Projects
Name
Role of c-Met in Cholangiocarcinoma Invasion
University
Thailand (IFMSA-Thailand) - Faculty of Medicine, Vajira Hospital, Navamindradhiraj University, Bangkok
Domain
Biochemistry
Departement
Basic Medical Science
Head
Assist. Prof. Sumet Vajanarodjana, Ph.D
Tutor
Apaporn Menakongka, Ph.D.
Languages
English
Duration
4 weeks
Availability
Cities/Months Jan Feb Mar Apr May Jun Jul Augt Sep Oct Nov Dec
Yes No No No No No Yes No No No No No
Type of Research Project
- Basic science
What is the background of the project?
Cholangiocarcinoma (CCA) is the major health problem in Thailand. c-Met (hepatocyte growth factor receptor) overexpression is associated with poor prognosis in CCA patient. Moreover, our previous study showed the overexpression of c-Met in CCA cell lines and the role of HGF/c-Met signaling in CCA cell invasion. Since CCA responds poorly to chemotherapy, more effective therapeutic protocol, such as targeted drug therapy, is needed for this cancer. c-Met becomes an attractive target for cancer therapy because of its pivotal role in cancer metastasis, c-Met inhibitor might be one of the drug that specific for CCA. The aim of the project is to 1. Determine c-Met expression and activation in CCA cell lines by western blotting. 2. Examine cell invasive ability of various CCA cell lines with different level of c-Met expression by wound healing assay 3. Correlate the invasive ability with the levels of c-Met expression and activation. 4. Study the effect of c-Met inhibitor on inhibition of CCA cell invasion and correlate the sensitivity of the inhibitor with the levels of c-Met expression and activation. The techniques using in this project is 1. cell culture technique Methods for culturing human cholangiocarinoma cell lines -Routine Feeding and Maintenance -Subculture the cell -Cell counting -Cryopreservation 2. western blotting -seed cancer cell lines in culture plate -lysis of cell sample in appropriate lysis buffer (eg RIPA) -Determine protein concemtration -separate protein by SDS-PAGE -transfer proteins from the gel to membrane -probe the membrane with specific antibody -Detect band of protein on membrane 3. wound healing assay -seed cancer cells in 24-well plates and culture until confluent -using a pipette tip make a straight scratch, simulating a wound -imaging of both wound edges using the 10x objective -Grow cells for additional 6-24 h -imaging of cell culture again using the 10x objective -the gap distance can be quantitatively evaluated using software such as Photoshop or ImageJ.
What is the aim of the project?
1. Determine c-Met expression and activation in CCA cell lines by western blotting. 2. Examine cell invasive ability of various CCA cell lines with different level of c-Met expression by wound healing assay 3. Correlate the invasive ability with the levels of c-Met expression and activation. 4. Study the effect of c-Met inhibitor on inhibition of CCA cell invasion and correlate the sensitivity of the inhibitor with the levels of c-Met expression and activation.
What techniques and methods are used?
1. cell culture technique Methods for culturing human cholangiocarinoma cell lines -Routine Feeding and Maintenance -Subculture the cell -Cell counting -Cryopreservation 2. western blotting -seed cancer cell lines in culture plate -lysis of cell sample in appropriate lysis buffer (eg RIPA) -Determine protein concemtration -separate protein by SDS-PAGE -transfer proteins from the gel to membrane -probe the membrane with specific antibody -Detect band of protein on membrane 3. wound healing assay -seed cancer cells in 24-well plates and culture until confluent -using a pipette tip make a straight scratch, simulating a wound -imaging of both wound edges using the 10x objective -Grow cells for additional 6-24 h -imaging of cell culture again using the 10x objective -the gap distance can be quantitatively evaluated using software such as Photoshop or ImageJ
What is the role of the student?
- The student will observe the practical experiments but will be highly involved in the analysis of the results
What are the tasks expected to be accomplished by the student?
Student should be able to - perform in vitro culture of cholangiocarcinoma cell lines on his/her own: Including maintaining cultured cells, passaging cells (subculturing adherent cells), freezing and thawing cells, and seeding cells in culture plates for western blotting and wound healing experiments - perform western blotting and wound healing assay under supervision
Will there be any theoretical teaching provided (preliminary readings, lectures, courses, seminars etc)
1
What is expected from the student at the end of the research exchange? What will be the general outcome of the student?
- The student will prepare a presentation
What skills are required of the student? Is there any special knowledge or a certain level of studies needed?
In order to work with cell culture successfully, it depends on keeping the cells free from contamination by microorganisms such as bacterial, fungi, and viruses. In cell culture laboratory, the sterile technique for preventing contamination of your cell is required. Sterile techniques for cell culture are composed of a sterile work area, good personal hygiene, sterile reagents and media, and sterile handling. Sterile handling such as: Always wipe your hands and your work area with 70% ethanol. Wipe the outside of the containers, flasks, plates, and dishes with 70% ethanol before placing them in the cell culture hood. Use sterile glass or disposable plastic pipettes and a pipettor to work with liquids, and use each pipette only once to avoid cross contamination. Always cap the bottles and flasks after use and seal multi-well plates with tape or place them in resealable bags Use only sterile glassware and other equipment. Be careful not to talk, sing, or whistle when you are performing sterile procedures. Perform your experiments as rapidly as possible to minimize contamination. Learn more from this video: https://brightcove.hs.llnwd.net/e1/uds/pd/3663210762001/3663210762001_4419807926001_3971669648001.mp4?pubId=3663210762001&videoId=3971669648001
Are there any legal limitations in the student’s involvement
No
Hours
7
Type of students accepted
This project accepts:
- Medical students
- Graduated students (less than 6 months)
- Pre-Medical students from the American-British system
- Students in biomedical fields
Articles
- Menakongka A; Suthiphongchai T. Involvement of PI3K and ERK1/2 pathways in hepatocyte growth factor-induced cholangiocarcinoma cell invasion. World J Gastroenterol. 2010 Feb 14;16(6):713-22.
- Socoteanu MP; Mott F; Alpini G; Frankel AE. c-Met targeted therapy of cholangiocarcinoma. World J Gastroenterol. 2008 May 21;14(19):2990-4.
- Goral V. Cholangiocarcinoma: New Insights. Asian Pac J Cancer Prev. 2017 Jun 25;18(6):1469-1473.