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For the investigation of expansion of lung macrophage numbers post-hatch following in ovo CpG DNA delivery, we quantified the macrophages in lungs post-hatch using flow cytometry technique. We found that in ovo CpG DNA treatment significantly increased the macrophage numbers post-hatch when compared to that in non-CpG DNA treated controls chickens (Figure 4, p = 0.0041). Figure 4 In ovo delivery of CpG DNA expands macrophage AZD2014 numbers in lungs post-hatch. (a�Cd) indicate the representative fluorescence-activated cell sorting diagrams showing unstained control, iso type control, percentage of KUL01+ macrophages in lungs post-hatch ... 2.4. In Ovo Delivery of CpG DNA Induces Protection against ILTV Caused Morbidity and Mortality Reducing Viral Replication in the Respiratory Tract of Chickens Since we observed that in ovo delivered CpG DNA was capable of increasing lung macrophages post-hatch, we then hypothesized that the chickens that received in ovo CpG DNA leading to increased macrophages post-hatch may be able elicit protective response against ILTV infection encountered post-hatch. For this part of the study, we in ovo delivered either CpG DNA or PBS, allowed the eggs to hatch and then infected the hatched chickens intratracheally with ILTV on the day of hatch. We found that in ovo delivered CpG DNA (1) decreases clinical signs associated with post-hatch ILTV infection (p allobarbital decreases mortality associated with post-hatch ILTV infection (p JNK inhibitor cost reducing viral replication in the respiratory tract of chickens. SPF eggs were injected with CpG DNA or PBS in ovo, allowed to hatch and hatched chickens were infected ... 2.5. In Ovo Delivery of CpG DNA Does Not Affect the Hatchability of Eggs We recorded hatchability rates of 86% and 75% following in ovo delivery of CpG DNA and PBS, respectively. 3. Discussion Our current investigations led to three major findings. Firstly, CpG DNA decreases ILTV replication in lungs pre-hatch correlating with expansion of macrophage populations and increase in mRNA expression of IL-1�� in lungs. Secondly, we found that avian macrophages could be stimulated with CpG DNA in vitro to produce NO and increase in mRNA expression of iNOS and IL-1�� genes.