Protective Role of Mesenchymal MyD88 Signaling Under Homeostasis and Initiation of Inflammation in the Colon

Simple item page
dc.creatorUribe, Gabriela
dc.creator.orcid0000-0002-9478-4293
dc.date.accessioned2022-09-01T14:16:24Z
dc.date.available2022-09-01T14:16:24Z
dc.date.created2019-08
dc.date.submittedAugust 2019
dc.date.updated2022-09-01T14:16:25Z
dc.description.abstractIntestinal Mesenchymal stromal cells (fibroblasts, myofibroblasts and their progenitors) have recently emerged as key players in the interplay between the microbiota and professional immune cells under homeostasis and chronic inflammation. Initiation of microbial interactions with host cells, including stromal cells, requires myeloid differentiation factor 88 (MyD88). Overall, MyD88-dependent interactions between the microbiota and the host is reported to contribute to epithelial barrier restoration via repositioning of mesenchymal stromal cells producing COX-2/PGE2 during injury. However, whether MyD88 signaling in mesenchymal cells is involved in this process is not known. Moreover, mesenchymal cell MyD88 signaling is involved in mucosal tolerance via suppression of type 1 adaptive immune responses. However, the impact of MyD88-dependent signaling within mesenchymal stromal cells on professional innate immune cells such as macrophages within the gut mucosa is poorly understood. Therefore, in this project I evaluate the hypothesis that MyD88-mediated signaling in mesenchymal stromal cells is required for the control over inflammatory responses in the colon. To test this hypothesis, I used primary human mesenchymal cells in culture, as well generated several mesenchymal stromal cell-specific MyD88-inducible knockout mice under homeostasis and in a model of acute colitis using DSS. Histological analysis was used to determine inflammatory and fibrotic changes in the colonic mucosa. The intracellular and cellular responses were analyzed by using RNAseq, qRT-PCR, ELISA, western blot, cytokine/chemokine multiplex array, flow cytometry, and confocal microscopy. In our first aim, I found that deletion of MyD88 within mesenchymal stromal cells in vivo resulted in inflammatory changes and increased fibrosis within the colonic mucosa as well as moderately aggravated DSS induced acute colitis. In addition to IBD-type dysbiotic changes of the fecal microbiota in mice lacking MyD88 within mesenchymal stromal cells, RNAseq analysis of the colonic mucosa from these mice revealed changes in pathways controlling epithelial barrier maintenance and innate and adaptive cell inflammatory responses in DSS-treated and non-treated animals. In order to understand how stromal cell-intrinsic MyD88 signaling activated by the microbiota contributes to the regulation of intestinal homeostasis and how it is disrupted during the immunopathogenesis of IBD, we used Lactobacillus rhamnosus GG (LbGG) as a representative species of normal beneficial gut microbiota. I demonstrated that mesenchymal stromal cells isolated from the normal human colonic mucosa responded to LbGG with activation of the eicosanoid pathway resulting in increased expression of COX-2 mediated PGE2. Similar observation were made in vivo, using a murine model. I also show that in culture and in vivo this process requires MyD88 signaling within mesenchymal stromal cells. Taken together, these data suggest that mesenchymal stromal cells are among the major contributors to the increase of COX-2-mediated PGE2 in response to the normal microbiota, contributing to the maintenance of mucosal homeostasis in the colon via MyD88. In experiments performed in my second aim, the inflammatory changes observed in our initial RNAseq analysis were associated with increased infiltration of F4/80+CD11b+ macrophages. Increase in CX3CR1highCCR2+ macrophages producing TNF-α was also observed and depletion of the macrophages with clondronate resulted in a decrease in total TNF-α levels within the colonic mucosa. This suggests that there is increased chemotaxis of inflammatory macrophages to the colonic mucosa. In conclusion, my data from my work suggest that MyD88 signaling within mesenchymal stromal cells contributes to colonic mucosal homeostasis through the production of PGE2 and the suppression of the influx of TNF-α and IL-6 producing inflammatory macrophages.
dc.format.mimetypeapplication/pdf
dc.identifier.urihttps://hdl.handle.net/2152.3/11729
dc.subject5-LO: 5-lypoxygenase
dc.subjectAA: Arachidonic Acid
dc.subjectAJ: Adherens junctions
dc.subjectBM: Bone marrow
dc.subjectCD: Crohn’s Disease
dc.subjectCMFs: Colonic CD90+ (myo) fibroblasts
dc.subjectCOX-2: Cyclooxygenase 2
dc.subjectcPLA2: Cytosolic phospholipase A2
dc.subjectDSS: Dextran Sulfate Sodium
dc.subjectEB: Epithelial Barrier
dc.subjectECM: Extracellular matrix
dc.subjectER-TR7: anti-reticular fibroblasts and reticular fibres antibody
dc.subjectFib-MyD88 KO:Tamoxifen-inducible MyD88 floxed mice
dc.subjectspecific to Col1α2+ cells
dc.subjectGI: Gastrointestinal
dc.subjecti.p.: intraperitoneal injection
dc.subjectIEC: Intestinal Epithelial Cells
dc.subjectIFN-γ: Interferon gamma
dc.subjectIL-1: interleukin 1 receptors
dc.subjectIL-1: Interleukin-1
dc.subjectIL-10: Interleukin-10
dc.subjectIL-12: Interleukin-12
dc.subjectIL-6: Interleukin-6
dc.subjectILC: Innate lymphoid cells
dc.subjectIPA: Ingenuity pathway analysis
dc.subjectKO: Knockout
dc.subjectLbGG: Lactobacillus rhamnosus GG
dc.subjectMC: Micro-colitis
dc.subjectMF-MyD88 KO: Tamoxifen-inducible MyD88 floxed mice
dc.subjectspecific to α-SMA+ cells
dc.subjectMFs: Myofibroblasts and fibroblasts
dc.subjectMLN: Mesenteric lymph node
dc.subjectMRS: De Man, Rogosa and Sharpe
dc.subjectMSC: Mesenchymal stem cells
dc.subjectMyD88: Myeloid differentiation factor 88
dc.subjectPDGFRα: platelet-derived growth factor receptor A
dc.subjectPD-L1: Programed death ligand 1
dc.subjectPGE2: Prostaglandin E2
dc.subjectTh: T-Helper
dc.subjectTJ: Tight junctions
dc.subjectTLRs: Toll-like receptors
dc.subjectTMX: Tamoxifen
dc.subjectTNF- α: Tumor necrosis factor- α
dc.subjectTreg: Regulatory T cells
dc.subjectUC: Ulcerative Colitis
dc.subjectWB: Western blot
dc.subjectα-SMA: smooth muscle actin
dc.titleProtective Role of Mesenchymal MyD88 Signaling Under Homeostasis and Initiation of Inflammation in the Colon
dc.typeThesis
dc.type.materialtext
thesis.degree.departmentCell Biology
thesis.degree.grantorThe University of Texas Medical Branch at Galveston
thesis.degree.levelDoctoral
thesis.degree.nameCell Biology (Doctoral)

Files

Original bundle

Now showing 1 - 1 of 1
Thumbnail Image
Name:
URIBE-DISSERTATIONDOCTORAL-2019.pdf
Size:
2.84 MB
Format:
Adobe Portable Document Format
Download

License bundle

Now showing 1 - 1 of 1
No Thumbnail Available
Name:
LICENSE.txt
Size:
1.84 KB
Format:
Plain Text
Description:
Download

Collections

Published ETD Collection