Role of Human Cripto-1 in Tumor Angiogenesis

Tumor Growth Factor Section, Mammary Biology and Tumorigenesis Laboratory, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA.
Journal of the National Cancer Institute (Impact Factor: 12.58). 02/2005; 97(2):132-41. DOI: 10.1093/jnci/dji011
Source: PubMed


Human cripto-1 (CR-1) promotes cell transformation and increases migration and invasion of various mouse and human epithelial cell lines. We investigated whether CR-1 also stimulates angiogenesis.
We used human umbilical vein endothelial cells (HUVECs) to measure in vitro migration with fibronectin-coated Boyden chambers, invasion with Matrigel-coated Boyden chambers, proliferation with a tetrazolium salt, and differentiation with an in vitro Matrigel assay. We investigated new blood vessel formation in vivo by use of Matrigel-filled silicone cylinders implanted under the skin of nude mice and by use of a breast cancer xenograft model with CR-1-transfected or control Neo-transfected MCF-7 human breast cancer cells. We also used a blocking anti-CR-1 monoclonal antibody to investigate the role of CR-1 in angiogenesis in vivo and in vitro. All statistical tests were two-sided.
CR-1 stimulated HUVEC proliferation, migration, and invasion and induced HUVEC differentiation into vascular-like structures on Matrigel. In vivo, recombinant CR-1 protein induced microvessel formation in Matrigel-filled silicone cylinders, and microvessel formation was statistically significantly inhibited with a blocking anti-CR-1 monoclonal antibody (CR-1 and antibody = 127% of microvessel formation compared with that in untreated control cylinders and CR-1 alone = 259%; difference = 132%, 95% confidence interval [CI] = 123% to 140%; P<.001). Tumors formed by CR-1-transfected MCF-7 cells in the cleared mammary fat pad of nude mice had higher microvessel density than tumors formed by control Neo-transfected MCF-7 cells (CR-1-transfected cells = 4.66 vessels per field and Neo-transfected cells = 2.33 vessels per field; difference = 2.33 vessels per field, 95% CI = 1.2 to 2.8; P = .004).
CR-1 appears to have an important role in the multistep process of angiogenesis.

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    • "We believe that our findings could underscore the specific roles of trophoblast cells at the maternal-fetal interface. CRIPTO-1 signaling within tumor cells has previously been demonstrated to modulate cellular growth, survival, and invasion in several human cancers [30, 32, 33], and this could be especially relevant to the biology of trophoblast cells. In particular, extravillous cytotrophoblast cells are nonproliferative and exhibit a low apoptotic index during the late stages of gestation, which suggests the hypothesis that CRIPTO-1 contributes to trophoblast invasiveness or cell survival mechanisms [34]. "
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    ABSTRACT: CRIPTO-(CR)1 is a protein associated with tumorigenesis and metastasis. Here we demonstrate that CR-1 expression in normal and creta placentas is associated with various degrees of uterine invasion. Cytokeratin (CK) and CR-1 protein expression was visualized by immunohistochemical staining of formalin-fixed, paraffin-embedded placental specimens (control placentas, n = 9; accreta, n = 6; increta, n = 10; percreta, n = 15). The pattern of extravillous trophoblast (EVT) cell morphology was distinctive in creta placentas: densely-compacted cell columns and large star-shaped cells with a typically migratory phenotype, not common among third trimester control placentas. Quantification revealed higher CR-1 immunoreactivities in accreta (P = 0.001), increta (P = 0.0002), and percreta placentas (P = 0.001) than in controls. In contrast to controls, there was a significant positive relationship between CR-1 and CK reactivity in all creta placentas (accreta, P = 0.02; increta, P = 0.0001, and percreta, P = 0.025). This study demonstrated CR-1 expression in the placental bed, its increased expression in creta placentas, and EVT cells as the main CR-1-producing cell type. Morphological examination revealed an immature and invasive trophoblast profile in creta placentas, suggesting impairment of the trophoblast differentiation pathway. These findings provide important new insights into the pathophysiology of abnormal creta placentation and its gestational consequences.
    BioMed Research International 08/2014; 2014:892856. DOI:10.1155/2014/892856 · 3.17 Impact Factor
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    • "We performed endothelial cell tubule formation assays (2D assays) in accordance with the previous report 16. To incorporate a monolayer of tumor cells, we modified the 2D assays in the following manner. "
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    ABSTRACT: We have developed novel phenotypic fluorescent three-dimensional co-culture platforms that efficiently and economically screen anti-angiogenic/anti-metastatic drugs on a high-throughput scale. Individual cell populations can be identified and isolated for protein/gene expression profiling studies and cellular movement/interactions can be tracked by time-lapse cinematography. More importantly, these platforms closely parallel the in vivo angiogenic and metastatic outcomes of a given tumor xenograft in the nude mouse model but, unlike in vivo models, our co-culture platforms produce comparable results in five to nine days. Potentially, by incorporating cancer patient biopsies, the co-culture platforms should greatly improve the effectiveness and efficiency of personalized chemotherapy.
    Journal of Cancer 06/2013; 4(5):402-15. DOI:10.7150/jca.6780 · 3.27 Impact Factor
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    • "In an additional study, combined analysis of Cripto-1 and E-cadherin has significant value in evaluating the metastatic potential of gastric cancer and predicting patient prognosis[6]. In vitro and transgenic mice studies have shown that cripto-1 play an important oncogenic role during tumorigenesis by promoting cell proliferation, survival, migration and invasion, as well inducing epithelial-to-mesenchymal transition(EMT), transformation, branching morphogenesis and tumour angiogenesis[9,10]. "
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    ABSTRACT: Human Cripto-1, a member of the EGF-CFC family, is indispensable for early embryonic development. Cripto-1 plays an important oncogenic role during tumorigenesis and is overexpressed in a wide range of epithelial carcinomas, yet little is known about Cripto-1 in nasopharyngeal carcinoma (NPC). The aim of this study was to analyze the roles of Cripto-1 in the progression and clinical characteristics in NPC clinical samples and cell lines. The expression of Cripto-1 at mRNA level was detected by the reverse transcription-polymerase chain reaction (RT-PCR) and real time RT-PCR, and western blot was used to examine the protein expression. Cripto-1 expression and its clinical characteristics were investigated by performing immunohistochemical analysis on a total of 37 NPC clinical tissue samples. Lentiviral vectors were constructed to get an efficient expression of anti-Cripto-1 siRNA in CNE-2 and C666-1 cells, with invalid RNAi sequence as control. After the inhibition of the endogenous Cripto-1, the growth, cell cycle and invasion of cells were detected by MTT, FACS and Boyden chamber assay respectively. Moreover, in vivo, the proliferation of the tumor cells was evaluated in xenotransplant nude mice model with whole-body visualizing instrument. The results of real-time RT-PCR and western blot showed that the expression level of Cripto-1 was markedly higher in NPC cell lines than that in the immortalized nasopharyngeal epithelial cell at both mRNA and protein levels. RT-PCR of 17 NPC tissues showed a high expression rate in 76.5% (13/17) cases. In an immunohistochemical study, Cripto-1 was found to express in 54.1% (20/37) cases of NPC. In addition, Cripto-1 overexpression was significantly associated with N classification (p = 0.034), distant metastasis (p = 0.036), and clinical stage (p = 0.007). Inhibition of endogenous Cripto-1 by lentivirus-mediated RNAi silencing technique suppressed NPC cell growth and invasion in vitro. In vivo, the average weight (p = 0.026) and volume (p = 0.044) of tumor in CNE-2/GFP+/Cripto-1- xenotransplant mice group were significantly lower than those in the control group. The Ki67 index was obviously lower in Cripto-1 RNAi treated tumors (p < 0.01). Data of this study suggest that Cripto-1 overexpression is connected with the tumorigenesis and progression of NPC, lentivector-mediated RNAi might be feasible for the inhibition of the growth and invasion of NPC.
    BMC Cancer 10/2009; 9(1):315. DOI:10.1186/1471-2407-9-315 · 3.36 Impact Factor
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