Recording online processes in task-oriented reading with Read&Answer.

We present an application to study task-oriented reading processes called Read&Answer. The application mimics paper-and-pencil situations in which a reader interacts with one or more documents to perform a specific task, such as answering questions, writing an essay, or similar activities. Read&Answer presents documents and questions with a mask. The reader unmasks documents and questions so that only a piece of information is available at a time. This way the entire interaction between the reader and the documents on the task is recorded and can be analyzed. We describe Read&Answer and present its applications for research and assessment. Finally, we explain two studies that compare readers' performance on Read&Answer with students' reading times and comprehension levels on a paper-and-pencil task, and on a computer task recorded with eye-tracking. The use of Read&Answer produced similar comprehension scores, although it changed the pattern of reading times.

Targeted adriamycin delivery to MXT-B2 metastatic mammary carcinoma cells by transferrin liposomes: effect of adriamycin ADR-to-lipid ratio.

In the protein-targeted therapy for cancer, transferrin (Tf) is used to reach a selective and specific target in cancer cells. Tf is used conjugated to chemotherapeutic drugs, insulin, toxins, antibodies, polymers, nanoparticles, lipoplexes and liposomes. Using this latter approach, hydrophobically derivatized Tf was incorporated to liposomal bilayers. The biological activity of Tf-liposome was tested using MXT-B2 cells, a metastatic mammary carcinoma cell line. In Tf binding assays, the Scatchard analysis indicated 4.5x10(5) Tf receptors/cell. In cell growth assays, Tf-liposomes stimulated cell growth in a dose-dependent manner, up to a maximum of 32% of the total free Tf stimulation. Following this, we prepared Tf-liposomes encapsulating adriamycin (ADR) at two different ADR-to-lipid ratios. In vitro cytotoxicity assays against MXT-B2 cells gave IC(50) values 2.1-times lower for Tf-liposomal ADR in comparison to control liposomal ADR. However, similar IC(50) values were found for low ADR-to-lipid ratio Tf-liposomal ADR, as well as for control liposomal ADR. The free Tf added in excess increased the IC(50) value of Tf-liposomal ADR by 51%, while the IC(50) value of control liposomal ADR was unaffected, supporting a receptor-mediated mechanism of targeting by Tf. In addition, the lower IC(50) value is correlated with a higher total of ADR accumulation in the cells.

Overproduction of VEGF concomitantly expressed with its receptors promotes growth and survival of melanoma cells through MAPK and PI3K signaling.

Vascular endothelial growth factor (VEGF) is an important mediator of tumor-associated angiogenesis, and consequently it has been associated with metastasis. We report here that the overexpression of VEGF(165) in melanoma xenografts promotes an acceleration of tumor growth and an increase in angiogenesis as well as the spontaneous metastasis formation. In addition, VEGF receptors (VEGFR)1, VEGFR2 and neurophilin-1 are expressed in A375 melanoma cells. Forced overexpression of VEGF in these cells induces cell growth and triggers survival activity in serum-starved cultures, by a mechanism dependent on the mitogen-activating protein kinase signaling pathway. Furthermore, these effects are dependent MEK 1/2 activity. Kinase domain region-specific tyrosine kinase inhibitors dramatically reduced DNA synthesis to 20% with respect to the controls, although they did not completely suppress either the p44 or p42-phosphorylated forms of extracellular signal-regulated protein kinase. These inhibitors also provoked a decrease in Akt phosphorylation. We observed a dramatic reduction in survival after treatment with phosphatidylinositol 3'-kinase (PI3K)-specific inhibitor in the presence of specific tyrosinase inhibitors. We suggest that the overproduction of VEGF(165) concomitantly expressed with its receptors favors cell growth and survival of melanoma cells through MAPK and PI3K signaling pathways. These data support the involvement in melanoma growth and survival of a VEGF-dependent internal autocrine loop mechanism, at least in vitro.

Requirement of plasminogen binding to its cell-surface receptor α-enolase for efficient regeneration of normal and dystrophic skeletal muscle.

Adult regenerative myogenesis is central for restoring normal tissue structure and function after muscle damage. In muscle repair after injury, as in severe myopathies, damaged and necrotic fibers are removed by infiltrating inflammatory cells and then replaced by muscle stem cells or satellite cells, which will fuse to form new myofibers. Extracellular proteolysis mediated by uPA-generated plasmin plays a critical role in controlling inflammation and satellite-cell-dependent myogenesis. α-enolase has been described as plasminogen receptor in several cell types, where it acts concentrating plasmin proteolytic activity on the cell surface. In this study, we investigated whether α-enolase plasminogen receptor plays a regulatory role during the muscular repair process. Inhibitors of α-enolase/plasminogen binding: MAb11G1 (a monoclonal antibody against α-enolase) and ε-aminocaproic acid, EACA (a lysine analogue) inhibited the myogenic abilities of satellite cells-derived myoblasts. Furthermore, knockdown of α-enolase decreased myogenic fusion of myoblasts. Injured wild-type mice and dystrophic mdx mice were also treated with MAb11G1 and EACA. These treatments had negative impacts on muscle repair impairing satellite cell functions in vitro in agreement with blunted growth of new myofibers in vivo. Furthermore, both MAb11G1 and EACA treatments impaired adequate inflammatory cell infiltration and promoted extracellular matrix deposition in vivo, which resulted in persistent degeneration. These results demonstrate the novel requirement of α-enolase for restoring homeostasis of injured muscle tissue, by controlling the pericellular localization of plasmin activity.

Upregulation of MMP-9 in MDCK epithelial cell line in response to expression of the Snail transcription factor.

Overexpression of the transcription factor Snail in epithelial MDCK cells promotes the epithelial-mesenchymal transition (EMT) and the acquisition of an invasive phenotype. We report here that the expression of Snail is associated with an increase in the promoter activity and expression of the matrix metalloproteinase MMP-9. The effect of Snail silencing on MMP-9 expression corroborates this finding. Induced transcription of MMP-9 by Snail is driven by a mechanism dependent on the MAPK and phosphoinositide 3-kinase (PI3K) signalling pathways. Although other regions of the promoter were required for a complete stimulation by Snail, a minimal fragment (nucleotides -97 to +114) produces a response following an increased phosphorylation of Sp-1 and either Sp-1 or Ets-1 binding to the GC-box elements contained in this region. The expression of a dominant negative form of MEK decreased these complexes. A moderate increase in the binding of the nuclear factor kappaB (NFkappaB) to the upstream region (nucleotide -562) of the MMP-9 promoter was also observed in Snail-expressing cells. Interestingly, oncogenic H-Ras (RasV12) synergistically co-operates with Snail in the induction of MMP-9 transcription and expression. Altogether, these results indicate that MMP-9 transcription is activated in response to Snail expression and that it might explain, at least in part, the invasive properties of the Snail-expressing cells.