Seihachiro Niwayama

In vitro and in vivo antiviral activity of scopadulcic acid B from Scoparia dulcis, Scrophulariaceae, against herpes simplex virus type 1

The antiviral activity of five diterpenoids isolated from Scoparia dulcis L., Scrophulariaceae, was examined in vitro against herpes simplex virus type 1. Among these compounds, only scopadulcic acid B was found to inhibit the viral replication with the in vitro therapeutic index of 16.7. The action of scopadulcic acid B was not due to a direct virucidal effect or inhibition of virus attachment to host cells. Single-cycle replication experiments indicated that the compound interfered with considerably early events of virus growth. The influence of scopadulcic acid B on the course of the primary corneal herpes simplex virus infection was investigated by means of a hamster test model. When the treatment was initiated immediately after virus inoculation, scopadulcic acid B, when applied orally or intraperitoneally, effectively prolonged both the appearance of herpetic lesions and the survival time at the dose of 100 and 200 mg/kg per day.

Evaluation of the Efficacy of Split-Product Trivalent A(H1N1), A(H3N2), and B Influenza Vaccines: Reactogenicity, Immunogenicity and Persistence of Antibodies Following Two Doses of Vaccines

The reactogenicity and immunogenicity of Tween‐ether split trivalent A(H1N1), A (H3N2), and B influenza vaccine in primary school children aged seven to 12 years, and the persistence of antibodies following two doses of vaccine were studied during 1980–1984. Adverse reactions were infrequent, and, even when reported, were chiefly local ones, mild in nature and of short duration. Most of the reactions were less frequent after the second dose than after the first dose. Most of the systemic reactions occurred during the intervaccination period with almost equal frequency, indicating that careful consideration is required to judge whether they were induced by vaccination or not. This vaccine had induced adequate hemagglutination inhibiting (HAI) antibody because the geometric mean titers (GMTs) of the vaccinees were two‐to eightfold higher than those of the nonvaccinees to any of the vaccine antigens following two doses of vaccine. In general, the responses to A(H3N2) virus were the best among the vaccine antigens through the four vaccination seasons, but there was a tendency to show a poorer response to the same type (or subtype) of virus antigen as the causative one during a protracted epidemic. The antibodies induced by either vaccination or natural infection were shown to persist for less than a year, supporting the recommendation for annual vaccination.

Yucca leaf protein (YLP) stops the protein synthesis in HSV-infected cells and inhibits virus replication

Yucca leaf protein (YLP), an inhibitor of tobacco mosaic virus isolated from the leaves of Yucca recurvifolia Salisb., exhibited potent activity against herpes simplex virus type 1 (HSV-1) with no cytotoxicity below 300 micrograms/ml. The inhibitory dose was varied with the time of addition; 50% effective concentrations (ED50) of YLP were 3, 19 and 95 micrograms/ml when YLP exposure was begun 3 h before virus infection, 0 h and 3 h after infection, respectively. This protein also inhibited the multiplication of herpes simplex virus type 2 and human cytomegalovirus. YLP has been shown to have a weak virucidal activity at higher concentrations. Analysis of early events following infection showed that YLP affected viral penetration in HeLa cells but did not interfere with adsorption to the cells. YLP was found to exert strong inhibition of protein synthesis in virus-infected cells but not in uninfected cells. This selective effect can be considered to attribute mainly to the antiviral activity of YLP.

Influence of trifluoperazine on the late stage of influenza virus infection in MDCK cells.

We investigated the influence of the anticalmodulin drug, trifluoperazine (TFP) on influenza virus growth in MDCK cells. The inhibitory effect of TFP on virus growth was observed even when TFP was added at a late stage of infection. This inhibitory effect was concentration-dependent in the concentration range of 20-35 microM. At 35 microM, TFP caused a complete alteration in the distribution pattern of hemagglutinin (HA), concomitant with a decrease in the appearance of HA on the cell surface. After removal of the drug, the HA gradually began to show a normal distribution pattern and reappeared on the cell surface. The time course of rearrangement of HA was in accord with that of the recovery of cell supernatant infectivity. Scanning electron microscopic study revealed that the drug did not cause accumulation of the progeny viruses on the cell surface. The drug effect on the virus growth was reversed by the simultaneous presence of purified calmodulin (CaM). These data suggest that TFP acts as a reversible inhibitor of influenza virus morphogenesis, but not budding, by disturbing cellular CaM and/or CaM-dependent functions.

Inhibition of experimental pulmonary metastasis in mice by β-cyclodextrin-benzaldehyde

SummaryThe effect of β-cyclodextrin-benzaldehyde (CDBA) on experimental pulmonary metastasis in C3H/He mice was examined. In an in vitro assay, the growth of RCT(+) cells was inhibited by 1200 μg/ml CDBA using unrenewed media, and by 600 μg/ml CDBA in that using daily renewed media. When mice were treated daily with CDBA, 3 weeks later the number of lung nodules developing after i.v. injection of 1×106 RCT(+) cells was significantly decreased in a dose-dependent manner, i.e., 73.8%, 85.6%, and 95.7% inhibition was observed following 0.5, 5, and 25 mg CDBA/mouse per day p.o. administration, respectively. The same mice showed almost as much natural killer (NK) activity as normal mice. Therefore, experiments were designed to evaluate the effect of CDBA on the NK activity of tumor-free mice whose immunity had been suppressed by 5-fluorouracil (5FU). Injections of 5FU only suppressed this activity to about 50% of normal mice, but the combined treatment with CDBA negated the suppressive effect of 5FU on NK activity. The results suggested that the inhibition of experimental pulmonary metastasis might be induced by the possible combined effects of CDBA; that is, the direct inhibition of tumors and the augmentation of NK cell activity.

Evaluation of the efficacy of split-product trivalent A(H1N1), A(H3N2), and B influenza vaccines: protective efficacy.

A total of 1,995 primary school children (1,464 vaccinees and 531 nonvaccinees) were studied to evaluate the protective efficacy of Tween‐ether split trivalent A(H1N1), A(H3N2), and B influenza vaccines by comparison of the incidence of confirmed infection in two groups during 1980 to 1984. During the study period, epidemics caused by antigenically different influenza viruses, that is A(H1N1) epidemics in 1981 and 1984, a B epidemic in 1982 and an A(H3N2) epidemic in 1983, were experienced, and at the same time strains changed by antigenic drift were frequently isolated. In these epidemics, 61% to 87% of the children reported respiratory illnesses and 18% to 48% of the illnesses were influenza confirmed by seroconversion. Throughout these four epidemics, the incidence of confirmed infection among the vaccinees (7.8% to 33.8%) was 6.5% to 34.8% lower than that among the nonvaccinees (35.4% to 51.6%), demonstrating that the vaccine was effective (χ2=76.34, P<0.001). However, this effectiveness was not seen in an epidemic in one of the entrant schools in 1984, possibly caused by a strain with intense antigenic drift. On the basis of data on incidence of various symptoms, duration of fever and the number of days of absence from class, it was considered that clinical symptoms in the vaccinees were milder than those in the nonvaccinees. When the titers of hemagglutination‐inhibiting (HAI) antibody against the vaccine strains were measured, the protective level of HAI antibody giving ≦50% incidence of infection was 1:64, but it increased to 1:256 in the 1984 epidemic, reflecting the high rate of isolates with intense antigenic drift.

Effects of Gangliosides on the Growth of Herpes Simplex Virus Type 1-Infected Cells Derived from Neurons and on Viral Replication

Although it is well-known that herpes simplex virus can establish latent infections in neurons of sensory and sympathetic ganglia, little is known about the viral or cellular factors which regulate the latent state. Experiments were designed to elucidate the effects that can be produced by adding gangliosides, abundant components in neurons but not in most other cell types, to virus-infected cells from mouse trigeminal ganglia and from the neuroblastoma x glioma hybrid cell line NG108-15. The results obtained indicate that gangliosides, when used in combination with acyclovir, efficiently protect the infected cells from lysis in both cell systems, and that they can exert antiviral activity at least in part via suppressing protein kinase C activity.

Relation of H-2 expression on murine RCT(+) sarcoma cells to lung colonization and sensitivity to NK cells

SummaryMurine RCT(+) sarcoma cells were sorted using a fluorescence-activated cell sorter with regard to the expression of H-2 antigens and then an increased H-2-expressing subclone was established, and named RCT(+)H-2+. The experimental metastasis of RCT(+) cells was compared with that of RCT(+)H-2+ cells by counting pulmonary colonies on the 21st day after i.v. inoculation of tumor cells (5–10×104/mouse). When mice were inoculated with RCT(+) cells, mean numbers of pulmonary colonies were 2.1(range 0–6), 2.8(range 0–7) using 5×104 and 1×105 cells, respectively. On the other hand, in the mice inoculated with RCT(+)H-2+ cells, figures obtained were 7.0(range 4–16), 31.9(range 13–79), using 5×104 and 1×105 cells, respectively. The survival rate of RCT(+)H-2+ cells was higher than that of RCT(+) cells, when this was assayed in the early stage after i.v. injection of 51Cr-labeled cells (1×105 cells/mouse). In addition, RCT(+)H-2+ cells were more resistant than RCT(+) cells to lysis mediated by natural killer cells. These data suggest that an increase in metastatic ability was paralleled by an increase in the H-2 antigen expression and a decrease in sensitivity to the natural killer cells.

Augmentation of murine lymphokine-activated killer cell cytotoxicity by β-cyclodextrin-benzaldehyde

SummaryWe investigated the effect ofβ-cyclodextrin-benzaldehyde (CDBA) on lymphokine-activated killer (LAK) cell activity of spleen cells from normal or RCT(+)H-2+-sarcoma-bearing C3H/He mice. CDBA augmented the induction of LAK cytotoxicity in vitro against RCT(+)H-2+ tumor cells by IL-2, whereas the culture with CDBA alone did not. In a LAK cytotoxicity assay in vitro, the augmentative effect of CDBA was strongly exerted against spleen cells originating from 2-week-tumor-bearing mice, rather than those from normal mice or mice that had born tumors for 5 weeks. Such an augmentative effect was not observed against other tumor cells (YAC-1, D-6, Colon-26 and EL-4 cells) non-specifically. When the intravenous adoptive transfer of LAK cells was carried out in the mice, LAK cells from tumor-bearing mice induced by combined culture with interleukin-2 (IL-2) and CDBA markedly inhibited the pulmonary metastases of RCT(+)H-2+ tumor, while neither LAK cells from the same tumor-bearing mice induced by only IL-2 nor those from normal mice inhibited the pulmonary metastasis. The majority of LAK cells induced either by IL-2 plus CDBA or by IL-2 alone were found to be Thy1.2+ and asialoGM1+ cells by flow-cytometric analysis, but no obvious phenotypical difference was observed between them. However, the most significant effect of CDBA might be the maintenance of the Lyt-2+ cell level in the spleen cells from tumor-bearing mice. These results suggested that the costimulation of spleen cells with IL-2 and CDBA might induce cytotoxic T cells specific for syngeneic tumor cells.