Acinar ATP8b1/LPC pathway promotes macrophage efferocytosis and clearance of inflammation during chronic pancreatitis development.

Noninflammatory clearance of dying cells by professional phagocytes, termed efferocytosis, is fundamental in both homeostasis and inflammatory fibrosis disease but has not been confirmed to occur in chronic pancreatitis (CP). Here, we investigated whether efferocytosis constitutes a novel regulatory target in CP and its mechanisms. PRSS1 transgenic (PRSS1Tg) mice were treated with caerulein to mimic CP development. Phospholipid metabolite profiling and epigenetic assays were performed with PRSS1Tg CP models. The potential functions of Atp8b1 in CP model were clarified using Atp8b1-overexpressing adeno-associated virus, immunofluorescence, enzyme-linked immunosorbent assay(ELISA), and lipid metabolomic approaches. ATAC-seq combined with RNA-seq was then used to identify transcription factors binding to the Atp8b1 promoter, and ChIP-qPCR and luciferase assays were used to confirm that the identified transcription factor bound to the Atp8b1 promoter, and to identify the specific binding site. Flow cytometry was performed to analyze the proportion of pancreatic macrophages. Decreased efferocytosis with aggravated inflammation was identified in CP. The lysophosphatidylcholine (LPC) pathway was the most obviously dysregulated phospholipid pathway, and LPC and Atp8b1 expression gradually decreased during CP development. H3K27me3 ChIP-seq showed that increased Atp8b1 promoter methylation led to transcriptional inhibition. Atp8b1 complementation substantially increased the LPC concentration and improved CP outcomes. Bhlha15 was identified as a transcription factor that binds to the Atp8b1 promoter and regulates phospholipid metabolism. Our study indicates that the acinar Atp8b1/LPC pathway acts as an important "find-me" signal for macrophages and plays a protective role in CP, with Atp8b1 transcription promoted by the acinar cell-specific transcription factor Bhlha15. Bhlha15, Atp8b1, and LPC could be clinically translated into valuable therapeutic targets to overcome the limitations of current CP therapies.

Notch signaling downregulates MUC5AC expression in airway epithelial cells through Hes1-dependent mechanisms.

BACKGROUND: Mucus overproduction is one of the major pathological features of asthma, and MUC5AC is the major mucin component of airway mucus. However, whether Notch signaling is implicated in the regulation of MUC5AC expression in airway secretary cells is still undetermined. OBJECTIVE: The aim of this study is to examine whether Notch signaling can regulate MUC5AC expression and explore the molecular mechanisms. METHODS: Mouse mtCC1-2 cells and human NCI-H292 cells were transfected with NIC, and MUC5AC expression was examined. Using gene reporter assays, site-directed mutagenesis, and ChIP assays, the activity of both mouse and human MUC5AC promoter was analyzed. RESULTS: Notch signaling regulated MUC5AC expression both in mouse mtCC1-2 cells and in human NCI-H292 cells. Several Hes-binding site N-boxes were identified in the 5' region of both mouse and human MUC5AC promoters. Overexpression of NIC resulted in activation of the MUC5AC promoter. Site-directed mutagenesis report assays revealed that Hes proteins might repress both mouse and human MUC5AC promoter activity. Furthermore, ChIP assays confirmed that Hes1 binds to the MUC5AC promoter both in mouse mtCC1-2 cells and in human NCI-H292 cells. CONCLUSIONS: Notch signaling can directly downregulate MUC5AC promoter activity through Hes1-dependent mechanisms, which may be identified as possible targets for pharmacotherapy of airway mucus hypersecretion in asthma.