Thirty-eight-negative kinase 1 mediates trauma-induced intestinal injury and multi-organ failure
Milena Armacki, … , Alexander Kleger, Thomas Seufferlein
Milena Armacki, … , Alexander Kleger, Thomas Seufferlein
Published October 15, 2018
Citation Information: J Clin Invest. 2018;128(11):5056-5072. https://doi.org/10.1172/JCI97912.
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Research Article Gastroenterology Inflammation

Thirty-eight-negative kinase 1 mediates trauma-induced intestinal injury and multi-organ failure

Abstract

Dysregulated intestinal epithelial apoptosis initiates gut injury, alters the intestinal barrier, and can facilitate bacterial translocation leading to a systemic inflammatory response syndrome (SIRS) and/or multi-organ dysfunction syndrome (MODS). A variety of gastrointestinal disorders, including inflammatory bowel disease, have been linked to intestinal apoptosis. Similarly, intestinal hyperpermeability and gut failure occur in critically ill patients, putting the gut at the center of SIRS pathology. Regulation of apoptosis and immune-modulatory functions have been ascribed to Thirty-eight-negative kinase 1 (TNK1), whose activity is regulated merely by expression. We investigated the effect of TNK1 on intestinal integrity and its role in MODS. TNK1 expression induced crypt-specific apoptosis, leading to bacterial translocation, subsequent septic shock, and early death. Mechanistically, TNK1 expression in vivo resulted in STAT3 phosphorylation, nuclear translocation of p65, and release of IL-6 and TNF-α. A TNF-α neutralizing antibody partially blocked development of intestinal damage. Conversely, gut-specific deletion of TNK1 protected the intestinal mucosa from experimental colitis and prevented cytokine release in the gut. Finally, TNK1 was found to be deregulated in the gut in murine and porcine trauma models and human inflammatory bowel disease. Thus, TNK1 might be a target during MODS to prevent damage in several organs, notably the gut.

Authors

Milena Armacki, Anna Katharina Trugenberger, Ann K. Ellwanger, Tim Eiseler, Christiane Schwerdt, Lucas Bettac, Dominik Langgartner, Ninel Azoitei, Rebecca Halbgebauer, Rüdiger Groß, Tabea Barth, André Lechel, Benjamin M. Walter, Johann M. Kraus, Christoph Wiegreffe, Johannes Grimm, Annika Scheffold, Marlon R. Schneider, Kenneth Peuker, Sebastian Zeißig, Stefan Britsch, Stefan Rose-John, Sabine Vettorazzi, Eckhart Wolf, Andrea Tannapfel, Konrad Steinestel, Stefan O. Reber, Paul Walther, Hans A. Kestler, Peter Radermacher, Thomas F.E. Barth, Markus Huber-Lang, Alexander Kleger, Thomas Seufferlein

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Figure 3

Mice with forced TNK1 expression show massive apoptosis at the base of the intestinal crypts.

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Mice with forced TNK1 expression show massive apoptosis at the base of t...
(A–C) Graphs display elevated mRNA levels of proapoptotic (Bax) and prosurvival (Xiap, Bcl2) genes in the small and large intestine (n = 10 per group) of TNK1-expressing mice. (D) Western blots show expression of cC3 and its substrate cPARP in the small intestine and colon (n = 3). (E and F) Representative images of the murine small and large intestine show immunoreactivity for the apoptotic marker cC3 (arrowheads point to cC3-positive cells). Corresponding charts depict positional quantification of apoptosis (n = 6 per group), which was done according to Buczacki et al. (61) by counting of cells at the crypt (stem cell compartment) and villi. Cells were counted at the base of the crypt (from positions 0 to +4, counting from the bottom of the crypt to the transit-amplifying progenitor cells and the villus area). (G) Expression of the stem cell marker LGR5 in the small and large intestine is also reduced in Tnk1-expressing mice as indicated by quantitative reverse transcriptase PCR (RT-qPCR) analysis (n = 5 per group). (H) Representative images of the crypt-base stem cell compartment positive for cC3 (top panel: white arrows indicate stem and reserve stem [position +4] cells positive for apoptotic marker) or the Ki67 proliferative marker (bottom panel). (I) Crypt phenotype was quantified by counting of regenerating crypts, which are defined as containing at least 5 adjacent Ki67+ cells (arrows) contained within a crypt-like structure (34) (n = 10–12 per group). All analyses were performed 24 hours after doxycycline or saline treatment. Data are expressed as mean ± SEM. Differences were tested by parsametric 2-tailed, unpaired Student’s t tests. ANOVA test was applied for multiple-comparison analysis. The mean of each column was compared with the mean of a control column by Dunnett’s multiple-comparisons test. (*P = 0.01–0.05; **P = 0.001–0.01; ***P = 0.0001–0.001; ****P < 0.0001.) Scale bars: 50 (H); 100 μm (E and F).