Introduction: SMAD4 is a tumor suppressor known to be altered in patients with Colitis-Associated Carcinoma (CAC), a subtype of colorectal cancer. Previous studies have confirmed that loss of SMAD4 in mice with colitis led to upregulation of pro-inflammatory pathways and increased chemokine and cytokine levels. CCL20 was one such upregulated chemokine and it binds to the CCR6 receptor, the sole receptor it activates. Notably, B cells, T cells, and dendritic cells contain the CCR6 receptor, and there is evidence that CCL20 may particularly recruit dendritic cells to the gut epithelium to regulate the inflammatory environment. This led us to hypothesize that dendritic cell recruitment to the epithelium will increase in mice with a loss of SMAD4 and an elevated CCR6/CCL20 axis
Methods: First, cells were extracted from the bone marrow of mice and were differentiated into dendritic cells using GM-CSF and either IL-4 or ß-Mercaptoethanol. We found that either method induced abundant expression of dendritic cell markers as determined by RT-qPCR analysis. A co-culture experiment was performed assessing dendritic cell migration to SMAD4+ and SMAD4- epithelia from CCR6+ and CCR6- mice. RT-qPCR was performed to characterize the dendritic cells that migrated during the co-culture.We also performed immunohistochemical (IHC) staining of dendritic cells in colon tissue from CCR6+ and CCR6- mice with a loss of SMAD4.
Results: Results from IHC suggest that loss of CCR6 appears to have little effect on dendritic cell recruitment within the colon. Preliminary results from the co-culture experiment indicate that dendritic cells are able to migrate to the epithelium regardless of SMAD4 status. Results from the RT-qPCR performed ondendritic cells from the co-culture show that dendritic cells migrating to SMAD4- epithelia havehigher levels of a marker of dendritic cell activationwhen compared to those migrating to SMAD4+ epithelia.
Discussion: This early finding suggests that although dendritic cells are recruited to either SMAD4+ or SMAD4- epithelia, the dendritic cells may function differently in a SMAD4- epithelium. Additional studies will be performed to confirm this.Ultimately, exploring how Smad4 interacts with dendritic cells through this study will enhance our understanding of immune signaling in CAC pathogenesis and highlights the need for additional research in immune cell activity in the setting of colon inflammation.
.jpeg.jpg "Chimsom D. Agbim, MS photo")