Histomorphometric lung density evaluation of Immulina treatment using a murine influenza pneumonia model

Histomorphometric lung density measurements were used to evaluate the effects of Immulina on mouse pneumonia. Mice were intra-nasally exposed to H1N1 influenza virus at a dose of 5 × 10⁴ PFU/50 μL/mouse. Lung density was measured using the NIH ImageJ software program. Density values were compared to semiquantitative pneumonia severity scores. Lung photomicrographs were evaluated at 25-×, 40-× and 400-× magnification. The study included viral inoculated controls (IC) and non-inoculated controls (NC) and mice either treated or not treated with Immulina. Three doses of Immulina were included (25, 50 or 100 mg/kg) and administered using 3 protocols: prophylactic treatment (P), prodromal treatment (PD) and therapeutic treatment (TH) (note that in most of the evaluations of the data for the three treatment protocols were combined). Groups of mice were evaluated on days 3, 5, 7, 10 and 15 following exposure. The occurrence of "digital pneumonia" (DP) was defined as a density measurement above the 95% confidence limit of the corresponding NC values. A significant reduction in the occurrence of DP with Immulina treatment at the higher doses compared to IC was seen as early as day 3 and persisted up to day 15. There were also statistically significant dose-variable reductions in lung density in response to Immulina. The study suggests early administration of Immulina (P or PD protocols) may enhance resistance against influenza-induced viral pneumonia. A moderate correlation between pneumonia severity scores and lung density was observed for the 25-× and 40-× images (R = 0.56 and 0.53 respectively), and a strong correlation (R = 0.68) for 400-× images.