During the past year we have performed more experiments for the proposed studies. We have found that Tlr4 expression in alveolar type II cells was down regulated following bleomycin lung injury in a similar pattern to the damage and renewal of AEC2s after lung injury. Reduction of IL-6 production results from Tlr4 or Has2 deficiency on AEC2s. IL-6 recombinant protein treatment increased AEC2 proliferation, renewal, and reduced epithelium permeability of bleomycin injured Tlr4-/- mice in vivo. Exogenous HA (healon) promoted colony formation of AEC2 from SFTPC-Cre;Has2flox/flox mice and HA blocking peptide (pep 1) was able to block healon- promoted colony formation.
Furthermore, we have make good progress on the study of AEC2 stem cells in IPF patient lungs. we observed decreased HAS2 expression in IPF AEC2s. TLR4 expression of AEC2s from IPF lung and normal donor lungs were at similar level. High dose of exogenous HA was able to increase colony formation of IPF AEC2s. Our data indicate that there is failure of AEC2 stem cell renewal due to loss cell surface HA in IPF lung.