Microbiota-dependent signals are required to sustain TLR-mediated immune responses
Lehn K. Weaver, Danielle Minichino, Chhanda Biswas, Niansheng Chu, Jung-Jin Lee, Kyle Bittinger, Sabrin Albeituni, Kim E. Nichols, Edward M. Behrens
Lehn K. Weaver, Danielle Minichino, Chhanda Biswas, Niansheng Chu, Jung-Jin Lee, Kyle Bittinger, Sabrin Albeituni, Kim E. Nichols, Edward M. Behrens
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Research Article Inflammation

Microbiota-dependent signals are required to sustain TLR-mediated immune responses

Abstract

Host-commensal interactions are critical for the generation of robust inflammatory responses, yet the mechanisms leading to this effect remain poorly understood. Using a murine model of cytokine storm, we identified that host microbiota are required to sustain systemic TLR-driven immune responses. Mice treated with broad-spectrum antibiotics or raised in germ-free conditions responded normally to an initial TLR signal but failed to sustain production of proinflammatory cytokines following administration of repeated TLR signals in vivo. Mechanistically, host microbiota primed JAK signaling in myeloid progenitors to promote TLR-enhanced myelopoiesis, which is required for the accumulation of TLR-responsive monocytes. In the absence of TLR-enhanced monocytopoiesis, antibiotic-treated mice lost their ability to respond to repeated TLR stimuli and were protected from cytokine storm–induced immunopathology. These data reveal priming of TLR-enhanced myelopoiesis as a microbiota-dependent mechanism that regulates systemic inflammatory responses and highlight a role for host commensals in the pathogenesis of cytokine storm syndromes.

Authors

Lehn K. Weaver, Danielle Minichino, Chhanda Biswas, Niansheng Chu, Jung-Jin Lee, Kyle Bittinger, Sabrin Albeituni, Kim E. Nichols, Edward M. Behrens

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Figure 7

JAK signaling is altered in myeloid progenitors from antibiotic-treated mice.

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JAK signaling is altered in myeloid progenitors from antibiotic-treated ...
(A–D) C57BL/6 mice were treated with antibiotic (ABX) or control water for 3 weeks prior to sacrifice and extraction of whole bone marrow cells. Bone marrow CMPs, GMPs, and cMoPs were sorted to isolate RNA for microarray analysis. (A) Principle component analysis was used to visualize the variance between samples in a 3-dimensional PCA plot. (B) Unsupervised hierarchical clustering analysis was used to compare the transcriptomes of each sample of the microarray. (C) Gene set enrichment analysis plots demonstrate negative enrichment of type I (left) and type II (right) IFNs genes in CMPs from antibiotic-treated mice. (D) Ingenuity Pathway Analysis (IPA) of differentially expressed genes identified suppression of IRF7 as an upstream regulator of the transcriptional changes observed between myeloid progenitors from antibiotic-treated and control mice. Heatmaps of the 5 IRF7-regulated genes identified by IPA analysis from the microarray data set are displayed. (E) Whole bone marrow cells were isolated from mice treated with antibiotic or control water for 3 weeks. Baseline levels of pSTAT1 in myeloid progenitors were measured by flow cytometry. Data were compiled from 2 separate experiments, for a total of 9 mice per group, and analyzed by a Mann-Whitney U test. (F) CMPs were sorted from mice treated with antibiotic or control water in combination with every-other-day dosing of an isotype antibody or anti-mouse IFNAR1 antibody for 3 weeks (n = 4 mice/group). Sorted CMPs were cultured for 7 days in media containing M-CSF, GM-CSF, IL-3, and SCF before enumeration of mature myeloid cell progeny by flow cytometry. Data were analyzed by 2-way ANOVA (***P < 0.01, ****P < 0.0001, antibiotic-treated vs. control mice and antibiotic-treated vs. anti-IFNAR–treated mice).