ls. We therefore verified in control groups of mice that the number of eosinophils present 48 hrs after the last Ova aerosol remained significantly elevated in SP-A2/2 mice as compared to WT mice, as PubMed ID:http://www.ncbi.nlm.nih.gov/pubmed/22189597 has been previously published. The finding that eosinophils are decreased in the Ova only groups by day 28 of our model, as compared to the previously published findings on day 24, suggests that eosinophilia is resolving in the Ova only control groups of WT and SP-A2/2 mice. Lungs were harvested from three groups of WT and SP-A null mice: 1) saline treated control mice, 2) Ova sensitized/challenged mice, 3) and Ova sensitized/challenged and Mp infected mice. To determine if SP-A regulates eosinophil activation to Mp infection in the allergic mice, markers of eosinophils, such as IL-5 and eosinophil associated ribonuclease, were analyzed by RTPCR. In Ova allergic WT and SP-A2/2 mice, the levels of IL-5 and EAR RNA were dramatically but comparably increased over levels detected in samples taken from saline treated mice, suggesting similar numbers eosinophils were present in the lungs of WT and SP-A2/2 mice 5 days post Ova challenge. Interestingly, EAR expression, a marker typically associated with eosinophil activation, is significantly increased in the WT Ova+Mp compared to the WT Ova alone. This increase in EAR expression suggests that while the total eosinophil number is similar between the WT groups, the 
activation status of the eosinophils, as determined at the transcriptional level, in the WT Ova+Mp group is greater than in the WT Ova only. Additionally, when Ova allergic SP-A2/2 mice are infected with Mp, the levels of IL-5 and EAR RNA significantly increased over levels detected in the Ova allergic WT infected mice. This suggests that not only are more eosinophils present in the SP-A2/2 Ova+Mp mice, but also that SP-A may reduce the activation of eosinophils at the transcriptional level. Lung sections were analyzed from Ova sensitized/challenged Mp infected SP-A2/2 and WT mice by histochemical staining for cyanide resistant EPO activity. While the majority of EPO activity staining is observed in the lung parenchyma in the allergic infected WT mice, more staining was evident around the large airways of allergic infected SP-A2/2 mice, in close proximity to where we and others have shown Mp to colonize . Additionally, EPO activity was measured in both BAL and lung tissue from Mp-infected allergic mice. The total EPO activity measured in the infected allergic mice lacking SP-A was significantly greater than that in the control WT mice given the same treatment. Binding of SP-A to eosinophils specific cell surface markers, we found that there were significantly more macrophages, CD11b+ exudative macrophages, inflammatory monocytes, dendritic cells, neutrophils and eosinophils present in BAL of allergic/infected SP-A null mice as compared to WT mice, further suggesting an important regulatory role of SP-A in this allergic/10338-51-9 chemical information infectious model. Interestingly, we found no eosinophils in our previous studies examining Mp infection in a non-allergic airway lacking SP-A, although we did see increases in the other cell populations. Samples from the Ova+Mp experimental model were collected on day 28, the time at which airway physiology In order to assess the possibility that SP-A acts directly on eosinophils to modulate their responses, we sought to determine if SP-A binds to eosinophils. Eosinophils were purified from the blood of IL-5 transgenic mice
