Butyrate attenuates house dust mite-induced activation of 16HBE bronchial epithelial cells in association with modulation of gene expression

The airway epithelial barrier is the primary defense line of the lungs against allergens, such as house dust mite (HDM). A disrupted airway epithelial barrier plays a major role in asthma development. Short-chain fatty acids (SCFAs) are bacterial fermentation products that are associated with prevention against allergic inflammation. In this study, we aim to elucidate the direct preventative effects of SCFAs, particularly butyrate, on the HDM-induced bronchial epithelial cell (BEC) activation and barrier disruption in-vitro. Immortalized human 16HBE cells were exposed to HDM to simulate allergen-induced BEC inflammation. Pre-exposure of 1-1000 µM acetate, propionate or butyrate were studied for their ability to prevent epithelial activation. Histone deacetylase activity of cells was measured after pre-incubation with 100 or 200 µM butyrate and combined butyrate and HDM exposure. In a transwell, air-liquid interface (ALI)-culture, the effects of 100 µM butyrate on barrier integrity (transepithelial resistance (TEER) and 4 kDa FITC-dextran permeability) and inflammation were assessed in presence and absence of HDM. To gain broader insight in the effects of butyrate, RNA sequencing (RNAseq) was performed using 16HBE cells exposed to HDM, butyrate or both. Pre-incubation with butyrate protected against the production of HDM-induced IL-8 (1-100 µM), CCL5 (1-100-1000 µM) and CCL20 (1-10-100 µM) by solid phase cultured 16HBE-cells, although 1000 µM butyrate increased IL-8 and CCL20 release. Pre-incubation with acetate and propionate showed similar effects. 100 µM butyrate did not suppress HDAC activity, while a dose of 200 µM butyrate did. In ALI-culture, HDM increased barrier permeability, and pre-incubation with 100 µM butyrate tended to prevent this. Additionally, HDM-induced CCL20 production was significantly decreased by butyrate. RNAseq revealed 100 µM butyrate to significantly affect biological processes of the cells linked to gene expression, mitochondrial function, cell division and cell signaling. These effects were observed both in absence and presence of HDM-exposure. Butyrate, at physiologically relevant doses, modulates gene expression of 16HBE cells, affecting amongst others post-transcriptional and mitochondrial pathways. In ALI-cultures, butyrate prevented epithelial activation (CCL20) and tended to prevent HDM-induced barrier defects. These findings highlight the relevance of SCFAs such as butyrate in maintaining bronchial epithelial cell homeostasis relevant for prevention and control of allergic airway disease.