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

Codon ; Herpes Simplex ; virology ; Herpesvirus 1, Human ; genetics ; metabolism ; Humans ; Protein Binding ; Protein Biosynthesis ; Viral Proteins ; genetics ; metabolism

ATP-Binding Cassette Transporters ; immunology ; metabolism ; Animals ; Herpes Simplex ; immunology ; virology ; Herpesvirus 1, Human ; immunology ; metabolism ; Humans ; Immediate-Early Proteins ; immunology ; metabolism ; Viral Proteins ; immunology ; metabolism

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

Envelope proteins of herpes simplex virus (HSV) plays a vital role not only in the infection process of adsorption and invasion but also in the stimulation to the organism that gives rise to immune response. Among the envelope proteins, glycoprotein D (gD), which can induce specific immune response, are the primary targets of humoral and cellular immunity of the host. In order to analyze the antigenicity and immunogenicity of HSV-gD1, we chemically synthesized the extracellular domain fragment gene of gD1, cloned it into eucaryotic expression vector pCEP4, and transfected the HEK293 cells with the recombinant vector. Then we identified the recombinant protein by Western blotting, and detected antigenicity of the protein by ELISA. Finally, we used the purified gD1 protein to immunize Kunming mice in 1, 3, 5 weeks, and collected antiserum in 3, 5 and 7 weeks. We titrated the sera for the detection of anti gD1 using an ELISA assay. Gene sequencing analysis demonstrated that the recombinant plasmid pCEP4-gD1 was constructed successfully. Western blotting analysis indicated one major protein band, which molecular weights is approximate 46 kDa corresponding to the truncated forms of gD1 protein, was observed. ELISA assay showed that the expressed recombinant protein gD1 had good antigenicity. After the third immunization, antibody titer of the mouse anti-gD1 was at least 5 x10(3). The successful expression of the recombinant protein gD1, which can induce humoral immune response, lays a foundation for serological diagnosis and vaccine study of HSV.
Animals ; HEK293 Cells ; Herpesvirus 1, Human ; immunology ; metabolism ; Humans ; Immunization ; Mice ; Recombinant Proteins ; biosynthesis ; genetics ; immunology ; Transfection ; Viral Envelope Proteins ; biosynthesis ; genetics ; immunology

ADP-ribosyl Cyclase 1 ; metabolism ; Aged ; Epstein-Barr Virus Infections ; Herpesvirus 4, Human ; isolation & purification ; Humans ; Interferon Regulatory Factors ; metabolism ; Ki-67 Antigen ; metabolism ; Male ; Nasal Cavity ; Nose Neoplasms ; metabolism ; pathology ; surgery ; virology ; Plasmacytoma ; metabolism ; pathology ; surgery ; virology

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

Cyanovirin-N (CVN) is an 11 kDa anti-HIV protein originally isolated from extracts of a cyanobacterium, Nostoc ellipsosporum. The protein binds with high affinity to the viral envelope glycoprotein gp120 and irreversibly inactivates diverse HIV strains. A fusion gene consisting of cvn, sumo and 6xHis tag was synthesized by PCR according to the codon bias of Escherichia coli. The fusion protein is expressed in the cytoplasm of E. coli in a soluble form and up to 28% of the total protein. The recombinant CVN was purified to homogeneity by 2 rounds of Ni-NTA affinity chromatography and one round of SUMO protease cleavage. Bioactivity assay demonstrated that SUMO-CVN and CVN bound to gp120 with nanomolar concentration. In addition, CVN showed potent anti-HSV-1 and anti-HIV-1 activities in in vitro cellular assays. Therefore, the 6xHis SUMO fusion expression and purification system provides a better approach for large scale production of CVN for further microbicide development.
Antiviral Agents ; isolation & purification ; metabolism ; pharmacology ; Bacterial Proteins ; biosynthesis ; genetics ; pharmacology ; Carrier Proteins ; biosynthesis ; genetics ; pharmacology ; Escherichia coli ; genetics ; metabolism ; HIV-1 ; drug effects ; Herpesvirus 1, Human ; drug effects ; Recombinant Proteins ; biosynthesis ; genetics ; isolation & purification ; pharmacology

Herpes simplex virus type 1 (HSV-1) is a common human pathogen causing cold sores and even more serious diseases. It can establish a latent stage in sensory ganglia after primary epithelial infections, and reactivate in response to stress or sunlight. Previous studies have demonstrated that viral immediate-early protein ICP0 plays a key role in regulating the balance between lytic and latent infection. Recently, It has been determined that promyelocytic leukemia (PML) nuclear bodies (NBs), small nuclear sub-structures, contribute to the repression of HSV-1 infection in the absence of functional ICP0. In this review, we discuss the fundamentals of the interaction between ICP0 and PML NBs, suggesting a potential link between PML NBs and ICP0 in regulating lytic and latent infection of HSV-1.
Herpes Simplex ; virology ; Herpesvirus 1, Human ; genetics ; physiology ; Humans ; Immediate-Early Proteins ; metabolism ; Intranuclear Inclusion Bodies ; metabolism ; virology ; Leukemia, Promyelocytic, Acute ; metabolism ; Ubiquitin-Protein Ligases ; metabolism ; Virus Latency ; physiology

OBJECTIVETo observe the gene and protein expression of herpes simplex virus type I-thymidine kinase (HSV(1)-tk) in the ovarian tumor tissues and other organs after arterial infusion of HSV(1)-tk gene mediated by GE7 delivery system.

METHODSGE7-polylysine/pCMV-HSV(1)-tk/polylysine-HA20 complexes were constructed. Nine rats with induced ovarian tumor were divided into 3 groups, injecting the 4-element complexes or saline buffer through the ovarian artery and complexes through the tail vein, respectively. The ovarian tumors, hearts, livers, spleens, lungs and kidneys were obtained at 72 hours after injection. RT-PCR and Western Blot were preceeded to determine the expression of HSV(1)-tk gene and protein in the tumor tissues and other organs.

RESULTSIn the group of arterial injection with 4-element complexes, the HSV(1)-tk gene and protein were expressed strongly in the tumor tissues, while little or none was detected in other organs. In the group of arterial injection with saline buffer, no HSV(1)-tk gene and protein was detected in both tumor tissues and other organs. In the group of tail vein injection, none was detected in tumor tissues and only little was found in the livers, spleens, lungs and kidneys.

CONCLUSIONHigh target and gene transfer rates can be obtained when HSV(1)-tk gene is transferred via the artery route mediated by GE7 delivery system. HSV(1)-tk protein can be expressed after the gene transfer. The results may provide a new strategy for target killing of HSV(1)-tk/GCV system in ovarian tumors.

9,10-Dimethyl-1,2-benzanthracene ; Adenocarcinoma ; chemically induced ; genetics ; metabolism ; Animals ; Female ; Gene Transfer Techniques ; Herpesvirus 1, Human ; genetics ; Infusions, Intra-Arterial ; Ovarian Neoplasms ; chemically induced ; genetics ; metabolism ; RNA, Messenger ; metabolism ; Random Allocation ; Rats ; Rats, Wistar ; Thymidine Kinase ; biosynthesis ; genetics