Final Report, ECCO Grant for Celia Escudero-Hernández
Celia Escudero-Hernández, ECCO Grant Awardee
Intestinal epithelial cell stress modulates enteric fibroblastic and neuronal profiles in Inflammatory Bowel Disease
 C. Escudero-Hernández © C. Escudero-Hernández |
Background & aim of research
This project aimed to understand the effects that crucial IBD epithelial stress factors (i.e. ATG16L1 and XBP1 impairments) might have in enteric fibroblasts and neurons.
Methodology used in the research
We used intestinal epithelial organoids from mice lacking Atg16l1 or Xbp1 expression and implemented fibroblast and neuron primary cell cultures and organ-on-a-chip technology to model the gut in vitro. We also implemented in vitro models of fibrosis using biological (TGF-, EGF), chemical (bleomycin) and infectious (Salmonella) agents. We have now generated diverse tamoxifen-inducible knock-out mice that will serve to study specific effects during dextran sodium sulphate-induced colitis (Atg16l1 or Xbp1 depletion in neurons and Dnmt3a depletion in fibroblasts). We have also established in vivo transfection using adeno-associated viruses to label neurons in vivo based on a Nature protocol released in 2022 to facilitate enteric neuron isolation for single-nuclei RNA sequencing.
Main findings/results of the research
Organoids lacking Atg16l1 or Xbp1 expression did not exert any significant effect on their own on fibroblasts or neurons, which could partially have been due to rare cell subtype communication that is challenging to reproduce in vitro. However, we found that intestinal epithelial cells seem to signal fibroblasts via connective tissue growth factor (CTGF) using in vitro models of fibrosis. In addition, we identified TGF-1 and IL-11 as key signals for enteric fibroblast activation and development of fibrosis. As a key risk gene in IBD, we are exploring the effects that lacking DNA methyltransferase 3A (DNMT3A) exerts in fibroblasts in vitro, and eventually in mice during experimental colitis and fibrosis.
On the other hand, inflammatory insults to enteric neurons pointed to altered brain-derived neurotropic factor (BDNF) expression that could be key for gut dysmotility. Thus, we will directly study the effects of autophagy Atg16l1 and endoplasmic reticulum stress Xbp1 gene depletion in enteric neurons in vivo during experimental colitis.