Berberine Inhibition Of Tumor Cells Of Cyclin D1 Associated Signaling Pathway

At present, tumor morbidity still sticks at a high level in China even the world. It brings not only gigantic agony to the patients and their families, but also immense threat on the development and progress of society. Therefore, exploring safe, effective, low poisonous anti-tumor drugs is taken seriously by medical scientists. Developing this kind of drugs from natural compounds become the hot spot of researches. A common phenotype of all cancer cells is cell-cycle abnormal. The regulation and control of cell-cycle is related to intracellular signal pathways, therefore decrease the expression of up-regulators of cell-cycle through suppressing intracellular signal pathways is feasible for tumor therapy. This study verificatied the suppress effect of berberine on the cell-cycle progress and the expression of Cyclin D1, which was one of the most important up-regulators of cell-cycle. The effects of berberine on signal pathways related to Cyclin D1 were studied. The molecular mechanism of the suppress effect of berberine on Cyclin D1 expression was elucidated, so as to provided the theory basis to the anti-tumor application of berberine. Part I: The effect of berberine on the cell-cycle progress and the expression of Cyclin D1 in PG cells.1. MTT method was used to detected the effect of berberine on the proliferation of PG cells. Compared with control group, the proliferation of PG cells treated with different concentrations (10μg/ml, 20μg/ml, 40μg/ml) of berberine for 24 hours and 48 hours was suppressed significantly.2. The effect of berberine on cell-cycle progress of PG cells was detected by FACS. After treated with berberine, the proportion of PG cells at G0/G1 phase increased while cells at S phase and G2/M phase decreased.3. RT-PCR results demonstrated that the level of Cyclin D1 mRNA in PG cells decreased after treated with berberine for 24 hours. Berberine could inhibit the expression of Cyclin D1 in PG cells.Part II: The inhibitory effect of berberine on the signal pathways related to Cyclin D1 in PG cells. The molecular mechanism of the suppress effect of berberine on Cyclin D1 expression was elucidated.1. Reporter-gene results demonstrated that berberine could suppress AP-1 and Wnt signal pathway in PG cells while had no significant effect on NF-κB signal pathway. Berberine could significantly inhibit AP-1 signal pathway even while it activated by PDB+ionomysin.2. Western Blot results showed that the level of c-Jun in PG cells were decreased after treated with different concentration of berberine.3. EMSA results showed that berberine also could inhibit the combination of nuclear extract and oligonucleotide containing wild type AP-1 site. Berberine could inhibit the combination of nuclear extract and oligonucleotide from Cyclin D1 gene promoter region containing AP-1 site. The mute probes had no effect on the combination.Conclusion:1. Berberine blocks the cell-cycle progress to inhibit the proliferation of PG cells through suppressing the expression of Cyclin D1.2. Berberine inhibits the activity of AP-1 and Wnt pathway while has no significant effect on NF-κB pathway. Berberine significantly inhibits AP-1 signal pathway even while it activated by PDB+Ionomysin.3. Berberine inhibits c-Jun in PG cells which is one of the most important components of AP-1 transcription factors. Berberine suppressing the combination of AP-1 site on Cyclin D1 gene promoter region and AP-1 transcription factors is one of the molecular mechanisms which induce the expression of Cyclin D1 decreasing and the cell-cycle progress abnormal.