MicroRNAs are known to regulate gene function in many tissues and organs, but their expression and function, if any, in tooth development are elusive. We sought to identify them by microRNA screening analyses and reveal their overall roles by inactivating Dicer1 in the dental epithelium and mesenchyme. Discrete sets of microRNAs are expressed in molars compared with incisors as well as epithelium compared with mesenchyme. Conditional knockout (cKO) of Dicer1 (mature microRNAs) in the dental epithelium of the Pitx2-Cre mouse results in multiple and branched enamel-free incisors and cuspless molars, and change in incisor patterning and in incisor and molar size and shape. Analyses of differentiating dental epithelial markers reveal a defect in amelo-blast differentiation. Conversely, the cervical loop (stem cell niche) is expanded in Dicer1 cKO. These results demonstrate that tooth development is tightly controlled by microRNAs and that specific microRNAs regulate tooth epithelial stem cell differentiation.

Mesenchymal stem cells (MSCs) are well established to have promising therapeutic properties. TNF‐stimulated gene‐6 (TSG‐6), a potent tissue‐protective and anti‐inflammatory factor, has been demonstrated to be responsible for a significant part of the tissue‐protecting properties mediated by MSCs. Nevertheless, current knowledge about the biological function of TSG‐6 in MSCs is limited. Here, we demonstrated that TSG‐6 is a crucial factor that influences many functional properties of MSCs. The transcriptomic sequencing analysis of wild‐type (WT) and TSG‐6−/−‐MSCs shows that the loss of TSG‐6 expression leads to the perturbation of several transcription factors, cytokines, and other key biological pathways. TSG‐6−/−‐MSCs appeared morphologically different with dissimilar cytoskeleton organization, significantly reduced size of extracellular vesicles, decreased cell proliferative rate, and loss of differentiation abilities compared with the WT cells. These cellular effects may be due to TSG‐6‐mediated changes in the extracellular matrix (ECM) environment. The supplementation of ECM with exogenous TSG‐6, in fact, rescued cell proliferation and changes in morphology. Importantly, TSG‐6‐deficient MSCs displayed an increased capacity to release interleukin‐6 conferring pro‐inflammatory and pro‐tumorigenic properties to the MSCs. Overall, our data provide strong evidence that TSG‐6 is crucial for the maintenance of stemness and other biological properties of murine MSCs.

Background Sphingosine-1-phosphate receptor 2 (S1PR2) mediates pleiotropic functions encompassing cell proliferation, survival, and migration, which become collectively de-regulated in cancer. Information on whether S1PR2 participates in colorectal carcinogenesis/cancer is scanty, and we set out to fill the gap. Methods We screened expression changes of S1PR2 in human CRC and matched normal mucosa specimens [N = 76]. We compared CRC arising in inflammation-driven and genetically engineered models in wild-type (S1PR2+/+) and S1PR2 deficient (S1PR2−/−) mice. We reconstituted S1PR2 expression in RKO cells and assessed their growth in xenografts. Functionally, we mimicked the ablation of S1PR2 in normal mucosa by treating S1PR2+/+ organoids with JTE013 and characterized intestinal epithelial stem cells isolated from S1PR2−/−Lgr5-EGFP- mice. Results S1PR2 expression was lost in 33% of CRC; in 55%, it was significantly decreased, only 12% retaining expression comparable to normal mucosa. Both colitis-induced and genetic Apc+/min mouse models of CRC showed a higher incidence in size and number of carcinomas and/or high-grade adenomas, with increased cell proliferation in S1PR2−/− mice compared to S1PR2+/+ controls. Loss of S1PR2 impaired mucosal regeneration, ultimately promoting the expansion of intestinal stem cells. Whereas its overexpression attenuated cell cycle progression, it reduced the phosphorylation of AKT and augmented the levels of PTEN. Conclusions In normal colonic crypts, S1PR2 gains expression along with intestinal epithelial cells differentiation, but not in intestinal stem cells, and contrasts intestinal tumorigenesis by promoting epithelial differentiation, preventing the expansion of stem cells and braking their malignant transformation. Targeting of S1PR2 may be of therapeutic benefit for CRC expressing high Lgr5. Graphical Abstract. Schematic drawing of the role of S1PR2 in normal mucosa and colorectal cancer. In the normal mucosa, S1PR2 is highly expressed by differentiated cells at the upper region of both colon and intestinal crypts (S1PR2 ON), but not by the undifferentiated stem cell at the base of the crypts (S1PR2 OFF), in which acts as a negative proliferative regulator promoting epithelial differentiation. Its loss leads to the expansion of stem cells and reduced levels of PTEN and Axin-2, two negative regulators respectively of PI3K/AKT and Wnt signaling that control β-catenin signaling. The translocation of β-catenin into the nucleus promotes the transcription of target genes involved in the proliferation and malignant transformation. Thereby, S1PR2 works in the intestine as a tumor suppressor

Hyaluronan (HA) has proven to be beneficial in the treatment of several diseases. Recently, it has been shown that the local application of HA (IBD98E) improves endoscopic and clinical outcomes in subjects with active distal ulcerative colitis (UC). However, the mechanisms by which this polysaccharide exerts its beneficial effects are unclear. Here, we demonstrated that HA treatment in vitro and in vivo improved mucosal healing by accelerating intestinal epithelial regeneration. Indeed, mice treated with HA showed a faster recovery from colitis and reduced endoscopic signs of mucosal inflammation compared to those receiving saline. Furthermore, histological analysis revealed less ulcerated mucosa in mice treated with HA, characterized by re-epithelialized areas. TSG-6, the secreted product of TNF-stimulated gene-6, is an HA-binding protein shown previously to have tissue-protective properties and promote wound healing. Mucosal levels of TSG-6 increased in UC patients compared to the healthy controls and also after wounding in mice. TSG-6 deletion prevented the beneficial properties of HA in mucosal wound repair, suggesting that the interaction of HA with TSG-6 is crucial for intestinal epithelial regeneration. Overall these results are consistent with HA having a therapeutic effect via the promotion of mucosal healing in patients with ulcerative colitis.