Lgr4 is required for Paneth cell differentiation and maintenance of intestinal stem cells ex vivo

Gene inactivation of the orphan G protein-coupled receptor LGR4, a paralogue of the epithelial-stem-cell marker LGR5, results in a 50% decrease in epithelial cell proliferation and an 80% reduction in terminal differentiation of Paneth cells in postnatal mouse intestinal crypts. When cultured ex vivo, LGR4-deficient crypts or progenitors, but not LGR5-deficient progenitors, die rapidly with marked downregulation of stem-cell markers and Wnt target genes, including Lgr5. Partial rescue of this phenotype is achieved by addition of LiCl to the culture medium, but not Wnt agonists. Our results identify LGR4 as a permissive factor in the Wnt pathway in the intestine and, as such, as a potential target for intestinal cancer therapy.     

Notch signaling is required for normal prostatic epithelial cell proliferation and differentiation

Notch pathway is crucial for stem/progenitor cell maintenance, growth and differentiation in a variety of tissues. Using a transgenic cell ablation approach, we found in our previous study that cells expressing Notch1 are crucial for prostate early development and re-growth. Here, we further define the role of Notch signaling in regulating prostatic epithelial cell growth and differentiation using biochemical and genetic approaches in ex vivo or in vivo systems. Treatment of developing prostate grown in culture with inhibitors of gamma-secretase/presenilin, which is required for Notch cleavage and activation, caused a robust increase in proliferation of epithelial cells co-expressing cytokeratin 8 and 14, lack of luminal/basal layer segregation and dramatically reduced branching morphogenesis. Using conditional Notch1 gene deletion mouse models, we found that inactivation of Notch1 signaling resulted in profound prostatic alterations, including increased tufting, bridging and enhanced epithelial proliferation. Cells within these lesions co-expressed both luminal and basal cell markers, a feature of prostatic epithelial cells in predifferentiation developmental stages. Microarray analysis revealed that the gene expression in a number of genetic networks was altered following Notch1 gene deletion in prostate. Furthermore, expression of Notch1 and its effector Hey-1 gene in human prostate adenocarcinomas were found significantly down-regulated compared to normal control tissues. Taken together, these data suggest that Notch signaling is critical for normal cell proliferation and differentiation in the prostate, and deregulation of this pathway may facilitate prostatic tumorigenesis.     

The relation between cell proliferation,differentiation and ultrastructural development in rat intestinal epithelium

Summary The ultrastructural development of the principal cells in rat small intestine was studied by morphometric analyses in relation to the exact cell position along crypt and villus. From the bottom to the tip of the crypt, a gradual increase occurred in absolute size of the total cell, the cytoplasm, the terminal web and of nearly all cell organelles. Also, the relative size of the cytoplasm, mitochondria, microvilli and endoplasmic reticulum increased during crypt cell differentiation. No sudden changes in ultrastructure were observed in the so-called critical decision zone, normally located halfway up the crypt where the proliferative activity ceases. At the crypt-villous junction a 1.4–3 fold increase in cell size, cytoplasm, terminal web and of most organelles was noted. Expansion of the proliferative cell compartment over the total length of the crypt as occurs during recovery after a low X-irradiation dose (72 h after 400 R) does not affect the normal development of cellular ultrastructure. These findings are discussed in relation to biochemical and cell kinetic data.     

Pax9 is required for filiform papilla development and suppresses skin-specific differentiation of the mammalian tongue epithelium

The epidermis is a derivative of the surface ectoderm. It forms a protective barrier and specific appendages including hair, nails, and different eccrine glands. The surface ectoderm also forms the epithelium of the oral cavity and tongue, which develop a slightly different barrier and form different appendages such as teeth, filiform papillae, taste papillae, and salivary glands. How this region-specific differentiation is genetically controlled is largely unknown. We show here that Pax9, which is expressed in the epithelium of the tongue but not in skin, regulates several aspects of tongue-specific epithelial differentiation. In Pax9-deficient mice filiform papillae lack the anterior-posterior polarity, a defect that is associated with temporal-spatial changes in Hoxc13 expression. Barrier formation is disturbed in the mutant tongue and genome-wide expression profiling revealed that the expression of specific keratins (Krt), keratin-associated proteins, and members of the epidermal differentiation complex is significantly down-regulated. In situ hybridization demonstrated that several 'hard' keratins, Krt1-5, Krt1-24, and Krt2-16, are not expressed in the absence of Pax9. Notably, specific 'soft' keratins, Krt2-1 and Krt2-17, normally weakly expressed in the tongue but present at high levels in skin and in orthokeratinized oral dysplasia are up-regulated in the mutant tongue epithelium. This result indicates a partial trans-differentiation to an epithelium with skin-specific characteristics. Together, our findings show that Pax9 regulates appendage formation in the mammalian tongue and identify Pax9 as an important factor for the region-specific differentiation of the surface ectoderm.     

FGF10 is required for cell proliferation and gland formation in the stomach epithelium of the chicken embryo

The development of digestive organs in vertebrates involves active epithelial-mesenchymal interactions. In the chicken proventriculus (glandular stomach), the morphogenesis and cytodifferentiation of the epithelium are controlled by the inductive signaling factors that are secreted from the underlying mesenchyme. Previous studies have shown that Fgf10 is expressed in the developing chicken proventricular mesenchyme, whereas its receptors are present in the epithelium. In our present study, we show that FGF10 is an early mesenchymal signal that is critically associated with the developmental processes in the proventricular epithelium. Furthermore, virus-mediated Fgf10 overexpression in ovo results in a hypermorphic epithelial structure and an increase in epithelial cell number. In contrast, the overexpression of a secreted FGFR2b (sFGFR2b), an FGF10 antagonist, blocks cell proliferation and gland formation in the proventricular epithelium in ovo. This downregulation of proliferative activity was subsequently found to retard gland formation and also to delay differentiation of the epithelium. These results demonstrate that FGF10 signaling, mediated by FGFR1b and/or FGFR2b, is required for proliferation and gland formation in the epithelium in the developing chick embryo.     

Sox9 is essential for outer root sheath differentiation and the formation of the hair stem cell compartment

BACKGROUND: The mammalian hair represents an unparalleled model system to understand both developmental processes and stem cell biology. The hair follicle consists of several concentric epithelial sheaths with the outer root sheath (ORS) forming the outermost layer. Functionally, the ORS has been implicated in the migration of hair stem cells from the stem cell niche toward the hair bulb. However, factors required for the differentiation of this critical cell lineage remain to be identified. Here, we describe an unexpected role of the HMG-box-containing gene Sox9 in hair development. RESULTS: Sox9 expression can be first detected in the epithelial component of the hair placode but then becomes restricted to the outer root sheath (ORS) and the hair stem cell compartment (bulge). Using tissue-specific inactivation of Sox9, we demonstrate that this gene serves a crucial role in hair differentiation and that skin deleted for Sox9 lacks external hair. Strikingly, the ORS acquires epidermal characteristics with ectopic expression of GATA3. Moreover, Sox9 knock hair show severe proliferative defects and the stem cell niche never forms. Finally, we show that Sox9 expression depends on sonic hedgehog (Shh) signaling and demonstrate overexpression in skin tumors in mouse and man. CONCLUSIONS: We conclude that although Sox9 is dispensable for hair induction, it directs differentiation of the ORS and is required for the formation of the hair stem cell compartment. Our genetic analysis places Sox9 in a molecular cascade downstream of sonic hedgehog and suggests that this gene is involved in basal cell carcinoma.     

Nucleoplasmic calcium is required for cell proliferation

Ca(2+) signals regulate cell proliferation, but the spatial and temporal specificity of these signals is unknown. Here we use selective buffers of nucleoplasmic or cytoplasmic Ca(2+) to determine that cell proliferation depends upon Ca(2+) signals within the nucleus rather than in the cytoplasm. Nuclear Ca(2+) signals stimulate cell growth rather than inhibit apoptosis and specifically permit cells to advance through early prophase. Selective buffering of nuclear but not cytoplasmic Ca(2+) signals also impairs growth of tumors in vivo. These findings reveal a major physiological and potential pathophysiological role for nucleoplasmic Ca(2+) signals and suggest that this information can be used to design novel therapeutic strategies to regulate conditions of abnormal cell growth.     

c-Myc is required for the formation of intestinal crypts but dispensable for homeostasis of the adult intestinal epithelium

In self-renewing tissues such as the skin epidermis and the bone marrow, Myc proteins control differentiation of stem cells and proliferation of progenitor cell types. In the epithelium of the small intestine, we show that c-Myc and N-Myc are expressed in a differential manner. Whereas c-Myc is expressed in the proliferating transient-amplifying compartment of the crypts, N-Myc is restricted to the differentiated villus epithelium and a single cell located near the crypt base. c-Myc has been implicated as a critical target of the canonical Wnt pathway, which is essential for formation and maintenance of the intestinal mucosa. To genetically assess the role of c-Myc during development and homeostasis of the mammalian intestine we induced deletion of the c-myc(flox) allele in the villi and intestinal stem cell-bearing crypts of juvenile and adult mice, via tamoxifen-induced activation of the CreER(T2) recombinase, driven by the villin promoter. Absence of c-Myc activity in the juvenile mucosa at the onset of crypt morphogenesis leads to a failure to form normal numbers of crypts in the small intestine. However, all mice recover from this insult to form and maintain a normal epithelium in the absence of c-Myc activity and without apparent compensation by N-Myc or L-Myc. This study provides genetic and molecular evidence that proliferation and expansion of progenitors necessary to maintain the adult intestinal epithelium can unexpectedly occur in a Myc-independent manner.