Local and Regional Spread of Primary Conjunctival Squamous Cell Carcinoma.

Two cases of biopsy-proven conjunctival squamous cell carcinoma (SCC) that developed local and regional spread are described. The cases involved a 65-year-old woman and a 79-year-old man who were initially treated at outside institutions for SCC of the conjunctiva. The patients did not have a history of immune compromise. The female patient presented with direct extension into the lacrimal gland but deferred recommended exenteration. Despite eventual exenteration, she developed metastasis to a neck node 6 months later, which was treated with radiotherapy. The male patient presented with local recurrence and a parotid node metastasis treated with exenteration, parotidectomy, selective neck dissection, and postoperative radiotherapy. Review of the outside pathology of both cases revealed positive tumor margins at the time of original resection. Local control of conjunctival SCC is of critical importance to reduce the risk of orbital extension and regional spread.

Diagnostic Approaches to Metastatic Hepatocellular Carcinoma of the Orbit.

Orbital metastasis of hepatocellular carcinoma is exceedingly rare and caries a grave prognosis. Three cases of metastatic orbital hepatocellular carcinoma in which the primary tumor was initially unknown and the diagnostic challenges encountered are presented. With hepatocellular carcinoma, open biopsy and palliative tumor debulking has an increased bleeding risk due to the highly vascular nature of the tumor and coagulopathy associated with chronic liver disease. As an alternative, fine needle aspiration biopsy should be considered for hepatocellular carcinoma with a readily accessible mass and the availability of an experienced cytopathologist.

Efficient colonic mucosal wound repair requires Trem2 signaling.

The colonic epithelial lining undergoes constant replacement, driven by epithelial stem cells in crypts of Lieberkühn. Stem cells lost because of damage or disease can be replaced by adjacent crypts that undergo fission. The close proximity of an extraordinary number of luminal microbes creates a challenge for this repair process; infection must be prevented while immune system activation and epithelial stem cell genetic damage must be minimized. To understand the factors that modulate crypt/stem cell replacement in the mouse colon, we developed an in vivo acute injury system analogous to punch biopsy of the skin. In contrast to epidermal stem cells, colonic epithelial progenitors did not migrate over the wound bed. Instead, their proliferative expansion was confined to crypts adjacent to wound beds and was delayed to the latter phase of healing. This increased epithelial proliferation was coincident with the infiltration of Trem2 expressing macrophages and increased expression of IL-4 and IL-13 in the wound bed. Interestingly, Trem2(-/-) mice displayed slow and incomplete wound healing of colonic mucosal injuries. We found the latter phase of healing in Trem2(-/-) mice showed a diminished burst of epithelial proliferation, increased expression of IFN-gamma and TNF-alpha, diminished expression of IL-4 and IL-13, and increased markers of classical macrophage activation. Ablation of these cytokines in injured WT and Trem2(-/-) mice demonstrated that their expression ultimately determined the rate and nature of wound healing. These studies show that Trem2 signaling is an important pathway to promote healing of wounds in the colon where stem cell replacement is necessary.

Fgf9 signaling regulates small intestinal elongation and mesenchymal development.

Short bowel syndrome is an acquired condition in which the length of the small intestine is insufficient to perform its normal absorptive function. Current therapies are limited as the developmental mechanisms that normally regulate elongation of the small intestine are poorly understood. Here, we identify Fgf9 as an important epithelial-to-mesenchymal signal required for proper small intestinal morphogenesis. Mouse embryos that lack either Fgf9 or the mesenchymal receptors for Fgf9 contained a disproportionately shortened small intestine, decreased mesenchymal proliferation, premature differentiation of fibroblasts into myofibroblasts and significantly elevated Tgfbeta signaling. These findings suggest that Fgf9 normally functions to repress Tgfbeta signaling in these cells. In vivo, a small subset of mesenchymal cells expressed phospho-Erk and the secreted Tgfbeta inhibitors Fst and Fstl1 in an Fgf9-dependent fashion. The p-Erk/Fst/Fstl1-expressing cells were most consistent with intestinal mesenchymal stem cells (iMSCs). We found that isolated iMSCs expressed p-Erk, Fst and Fstl1, and could repress the differentiation of intestinal myofibroblasts in co-culture. These data suggest a model in which epithelial-derived Fgf9 stimulates iMSCs that in turn regulate underlying mesenchymal fibroblast proliferation and differentiation at least in part through inhibition of Tgfbeta signaling in the mesenchyme. Taken together, the interaction of FGF and TGFbeta signaling pathways in the intestinal mesenchyme could represent novel targets for future short bowel syndrome therapies.

Diverse adult stem cells share specific higher-order patterns of gene expression.

Adult tissue stem cells (SCs) share functional properties regardless of their tissue of residence. It had been thought that SCs might also share expression of certain "stemness" genes, although early investigations for such genes were unsuccessful. Here, we show that SCs from diverse tissues do preferentially express certain types of genes and that SCs resemble other SCs in terms of global gene expression more than they resemble the differentiated cells (DCs) of the tissues that they supply. Genes associated with nuclear function and RNA binding were over-represented in SCs. In contrast, DCs from diverse tissues shared enrichment in genes associated with extracellular space, signal transduction, and the plasma membrane. Further analysis showed that transit-amplifying cells could be distinguished from both SCs and DCs by heightened expression of cell division and DNA repair genes and decreased expression of apoptosis-related genes. This transit-amplifying cell-specific signature was confirmed by de novo generation of a global expression profile of a cell population highly enriched for transit-amplifying cells: colonic crypt-base columnar cells responding to mucosal injury. Thus, progenitor cells preferentially express intracellular or biosynthetic genes, and differentiation correlates with increased expression of genes for interacting with other cells or the microenvironment. The higher-order, Gene Ontology term-based analysis we use to distinguish SC- and DC-associated gene expression patterns can also be used to identify intermediate differentiation states (e.g., that of transit-amplifying cells) and, potentially, any biological state that is reflected in changes in global gene expression patterns. Disclosure of potential conflicts of interest is found at the end of this article.

An antibiotic-responsive mouse model of fulminant ulcerative colitis.

BACKGROUND: The constellation of human inflammatory bowel disease (IBD) includes ulcerative colitis and Crohn's disease, which both display a wide spectrum in the severity of pathology. One theory is that multiple genetic hits to the host immune system may contribute to the susceptibility and severity of IBD. However, experimental proof of this concept is still lacking. Several genetic mouse models that each recapitulate some aspects of human IBD have utilized a single gene defect to induce colitis. However, none have produced pathology clearly distinguishable as either ulcerative colitis or Crohn's disease, in part because none of them reproduce the most severe forms of disease that are observed in human patients. This lack of severe IBD models has posed a challenge for research into pathogenic mechanisms and development of new treatments. We hypothesized that multiple genetic hits to the regulatory machinery that normally inhibits immune activation in the intestine would generate more severe, reproducible pathology that would mimic either ulcerative colitis or Crohn's disease. METHODS AND FINDINGS: We generated a novel mouse line (dnKO) that possessed defects in both TGFbetaRII and IL-10R2 signaling. These mice rapidly and reproducibly developed a disease resembling fulminant human ulcerative colitis that was quite distinct from the much longer and more variable course of pathology observed previously in mice possessing only single defects. Pathogenesis was driven by uncontrolled production of proinflammatory cytokines resulting in large part from T cell activation. The disease process could be significantly ameliorated by administration of antibodies against IFNgamma and TNFalpha and was completely inhibited by a combination of broad-spectrum antibiotics. CONCLUSIONS: Here, we develop to our knowledge the first mouse model of fulminant ulcerative colitis by combining multiple genetic hits in immune regulation and demonstrate that the resulting disease is sensitive to both anticytokine therapy and broad-spectrum antibiotics. These findings indicated the IL-10 and TGFbeta pathways synergize to inhibit microbially induced production of proinflammatory cytokines, including IFNgamma and TNFalpha, which are known to play a role in the pathogenesis of human ulcerative colitis. Our findings also provide evidence that broad-spectrum antibiotics may have an application in the treatment of patients with ulcerative colitis. This model system will be useful in the future to explore the microbial factors that induce immune activation and characterize how these interactions produce disease.

Myd88-dependent positioning of Ptgs2-expressing stromal cells maintains colonic epithelial proliferation during injury.

We identified cellular and molecular mechanisms within the stem cell niche that control the activity of colonic epithelial progenitors (ColEPs) during injury. Here, we show that while WT mice maintained ColEP proliferation in the rectum following injury with dextran sodium sulfate, similarly treated Myd88(-/-) (TLR signaling-deficient) and prostaglandin-endoperoxide synthase 2(-/-) (Ptgs2(-/-)) mice exhibited a profound inhibition of epithelial proliferation and cellular organization within rectal crypts. Exogenous addition of 16,16-dimethyl PGE(2) (dmPGE(2)) rescued the effects of this injury in both knockout mouse strains, indicating that Myd88 signaling is upstream of Ptgs2 and PGE(2). In WT and Myd88(-/-) mice, Ptgs2 was expressed in scattered mesenchymal cells. Surprisingly, Ptgs2 expression was not regulated by injury. Rather, in WT mice, the combination of injury and Myd88 signaling led to the repositioning of a subset of the Ptgs2-expressing stromal cells from the mesenchyme surrounding the middle and upper crypts to an area surrounding the crypt base adjacent to ColEPs. These findings demonstrate that Myd88 and prostaglandin signaling pathways interact to preserve epithelial proliferation during injury using what we believe to be a previously undescribed mechanism requiring proper cellular mobilization within the crypt niche.