Reduced tumorigenicity and drug resistance through the downregulation of octamer-binding protein 4 and Nanog transcriptional factor expression in human breast stem cells.

Breast cancer is the most common type of malignancy among females. Previous studies examining breast cancer tissue have demonstrated the presence of stem cells, and have detected octamer­binding protein 4 (Oct4) and Nanog transcription factor expression. In the present study, breast cancer stem cells (CSCs) were isolated and enriched from MDA­MB­231 breast cancer cell lines, and were defined as MDA­MB­231 stem cells using flow cytometry. The expression of Oct4 and Nanog in breast CSCs were detected by quantitative polymerase chain reaction and western blotting. RNA interference (RNAi) was used in order to downregulate the expression of Oct4 and Nanog. Drug resistance and tumor­initiating capability following in vivo injection of MDA­MB­231 stem cells trans-duced with negative RNAi, Oct4 RNAi and Nanog RNAi were compared with that of MDA­MB­231 stem cells without siRNA transfection as a control group. In addition the capability of MDA­MB­231 breast cancer cells to initiate tumor formation in mice was compared with that of MDA­MB­231 stem cells. A paclitaxel inhibition test was also conducted in order to detect resistance of MDA­MB­231 breast cancer stem cells to this treatment. The MDA­MB­231 stem cells were revealed to exhibit elevated percentages of the cluster of differentiation (CD)44+CD24­/low subset, high tumorigenicity and resistance to chemotherapy, all of which are characteristic stem cell properties. In addition, the MDA­MB­231 stem cells were more tumorigenic in vivo. Furthermore, the breast CSCs also expressed high levels of the Oct4 and Nanog transcription factors. Therefore, downregulation of Oct4 or Nanog expression may reduce chemotherapeutic drug resistance and tumorigenicity in breast CSCs. In conclusion, Oct4 and Nanog expression may be a key factor in the development of resistance to chemotherapy and tumor growth of breast CSCs. This finding indicates that Oct4 or Nanog­targeted therapy may be a promising means of overcoming resistance to chemotherapy and inhibiting tumor growth in breast cancer treatment.

[The clinical significance of preoperative enteral immune nutrition in patients with malignant gastrointestinal tumors].

OBJECTIVE: To study the impact of preoperative enteral immune nutrition on patients with malignant gastrointestinal tumors. METHODS: 82 patients with malignant gastrointestinal tumors were divided equally into 2 groups:enteral nutrition group (EN) and normal diet group (Control). Enteral Nutritional Emulsion (TPF-T) served as nasogastically-fed liquid diet for the patients in EN group over a period of 7 days prior to surgery. Normal diet was given to the patients in control group under the same condition as those in EN group in terms of calories and nitrogen contents. Enzyme linked immunosorbent assay (ELISA) was performed to determine the quantity of serum albumin (ALB), transferrin protein (TRF), pre-albumin (PA) and retinol binding protein (RBP). Flow cytometry (FCM) was performed to determine T cell subsets. Postoperative complications, resumption of peristalsis, length of hospital stay, and nutritional costs were also recorded. RESULTS: TRF, PA and RBP increased significantly in the patients in EN group compared with those in control group (P < 0.05). The patients in EN group had significantly higher proportions of CD3+, CD4+/CD8+ higher than those of control (P < 0.05). No serious complications (eg. death or gastrointestinal fistula) were found in the patients. The total nutritional cost for the patients in EN group was similar to that of the controls (P > 0.05). The patients in EN group had less postoperative complications, quicker resumption of peristalsis, shorter hospital stay and lower level of postoperative nutrition cost compared with those of controls (P < 0.05). CONCLUSION: Enteral nutrition support can improve the nutritional status and immunity of patients with malignant gastrointestinal tumors, which has both pre-operative and post-operative benefits for the patients.

[Cytosine deaminase and thymidine kinase double suicide gene system driven by carcinoembryonic antigen promoter for the treatment of colorectal carcinoma xenograft in nude mice].

OBJECTIVE: To explore the therapeutic efficacy of double suicide gene system driven by carcinoembryonic antigen (CEA) promoter (Cp-CDglyTK) on colorectal carcinoma xenograft in nude mice. METHODS: The plasmid pcDNA3.1(-)Cp-CDglyTK was transfected into the CEA-positive SW480 and CEA-negative HeLa cells respectively. The expression of suicide gene was detected by RT-PCR. And the transfected cells were treated with 5-fluorocytosine (5-FC) and ganciclovir (GCV) at different concentrations and the cell-killing and bystander effects assayed by methyl thiazolyl tetrazolium (MTT). By a transplantation of cultivated cells, SW480 or HeLa cell lines were injected subcutaneously into right axillary of nude mice to establish 96 SW480 and 72 HeLa tumor animal models. Nude mice were completely randomized with statistical software according to tumor volume. For prodrug therapy, 48 SW480-bearing mice were divided equally into 4 groups of I-IV. At the same time, 48 HeLa-bearing mice were divided equally into 4 groups of V-VIII. Groups I & V received an intratumoral injection of PBS, groups II & VIGCV and 5-FC intratumorally, groups III & VII PBS intraperitoneally and groups IV & VIII GCV and 5-FC intraperitoneally. Forty-eight SW480-bearing mice were divided equally into 4 groups of IX∼XII and 24 Hela-bearing ones into groups of & in therapy experiment by suicide gene plus prodrug. Six groups received an intratumoral injection of liposome Lipofectamine and plasmid CP-CDglyTK and then an intraperitoneal injection of drug. The groups of IX and received an injection of PBS, group X GCV, group XI 5-FC and groups XII & GCV and 5-FC. The observation parameters included tumor bulk, tumor weight, survival time and treatment effect in each group. RESULTS: SW480 cells transfected by plasmid pcDNA3.1(-)Cp- CDglyTK expressed CDglyTK gene. The inhibition rates of GCV and 5-FC were significantly higher than those of HeLa cells (59.87% ± 0.21% vs 9.90% ± 0.09%, P < 0.01). And higher inhibition rates and stronger bystander effect existed in double versus single produg (all P < 0.05). Tumor size, final tumor weight and survival time of nude mice in groups ofII, IV, VI & VIII had no significant difference with groups ofI, III, V & VII (all P < 0.05). Final tumor size and weight of group XII was significantly smaller than those of groups of IX, X and XI ((150.0 ± 3.2) vs (522.5 ± 1.9) and (256.8 ± 10.4) and (260.7 ± 2.2) mm(3), (54.1 ± 10.4) vs (682.0 ± 12.0) and (251.8 ± 15.1) and (271.6 ± 17.7) mg, all P < 0.05). Meanwhile, the tumor inhibition rate and survival time of group XII(92.1% and (25.7 ± 0.8)d) were significant higher and longer than group X (63.1% and (21.8 ± 0.5) d) and group XI (60.2% and (18.0 ± 0.9) d) (all P < 0.05). However, no significant difference existed in tumor size, final tumor weight and survival time between groups and (all P > 0.05). The inhibition rate of group was merely 0.9%. CONCLUSION: CDglyTK double suicide gene system driven by CEA promoter may inhibit CEA positive colorectal cancer xenograft in prodrug-treated nude mice.