IDIES Project

 The IDIES project began in 2014 at CEBAS-CSIC. The main objective of this project is the initiation of high school students (research modality) to scientific research participating and collaborating in projects led by researchers of recognized national and international prestige.

This year, under the supervision of Dr. Giménez-Bastida, Dr. González-Sarrías and Prof. Juan Carlos Espín, students of the IES Domingo Valdivieso (Mazarrón, Murcia) will collaborate in the PolyBiota project. The students will work on the "In vitro evaluation of the effect of dietary polyphenols against angiogenesis in human aortic endothelial cells". The scientific activities will be mainly carried out at the University of Murcia (Cell culture service) and CEBAS-CSIC, where they will have the opportunity to learn cellular and molecular techniques (cell culture, cytotoxicity, endothelial cell migration or western blot) via hands-on activities and experiments in the laboratory followed by the analysis of the data collected. The results will be the basis for the preparation of an oral presentation and a poster that will be presented in the VIII IDIES Conference that will be held at the University of Murcia in June 2022.

INITIAL MEETING                                                                                                                                  

As in previous years, we started the IDIES project 2021 with an initial (on line) meeting in which we had the opportunity to get know each other. In this initial meeting, we explained to the students the design of the project, important scientific concepts that they should know as well as the different scientific activities to be performed in each visit to CEBAS-CSIC. 

 

FIRST SESSION: INTRODUCTION TO THE LABORATORY AND CELL CULTURE                

The Cell Culture Service at the University of Murcia was the place where we performed all the activities planned for this day. In the first activity, we explained to the students the general laboratory safety rules. We also showed and explained the equipment they can found in a laboratory, including centrifuges, hoods, incubators, etc.
The first activity they worked on was the preparation of the compounds we want to investigate. This activity allowed them to learn important concepts related to the preparation of solutions such as molarity. Besides, the students also used the precision balance, micropipettes, multichannel pipettes and practiced the sterilization of compounds that will be used to treat cells under sterile conditions.

In a second set of experiments, the students learned about different types of culture medium, they used the microscope to observe cells, changed culture medium, participated in cell subculturing and counted them using the trypan blue dye exclusion technique.

It was a long day but it was worth it!!!! See you in the second session.

SECOND SESSION: CYTOTOXYC ACTIVITY                                                                                  

"The dose makes the poison" is an adage credited to Paracelsus. This expression refers to the capacity of compounds present in nature such as natural molecules to exert toxic effects. Our objective today was to determine the capacity (or a lack of it) of a group of dietary phenolic compounds to exert cytotoxic activity against an in vitro cellular model.

Our students learned the MTT assay, a widely used technique that allows determining whether a molecule, at the concentration investigated, is(are) able to exert cytotoxic effects on a cell line. This assay is based on the ability of the cells to reduce the reagent 3-(4,5-dimethylthiazol-2-yl) to formazan (purple). Thus, the higher purple color formed the lower toxicity.

The students prepared different concentrations of a stock solution of dietary phenolic compounds (prepared in the first session) both in organic solvents and culture medium. They also treated the cells (seeded in 96-well plates) with the compounds, at different concentrations, and incubated them for 4 h. Next, they washed the cells with phosphate-buffered saline and added the reagent 3-(4,5-dimethylthiazol-2-yl (aka MTT). After 4 h of incubation, the MTT was removed and the formazan generated diluted in DMSO.

Our students quickly developed a clear understanding of the technique and worked hard to obtain accurate data. As shown in the next figure, the nice results obtained leave no room for doubt that they performed a great job (even after a hard morning of classes).

Next session is coming soon. See you there!

THIRD SESSION: MIGRATION AND TUBULOGENESIS OF ENDOTHELIAL CELLS             

Learning new techniques is a "must do" for our students, and today was not an exception. We worked on two different techniques, which are widely approached to investigate important physiological (and pathophysiological) processes such as angiogenesis. One of the assays was the technique known as wound healing. This assay allows investigating how a molecule (i.e., phenolic compounds) is able to modulate, promoting or inhibiting, the capacity of the endothelial cells to migrate. Another technique learned was tubulogenesis. Here, we tested whether the phenolic compounds investigated had any effect on the tubulogenic capacity of the cells. Finally, our students analyzed the images taken using the software ImageJ (and the Angiogenesis Analyzer plug-in) and started the discuss the results obtained. Are you curious about the possible relevance of our results in processes like tissue repair and(or) tumor growth? Find answer to this question in the upcoming IDIES Conference. You cannot miss it!!

FOUR SESSION: WESTERN BLOT AND PROTEIN ANALYSIS                                                     

Our students not only learn what effects are exerted by the phenolic compounds investigated, but also how these compounds modulate the cellular response. The western blot is an essential technique to determine how the activation (or alteration of this activation) of key proteins are involved regarding the effects of the phenolic compounds. The JAEintro fellowship Irene explained how to elaborate the acrylamide gels and the role of each element used in its preparation. A string of steps such as electrophoresis and transfer buffers preparation, samples loading, electrophoresis or membrane blocking (among others) were followed prior to the analysis of the proteins. Did we get a nice picture of the proteins? Find out in the upcoming IDIES Conference!