Mechanisms Of An Acute Otitis Media Model Induced By Intranasal With Streptococcus Pneumoniae And Secondary Infection With Non-typeable Haemophilus Influenzae

ObjectiveEstablish an acute otitis media model with the method of inoculateStreptococcus pneumoniae by intranasal first, then inject the Nontypeablehaemophilus influenzae into the middle ear cavity. And study themechanism of secondary infection in this model.Methods1. Take the C57BL/6J mice inoculated NTHi of different concentration(106,107,108CFU/ear) in middle ear cavity via tympanic membrane as theexperiment group, the others inoculated with PBS were treated as the groupcontrol. After inoculation, weigh the mice and observe them daliy. Micewere sacrificed at the first, third, fifth and seventh day post inoculation fortheir blood and middle ear lavage fluids. The cell from MELF werecollected for flow cytometry. HE staining of tissue sections with mice weremonitored. 2. Inoculate the S.pn19F into C57BL/6J by intranasal for colonizationof S.pn in nasal. Then inject the NTHi into the middle ear cavity throughtympanic membrane. After completing the secondary infection model,observe the mice weight, bacteria loads, cell count and the change ofcytokines level.Results1. Weight: the PBS group loss their weight at Day1, then grow upgradually. Meanwhile the grow up of the group NTHi’s weight start at Day3. Neutrophils occupy even99%of the cell counts in the MELF by flowcytometry, and the cell’s number of group control is lower. We also canobserve the inflammatory cell’s infiltration in the middle ear cavity and thebone wall’s destruction there where on the middle ear tissue sections.Cytokines in serum and MELF: the cytokines which include IL-6andTNF-alpha taken from group NTHi’s MELF is higher. But the level ofcytokines in the serum keeps the same.2. Weight: the group coIF has a remarkable loss weight. But this doesnot do a significant affect to the weight loss post inoculate. The clearanceof the bacterial has no difference between two groups. But the group coIFrecruit more neutrophils to the middle ear cavity while the expression ofIL-1beta and TNF-alpha is higher than group NTHi, too. C onclusionWe established the mice AOM inoculated by NTHi and the AOM ofsecondary infection inoculated by S.pn and NTHi, determined the level ofcytokines in both the models as well. This lay a foundation for study of theinteraction between microbe to microbe, multi-microbe to host.