Probes for TLR2

The pathogenesis of gastroesophageal reflux disease (GERD) is not fully understood. It involves the activation of mucosal immune-mediated and inflammatory responses. Toll-like receptors (TLR) 2 and TLR4 are pattern-recognition receptors of the innate immune system; they recognize microbial and endogenous ligands. Farnesoid X receptor (FXR) is a bile acid receptor that regulates the inflammatory response. We aimed to evaluate TLR2, TLR4 and FXR expression patterns in GERD. We re-evaluated 84 oesophageal biopsy samples according to the global severity (GS) score, including 26 cases with histologically normal oesophagus, 28 with histologically mild oesophagitis and 30 with severe oesophagitis. We used immunohistochemistry and in situ hybridization to assess the expression patterns of TLR2, TLR4 and FXR in oesophageal squamous cells. Immunohistochemistry showed that nuclear and cytoplasmic TLR2 was expressed predominantly in the basal layer of normal oesophageal epithelium. In oesophagitis, TLR2 expression increased throughout the epithelium, and the superficial expression was significantly more intensive compared to normal epithelium, p <0.01. Nuclear and cytoplasmic TLR4 was expressed throughout the thickness of squamous epithelium, with no change in oesophagitis. FXR was expressed in the nuclei of squamous cells, and the intensity of the expression increased significantly in oesophagitis (p <0.05). FXR expression correlated with basal TLR2. In situ hybridization confirmed the immunohistochemical expression patterns of TLR2 and TLR4. In GERD, TLR2, but not TLR4, expression was upregulated which indicates that innate immunity is activated according to a specific pattern in GERD. FXR expression was increased in GERD and might have a regulatory connection to TLR2.

Sudden, unpredictable, severe acute pain episodes are the most common sickle cell disease (SCD) complication. Some SCD patients experience frequent pain episodes while others experience rare episodes. Knowledge of the biology driving this variability is limited. Using gene transcription analyses, we previously showed an elevated inflammatory response is associated with increased SCD pain episode frequency. We sought to replicate these findings in a larger SCD cohort and identify hub genes closely associated with increased pain frequency. We conducted plasma-induced transcription analyses in 132 SCD patients (baseline health) and 60 Black controls (4-21 years, both groups). 3028 differentially expressed genes between SCD patients and controls were retained for subsequent analyses with Weighted Gene Co-Expression Network Analysis (WGCNA). WGCNA was used to define modules (functionally grouped genes) and we correlated these modules with number of pain episodes requiring health care utilization in prior three years. Of 11 identified modules, four showed significant correlation with number of pain episodes. Database for Annotation, Visualization, and Integrated Discovery (DAVID) was used for ontological analysis of the four significant modules and key biological processes identified were inflammatory response and cellular response to lipopolysaccharide. Cytoscape was used to construct a protein-protein interaction network and the 10 top hub genes identified in hierarchical order were: TNF, CCR5, CCR1, CCL2, CXCL2, ITGAM, CCL7, CXCL3, TLR2 and MMP9. These genes, as part of the inflammatory response, support the immune system contributes to increased pain episode frequency. Identified hub genes may be leveraged as therapeutic targets for reducing SCD pain episodes. 1R61NS114954-01.