Inhalational exposures to pathogenic microbes and microbial byproducts shape the pulmonary immune responses. Although the memory-based acquired immune responses play a pivotal role, the immunologic memory developed by innate immune cells have also been shown to contribute to the host-beneficial ?trained? responses. We propose that the airway epithelial cells (AECs), crucial lung innate immune cells, also undergo functional reprograming and contribute to this mucosal immunity. The overreaching goal of these studies is to understand the genetic and epigenetic regulation of the airway hyperreactivity observed in allergic asthma by focusing on the mucous responses. We developed and characterized the allergic asthma relevant 3D in-vitro model of the hyperreactive mucous response using differentiated human AECs. There was an augmented and rapid mucous response within the 24 h of the secondary challenge with significantly elevated levels of secretory mucin and the mucin regulatory transcriptional factors. To help understand the changes in genetic mechanism(s) responsible for this mucous hyperreactivity, a high throughput RNA sequencing analyses was conducted. We identified novel putative long non-coding RNAs (lncRNAs) that were significantly regulated following challenge. Based on the expression and the potential in gene regulation and modulating airway inflammation, three lncRNAs were selected for further analyses. One the identified lncRNA was also upregulated in asthmatics AECs compared to controls. Recent findings have highlighted the emerging role of noncoding RNAs in the regulation of airway inflammation and remodeling. However, there are still a handful of investigations on the lncRNA- associated regulatory mechanism in asthma and the characterization of these newly identified lncRNAs in the proposed studies could help pioneer this field of investigation. Our preliminary data suggest that one of the identified lncRNA might be an important regulator of airway mucous response that could conceivably modulate the airway inflammation and play a role in allergic asthma development. Here, we propose to characterize these novel lncRNAs in human AECs from asthmatic and control subjects using an asthma-relevant experimental model.
In Aim 1, we propose to validate and characterize these putative lncRNAs and the interacting factors will be analyzed by pull-down assays and the functional significance of these interactions will be determined by genetic editing.
In Aim 2, we will test whether the hyperreactive mucous response of asthmatic AECs can be regulated by modulating these lncRNAs using gain- and loss-of-function studies. The research outlined in this proposal will thus provide a key foundation toward understanding the molecular mechanisms by which lncRNAs modulate airway epithelial responses and probably help develop the novel strategies to treat or diagnose diseases characterized by hyperreactive mucous responses.
Allergic asthmatics suffer from hyperreactive mucous responses that could block airways and affect lung functions resulting in associated comorbidities and mortalities. The proposed studies are based on the novel mechanistic findings that the airway epithelial cells undergo a functional reprograming to modulate mucous responses and these mechanisms are dysregulated in asthmatics. Therefore, the proposed research could help provide a rationale for developing novel strategies that can be exploited to alleviate mucus hyperreactivity and asthmatic exacerbations.
