SESSION TITLE: Cytokines/Cellular Interactions Posters
SESSION TYPE: Original Investigation Poster
PRESENTED ON: Wednesday, October 30, 2013 at 01:30 PM - 02:30 PM
PURPOSE: Acute lung injury (ALI) is characterized by acute lung inflammation and disruption of the alveolar epithelial barrier. The integrity of the barrier is dependent on cell adhesion proteins, mainly tight junctions.PKC-ζ, an atypical protein kinase, which is abundant in the lung, has been shown to affect alveolar epithelial tight junction integrity. Therefore, we hypothesize that PKC-ζ is activated in ALI and its inhibition will preserve the alveolar epithelial barrier integrity and promote lung repair
METHODS: To test our hypothesis we instilled C57/Bl6 mice with bleomycin (4U/kg) and used a myristoylated PKC-ζ pseudosubstrate (ps) inhibitor to block PKC-ζ activation. Mice were harvested at one, three and six weeks. Bronchial alveolar lavage (BAL) was performed from which we determined cell count and diff, IL-1β, and protein concentration. Immunofluorescence staining and western blot were performed on lung tissues to detect PKC-ζ activation. Tissue samples were harvested to measure hydroxyproline as well as perform H&E and massons staining. Lung mechanics were assessed at 1 week, to measure static compliance and resistance. Computed micro-tomography (micro-CT) was performed at 6 weeks to quantify the amount of fibrosis in the bleomycin and bleomycin/PKC-ζ ps cohorts. In addition, primary rat alveolar epithelial cells were harvested and treated with bleomycin and PKC-ζ inhibitor in an in vitro model. Transepithelial electrical conductance, cell death and western blot analysis were performed in these conditions.
RESULTS: Bleomycin activates PKC-ζ in vivo and in cell culture. Treatment with PKC-ζ inhibitor prevented the activation of PKC-ζ. Both bleomycin and bleomycin/PKC-ζ ps groups induced inflammation; however PKC-ζ inhibitor prevented bleomycin increase in BAL protein concentration. This reflects preservation of the barrier. In the bleomycin group, fibrosis was detected by H&E stains and increased collagen deposition using massons stain. Hydroxyproline and TGF-β were also increased. Although mice treated with bleomycin and PKC-ζ pseudosubstrate showed a similar increase in fibrosis at three weeks , there was complete fibrosis resolution by six weeks in the latter group compared to bleomycin alone.
CONCLUSIONS: Inhibition of PKC-ζ preserves lung epithelial barrier and accelerates lung repair
CLINICAL IMPLICATIONS: PKC-ζ plays an important role in the disruption of the alveolar epithelial barrier and is a potential therapeutic target in patients with ALI.
DISCLOSURE: The following authors have nothing to disclose: Mosaab Bagegni, Jennifer Borcherding, Jessica Sieren, Luis Vargas, Alejandro Comellas
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