low dose group were both lower than that in virus control group at same time point. Relative VP16 mRNA expression in low dose group decreased at 21st day and relative VP16 mRNA expression in high dose group decreased continuously. Relative expression of IFN-gamma mRNA in high dose and low dose groups were both higher than that in virus control group at all time point except the 4th day. IFN-gamma mRNA in low dose group increased from the 4th day till the 14th day, and after the 14th day, the expression decreased slightly. Relative IFN-gamma mRNA in high dose group maintained increasing from 4th day till 21st day. Base on these results, we conclude that Gardenia extracts-T9 might exert the inhibition effect of viral replication by upregulating expression of IFN-gamma mRNA.
Animals ; Brain ; metabolism ; virology ; Female ; Gardenia ; chemistry ; Gene Expression Regulation, Viral ; drug effects ; genetics ; Herpesvirus 1, Human ; drug effects ; growth & development ; pathogenicity ; Interferon-gamma ; genetics ; Mice ; Mice, Inbred BALB C ; Plant Extracts ; administration & dosage ; pharmacology ; Polymerase Chain Reaction ; RNA, Messenger ; genetics ; Virus Replication ; drug effects

Geldanamycin (GA), an ansamycin antibiotic specifically binding heat shock protein 90 (Hsp90), exhibits a broad-spectrum antiviral effect. Herpes simplex virus type 1(HSV-1) replication in vitro was significantly inhibited by GA treatment. To explore the antiviral mechanism of GA against HSV-1, the 7267-spot human long oligonucleotide microarrays were applied to investigate the genes which might involved in the antiviral activity of GA in HeLa cells infected by HSV-1. Meanwhile, the reverse regulation of GA and HSV-1 on ACTG1, RAN, SOD1, HYAL1 were validated by using semi-quantitative RT-PCR. It is the first report of gene expression profile in cells infected by virus with GA treatment. The general impact of GA on cellular transcription may help to gain an insight into mechanism of its antiviral effect.
Antiviral Agents ; pharmacology ; Benzoquinones ; pharmacology ; Gene Expression Profiling ; Gene Expression Regulation ; drug effects ; HeLa Cells ; Herpesvirus 1, Human ; drug effects ; genetics ; growth & development ; Humans ; Lactams, Macrocyclic ; pharmacology ; Oligonucleotide Array Sequence Analysis ; Reverse Transcriptase Polymerase Chain Reaction ; Virus Replication ; drug effects ; genetics

GITR (glucocorticoid-induced TNF receptor) is a recently identified member of the TNF receptor superfamily. The receptor is preferentially expressed on CD4+CD25+ regulatory T cells and GITR signals break the suppressive activity of the subset. In this study, we wanted to reveal the in vivo function of GITR in herpes simplex virus type 1 (HSV-1) infection. A single injection of anti-GITR mAb (DTA-1) immediately after viral infection significantly increased the number of CD4+ and CD8+ T cells expressing CD25, an activation surface marker, and secreting IFN-gamma. We confirmed these in vivo observations by showing ex vivo that re-stimulation of CD4+ or CD8+ T cells with a CD4+ or CD8+ T-cell-specific HSV-1 peptide, respectively, induced a significant elevation in cell proliferation and in IFN-gamma secretion. Our results indicate that GITR signals play a critical role in the T-cell immunity to HSV-1.
Animals ; Antibodies, Monoclonal/pharmacology ; CD4-Positive T-Lymphocytes/immunology ; CD8-Positive T-Lymphocytes/immunology ; Cell Proliferation ; Female ; Glucocorticoids/*pharmacology ; Herpes Simplex/*immunology ; Herpesvirus 1, Human/pathogenicity ; *Immunity, Cellular ; Interferon Type II/secretion ; *Lymphocyte Activation ; Mice ; Mice, Inbred BALB C ; Peptide Fragments/metabolism ; Receptors, Interleukin-2/metabolism ; Receptors, Nerve Growth Factor/genetics/immunology/*metabolism ; Receptors, Tumor Necrosis Factor/genetics/immunology/*metabolism ; Research Support, Non-U.S. Gov't ; T-Lymphocytes/*immunology/metabolism/virology