Following OVA concern, all mice developed asthma-like disease. was itself immunogenic, inducing a uricase-specific adaptive immune response that occurred even when the enzymatic activity of uricase had been inactivated. Inhibition of the OVA-specific response was not due to the capacity of uricase to inhibit OVA uptake or processing and demonstration by dendritic cells, but at a later on step that inhibited OVA-specific CD4+ T cell proliferation and cytokine production. Whereas blocking uric acid formation by allopurinol did not affect results, administration of ultra-clean human being serum albumin at protein concentrations equivalent to that of uricase inhibited NO2-advertised sensitive airway disease. These results implicate that whereas uric acid levels are elevated in the airways of NO2-revealed mice, the powerful inhibitory effect of uricase administration on sensitive sensitization is definitely mediated more through antigen-specific immune deviation than on suppression of sensitive sensitization, a mechanism to be considered in the interpretation of results from additional experimental systems. NO2 exposure and NO2-advertised sensitive sensitization and concern For NO2 exposure, a single 1-hour dose of 15 ppm NO2 was given (12) and mice were analyzed at several times thereafter. Comparisons were made between mice exposed to NO2 or subjected to time in a similar exposure chamber through which HEPA-filtered space air flow was flowed. For NO2-advertised allergic sensitization, a single 1-hour exposure to 15ppm of NO2 on day time 1 was followed by 30 minutes of nebulized 1% OVA, Portion V (Sigma-Aldrich, St. Louis, MO) in saline, on days 1, 2, and 3 (29). All mice were OVA-challenged on days 14, 15, and 16, as explained (30). Analyses were performed at 48 hours after the final OVA challenge, on day time 18. Uricase, Galidesivir hydrochloride human being serum albumin, and allopurinol doses and delivery methods Recombinant uricase produced in was purchased from Sigma-Aldrich and delivered intranasally to isoflurane-anesthetized mice at 10 U per mouse in 40 l sterile saline. For some studies, uricase was inactivated by exposing 40 l/tube of 250 U/ml solutions in sterile saline to 254 nm UV light generated by a UV crosslinker (Stratalinker 1800, Stratagene, San Diego, CA) at a distance of 18 cm for 180 moments. Human being serum albumin (RMBIO, Missoula, MT) was delivered intranasally to isoflurane-anesthetized mice at 2 mg per mouse (equivalent to the protein content material of 10 U uricase) in 40 l sterile saline. Allopurinol (Sigma-Aldrich) was freshly dissolved in saline at 2.5 mg/ml and delivered sub-cutaneously at 25 mg/kg (31) 1 hour before and at 6 and 24, as well as with the allergic airway disease study at 48 hours, after NO2 inhalation. Assessment of airway responsiveness to methacholine Mice were anaesthetized with i.p. sodium pentobarbital (90 mg/kg), the trachea was cannulated, the mice were connected to a flexiVent? computer controlled small animal ventilator (SCIREQ, QC, Canada), and the mice were ventilated at 200 breaths/minute having a 0.25 ml Galidesivir hydrochloride tidal volume. Next, the mice were paralyzed with an i.p. injection of pancuronium bromide (0.8 g/kg). The animals were stabilized over about ten minutes of regular air flow at a positive end-expiratory pressure (PEEP) of 3 cmH2O. A standard lung volume Mela history was then founded by delivering two total lung capacity maneuvers (TLC) to a pressure limit of 25 cmH2O and holding for three mere seconds. Next, two baseline measurements of Galidesivir hydrochloride respiratory input impedance (Zrs) were obtained, followed by an inhalation of aerosolized PBS (control) for 10 mere seconds, achieved by an in-line piezo electric nebulizer (Aeroneb, Aerogen, Galway, Ireland). Zrs was then measured every 10 mere seconds for 3 minutes (18 measurements of Zrs in total). This total sequence of maneuvers and measurements was then repeated for aerosol exposures to four ascending doses of aerosolized methacholine (12.5, 25, 50, and 100 mg/ml). Data were fit.
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