Terminal differentiation of intestinal goblet cells is affected in agr2 morphants.

Mammalian Anterior Gradient 2 (AGR2) is a protein disulfide isomerase that is required for the production of intestinal mucus and Paneth and goblet cell homeostasis. However, whether increased endoplasmic reticulum (ER) stress
occurs in Agr2-/- mice remains a controversial issue. We characterized the function of zebrafish agr2 by both morpholino antisense oligomermediated knockdown and agr2 mRNA overexpression. Fluorescent whole-mount double in situ hybridization indicated that in the intestine, agr2 was only expressed in goblet cells. Significantly increased numbers of immature Alcian blue-stained goblet cells were observed in the intestines of 104- and 120-hours post fertilization (hpf) agr2 morphants. Transmission electron microscopy analyses further confirmed the existence of immature pre-goblet cells containing few mucous granules in the mid-intestines of 104- and 120-hpf agr2 morphants. agr2 expression was not significantly induced by an ER stress inducer, tunicamycin. Expression of the ER chaperone gene hspa5, the spliced form of xbp1s, c/enhancer binding protein homologous protein chop, and the activating transcription factor 4b1 atf4b1 were not significantly induced in either 104-hpf agr2 morphants or agr2-overexpressed embryos. Similar percentages of P-Histone H3-stained M phase cells were identified in intestines of 104-hpf agr2 morphants and control embryos. Our study demonstrates that in contrast to mouse AGR2, zebrafish Agr2 is expressed in only one intestinal secretory cell type - the goblet cells. Agr2 is essential for terminal differentiation of intestinal goblet cells in zebrafish embryos. Either knockdown of agr2 function or agr2 overexpression could not extensively induce expression of members of the unfolded protein response pathway.

Glycosphingolipids (GSLs) are ubiquitous components of cell membranes that can act as mediators of cell adhesion and signal transduction and can possibly be used as cell type-specific markers. Our previous study indicated that there was a striking switch in the core structures of GSLs during differentiation of human embryonic stem cells (hESCs) into embryoid body (EB), suggesting a close association of GSLs with cell

Cancer-initiating Cells in Kras^G12D-induced lung Adenocarcinoma (Cancer Research 71:7250 2011)

We established an inducible Kras^G12D-driven lung adenocarcinoma in CCSP-rtTA/TetO-Cre/LSL-Kras^G12D mice that enable pursuits of the cellular and molecular processes involved in Kras-induced tumorigenesis. To investigate the cellular origin of this cancer, we first report a strategy using FACS fractionation that could highly enrich bronchiolar Clara and alveolar type II cells, respectively. The EpCAM⁺MHCII^- cells (bronchiolar origin) were more enriched with tumorigenic cells in generating secondary tumors than EpCAM⁺MHCII⁺ cells (alveolar origin) in primary tumors that had been already initiated with oncogenic Kras activation. In addition, secondary tumors derived from EpCAM⁺MHCII^- cells showed diversity of tumor locations compared to those derived from EpCAM⁺MHCII⁺ cells . In the alveolar region, secondary tumors from EpCAM⁺MHCII^- cells expressed not only bronchiolar epithelial marker, panCK, but also differentiation marker, proSPC, consistent with the notion that cancer-initiating cells display not only the abilities for self-renewal but also the features of differentiation to generate heterogeneous tumors with phenotypic diversity. Furthermore, high level of ERK1/2 activation and colony-forming ability as well as lack of Sprouty-2 expression were also observed in EpCAM⁺MHCII^- cells ; these signaling pathways may account for the mechanism of Kras^G12D-induced tumorigenesis.