The Role of Transforming Growth Factor Beta (TGFbeta) in Promoting Allergic Inflammation

National Institute Of Allergy And Infectious Diseases

Investigators

Linked publications, trials & patents

Abstract

Although allergic diseases are among the most pervasive medical conditions to affect humans in the world, surprisingly little is known regarding the basic pathophysiologic mechanisms involved. In FY23, we used a mouse model of LDS to make significant strides in understanding the mechanisms by which impaired TGFb signaling promotes eosinophilic esophagitis (EoE). EoE is a chronic allergic disease characterized by an excessive accumulation of eosinophils in the esophagus that is a common cause of feeding problems in children and food impaction and dysphagia in adults. We found that 100% of LDS mice spontaneously develop EoE that closely recapitulates human EoE including the presence of a mixed inflammatory infiltrate (prominent eosinophils, mast cells, type 2 innate lymphoid cells ILC2s, T cells, and myeloid antigen presenting cells), basal cell hyperplasia, and food impaction. Recent work suggests that EoE in patients can be subclassified into three endotypes (EoEe1, 2, 3). Based on the esophageal transcriptome (including elevated expression of Tslp and Tnfaip6), early age of disease onset (evident at weaning), and lack of excess collagen deposition, EoE in LDS mice appears to most closely recapitulate EoEe2 in humans, which is usually pediatric-onset, lacks fibrostenosis, and is distinguished by high expression of TSLP and TNFAIP6. Thus, impaired TGFb signaling is sufficient to drive the spontaneous development of EoE that closely recapitulates the human disease clinically, histologically, immunologically and transcriptionally, and therefore this model presents a unique opportunity to elucidate the pathogenesis of EoE. Although TGFb is well known to regulate T cell differentiation and function, and T cells are posited to be the primary drivers of inflammation in EoE, the severity of EoE in LDS mice was not affected by the complete absence of all T and B cells. Furthermore, mixed bone marrow chimeras revealed that defects in non-hematopoietic cells were not only necessary but sufficient to induce EoE. Further histologic evaluation revealed that inflammation in LDS mice was strictly localized to tissues lined by stratified squamous epithelium as is found in the esophagus of both mice and humans. Despite the extensive inflammation, esophageal barrier function remained intact in LDS mice. No difference in transepithelial electrical resistance (TEER) or in the paracellular transport of molecules comparable in size to common food antigens was observed between esophagi of WT and LDS mice. However, the differentiation of esophageal epithelial cells was severely disrupted in LDS. Genes typically expressed by immature epithelial cells were enriched in the LDS esophageal transcriptome, and we found greater incorporation of the thymidine analog, bromodeoxyuridine (BrdU), in LDS epithelial cells compared to wild type (WT) controls. Collectively, these data demonstrate that LDS mutations lead to enhanced squamous epithelial proliferation and impaired differentiation without compromising barrier function. While basal cell hyperplasia is known to be a key feature of EoE, our findings show that disruption in TGFb signaling may be a key driver of this phenotype. Expression of several genes that promote secretion and activation of IL-1 were notably increased in esophagi of LDS mice. Release of IL-1 is a known innate immune mechanism by which distressed epithelial cells can initiate an inflammatory cascade at mucosal surfaces, a process that is amplified by TNF-a. The esophageal transcriptome of LDS mice revealed enhanced expression of both IL-1 and TNF-a responsive genes, which included a broad range of Damage-Associated Molecular Patterns (DAMPs), keratinocyte growth factors, complement components, cytokines, chemokines, and adhesion molecules that promote the activation and recruitment of Th2 cells, ILC2s, eosinophils, and mast cells. Thus, impaired epithelial development as a result of reduced TGFb signaling leads to overexpression of numerous mediators that promote allergic inflammation and epithelial proliferation - hallmark findings in EoE. To confirm that epithelial-cell intrinsic defects were sufficient to drive eosinophilic inflammation in the absence of immune cells or other external stimuli, organoids were generated from esophagi of LDS mice and WT littermates. Similar to our observations in intact esophagi, LDS organoids exhibited enhanced basal cell proliferation and overexpression of Tslp, Tnfaip6, Il1a, Il1b, and Tnf compared to WT organoids. Furthermore, supernatants from LDS but not WT organoids were able to promote eosinophil recruitment in a transwell assay, suggesting LDS organoids spontaneously secreted eosinophil chemoattractants. In summary, our work demonstrates that TGFb plays a fundamental, non-redundant, epithelial cell-intrinsic role in controlling tissue-specific allergic inflammation independent of its role in adaptive immunity that manifests clinically as EoE.