Expression of Angiogenic Factors in Invasive Retinoblastoma Tumors Is Associated With Increase in Tumor Cells Expressing Stem Cell Marker Sox2.

CONTEXT: Progression of retinoblastoma is associated with increased tumor angiogenesis. However, a clear relationship between the expression of angiogenic markers in specific regions of the tumor and tumor progression has not been established. This study investigates the association between angiogenic factors in retinoblastomas with choroidal and/or optic nerve invasion (high-risk/invasive retinoblastoma) and expression of Sox2, a stem cell marker. OBJECTIVE: To investigate the association between the expression of angiogenic factors and markers of tumor invasiveness, such as the stem cell marker Sox2, in retinoblastoma tissues. DESIGN: Immunohistochemistry was used to evaluate coexpression of the angiogenic growth factors vascular endothelial growth factor A (VEGF-A), VEGF receptor 2 (VEGFR-2), and endoglin (CD105); markers of glial differentiation (vimentin and glial fibrillary acidic protein); and a neural stem cell marker (Sox2). Expression was assessed in nonneoplastic and neoplastic ocular tissues collected from enucleated eyes of patients with retinoblastoma. During qualitative data interpretation, evaluating pathologists were masked to patient grouping. RESULTS: Expression of VEGF-A and VEGFR-2 in noninvasive (non-high-risk feature) retinoblastoma tumors was lower than in the invasive, or high-risk feature tumors. Moreover, our data indicate that the tumor cells, and not the surrounding stroma, secrete VEGF-A and that angiogenesis is mostly localized to the iris. Finally, our data showed that the expression of the neural stem cell marker Sox2 is associated with eyes with increased VEGF-A expression and tumor invasiveness. CONCLUSIONS: Increased expression of angiogenic factors, with a concomitant increase in expression of the stem cell marker Sox2 observed in retinoblastoma tissues, may partially explain the aggressiveness of these tumors. The complex interaction of angiogenic and stem cell-related pathways in these tumors, especially in high-risk feature retinoblastoma, suggests that targeting tumor cells capable of secreting vasculogenic factors, as well as proangiogenic genes and signaling pathways, may be necessary for development of effective antimetastatic retinoblastoma drugs.

Establishment and propagation of human retinoblastoma tumors in immune deficient mice.

Culturing retinoblastoma tumor cells in defined stem cell media gives rise to primary tumorspheres that can be grown and maintained for only a limited time. These cultured tumorspheres may exhibit markedly different cellular phenotypes when compared to the original tumors. Demonstration that cultured cells have the capability of forming new tumors is important to ensure that cultured cells model the biology of the original tumor. Here we present a protocol for propagating human retinoblastoma tumors in vivo using Rag2(-/-) immune deficient mice. Cultured human retinoblastoma tumorspheres of low passage or cells obtained from freshly harvested human retinoblastoma tumors injected directly into the vitreous cavity of murine eyes form tumors within 2-4 weeks. These tumors can be harvested and either further passaged into murine eyes in vivo or grown as tumorspheres in vitro. Propagation has been successfully carried out for at least three passages thus establishing a continuing source of human retinoblastoma tissue for further experimentation.

The prostate-specific G-protein coupled receptors PSGR and PSGR2 are prostate cancer biomarkers that are complementary to alpha-methylacyl-CoA racemase.

BACKGROUND: Immunohistochemistry (IHC) to detect alpha-methylacyl-CoA racemase (AMACR) expression can be useful in the diagnosis of small foci of prostate cancer on needle biopsy specimens, although it still has limitations in terms of both sensitivity and specificity. We have previously described the increased expression of two prostate-specific G-protein coupled receptors (PSGR and PSGR2) in human prostate cancer. To examine their potential usefulness as cancer biomarkers, we have evaluated their expression relative to AMACR in prostate cancer tissues. METHODS: Expression of PSGR, PSGR2, and AMACR were examined by quantitative reverse-transcriptase PCR in mRNAs from benign prostate and prostate cancer tissues. Expression of PSGR2 and AMACR was also examined by in situ hybridization using a prostate cancer tissue microarray. RESULTS: By in situ hybridization, 24 of 40 prostate cancer cases showed concordant expression of PSGR2 and AMACR. However, in 16 cases there was significant discordance between expression levels of these two markers. By quantitative RT-PCR all three markers were substantially increased in cancer, with AMACR the most overexpressed (30-fold), followed by PSGR2 (13-fold) and PSGR (10-fold). AMACR was the best single marker of prostate cancer but in 7 of the 59 total cases the expression of AMACR was not significantly elevated while PSGR and/or PSGR2 were substantially elevated. CONCLUSION: All three biomarkers are increased in prostate cancer but their expression is not completely concordant. There is a subset of cases in which analysis of expression of PSGR and/or PSGR2, in addition to AMACR, would be diagnostically useful.

Increased expression of the metastasis-associated gene Ehm2 in prostate cancer.

BACKGROUND: Alterations of fibroblast growth factors and their receptors contribute to prostate cancer progression by enhancing cell survival, motility, and proliferation. The expression of the FGFR-4 Arg(388) variant is correlated with the occurrence of pelvic lymph node metastasis and biochemical (PSA) recurrence in men undergoing radical prostatectomy. Ehm2 is an androgen-regulated gene that has been associated with metastasis in other systems, so we sought to determine if it is expressed in prostate cancer and if the FGFR-4 Arg(388) variant can increase its expression. METHODS: Expression of Ehm2 was examined by quantitative RT-PCR and Western blotting in prostate cell lines and by quantitative RT-PCR, in situ hybridization, and immunohistochemistry in prostate tissues. The effect of Ehm2 expression on collagen IV adhesion was tested by transient overexpression and RNA interference. RESULTS: Ehm2 expression is upregulated in prostate cancer cell lines and prostate cancer tissues. Expression of the FGFR-4 Arg(388) variant results in increased expression of Ehm2. Increased expression of Ehm2 leads to decreased adhesion to collagen IV, which has been associated with metastasis in cancers. Analysis of tissue microarrays revealed that increased Ehm2 expression is associated with biochemical recurrence after radical prostatectomy, which is indicative of more aggressive disease. CONCLUSIONS: Ehm2 is overexpressed in prostate cancer and may enhance disease progression and metastasis.

Effects of Helicobacter pylori infection on the link between regenerating gene expression and serum gastrin levels in Mongolian gerbils.

Although regenerating gene (Reg) protein is reported to have a trophic effect on gastric epithelial cells, its involvement in human gastric diseases is not clear. We have recently shown that both gastrin and gastric mucosal inflammation enhance Reg gene expression in the fundic mucosa in rats. This study was designed to clarify whether Reg protein is involved in Helicobacter pylori-induced gastritis and whether Reg gene expression is linked to serum gastrin levels in this condition. Mongolian gerbils were inoculated with an H. pylori strain isolated from a gastric cancer patient. Four weeks later, some of the gerbils with H. pylori infection were eradicated by lansoprazole, amoxicillin, and clarithromycin. The time courses of changes in Reg gene expression, serum gastrin levels, gastric acidity, and histopathologic factors were examined. Four weeks after H. pylori infection, gastritis started spreading to the fundic mucosa, and gastric acidity started reducing. Serum gastrin levels and Reg mRNA expression in the fundus were significantly increased 6 weeks after infection. Reg mRNA expression in the fundus correlated significantly with both serum gastrin levels and the severity of fundic mucosal inflammation. After H. pylori eradication, serum gastrin levels and fundic mucosal inflammation were normalized, and the increase in Reg mRNA expression was abolished. The Reg gene is associated with hypergastrinemia and fundic mucosal inflammation and may be involved in H. pylori-induced gastritis.