Masanori Terajima

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.

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.

Effect of Genetic Risk Factors and Disease Progression on the Cerebrospinal Fluid Tau Levels in Alzheimer's Disease

OBJECTIVE: This study was undertaken to gain insights into the clinical utility of measuring cerebrospinal fluid tau protein (CSF‐tau) to aid in the diagnosis of Alzheimers disease (AD).

Pupil dilatation assay by tropicamide is modulated by apolipoprotein E epsilon 4 allele dosage in Alzheimer's disease.

The mydriatic response to dilute tropicamide was studied in 25 Japanese patients with Alzheimers disease (AD), 13 patients with non-AD neurological diseases (non-AD) and 11 normal elderly‘ subjects (control). Although the changes in resting pupil diameter and area over baseline were significantly greater (p < 0.05) in AD patients than in non-AD patients and controls, there was considerable overlap between the three groups. The change in resting pupil area was significantly greater (p < 0.05) in AD patients homozygous for ApoE 4 than in AD patients heterozygous for or without this allele. Despite a limited sample size in the present study, our results indicate that the pupil dilation assay by tropicamide is not an effective diagnostic tool for AD, and it may be modulated by different gene dosage of ApoE 4.

ELEVATED CEREBROSPINAL FLUID TAU LEVELS: IMPLICATIONS FOR THE EARLY DIAGNOSIS OF ALZHEIMER'S DISEASE

Hiroyuki Arai, MD, PhD

Dipeptidase inhibitor and epithelial removal potentiate leukotriene D4-induced human tracheal smooth muscle contraction

We investigated the role of epithelium in smooth muscle contraction induced by leukotriene D4 (LTD4) in isolated human trachea. The contractile response to LTD4 was potentiated by an inhibitor of dipeptidases L-cysteine and by removal of the epithelium. Both L-cysteine (3 x 10(-3) M) and removal of the epithelium shifted the concentration-response curves to LTD4, to lower concentrations by 0.7 and 0.6 log units, respectively. Incubation of cultured or isolated human tracheal epithelial cells with LTD4 resulted in the formation of LTE4, which was completely blocked by pretreatment with L-cysteine (3 x 10(-3) M). The isolated and cultured human tracheal epithelial cells contained microsomal dipeptidase (MDP) activity. Immunohistochemical study indicated MDP protein was present in the epithelium and endothelial cells of submucosal microvessels in the human trachea. These results suggest that the epithelium modulates the contractile response to LTD4 in human trachea by dipeptidases degrading LTD4.

Regulatory Function of Delta/YY-1 on the Locus Control Region-like Sequence of Mouse Glycophorin Gene in Erythroleukemia Cells

The far upstream region (−1.2-0.9 kilobase pairs) of the mouse glycophorin gene contains the locus control region (LCR)-like region, which acts as an erythroid-specific enhancer dependent on chromosomal integration in murine erythroleukemia (MEL) cells. In the present study, we demonstrated that this region binds six nuclear factors. The binding of GATA-1 to corresponding sites did not show any change before or after induction with dimethyl sulfoxide, but the binding of Spi-1/PU.l and an unidentified factor called glycophorin regulatory element binding factor (GRBF) showed a change during induction. While binding activity of Spi-l/PU.l dropped soon after induction, the GRBF activity increased after induction when expression of the glycophorin gene began. After identification of the consensus binding site of GRBF, we cloned cDNA for that factor by Southwestern method, and it was identified as a previously reported transcription factor, delta, a murine form of YY-l which is a versatile transcription factor. Mutation analysis in the delta/YY-1 binding site within the LCR-like region indicated that delta/YY-1 acts as a regulatory protein in combination with the E-box-binding protein that binds to the neighboring sequence.