Graft-versus-host disease propagation depends on increased intestinal epithelial tight junction permeability
Sam C. Nalle, … , Peter A. Savage, Jerrold R. Turner
Sam C. Nalle, … , Peter A. Savage, Jerrold R. Turner
Published February 1, 2019; First published January 22, 2019
Citation Information: J Clin Invest. 2019;129(2):902-914. https://doi.org/10.1172/JCI98554.
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Research Article Gastroenterology Oncology

Graft-versus-host disease propagation depends on increased intestinal epithelial tight junction permeability

Abstract

Graft-versus-host disease (GVHD) is a complication of hematopoietic stem cell transplantation (HSCT) that affects multiple organs. GVHD-associated intestinal damage can be separated into two distinct phases, initiation and propagation, which correspond to conditioning-induced damage and effector T cell activation and infiltration, respectively. Substantial evidence indicates that intestinal damage induced by pretransplant conditioning is a key driver of GVHD initiation. Here, we aimed to determine the impact of dysregulated intestinal permeability on the subsequent GVHD propagation phase. The initiation phase of GVHD was unchanged in mice lacking long MLCK (MLCK210), an established regulator of epithelial tight junction permeability. However, MLCK210-deficient mice were protected from sustained barrier loss and exhibited limited GVHD propagation, as indicated by reduced histopathology, fewer CD8+ effector T cells in the gut, and improved overall survival. Consistent with these findings, intestinal epithelial MLCK210 expression and enzymatic activity were similarly increased in human and mouse GVHD biopsies. Intestinal epithelial barrier loss mediated by MLCK210 is therefore a key driver of the GVHD propagation. These data suggest that inhibition of MLCK210-dependent barrier regulation may be an effective approach to limiting GVHD progression.

Authors

Sam C. Nalle, Li Zuo, Ma. Lora Drizella M. Ong, Gurminder Singh, Alicia M. Worthylake, Wangsun Choi, Mario Cabrero Manresa, Anna P. Southworth, Karen L. Edelblum, Gregory J. Baker, Nora E. Joseph, Peter A. Savage, Jerrold R. Turner

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Figure 3

Allogeneic BMT fails to induce barrier dysfunction in MLCK–/– mice.

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Allogeneic BMT fails to induce barrier dysfunction in MLCK–/– mice. (A) ...
(A) Probes used to measure MLCK210-dependent barrier loss. (B) Intestinal permeability to fluorescein was evaluated on d14 after syngeneic (B6→WT) or allogeneic (129→WT or MLCK–/–) BMT. Values are normalized to the mean of WT mice that did not receive BMT. Each point represents an individual mouse. *P < 0.05, 2-tailed t test. (C and D) Intestinal permeability to 4-kDa dextran on d14 (C) or d35 (D) after BMT, normalized to the mean of WT mice without BMT. Each point represents an individual mouse. *P < 0.05, 2-tailed t test. (E and F) Jejunum harvested on d35 and immunostained for phosphorylated myosin light chain (pMLC, green) and E-cadherin (ECAD, red). Arrowheads denote perijunctional actomyosin ring. Note the preservation of myosin light chain phosphorylation within villus smooth muscle in MLCK–/– mice. Each point represents the average of 4 fields from one segment of tissue; 2 segments were analyzed per mouse. Data are normalized to the mean of WT mice without BMT. Scale bars: 50 μm (top), 10 μm (bottom). **P < 0.01, ANOVA with Bonferroni’s correction (vs. all other conditions). (G and H) 35 days after BMT, mice were injected with Alexa Fluor 647–BSA and sacrificed 30 minutes later. Confocal imaging demonstrates bright intravascular signal, indicative of retained dye, in WT and MLCK–/– mice without BMT or after syngeneic (B6→B6) BMT. In contrast, intravascular signal is markedly diminished in both WT and MLCK–/– mice after allogeneic (129→B6) BMT. Note the sharp paracellular pattern in MLCK–/– mice, consistent with preservation of the tight junction barrier, relative to the diffuse signal over the epithelium of WT mice (arrowheads). Scale bar: 25 μm. Each point represents an average of 6 regions of lamina propria from one mouse. Data are normalized to mean of WT mice without BMT. **P < 0.01, ANOVA with Bonferroni’s correction (vs. other conditions for each genotype; matched conditions across genotypes were not significantly different).