Transcriptional activation of NF-kappa B activity by Epstein-Barr virus (EBV) LMP1 as a selective therapeutic strategy for EBV-associated diseases

Epstein-Barr virus (EBV) has been known to be associated with many malignant tumors, including nasopharyngeal carcinoma (NPC). Previous studies have indicated that an EBV-encoded oncoprotein, latent membrane protein 1 (LMP1), is expressed in many NPC tissues. LMP1 has been shown to stimulate HIV LTR through the two NF-kappa B binding sites within this promoter. In this study, we examined the feasibility of using this property of LMP1 as a therapeutic strategy for the treatment of NPC. This therapy consists of the preferential killing of the LMP1-expressing cells by gene transfer using the NF-kappa B-mediated herpes simplex virus thymidine kinase (HSVtk)/ganciclovir (GCV) system. The 800-bp HIV-LTR, which contains two NF-kappa B binding sites, was used to drive the HSVtk gene. Stable C33A cell clones expressing the LMP1 and the HSVtk genes were subjected to the GCV sensitivity test. Results showed that cells expressing both the LMP1 and the HSVtk genes were highly sensitive to GCV treatment. These cells were introduced into nude mice subcutaneously and tumors became palpable within 2 weeks. GCV was then introduced intraperitoneally to these mice and the sizes of the tumors were measured daily. Results showed that the tumors regressed in the group of mice carrying cells that stably expressed both the LMP1 and the HSVtk genes, but not in mice carrying cells containing LMP1 or HSVtk alone. Our data indicate that the HSVtk gene expressed from a NF-kappa B-binding motif-containing promoter that is regulated by LMP1 may be used as an in vivo gene therapy strategy of EBV LMP1-expressing cancers such as NPC.     

Inactivation of respiratory syncytial virus by detergents and disinfectants

The activity of a number of detergents and disinfectants against respiratory syncytial virus (RSV) was evaluated in an in vitro assay system. Equal volumes of RSV and serial 10-fold dilutions of the test agents were mixed at 4 degrees C for 5 minutes. The RSV titer in each mixture was compared with that of untreated RSV alone. In 14 experiments with input RSV titers ranging from 2.6 x 10(3) to 2 x 10(7) plaque-forming units/ml, a 10-fold dilution of 5.25% sodium hypochlorite (generic bleach) inactivated (> or = 3-log reduction in titer) the virus. With lower RSV titers inactivation was also observed at a 100-fold dilution of bleach. Fetal calf serum concentrations up to 50% as an organic load did not diminish the bleach effect. The degree of RSV inactivation was also defined for Lysol, povidone-iodine, Amphyl, Hibiclens, Osyl, ethanol and Listermint. The short contact time, the reproducible nature of the findings and the continued effectiveness with increasing organic loads all suggest that detergents and disinfectants can potentially play an important role in decreasing the spread of RSV infection.     

Recombinant human respiratory syncytial virus (RSV) from cDNA and construction of subgroup A and B chimeric RSV

OBJECTIVE: The present study was performed to discriminate between central and peripheral effects of noradrenaline (NA) in normotensive, non-obese, type 2 diabetic patients. METHODS: Study I: In 10 patients and 10 healthy volunteer (HV) cumulative doses of NA were infused intravenously until mean arterial pressure (MAP) rose with 20 mmHg, and subsequently the effects on the forearm blood flow (FBF) was measured. Also, the FBF response to intra-arterial NA (0.025, 0.1, 0.4 micrograms min-1) was measured. Study II: In 13 patients and 14 HV the venous constrictor response to a cumulative local infusion of NA in a dorsal hand vein was determined. RESULTS: In study I the circulating plasma NA concentrations inducing a rise in MAP of 20 mmHg, were lower in the type 2 patients relative to the HV (p < 0.01). The relationship between changes in pressure and changes in heart rate were similar in both groups. Moreover, FBF responsiveness to intra-arterial NA was not different between the two groups. The slopes of the delta MAP/NA regression lines were correlated with basal insulin levels and relative insulin resistance in the healthy volunteers (R = 0.77, p < 0.01, and R = 0.83, p < 0.01), but not in the type 2 diabetic patients. In study II no differences were observed in the dose generating half maximum (ED50) and the maximum (Emax) response to NA between the type 2 patients and the HV. CONCLUSIONS: Non-obese normotensive type 2 patients have an increased pressor response to NA, which is not based upon a defect in skeletal muscle resistance arterioles, peripheral veins, or a defect in the baroreceptor system. Therefore, in type 2 diabetes the noradrenergic responsiveness of other vascular beds, such as the splanchnic or renal, must be enhanced.     

Incidence of acute otitis media associated with group A and B respiratory syncytial virus infections

The comparative association of respiratory syncytial virus group A and B infections with acute otitis media was determined by analysing the hospital records of children with community-acquired respiratory syncytial virus infection during three successive outbreaks from 1987 to 1992. Of 326 episodes analysed, 192 (59%) were caused by group A and 134 (41%) by group B infections. Acute otitis media was diagnosed in 101 (75%) children with group B infection, compared with 119 (62%) with group A infection (p = 0.01). Group A infections were more often associated with wheezing (71% versus 59% in group B; p = 0.02) and oxygen therapy in inpatients (48% versus 31%, respectively; p = 0.008). The higher incidence of acute otitis media associated with group B infections was observed both after adjustment for potential confounding variables and during each outbreak.     

Preparation of respiratory syncytial virus subgroup A and B antigens for enzyme immunoassay antibody detection

A simplified method was described for purification of respiratory syncytial virus (RSV) subgroup A and B aimed to be used as antigens in enzyme immunoassay (EIA). The titer of each RSV subgroup and the amount of protein was determined from the visible band in 45% sucrose gradient. The quality of prepared RSV subgroup antigens for EIA was described in terms of the achievable final titer, the amount of protein, and EIA criss-cross titration. The RSV subgroup A and B antigens, diluted as 1:100 (low opalescent band in 45% sucrose layer) or 1:800 (high opalescent band in 45% sucrose layer) produced a positive reaction in EIA criss-cross titration with IgG antibodies from the patient's serum (convalescent phase) diluted as 1:25,600 (for RSV A) and 1:6,400 (for RSV B). This method offers shorter and more simplified steps of viral antigen purification, and provides acceptable quantity and quality of viral antigens appropriate for use in EIA.     

Respiratory syncytial virus infection of human respiratory epithelial cells up-regulates class I MHC expression through the induction of IFN-beta and IL-1 alpha

CD8+ T cells mediate some of the damage to the lung epithelium following respiratory syncytial virus (RSV) infection. Since CD8+ T cells recognize antigen-laden class I MHC molecules on the target cells, we examined in this study the expression of class I MHC by RSV-infected respiratory epithelial cells. Respiratory epithelial cell lines and bronchial epithelial cells from normal human tissue responded to RSV infection with an increased expression of class I MHC as determined by flow cytometry and immunoprecipitation of class I MHC from metabolically radiolabeled cells. The increase in class I MHC expression was dependent on infectious, replicating virus. UV-irradiated culture supernatants from RSV-infected A549 cells, when added to fresh A549 cell cultures, induced an increase in class I MHC expression by those cells. The class I MHC increasing activity within supernatants from A549 cells was due, in large part, to IFN-beta, and to a lesser extent to IL-1 alpha. The addition of neutralizing Abs to both cytokines completely blocked the increase in class I MHC expression by cells treated with the above-mentioned supernatants. These results demonstrate that RSV infection elicits IFN-beta production by respiratory epithelial cells, which in turn leads to an increase in their synthesis of class I MHC, which would facilitate their recognition and lysis by RSV-specific CD8+ T cells.     

Inhibition of sodium selenite on transformation of human B lymphocytes by Epstein-Barr virus

We detected the effect of non-toxic doses of sodium selenite (Se) on the expressing effect of nuclear antigen (EBNA) of Epstein-Barr virus (EBV) in Raji cell line by microfluorescence spectrophoto-quantitation (MFS) and studied the inhibing transformation in B lymphocytes from human umbilical cord blood (BL) with EBV in vitro by Se. We found that Se (0.01-0.50 microgram/ml) can lower EBNA-MFS value 8.4%-21.8% in Raji cell line. The transformation effect of BL with EBV was markedly inhibited by Se (0.1-1.0 microgram/ml) in pretreatment and cotreatment. The inhibition rates of BL transformation were 73.4%-92.1% and 60.3%-77.2% respectively. EBV is associated with nasopharyngeal carcinoma (NPC). The Se levels of serum and hair in NPC patients were markedly lower than those in healthy persons. The results suggest that supplement Se to persons for active expression of EBV gene and NPC patients may help protect againt and treat NPC and diseases associated with EBV.     

Effect of compounds with antibacterial activities in human milk on respiratory syncytial virus and cytomegalovirus in vitro

In differential diagnosis of a delir also adverse effects of medicaments have to be taken into account beside other causes. We report a case of an agitated delir with nocturnal disturbance of consciousness, confusion, restlessness and sleeplessness. This delir existed exclusively during the therapy of a cutaneous herpes zoster with zovirax-pills which can only be explained by a causal connection--after exclusion of other causes. As a so far undescribed predisposition for neurotoxicity of oral therapy with acyclovir signs of vascular encephalopathy were found in the patient's cranial magnetic resonance imaging. The central nervous side effects of acyclovir were summarized shortly.     

Cross-reactive and group-specific immune responses to a neutralizing epitope of the human respiratory syncytial virus fusion protein

Immune responses to a synthetic peptide corresponding to amino-acids 205-225 of the fusion protein from group B respiratory syncytial (RS) virus, were studied in mice and rabbits, and compared to a similar peptide from group A RS virus. Peptide 205-225 (B) was recognized by monoclonal antibody RS-348, and was immunogenic in both mice and rabbits, as was peptide 205-225 from the fusion protein of a group A strain. Peptide 205-225 (B) induced a proliferative T-cell response, demonstrating the existence of a T-cell epitope in this region of the fusion protein of group B viruses. Both peptides were able to induce a T-cell cross-reactive proliferation when mice were primed with either the homologous or the heterologous peptide. ELISA were performed using synthetic peptides or whole virus (from group A and B) as antigens. Mice anti-peptide sera recognized both homologous and heterologous peptides. A similar pattern was observed with RS virus strains. In indirect immunofluorescence assays, both anti-peptide rabbit sera recognized human nasal epithelial cells infected with A or B strains of RS virus. In contrast, while anti-peptide 205-225 rabbit serum from group A neutralized group A and B strains of RS virus, anti-peptide 205-225 rabbit serum from group B was unable to neutralize a group A virus, although it neutralized a group B strain. These results are similar to the immune response observed in children following primary RS virus infection.     

Distinct patterns of T- and B-cell immunity to respiratory syncytial virus induced by individual viral proteins

The purpose of this study was to determine the safety of 4 week re-use of vinyl urinary leg and bed bags in the acute rehabilitation setting when decontaminated daily with a dilute bleach (sodium hypochlorite) rinse. Patients requiring urinary bags (n = 54) were randomly assigned to Control (C) and Experimental (E) groups. C's bags were replaced weekly; E's only after four weeks. Both groups received identical daily bag decontamination and weekly urine and bag cultures. No significant differences were found between groups with ANCOVA, controlling for baseline urine cultures, age, number of days catheterized, and use of antibiotics. Thirty different organisms were cultured in urine and bags; when the procedure was done daily, all bag cultures showed only minimal contamination (0-100CFU/mL). Bacterial growth (4.4% of leg bags) > 100CFU/mL was found only when the daily decontamination procedure had been omitted. In fact, 57% of leg bags and 76.5% of bed bags returned with no growth. We conclude that it is safe and cost effective to reuse vinyl urinary leg and bed bags for four weeks.