Yoshio Numazaki

Rhinovirus infection of primary cultures of human tracheal epithelium: role of ICAM-1 and IL-1β

Exacerbations of asthma are often associated with respiratory infection caused by rhinoviruses. To study the effects of rhinovirus infection on respiratory epithelium, a primary target for respiratory viruses, human rhinovirus (HRV)-2 and HRV-14 were infected to primary cultures of human tracheal epithelial cells. Viral infection was confirmed by showing that viral titers of supernatants and lysates from infected cells increased with time and by polymerase chain reaction. HRV-2 and HRV-14 infections upregulated the expression of intercellular adhesion molecule-1 (ICAM-1) mRNA, the major rhinovirus receptor, on epithelial cells, and they increased the production of interleukin (IL)-1β, IL-6, IL-8, and tumor necrosis factor (TNF)-α in supernatants. Antibodies to ICAM-1 inhibited HRV-14 infection of epithelial cells and decreased the production of cytokines after HRV-14 infection, but they did not alter HRV-2 infection-induced production of cytokines. IL-1β upregulated ICAM-1 mRNA expression and increased susceptibility to HRV-14 infection, whereas other cytokines failed to alter ICAM-1 mRNA expression. Furthermore, a neutralizing antibody to IL-1β significantly decreased viral titers of supernatants and ICAM-1 mRNA expression after HRV-14 infection, but a neutralizing antibody to TNF-α was without effect. Immunohistochemical studies revealed that both HRV-14 infection and IL-1β increased ICAM-1 expression on cultured epithelial cells. These findings imply that HRV-14 infection upregulated ICAM-1 expression on epithelial cells through increased production of IL-1β, thereby increasing susceptibility to infection. These events may be important for amplification of airway inflammation after viral infection in asthma.

A Microplate Method for Isolation of Viruses from Infants and Children with Acute Respiratory Infections

Between December 1984 and December 1986, a microplate technique was adopted for isolation of viruses from infants and children with acute respiratory infections. By using two kinds of tissue culture microplates, i.e., the HHVM plate, containing human embryonic fibroblast (HEF), HEp‐2, Vero and MDCK cells, and the MK plate which contains secondary monkey kidney cells, 1,080 field viruses were isolated from 1,061 (24.9%) out of 4,254 throat swabs. Of these 1,080 isolates, 1,003 (92.9%) were recovered in the HHVM plates and the remaining 77 (7.1%) in the MK plates. With the HHVM plate, influenza A and B viruses were cultivated in MDCK, RS virus in HEp‐2, parainfluenza and mumps viruses in Vero, adenoviruses in both HEF and HEp‐2, polioviruses in HEF, HEp‐2 and Vero, coxsackie B viruses in both HEp‐2 and Vero, rhino and echo viruses in HEF, herpes simplex virus in both HEF and HEp‐2, and cytomegalovirus in HEF, although MK were more sensitive than Vero to parainfluenza and coxsackie B viruses. There was no difference in the rate of isolation of viruses between the microplate and ordinary tube methods. Cross contamination in the microplates was negligible for routine work.

Acute hemorrhagic cystitis in children. Isolation of adenovirus type 11.

Abstract A viral etiology of acute hemorrhagic cystitis was sought in 11 children seven to 15 years of age. Type 11 adenovirus was recovered from the urine of nine of these, and a significant rise in neutralizing antibody titer against Type 11 adenovirus was demonstrated in all. No virus was isolated from the urine of normal children or children with other diseases. Type 11 adenovirus is proposed as an etiologic agent of acute hemorrhagic cystitis.

Interspecies transmission of influenza C virus between humans and pigs

The antigenic and genetic characteristics of the 18 human strains of influenza C virus isolated in Yamagata and Sendai Cities, Japan between January 1991 and February 1993 were investigated. Antigenic analysis with monoclonal antibodies to the hemagglutinin-esterase glycoprotein showed that the isolates could be divided into three distinct groups closely related to C/Yamagata/26/81, C/Aichi/1/81 and C/Mississippi/80, respectively. T1-oligonucleotide fingerprinting of total vRNA revealed that the six isolates belonging to the C/Yamagata/26/81 virus group had the genomes greatly similar to one another but considerably different from those of the 1988/1990 isolates (except C/Yamagata/10/89) of the same antigenic group. Comparison of total or partial nucleotide sequences of the seven RNA segments of the three strains (C/Miyagi/3/91, C/Miyagi/9/91 and C/Miyagi/2/92) representative of the 1991/1993 strains of the C/Yamagata/26/81 virus group with those of the previous influenza C isolates obtained from humans and pigs during 1980/1989 showed that the 1991/1993 strains, like C/Yamagata/10/89, are more closely related to viruses isolated from pigs in Beijing, China in 1981/1982 than to any of the isolates from humans. This observation suggests strongly that interspecies transmission of influenza C virus between humans and pigs has occurred in nature, although it is not known whether the virus has been transmitted from pigs to humans or from humans to pigs.

Further Study on Acute Hemorrhagic Cystitis Due to Adenovirus Type 11

Abstract To establish the viral origin of acute hemorrhagic cystitis in children as well as adults, 28 patients (23 children and five adults) were examined virologically or serologically during the...

Neutralizing Mechanisms of Two Human Monoclonal Antibodies Against Human Cytomegalovirus Glycoprotein 130/55

The neutralization of human cytomegalovirus (HCMV) after adsorption to the cell surface at 4 degrees C was studied using two neutralizing monoclonal antibodies (C-23 and C-41) recognizing glycoprotein 130/55. HCMV adsorbed to cells was neutralized by C-23 (complement-independent), but not by C-41 (complement-dependent). Furthermore, the virus remained sensitive to C-23 for 120 min after shifting up from 4 degrees C to 37 degrees C, suggesting that C-23 might block an early stage of virus penetration into cells, and also that transition from virus attachment to virus penetration might be quite slow. The cell-to-cell infection of HCMV was also blocked only by C-23, and not by C-41. On the basis of the results presented here, we suggest that C-41 blocks the attachment of virus to the cell surface whereas C-23 prevents the penetration of virus into the cell.

Infection of human respiratory submucosal glands with rhinovirus: effects on cytokine and ICAM-1 production.

To further understand the early biochemical events that occur in infected surface epithelium, we developed for the first time a model in which a respiratory submucosal gland cell population can be infected with rhinovirus (RV). Viral infection was confirmed by demonstrating with PCR that viral titers in supernatants and lysates from infected cells increased with time. Infection by RV14 upregulated the expression of intercellular adhesion molecule-1 (ICAM-1) mRNA, the major RV receptor, on submucosal gland cells, and it increased production of interleukin (IL)-1α, IL-1β, IL-6, IL-8, tumor necrosis factor-α, and granulocyte-macrophage colony-stimulating factor in supernatants. Antibodies to ICAM-1 inhibited RV infection of submucosal gland cells and decreased the production of cytokines after RV infection. Both IL-1α and IL-1β upregulated ICAM-1 mRNA expression and increased susceptibility to RV infection, whereas other cytokines failed to alter ICAM-1 mRNA expression. Furthermore, neutralizing antibodies to IL-1α and IL-1β significantly decreased the viral titers in supernatants and ICAM-1 mRNA expression after RV infection, but a neutralizing antibody to tumor necrosis factor-α was without effect. These findings suggest that respiratory submucosal gland cells play an important role in the initial stages of inflammation and provide useful insights into the pathogenesis of RV infection.To further understand the early biochemical events that occur in infected surface epithelium, we developed for the first time a model in which a respiratory submucosal gland cell population can be infected with rhinovirus (RV). Viral infection was confirmed by demonstrating with PCR that viral titers in supernatants and lysates from infected cells increased with time. Infection by RV14 upregulated the expression of intercellular adhesion molecule-1 (ICAM-1) mRNA, the major RV receptor, on submucosal gland cells, and it increased production of interleukin (IL)-1alpha, IL-1beta, IL-6, IL-8, tumor necrosis factor-alpha, and granulocyte-macrophage colony-stimulating factor in supernatants. Antibodies to ICAM-1 inhibited RV infection of submucosal gland cells and decreased the production of cytokines after RV infection. Both IL-1alpha and IL-1beta upregulated ICAM-1 mRNA expression and increased susceptibility to RV infection, whereas other cytokines failed to alter ICAM-1 mRNA expression. Furthermore, neutralizing antibodies to IL-1alpha and IL-1beta significantly decreased the viral titers in supernatants and ICAM-1 mRNA expression after RV infection, but a neutralizing antibody to tumor necrosis factor-alpha was without effect. These findings suggest that respiratory submucosal gland cells play an important role in the initial stages of inflammation and provide useful insights into the pathogenesis of RV infection.

FREQUENT OCCURRENCE OF GENETIC REASSORTMENT BETWEEN INFLUENZA C VIRUS STRAINS IN NATURE

Previous studies of the haemagglutinin-esterase (HE) genes of various influenza C isolates suggested the existence of three distinct virus lineages (C/Yamagata/26/81-, C/Aichi/1/81- and C/Mississippi/80-related lineages) in Japan in the 1980s. Here we analysed the genetic properties of three strains (C/Yamagata/5/92, C/Miyagi/3/93 and C/Miyagi/4/93) isolated in Yamagata and Sendai Cities, Japan, in 1992/1993. Comparison of total or partial nucleotide sequences of the seven RNA segments of C/Yamagata/5/92 with those of 11 previous isolates suggested that the 1992 strain is a reassortant which inherited HE, P3, NP and M genes from a C/Mississippi/80-like virus and PB2, PB1 and NS genes from a C/pig/Beijing/115/81-like virus. Furthermore, it became evident that at least two (C/England/83 and C/Yamagata/9/88) of the 11 reference strains are also reassortants.

Measles case fatality by sex, vaccination status, and HIV-1 antibody in Zambian children.

Measles is more severe in HIV-1-infected children. However the long-term protective efficacy and safety of the measles vaccine in HIV-infected African children is unknown. A study was done at the University Teaching Hospital Lusaka Zambia to determine the impact of HIV-1 infection on the case fatality rate (CFR) in patients in hospital with measles. Children 9-59 months old with measles were studied between January 1993 and June 1995. History of measles vaccination was obtained from the childrens immunization cards. Serum samples were collected on admission and tested by ELISA and particle agglutination test. 356 patients (152 girls and 204 boys) were included in the analysis. The HIV-1 antibody-positive rate was 19% (68/356) and the overall CFR was 12% (43/356). CFRs were compared between vaccinated and unvaccinated children in each category stratified by sex and HIV-1 antibody status. In HIV-1 seronegative boys the CFR was significantly lower in vaccinated than in unvaccinated cases (odds ratio [OR] 0-13 p < 0.05). However no significant CFR reduction in vaccinated children was seen in HIV-1 seropositive boys (OR 0.68 p= 0.58) or HIV-1 seronegative girls (OR 0.92 p = 0.92). Surprisingly in HIV-1 seropositive girls the CFR was higher in the vaccinated than in the unvaccinated groups (OR 3.57 p = 0.18). Why HIV-1 seropositive and vaccinated girls had the highest CFR is not clear. However a prospective study in Senegal also showed a higher mortality rate in girls after high-titer measles vaccination and it has been suggested that high-titer measles vaccination induces immunological changes especially in girls. Although a trial of high-titer measles vaccine has never been done in Zambia the single-dose standard-titer Biken-CAM vaccine is generally given at the age of 9 months. Even a standard-titer measles vaccine might cause such immunological changes in HIV-1-infected children. Because CFR due to measles infection was examined a direct mechanism such as an adverse autoimmune response may be involved.

Rotavirus gastro-enteritis in hospitalized children with acute diarrhoea in Zambia

The clinical and epidemiological aspects of rotavirus diarrhoea were studied in hospitalized children with acute diarrhoea in Lusaka, Zambia. Two hundred and fifty-six (24.0%) of 1069 children admitted to the study were shedding rotavirus. The rotavirus-positive rate was highest in children less than 1 year of age (37.0%) and it was also high in those less than 6 months old. Rotavirus diarrhoea was seen throughout the year with a higher rotavirus-positive rate in the dry season. In rotavirus-positive diarrhoea patients, more children were dehydrated (82.4%) than in the rotavirus-negative group (56.2%). Rotavirus infection was more common in the children with normal nutritional status (27.6%, 162/588) than in those with malnutrition (19.3%, 93/482). The associated case fatality rate in the rotavirus-positive group was 6.4%, significantly less than in the rotavirus-negative group (OR 0.44, 95% CI 0.24-0.79), and mortality cases were seen only in children less than 2 years old.