Results

During the 6 week visit to dr Jiri Zavadil's lab I have been introduced to the novel laboratory techinques used within the field of functional genomics, some of which we have performed on the Croatian samples. I gained knowledge about the mechanisms underlying cancer development. I had the opportunity to meet top scientits working in this field. Also, I had the open access to nearly all biomedical journals and had the opportunity to attend meetings and symposia on cancer research and system biology.

MicroRNA profiling analyis was done on the paired samples (unaffected and cancer RNA) of the patients operated from upper urothelial cancer associated with endemic nephropathy. The following steps were performed:

1. quality check using Nanodrop 2000 and Agilent 2100 Expert Bioanalyzer

2. selection of appropriate samples

3. RT & qPCR using Applied Biosystems megaplex RT primer pools for human miRNAs, analyzed by the microfluidics Taqman Low Density Arrays

4. obtaining raw data

5. data analysis using Gene Spring, Target Scan, Ingenuity Pathway Analysis, DAVID bioinformatics database.

We have found a signature of more than 100 miRNAs differentially modulated in tumor vs. unaffected urothelial tissue (Figure 1). Within the 111 miRs signature we found cancer related miRNAs to be upregulated and downregulated antimetastatic microRNAs, i.e. miRNAs targeting cellular movement, migration and invasion.

These results are to be integrated with the gene chip mRNA analysis which will presumably give us clues on the mechanisms involved in aristolochic acid induced carcinogenesis.

Figure 1. A 111 microRNAs differentialy expressed in the tumor vs unaffected urothelial tissue