Aiming to understand the mechanisms underlying the different responses to berberine among different cell lines

Berberine, a bioactive natural isoquinoline alkaloid, is known to generate a variety of pharmacological effects in different cell types. Because of its ability to arrest the cell cycle and cause apoptosis in several malignant cell lines, berberine has received attention as a potential anticancer agent. To investigate the mechanisms underlying the different responses to berberine, we started to analyze its dose-dependent and time-dependent intracellular localization in two human tumor cell lines: MIA PaCa- 2 (from pancreatic carcinoma), U343 (from glioblastoma). Human dermal fibroblasts (HDF) were used as a non-tumor control. Berberine presents natural green fluorescence, which allows identification of the intracellular site of accumulation in living cells. We found that the alkaloid may accumulate in different cell compartments, with a dynamic dose-dependent and time-dependent pattern of localization. The results revealed different localization of berberine in cytoplasm and mitochondria and/or nuclei in cancer cells with respect to non-tumor cells. Moreover, berberine treatments reduced cell viability in a cell line-specific manner. To further investigate the effects of berberine, the expression profile of genes involved at different levels in fundamental biological processes, was analyzed. As tumor suppressor genes are often methylated in the process of carcinogenesis, we evaluated DNMT1 and DNMT3B coding for maintenance and de novo methyltransferases, respectively. Additionally, MGMT, a geneencoding O6-methylguanine-DNA methyltransferase, and recognized to play a crucial role in the defense against chemotherapy alkylating agents, was analyzed. As literature data demonstrate that berberine may induce apoptosis in cancer cells but not in normal cells and this observation is valuable for development of new anti-cancer therapies, we also compared the variation in the gene expression level of cysteine- aspartic acid protease 3 (CASP3) and the activity of this enzyme in MIA PaCa-2, U343 and HDF cell lines. On the whole the results indicate that berberine differentially affects the behaviour of MIA PaCa-2, U343, and non-tumor HDF cells.