| Ovarian cancer is considered the most fatal cancer of all gynecologic malignancies. In China, the morbidity of ovarian carcinoma is high, only after that of breast cancer and cervical cancer. The mortality of ovarian carcinoma is the highest among these three gynecological malignancies, impairing women's health seriously. About 70% of ovarian carcinoma cases are in advanced grade at the time of diagnosis, losing the opportunity of radical excision of the diseased organ. Chemotherapy is the important assistance to the treatment of ovarian carcinoma, including those that can't be radically excised. For these cases, combined chemotherapy after tumor reduction is a critical measure for improving the prognosis of ovarian carcinoma patients. Chemoresistance exerts profound impact on prognosis of ovarian carcinoma. Among the various chemical pharmaceutics for treatment of ovarian carcinoma, platinum compounds are most effective. The effective power may reach 29%-35%. Therapeutic effect of Cisplatin (DDP) is dose-dependent. The dose-dependent toxic effects and drug resistance decrease its anti-carcinoma effects. The five year survival rate of ovarian cancer patients is low because of drug resistance of DDP. There is no satisfactory strategy to solve this contradiction.Establishing and culturing specific cell lines to study their behavior changes under different biological environment is fundamental for the understanding of chemoressitance. SKOV3 is a cell line isolated from ovarian carcinoma patient of serous papillary cystadenocarcinoma, the most aggressive and lethal type of human ovarian carcinoma. In the present study, we developed a SKOV3 cell line resistant to chemotherapeutic drug cisplatin using intermittent induction. This cisplatin-resistant ovarian carcinoma cell line served a convenient research subject for understanding the mechanisms of drug resistance and for exploring new approach to increase the sensitivity of chemoresistant ovarian carcinoma.MSCs are known to possess multi-lineage differentiation potentia. They can be directed to differentiate into specific cell lineages such as fat, bone and cartilage tissues under different micro-environmental conditions. They also exert therapeutic effects through endocrine and paracrine. These characteristics make the cells unique for various therapeutic potentials such as supporting tissue regeneration, correcting inherited disorders, inhibiting chronic inflammation and releasing biological agents. Therefore, administration of MSCs may be a promising new strategy to treat a variety of diseases.The current research aimed to determine the feasibility of treating ovarian carcinoma resistant to chemotherapeutic cisplatin with bone marrow-derived mesenchymal stem cells. It has been reported that tumor entity possesses the unique ability to attract self bone marrow-derived mesenchymal stem cells to the local tumor. Based upon this chemoattraction of tumor tissues, we proposed that intravenously-injected MSCs would selectively locate in the capillaries malignant tissues. As a result, MSCs decreased blood supply and changed cytokine environment of targeted malignancy mass, offering a new strategy for cancer therapy independent of chemoresistant mechanisms. The current study includes the following three parts: Part 1:Development of ovarian carcinoma cell line resistant to chemotherapeutic cisplatin. In this study, we performed intermittent induction method to render human ovarian carcinoma cell line SKOV3 refractory to cisplatin. The resistant phenotype of the resistant cell line is very stable. The sensitive cells were gradually depleted after repeat incubation in drug-containing medium until the cells survived the cytotoxicity of significantly overdose cisplatin. The cisplatin-resistant ovarian carcinoma cells were morphologically different from cells sensitive to cisplatin. The latter were larger than the resistant cells. The resistant cells displayed the typical morphological features of stem cells. Besides, using flow cytometry, we found 99.3% of the resistant cells were positive to stem cell marker CD44 while only 0.04% of the sensitive cells were CD44-positive. Therefore, we successfully developed a cisplatin-resistant cell lines positive to stem cell marker CD44 for subsequent experimental studies.Part 2:Observation of in vitro biological behaviors of ovarian carcinoma cells SKOV3 with chemoresistance to cisplatin. Through cell proliferation and adhesion experiments, we found that after addition of cisplatin into culture medium, ciplatin-resistant SKOV3 proliferated more quickly than its counterpart SKOV3 sensitive to cisplatin under the same culture circumstances. In addition, the adhesion rate of resistant SKOV3 cells to matrix protein Matrigel was significantly lower than that of non-resistant cells, indicating the resistant-cells were less adherent and therefore easier to migrate. The percentage of resistant SKOV3 cells in G1 phase of cell cycle increased compared to correspondent non-resistant cells, indicating the resistant cells were less sensitive to chemotherapy.Part 3:Study on the effects of bone marrow-derived mesenchymal stem cells on transplanted ovarian carcinoma cells. We established an athymic mouse model burdened with ovarian carcinoma via subcutaneous injection of our developed cisplatin-resistant cells. We then injected primary bone marrow-derived mesenchymal stem cells into mice burdened with ovarian cancer via tail vein using the mice injected with PBS as control. Observation and measurement of all mass indicated that tumors in the MSCs-injected mice were smaller than those in the control mice, showing that MSCs-treated animals had a trend of decrease in tumor size compared to those without MSC administration. The result suggested that MSCs decreased the proliferation and growth of ovarian carcinoma resistant to cisplatin although the precise mechanism remained to be identified.In conclusion, we established a human ovarian carcinoma cell line which is resistant to cisplatin. The resistant cells were significantly different in morphology. cell adhesion and proliferation compared to drug sensitive cells. Our experiment also showed that bone marrow-derived mesenchymal stem cells could suppress the growth of ovarian carcinoma resistant to cisplatin in vito. |
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