Selective activation of T helper cell (Th) subsets affects disease severity in RSV infection. Animal studies have shown by Northern blot analysis that Th type 2 response to formalin-inactivated RSV vaccine is responsible for enhanced pathology in RSV disease. We developed a quantitative RT-PCR to characterize the cytokine response following primary and secondary RSV infection. Methods: Retired breeder BALB/c mice were intranasally inoculated with 106.5 plaque-forming units of RSV A2 on days 0 and +28. At various times after secondary exposure, RNA was extracted from the left lung of 2 animals and cytokine levels determined by RT-PCR. Lung RNA was reverse transcribed to complimentary DNA (cDNA) using oligo-dT primers and PCR amplified with cytokine-specific primers in the presence of increasing concentrations of primer-specific competitive fragment (CF). cDNA levels were determined from the concentration of CF at the point of equivalence of cDNA and CF on ethidium bromide stained agarose gel. Results from each time point were averaged. All samples were normalized to β-actin prior to cytokine quantitation. Results: γ-IFN was detected after primary and secondary infection, the amount increasing with time after secondary exposure. No or trace amounts of IL-2 or IL-4 was detected by this sensitive technique. Conclusion: Using this model, cytokine response to RSV infection and reinfection was quantified. As expected, primary and secondary RSV infection were associated with Th type 1 responses. Our model may be used to assess vaccines, antiviral and antiinflammatory agents for the treatment and prevention of RSV disease. Table

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