| Breast cancer is a common malignancy among women, and is one of the important threats for women health. It is estimated that around 500,000 women die of breast cancereach year. Of which 69% arefrom developing countries. With the increase of average life span and the merge of westernized life style, breast cancer incidence in developing countries has increased year by year. The statistics show that Chinahas 1.1 million new cases of breast cancer each year, the increase rate is2-3% annually. Due to the fast proliferation of the tumor cells, DNA replication is much more active in tumor cells than in normal cells. Researchers have developed various type of drugs totarget theactive DNA replication. In the clinical treatment of breast cancer, there are some drugs to inhibit the DNA replication, such as doxorubicin,cislpatin,cyclophosphamide and carboplatin. However, these drugs could have toxicityin normal cells of the patients, especially in stem cellswhich have activeDNA proliferation. The toxicityfor normal cells could result in serious side effects. Thus, it is urgent to study the way to reduce theside effects of drugs in clinical pratice.The Puer tea has a long history to be a functional drink for health care, and has been recorded in Chinses tradional medcine books long time ago. The modern medical studies have shown that the Puer tea could have anti-inflammatory, lipid-lowering, antihypertensiveeffectsother than the traditional health effects. Meanwhile, the previous datafrom our lab have been shown that the mouse tumor cells were sensitized for DNA toxic doxorubicin after joint treatment of Puer tea and doxorubicin. At the same time, the joint treatment was able to protect normal mousecells from killing ofdoxorubicin. Toverifythephenomenon that Puer tea joint chemotherapy drugs could kill the tumor cell, but not normal cell in hunman cells. This studyaims to understand the act of Puer tea joint chemotherapy drugs in human cells and its possible mechanism. Our study will provide new ideas for the treatment of cancer, anda solid theoretical foundation for the application of Puer tea inclinically.We used breast cancer cells as the main objects, and study the effect of Puer tea combined with cisplatin. Firstly, we tseted human breast cancer MDA-MB-468 bearing a hot mutaion R273H of p53 and set control group, Puer tea treatment group, cisplatin treatment group and joint group. We detected the cell proliferation by SRB colorimetric assay. The results showed that the joint treatment could sensitize the tumor cell more to cisplatin compared with the treatment of cisplatin only. As the same time, even the low-dose cisplatin (0.1μM) combined with Puer tea have already markedlyreduce the the tumor cells’ proliferation. Meanwhile, pulsed field gelelectrophoresis (PFGE) assay revealed thatthe double strand breaksDNA damge become more severe by the treatment of puer combined with 0.1μM cisplatin than treated by cisplatin only or Puer tea only.. Morever, treated by Puer tea and cisplain could increase the expression level ofγ-H2AX, which is one of DNA damage stress responseproteins.More cleaved PARP were detected, which is relative to increased apotoisis.Heat shock protein HSP70/90 decreased compared with the treatment of cisplatin only or Puer tea only.Secondly, we tested another human breast cancer cell T47D which carrys a non-hot mutation L194F of p53 andthe methylation of p16 gene in the promoter region. The same treatments were done as with MDA-MB-468 cells. The results showed that T47D was sensitized by the joint treatment of Puer tea and cisplatin compared with cisplatin only or Puer tea only. And showed in PFGE assay, T47D, treated by puer and cisplatin, could result in more serious DNA damage compared with cisplatin only or Puer tea only. Like in MDA-MB-468, the expression level ofy-H2AX and p-p53 increased more than other treatments (cisplatin only or puer only). However, when treated by Puer tea and cisplatin, the expression of HSP70/90 did not decline. In fact, their expression was stable in T47D cell.Interestingly, by RT-PCR and Western Blot assaya, we found that p16 expression increased after treated by Puer tea only or Puer tea and cisplatin. These datasuggested that the status of the methlytion in p16 promoter region had been changed. In another word, Puer tea treatment could demethylate p16. Then, the senescencetest throgh via β-galactosidaseactivity assay showed that the senescent cell rate was higher in T47D cellstreated with Puer tea and cisplatin than with other treatments (cisplatin only or Puer tea only).From what has been discussed above, we found that joint use ofPuer tea and cisplatin could not only sensitized human breast cancer cell (MDA-MB-468 and T47D), but also not harm the normal breast cells (MCF-10A). Ourstudy also showed that the treatment of Puer tea and cisplatin could result in severs DNAdamage in the breast cancer cells T47D and MDA-MB-468 via. However, the molecular mechanism was not quite the same. The changeof HSP70/90 expression level after Puer tea and cisplatin treatment varied,was stable in in T47D cells butdeclined in MDA-MB-468 cells. This indicates there maybe different pathwaysfor killing different tumorcelis. The further manchanism should be studied.p16 methlyation is broadly found in many of human tumors, Puer tea could be a wild spectrum anti-tumor product. |
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