Biology subjects

Hough, R.

Publications and source records attributed to Hough, R..

2 recordsLinked to original sources

bioRxiv2019

Macrophage-specific NF-kappa B activation dynamics can segregate inflammatory bowel disease patients

The heterogeneous nature of inflammatory bowel disease (IBD) presents challenges, particularly when choosing therapy. Activation of the NF-{kappa}B transcription factor is a highly-regulated, dynamic event in IBD pathogenesis. We expressed the human NF-{kappa}B/p65 subunit in blood-derived macrophages, using lentivirus. Confocal imaging of p65 activation revealed that a higher proportion of macrophages from Crohns patients responded to lipid-A compared to controls. In contrast, cells from ulcerative colitis (UC) patients exhibited a shorter duration of p65 nuclear localisation compared to healthy controls and Crohns donors. Using a similar lentivirus approach, NF-{kappa}B-regulated luciferase was expressed in patient macrophages, isolated from frozen peripheral blood mononuclear cell samples. Following activation, samples could be segregated into three clusters based on the NF-{kappa}B-regulated luciferase response. The majority of UC samples appeared in hypo-responsive cluster 1, with Crohns patients representing the majority of hyper-responsive cluster 3. A positive correlation was seen between NF-{kappa}B-induced luciferase activity and cytokine levels released to medium from stimulated macrophages, but not in serum or biopsy. Analysis of macrophage cytokine responses and patient metadata revealed a strong correlation between Crohns patients who smoked and hyper-activation of p65. These in vitro dynamic assays of NF-{kappa}B activation in blood-derived macrophages segregate IBD patients into groups with different phenotypes and therefore may help determine response to therapy. Significance statementThis manuscript describes two dynamic assays of NF-{kappa}B activation in blood-derived macrophages that can segregate IBD patients into groups with different phenotypes. For the first time we introduce the use of dynamic measurements of a transcription factor activation as a method to stratify patients and we are confident that our approach will lead in future to early patient stratification and prediction of treatment outcome.

immunology↗

bioRxiv2017

Mice lacking NF-κB1 exhibit marked DNA damage responses and more severe gastric pathology in response to parenteral tamoxifen administration.

BackgroundTamoxifen (TAM) has recently been shown to cause acute gastric atrophy and metaplasia in mice. We have previously demonstrated that the outcome of Helicobacter felis infection, which induces similar gastric lesions in mice, is altered by deletion of specific NF-{kappa}B subunits. Nfkb1-/- mice developed more severe gastric atrophy than wild-type (WT) mice 6 weeks after H. felis infection. In contrast, Nfkb2-/- mice were protected from this pathology. We therefore hypothesized that gastric lesions induced by TAM may be similarly regulated by signaling via NF-{kappa}B subunits.\n\nMethodsGroups of 5 female C57BL/6 (WT), Nfkb1-/-, Nfkb2-/- and c-Rel-/- mice were administered 150mg/kg TAM by IP injection. 72 hours later, gastric corpus tissues were taken for quantitative histological assessment. In addition, groups of 6 female WT and Nfkb1-/- mice were exposed to 12Gy {gamma}-irradiation. Gastric epithelial apoptosis was quantified 6 and 48 hours after irradiation.\n\nResultsTAM induced gastric epithelial lesions in all strains of mice, but this was more severe in Nfkb1-/- mice than WT mice. Nfkb1-/- mice exhibited more severe parietal cell loss than WT mice, had increased gastric epithelial expression of Ki67 and had an exaggerated gastric epithelial DNA damage response as quantified by {gamma}H2AX. To determine investigate whether the difference in gastric epithelial DNA damage response of Nfkb1-/- mice was unique to TAM induced DNA damage, or a generic consequence of DNA damage, we also assessed gastric epithelial apoptosis following {gamma}-irradiation. 6 hours after {gamma}-irradiation, gastric epithelial apoptosis was increased in the gastric corpus and antrum of Nfkb1-/- mice.\n\nConclusionsNF-{kappa}B1 mediated signaling regulates the development of gastric mucosal pathology following TAM administration. This is associated with an exaggerated gastric epithelial DNA damage response. This aberrant response appears to reflect a more generic sensitization of the gastric mucosa of Nfkb1-/- mice to DNA damage.

immunology↗