Helicobacter pylori infection is the major risk factor for gastric adenocarcinoma. The link with gastric adenocarcinoma is partly due to the H. pylori CagA oncoprotein. CagA is responsible for a particular cell phenotype in vitro, the 'hummingbird' phenotype, that corresponds to an elongation of the cells, mimicking an epithelial-mesenchymal transition (EMT). EMT participates in the carcinogenesis process, and is involved in the generation of cancer stem cells (CSCs). However, its involvement in gastric carcinogenesis has yet not been studied. Therefore, the aim of this study was to determine the role of H. pylori in EMT and in the emergence of gastric CSCs. For this purpose, gastric epithelial cells were cocultured with a cagA-positive H. pylori strain or its isogenic-deleted mutants or were transfected with CagA expression vectors. Study of the expression of epithelial and mesenchymal markers showed that H. pylori, via CagA, is responsible for an EMT phenotype associated with an increase in mesenchymal markers as well as CD44 expression, a known gastric CSC marker. Moreover, infection led to an increased ability to migrate, to invade and to form tumorspheres. Cell sorting experiments showed that only the CD44(high) cells induced by H. pylori infection displayed the mesenchymal phenotype and CSC properties in vitro, and had higher tumorigenic properties than CD44(low) cells in xenografted mice. Immunohistochemistry analyses on human and mouse gastric mucosa tissue samples confirmed a high expression of CD44 and mesenchymal markers in H. pylori-infected cases, and in gastric dysplasia and carcinoma. All of these data suggest that H. pylori, via CagA, unveils CSC-like properties by induction of EMT-like changes in gastric epithelial cells.

Gastric carcinomas are heterogeneous, and the current therapy remains essentially based on surgery with conventional chemotherapy and radiotherapy. This study aimed to characterize biomarkers allowing the detection of cancer stem cells (CSC) in human gastric carcinoma of different histologic types. The primary tumors from 37 patients with intestinal- or diffuse-type noncardia gastric carcinoma were studied, and patient-derived tumor xenograft (PDX) models in immunodeficient mice were developed. The expressions of 10 putative cell surface markers of CSCs, as well as aldehyde dehydrogenase (ALDH) activity, were studied, and the tumorigenic properties of cells were evaluated by tumorsphere assays and xenografts by limiting dilution assays. We found that a subpopulation of gastric carcinoma cells expressing EPCAM, CD133, CD166, CD44, and a high ALDH activity presented the properties to generate new heterogeneous tumorspheres and tumors CD44 and CD166 were coexpressed, representing 6.1% to 37.5% of the cells; ALDH activity was detected in 1.6% to 15.4% of the cells; and the ALDH cells represented a core within the CD44/CD166 subpopulation that contained the highest frequency of tumorigenic CSCs The ALDH cells possessed drug efflux properties and were more resistant to standard chemotherapy than the ALDH cells, a process that was partially reversed by verapamil treatment. CD44 and ALDH are the most specific biomarkers to detect and isolate tumorigenic and chemoresistant gastric CSCs in noncardia gastric carcinomas independently of the histologic classification of the tumor. .

The number and status of lymph nodes examined is crucial for tumor staging. Impact of preoperative chemoradiotherapy on lymph nodes status and survival is still controversial in rectal carcinoma. The aim of this study was (i) to define the impact of preoperative chemoradiotherapy on the number of both retrieved and positive lymph nodes in rectal cancer specimen, (ii) to evaluate the influence of the number of lymph nodes retrieved on survival in patients treated by preoperative chemoradiotherapy. From 1994 to 2004, 495 patients underwent rectal excision for cancer, of which 332 received long course preoperative radiotherapy. Surgery and pathologic assessment were standardized. Multivariate analysis evaluated the influence of clinical and pathologic variables on the number of both retrieved and positive lymph nodes. Kaplan-Meier method and log-rank test assessed the relation between survival and the number of lymph nodes retrieved in patients treated by preoperative chemoradiotherapy. Compared with surgery alone, preoperative chemoradiotherapy decreased both the mean number of lymph nodes retrieved (17 vs. 13; P<0.001) and the mean number of positive lymph nodes (2.3 vs. 1.2; P=0.001). Multivariate analysis confirmed the independent impact of preoperative chemoradiotherapy on retrieved and positive lymph nodes. In patients treated by preoperative chemoradiotherapy, the 5-year overall (71%) and disease-free (60%) survival was not associated with the number of lymph nodes retrieved. Although long course preoperative chemoradiotherapy decreases by 24%, the mean number of lymph nodes retrieved and by 48% the mean number of positive lymph nodes, survival was not influenced by the number of lymph nodes retrieved in irradiated rectal specimen.

Gastric carcinoma is the third leading cause of cancer-related death worldwide. This cancer, most of the time metastatic, is essentially treated by surgery associated with conventional chemotherapy, and has a poor prognosis. The existence of cancer stem cells (CSC) expressing CD44 and a high aldehyde dehydrogenase (ALDH) activity has recently been demonstrated in gastric carcinoma and has opened new perspectives to develop targeted therapy. In this study, we evaluated the effects of all-trans-retinoic acid (ATRA) on CSCs in human gastric carcinoma. ATRA effects were evaluated on the proliferation and tumorigenic properties of gastric carcinoma cells from patient-derived tumors and cell lines in conventional 2D cultures, in 3D culture systems (tumorsphere assay) and in mouse xenograft models. ATRA inhibited both tumorspheres initiation and growth in vitro, which was associated with a cell-cycle arrest through the upregulation of cyclin-dependent kinase (CDK) inhibitors and the downregulation of cell-cycle progression activators. More importantly, ATRA downregulated the expression of the CSC markers CD44 and ALDH as well as stemness genes such as Klf4 and Sox2 and induced differentiation of tumorspheres. Finally, 2 weeks of daily ATRA treatment were sufficient to inhibit gastric tumor progression in vivo, which was associated with a decrease in CD44, ALDH1, Ki67 and PCNA expression in the remaining tumor cells. Administration of ATRA appears to be a potent strategy to efficiently inhibit tumor growth and more importantly to target gastric CSCs in both intestinal and diffuse types of gastric carcinoma.

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