Date of Award

9-21-2009

Document Type

Thesis

Degree Name

Medical Doctor (MD)

First Advisor

Jeffrey S. Kahn

Second Advisor

Richard Martinello

Third Advisor

Jack Elias

Abstract

DIFFERENTIAL INDUCTION OF INTERLEUKIN-6 BY CLINICAL ISOLATES OF RESPIRATORY SYNCYTIAL VIRUS Rachel L. Wattier, Isaac Lazar, Richard A. Martinello, Carla Weibel, Jeffrey S. Kahn. Division of Infectious Diseases, Department of Pediatrics, Department of Epidemiology and Public Health, Yale University School of Medicine, New Haven, CT. This study was conducted to determine whether genetic variation between naturally circulating respiratory syncytial virus (RSV) isolates results in different capacities to elicit innate immune responses in human pulmonary epithelial cells. We hypothesize that genetically distinct RSV isolates interact with the immune regulatory mechanisms in human pulmonary epithelial cells to elicit different patterns of cytokine expression. Human pulmonary epithelial cells (A549) were infected with 36 genetically distinct RSV clinical isolates. Induction of interleukin (IL)-1α, IL-6 and tumor necrosis factor (TNF)-α was measured by ELISA up to 48 hours post-infection. Cytokine mRNA expression and replication kinetics were measured by quantitative RT-PCR. Two genetically distinct but closely related isolates were selected for further investigation, based on differences between these isolates upon initial comparison of cytokine induction. These isolates were found to induce markedly different levels of IL-6 beginning at 6 hours post-infection and continuing up to 48 hours post-infection. Isolate NH/GB4/1125/01-02 induced up to 13-fold higher levels of IL-6 at 24 hours post-infection than did isolate NH/GB1/1067/01-02. These differences in IL-6 secretion were accompanied by significant differences in IL-6 mRNA expression. The two isolates had similar replication kinetics. Neither isolate induced IL-1α or TNF-α when A549 cells were infected with purified viral preparations. We conclude that RSV clinical isolates differ from one another in their capacity to induce IL-6. The different levels of IL-6 secretion are due in part to differences in transcriptional regulation. Different levels of IL-6 induction cannot be attributed to differences in replication kinetics. Phenotypic differences in the capacity to induce innate immune responses may account for the wide variation in clinical manifestations of RSV infection in children. Further investigation of these and other RSV isolates with phenotypic differences will identify genetic markers related to cytokine induction by RSV. This will enhance understanding of RSV virulence factors and may identify novel therapeutic targets for RSV infection.

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