ABSTRACT
BACKGROUND:Previous studies have found that the expression level of miR-486 in glioma stem cels (CD133+) is significantly down-regulated compared with that in glioma non-stem cels (CD133-), but the effect of down-regulation of miR-486 on CD133+ cels remains unclear . OBJECTIVE: To explore the effect of miR-486 on CD133+ cels. METHODS:CD133+ glioma stem cels and CD133- glioma cels were separated from U87 cels by flow cytometer. miR-486 overexpression glioma stem cels were constructed by lipofection transfection. RESULTS AND CONCLUSION:After sorting and purification, the content of the CD133+ fraction was enriched up to 83.5%. The expression level of miR-468 in CD133+ glioma stem cels was obviously down-regulated compared with that in CD133- glioma cels. CD133+ glioma stem cels overexpressing miR-486 were fabricated successfuly. Results from in vitro experiments showed that miR-486 overexpression could dramaticaly decrease the proliferation of glioma stem cels, induce a cel cycle arrest in G1/S phase for CD133+ glioma stem cels and promote cel apoptosis. These findings suggest that miR-486 can be a suppressor of glioma stem cels, which offers a novel potential therapeutic target for glioma stem cels and human glioma.
ABSTRACT
BACKGROUND:Tumor cels are resistant to chemotherapeutic drugs, and drug resistance is closely correlated with tumor stem cels. Therefore, how to kil tumor stem cels wil become the key to the treatment of oral squamous cel carcinoma. OBJECTIVE:To study the effect of 5-fluorouracil on biological characteristics of tongue squamous cel carcinoma Tca8113 cels. METHODS:Viability of Tca8113 cels treated with different concentrations of 5-fluorouracil was determined by cel counting kit-8, and the best drug concentration and time were screened for subsequent experiments. Tca8113 cels without 5-fluorouracil acted as control group. Then the cel cycle and percentage of the side population cels in Tca8113 cels were determined by flow cytometry. Scratch test was used to determine the migration ability of Tca8113 cels. RESULTS AND CONCLUSION:Results from cel counting kit-8 showed that 5-fluorouracil inhibited the viability of Tca8113 cels positively in a time- and dose-dependent manner. Tca8113 cels under intervention with 50 mg/L 5-fluorouracil for 48 hours showed lowest cel viability. Flow cytometry results showed that in the experimental group, G0/G1 phase cels increased significantly compared with the control group (P=0.01), S phase cels decreased significantly compared with the control group (P=0.244), and G2/M phase cels disappeared completely. After treatment with 5-fluorouracil, the percentage of side population cels was increased significantly (P=0.00). The scratch test showed that in the experimental group, the cels had better ability of wound healing than those in the control group. In conclusion, 5-fluorouracil can enrich the cancer stem cel population in Tca8113 cels.
ABSTRACT
BACKGROUND:In the early development of zebrafish embryos, cels divide and proliferate rapidly, but low concentration of deguelin can delay the development of zebrafish embryos. OBJECTIVE:To observe the effect of different concentrations of deguelin on cyclin D1 gene expression in zebrafish embryos. METHODS:Though normal fertilization, zebrafish embryos that incubated to the 2-cel stage (about 0.75 hour after fertilization) and shield stage (6 hours after fertilization) were colected and put into 12-wel plates treated with 100, 200, 400 nmol/L deguelin at 28.5℃in an incubator til the shield period and 24 hours after fertilization, respectively. Simultaneously embroys treated with 1% dimethyl sulfoxide solution were as a control group, cultured in the same conditions. Cyclin D1 RNA probes were prepared for the whole mountin situhybridization, observing staining by an upright fluorescent microscope camera to detect the effect of deguelin on cyclin D1 expression in zebrafish embryos. RESULTS AND CONCLUSION:Deguelin showed significant negative regulation on the expression of cyclin D1 gene in zebrafish embryos. Cyclin D1 expressed in outsourcing cels in embryos of shield stage, and a significant reduction in the expression of cyclin D1 came up with the increasing concentrations of deguelin. In the 400 nmol/L deguelin treatment group, there was nearly no expression of cyclin D1. Cyclin D1 expressed in the brain, central nervous system, immature eye, somites, trunk, and tail of embryos at 24 hours after fertilization, and reduced significantly in the 100 nmol/L deguelin treatment group, especialy in the proliferative area. In the 200 and 400 nmol/L treatment groups, the embryonic development slowed down signficantly, and cyclin D1 gene mainly expressed in the dorsal ectoderm cels.
ABSTRACT
BACKGROUND:Studies have shown that stem cels transfected with human telomerase reverse transcriptase (hTERT) gene can stably express high-level telomerase activity and strengthen cel proliferation, which lays the foundation to establish geneticaly engineered immortalized stem cel lines. OBJECTIVE:To explore the effects of hTERT transfection on proliferation and cel cycle of rat fetal liver stem cels in vitro. METHODS:Rat fetal liver stem cels culturedin vitro were transfected by recombinant adeno-associated virus carrying hTERT genes. RT-PCR and western blot assay were used to detect the expression of hTERT mRNA and protein, respectively. Cel Counting Kit-8 method and cel growth curve were used to detect cel growth and proliferation. Changes in cel cycle distribution were determined by flow cytometry. RESULTS AND CONCLUSION:Compared with the control group and empty virus group, in the hTERT infection group, hTERT expressed at gene and protein levels, the growth rate of the cels increased, and the number of cels at G0/G1 phase and S phase was decreased and increased, respectively. The results show that recombinant adeno-associated virus as a vector of hTERT gene used for gene transfection can promote the in vitro proliferation of rat fetal liver stem cels and play an optimal role in cel culture.