Loss of the SxxSS Motif in a Human T-Cell Factor-4 Isoform Confers Hypoxia Resistance to Liver Cancer: An Oncogenic Switch in Wnt Signaling

PURPOSE: Aberrantly activated Wnt/β-catenin signaling is important in hepatocellular carcinoma (HCC) development. Downstream gene expressions involving the Wnt/β-catenin cascade occur through T-cell factor (TCF) proteins. Here, we show the oncogenic potential of human TCF-4 isoforms based on the expression of a single conserved SxxSS motif. METHODS: We investigated the TCF-4J and K isoform pair characterized by the presence (K) or absence (J) of the SxxSS motif. The mRNA expression profiles were examined in 47 pairs of human HCCs and adjacent non-cancerous liver tissues by RT-PCR. Proliferation, sphere assays and immunoblot analysis were performed under normoxia and hypoxia conditions. The ability of HCC cells overexpressing TCF-4J (J cells) and K (K cells) to grow as solid tumors in nude mice was explored. RESULTS: TCF-4J expression was significantly upregulated in HCC tumors compared to corresponding peritumor and normal liver and was preferentially expressed in poorly differentiated HCCs. In contrast, TCF-4K was downregulated in those same HCC tumors. TCF-4J-overexpressing HCC cells (J cells) revealed a survival advantage under hypoxic conditions, high proliferation rate and formation of aggregates/spheres compared to overexpression of TCF-4K (K cells). The hypoxic J cells had high expression levels of HIF-2α and EGFR as possible mechanisms to promote tumorigenesis. Increased stability of HIF-2α under hypoxia in J cells was associated with a decreased level of von Hippel-Lindau (VHL) protein, a known E3 ligase for HIF-αs. In a xenograft model, the J cells rapidly developed tumors compared to K cells. Tumor tissues derived from J cells exhibited high expression levels of HIF-2α and EGFR compared to the slow developing and small K cell derived tumors. CONCLUSIONS: Our results suggest that the specific TCF-4J isoform, which lacks a regulatory SxxSS motif, has robust tumor-initiating potential under hypoxic conditions

Inter-comparison of MAX-DOAS measurements of tropospheric HONO slant column densities and vertical profiles during the CINDI-2 campaign

We present the inter-comparison of delta slant column densities (SCDs) and vertical profiles of nitrous acid (HONO) derived from measurements of different multi-axis differential optical absorption spectroscopy (MAX-DOAS) instruments and using different inversion algorithms during the Second Cabauw Inter-comparison campaign for Nitrogen Dioxide measuring Instruments (CINDI-2) in September 2016 at Cabauw, the Netherlands (51.97∘ N, 4.93∘ E). The HONO vertical profiles, vertical column densities (VCDs), and near-surface volume mixing ratios are compared between different MAX-DOAS instruments and profile inversion algorithms for the first time. Systematic and random discrepancies of the HONO results are derived from the comparisons of all data sets against their median values. Systematic discrepancies of HONO delta SCDs are observed in the range of ±0.3×1015 molec. cm−2, which is half of the typical random discrepancy of 0.6×1015 molec. cm−2. For a typical high HONO delta SCD of 2×1015 molec. cm−2, the relative systematic and random discrepancies are about 15 % and 30 %, respectively. The inter-comparison of HONO profiles shows that both systematic and random discrepancies of HONO VCDs and near-surface volume mixing ratios (VMRs) are mostly in the range of ∼±0.5×1014 molec. cm−2 and ∼±0.1 ppb (typically ∼20 %). Further we find that the discrepancies of the retrieved HONO profiles are dominated by discrepancies of the HONO delta SCDs. The profile retrievals only contribute to the discrepancies of the HONO profiles by ∼5 %. However, some data sets with substantially larger discrepancies than the typical values indicate that inappropriate implementations of profile inversion algorithms and configurations of radiative transfer models in the profile retrievals can also be an important uncertainty source. In addition, estimations of measurement uncertainties of HONO dSCDs, which can significantly impact profile retrievals using the optimal estimation method, need to consider not only DOAS fit errors, but also atmospheric variability, especially for an instrument with a DOAS fit error lower than ∼3×1014 molec. cm−2. The MAX-DOAS results during the CINDI-2 campaign indicate that the peak HONO levels (e.g. near-surface VMRs of ∼0.4 ppb) often appeared in the early morning and below 0.2 km. The near-surface VMRs retrieved from the MAX-DOAS observations are compared with those measured using a co-located long-path DOAS instrument. The systematic differences are smaller than 0.15 and 0.07 ppb during early morning and around noon, respectively. Since true HONO values at high altitudes are not known in the absence of real measurements, in order to evaluate the abilities of profile inversion algorithms to respond to different HONO profile shapes, we performed sensitivity studies using synthetic HONO delta SCDs simulated by a radiative transfer model with assumed HONO profiles. The tests indicate that the profile inversion algorithms based on the optimal estimation method with proper configurations can reproduce the different HONO profile shapes well. Therefore we conclude that the features of HONO accumulated near the surface derived from MAX-DOAS measurements are expected to represent the ambient HONO profiles well

Cellular Phenotype-Dependent and -Independent Effects of Vitamin C on the Renewal and Gene Expression of Mouse Embryonic Fibroblasts

Vitamin C has been shown to delay the cellular senescence and was considered a candidate for chemoprevention and cancer therapy. To understand the reported contrasting roles of vitamin C: growth-promoting in the primary cells and growth-inhibiting in cancer cells, primary mouse embryonic fibroblasts (MEF) and their isogenic spontaneously immortalized fibroblasts with unlimited cell division potential were used as the model pair. We used microarray gene expression profiling to show that the immortalized MEF possess human cancer gene expression fingerprints including a pattern of up-regulation of inflammatory response-related genes. Using the MEF model, we found that a physiological treatment level of vitamin C (10−5 M), but not other unrelated antioxidants, enhanced cell growth. The growth-promoting effect was associated with a pattern of enhanced expression of cell cycle- and cell division-related genes in both primary and immortalized cells. In the immortalized MEF, physiological treatment levels of vitamin C also enhanced the expression of immortalization-associated genes including a down-regulation of genes in the extracellular matrix functional category. In contrast, confocal immunofluorescence imaging of the primary MEF suggested an increase in collagen IV protein upon vitamin C treatment. Similar to the cancer cells, the growth-inhibitory effect of the redox-active form of vitamin C was preferentially observed in immortalized MEF. All effects of vitamin C required its intracellular presence since the transporter-deficient SVCT2−/− MEF did not respond to vitamin C. SVCT2−/− MEF divided and became immortalized readily indicating little dependence on vitamin C for the cell division. Immortalized SVCT2−/− MEF required higher concentration of vitamin C for the growth inhibition compared to the immortalized wildtype MEF suggesting an intracellular vitamin C toxicity. The relevance of our observation in aging and human cancer prevention was discussed

Crosstalk between Medulloblastoma Cells and Endothelium Triggers a Strong Chemotactic Signal Recruiting T Lymphocytes to the Tumor Microenvironment

Cancer cells can live and grow if they succeed in creating a favorable niche that often includes elements from the immune system. While T lymphocytes play an important role in the host response to tumor growth, the mechanism of their trafficking to the tumor remains poorly understood. We show here that T lymphocytes consistently infiltrate the primary brain cancer, medulloblastoma. We demonstrate, both in vitro and in vivo, that these T lymphocytes are attracted to tumor deposits only after the tumor cells have interacted with tumor vascular endothelium. Macrophage Migration Inhibitory Factor (MIF)” is the key chemokine molecule secreted by tumor cells which induces the tumor vascular endothelial cells to secrete the potent T lymphocyte attractant “Regulated upon Activation, Normal T-cell Expressed, and Secreted (RANTES).” This in turn creates a chemotactic gradient for RANTES-receptor bearing T lymphocytes. Manipulation of this pathway could have important therapeutic implications

Imaging of Angiotropism/Vascular Co-Option in a Murine Model of Brain Melanoma:Implications for Melanoma Progression along Extravascular Pathways

Angiotropism/pericytic mimicry and vascular co-option involve tumor cell interactions with the abluminal vascular surface. These two phenomena may be closely related. However, investigations of the two processes have developed in an independent fashion and different explanations offered as to their biological nature. Angiotropism describes the propensity of tumor cells to spread distantly via continuous migration along abluminal vascular surfaces, or extravascular migratory metastasis (EVMM). Vascular co-option has been proposed as an alternative mechanism by which tumors cells may gain access to a blood supply. We have used a murine brain melanoma model to analyze the interactions of GFP human melanoma cells injected into the mouse brain with red fluorescent lectin-labeled microvascular channels. Results have shown a striking spread of melanoma cells along preexisting microvascular channels and features of both vascular co-option and angiotropism/pericytic mimicry. This study has also documented the perivascular expression of Serpin B2 by angiotropic melanoma cells in the murine brain and in human melanoma brain metastases. Our findings suggest that vascular co-option and angiotropism/pericytic mimicry are closely related if not identical processes. Further studies are needed in order to establish whether EVMM is an alternative form of cancer metastasis in addition to intravascular cancer dissemination