OBJECTIVE: To study the effects of oligodeoxynucleotides complementary Stat3 on apoptosis in laryngeal carcinoma Hep-2 cell. METHOD: Oligodeoxynucleotides complementary Stat3 was designed, which was transferred into laryngeal carcinoma Hep-2 cell by lipofection. Expression of Stat3 and p-Stat3 were detected by Western blot and PCR. MTT was used to observe the growth-inhibiting ratio. DNA ladder, AO/EB and FCM were used to observe the apoptosis of laryngeal carcinoma cell Hep-2 in vitro. RESULT: Western blot and PCR results demonstrated that oligodeoxynucleotides complementary Stat3 could significantly inhibit the expression of Stat3 and p-Stat3 in Hep-2 cell. MTT results showed that it could significantly suppress the growth of Hep-2 cell. The DNA ladder, AO/EB and FCM results showed it could inhibit the expression of Stat3 and induce the apoptosis of Hep-2 cell in a concentration-dependent manner. CONCLUSION Oligodeoxynucleotides complementary Stat3 could induce the apoptosis and suppress cell proliferation in laryngeal carcinoma Hep-2 cell.
Apoptosis ; drug effects ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Humans ; Oligodeoxyribonucleotides ; genetics ; pharmacology ; STAT3 Transcription Factor ; genetics ; Transfection

To explore the biological function and possible underlying mechanism of stathmin gene during hepatocarcinogenesis. Three pairs of chemically synthesized small interfering RNA (siRNA) targeting on stathmin were transfected into HCCLM3 by LipofectamineTM 2000. After confirming the interfering effects of stathmin siRNAs through reverse transcription PCR and Western blotting, the HCCLM3 cells proliferation and apoptosis were detected by cell count kit-8 (CCK-8) and flow cytometry analysis, and the expressions of tumor-related genes (c-myc, c-fos, p53, etc) were observed by real-time PCR. Stathmin expression was effectively inhibited up to 90% by stathmin silencing in HCCLM3 cells (P is less than to 0.05) . By using CCK8 assay, it was shown that HCCLM3 cells proliferation were obviously depressed by 13.04%+/-0.10%, 28.10%+/-0.41% and 37.36%+/-2.15% at the time point of 24 h, 48 h and 72 h with the comparison to Mock group (F = 4.21, P is less than to 0.05). The results of flow cytometry demonstrated that the percentage of apoptotic cells was increased to 25.11%+/-1.62% in RNAi group, compared with 9.20 %+/-0.64 % in Mock group (F = 44.67, P is less than to 0.01). The results of real-time PCR showed that oncogenes c-myc and c-fos expressions were repressed, proliferation-associated gene ki-67 was down-regulated, and apoptosis-promoting gene caspase-3, bax and p53 were induced (P is less than to 0.05). Stathmin may promote cell proliferation, inhibit cell apoptosis and induce malignant transformation of hepatocytes by regulating some tumor-related genes expressions.
Apoptosis ; drug effects ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Humans ; RNA Interference ; RNA, Small Interfering ; genetics ; Stathmin

OBJECTIVETo study the effect of safflower injection on the proliferation and apoptosis of human leukemia cell line HEL and the relevant molecular mechanisms.

METHODSHEL cells were treated with different concentrations of safflower injection. HEL cells without safflower injection treatment were used as the control group. MTT method was used to detect the inhibitory rate of the HEL cells at 24, 48 and 72 hours after various concentrations of safflower injection treatment (10, 20, 30, 40 and 50 mg/mL). The cell cycle and apoptosis were detected using flow cytometry and the HOXB3-mRNA expression was measured by RT-PCR at 48 hours after safflower injection treatment (10, 20 and 30 mg/mL).

RESULTSCompared with the control group, various concentrations of safflower injection inhibited HEL cell proliferation in a dose-dependent manner (P<0.05). At 48 hours after various concentrations of safflower injection treatment, the number of treated cells in the G2/M phase increased, but that in the S phase decreased, and the apoptosis rate was significantly higher than that in the control group, with a dose-dependent manner (P<0.05). The expression of HOXB3-mRNA in safflower injection-treated cells decreased in a dose-dependent manner compared with the control group (P<0.05).

CONCLUSIONSSafflower injection can inhibit proliferation and induce apoptosis of HEL cells in vitro, and its underlying mechanisms may involve down-regulation of the HOXB3-mRNA expression.

Apoptosis ; drug effects ; Carthamus tinctorius ; Cell Cycle ; drug effects ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Homeodomain Proteins ; genetics ; Humans ; Injections ; Leukemia ; drug therapy ; metabolism ; pathology

OBJECTIVE: To study the impacts of RelA antisense oligonucleotides on proliferation in laryngeal carcinoma Hep-2 cell. METHOD: RelA antisense oligonucleotides was designed, which was transferred into laryngeal carcinoma Hep-2 cell. MTT was used to detect the growth-inhibiting ratio at different transferred timepoints. Hep-2 cell which was transferred 48 h was used to do colony assay, and expression of RelA was detected by Reverse Transcription PCR and Western blot. RESULT: MTT results showed that RelA antisense oligonucleotides could significantly suppress the proliferation of Hep-2 cell, and the suppression-ratio elevated with time. There were statistical difference compared with control groups. The number of cells colony was reduced in RelA antisense oligonucleotides group compared with control groups, which had statistic significance. RT-PCR and Western blot results demonstrated that RelA antisense oligonucleotides could significantly inhibit the expression of messenger RNA and protein in Hep-2 cell. CONCLUSION RelA antisense oligonucleotides can inhibit the expression of messenger RNA and protein, and induce the cell proliferation and increase the number of cells colony in Hep-2 cell.
Cell Line, Tumor ; Cell Proliferation ; drug effects ; Humans ; Laryngeal Neoplasms ; genetics ; pathology ; Oligonucleotides, Antisense ; genetics ; pharmacology ; Transcription Factor RelA ; genetics

The objective of this study was to establish an RNAi approach that can specifically target aqp1 gene sequence in vitro, and to assess the inhibitory effect of this siRNA on K562 cells. The siRNA targeting aqp1-mRNA was designed and transfected into K562 cells by using Lipofectamine(TM) 2000 reagent. Phase-contrast microscopy was used to analyze morphology changes of K562 cells. Cell viability was determined by MTT assay, and flow cytometry and DNA ladder analysis were carried out to identify siRNA-induced apoptosis. The expression levels of aqp1-mRNA at different transfection time were detected by RT-PCR. The results showed that the siRNA was successful by established. The transfected K562 cells displayed the significant apoptosis. The aqp1-siRNAs could obviously inhibit the activity of K562 cells. Cellular DNA fragmentation was observed in the siRNA group after transfection for 48 hours, the apoptosis rates at 24, 48 and 72 hours after transfection were 24.2%, 36.1% and 42.9% respectively. The aqp1-mRNA expression in the cells treated by aqp1-siRNA for 24, 48 and 72 hours were significantly reduced by 33%, 46% and 57% respectively. It is concluded that the aqp1-siRNA can efficiently and specifically inhibited the proliferation and inducing apoptosis of K562 cells. Gene aqp1 can be a potential target point for therapy of malignant tumor.
Apoptosis ; drug effects ; Aquaporin 1 ; genetics ; Cell Proliferation ; drug effects ; Cell Survival ; drug effects ; Gene Targeting ; Humans ; K562 Cells ; RNA Interference ; RNA, Small Interfering ; genetics ; pharmacology

The objective of this study was to investigate the effect of hexamethylene bisacetamide (HMBA) on differentiation of HL-60 cells and its possible molecular mechanism. HL-60 cells were co-cultured with different concentrations of HMBA (0.5, 1, 2 mmol/L) for 4 days, then the proliferation was assayed by MTT at different time points. Wright-Giemsa staining was used to observe the change in morphology. Cell differentiation antigen CD11b expression and the altered distribution of cell cycle in HL-60 induced by HMBA were analyzed by flow cytometry. The expressions of c-myc, mad1, p21, p27, hTERT and HDAC1 mRNA were detected by RT-PCR. The results indicated that the proliferation of HL-60 cells was inhibited by HMBA in a time-and-dose-dependent manner. Upon 2 mmol/L HMBA treatment, the HL-60 cells arrested at G(0)/G(1) phase and differentiated into granular line in morphology, with the up-regulation of CD11b expression. The expression of c-myc and hTERT mRNA obviously down-regulated, the expression of p21, p27 and mad1 mRNA up-regulated, while there was no change of the expression of hTERT mRNA. It is concluded that effect of HMBA on the differentiation of HL-60 cells may partly contribute to switch from c-myc to mad1 expression, to up-regulate expressions of p21 and p27 mRNA, and down-regulate hTERT mRNA expression, while there is no relation with the expression of HDAC1 mRNA.
Acetamides ; pharmacology ; Antineoplastic Agents ; pharmacology ; Cell Cycle ; drug effects ; Cell Differentiation ; drug effects ; genetics ; Cell Proliferation ; drug effects ; HL-60 Cells ; Humans

OBJECTIVETo investigate the effect of the RNAi and the chemotherapy drugs nolatrxed on the expression of thymidylate synthase(TS) and the growth of the colorectal carcinoma LOVO cells.

METHODSThe siRNA was constructed targeting the human TS gene, and then transfected into the human colorectal cancer LOVO cells. RT-PCR and Western blot technique were used to observe the TS gene and protein expression levels, and MTT was used to detect cell proliferation after silencing the TS gene. In addition, siRNA and nolatrxed were applied to the LOVO cells to observe the TS protein expression and cell growth.

RESULTSTS siRNA significantly reduced the expression of TS gene and protein in LOVO cells, and inhibited cell growth. The IC50 value of LOVO cells was (1.46±0.25) μmol/L in TS siRNA combined with nolatrexed group, (6.81±0.31) μmol/L in the negative control group, and (6.47±0.43) μmol/L in the single nolatrexed group. After treatment of TS siRNA combined with nolatrexed on LOVO cells for 36 hours, the apoptosis index was higher than that in single TS siRNA and nolatrexed[(62.12±0.89)% vs.(21.56±0.67)% and(40.51±0.83)%, both P<0.05].

CONCLUSIONTS siRNA can partly suppress the expression of TS gene in LOVO cells, inhibit cell proliferation, promote cell apoptosis and enhance cell sensitivity to apoptosis induced by nolatrexd.

Apoptosis ; drug effects ; Cell Cycle ; drug effects ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Colorectal Neoplasms ; enzymology ; pathology ; Humans ; Quinazolines ; pharmacology ; RNA Interference ; Thymidylate Synthase ; genetics ; metabolism

OBJECTIVETo study the effect of laminarin sulphate (LAMS) on the expressions of PTEN and P27kip1 in androgen-independent prostate cancer PC-3 cells in vitro, and investigate the mechanism of its anti-tumor action.

METHODSThe inhibitory effects of different concentrations of LAMS (0, 50, 100, 200 microg/ml) on androgen-independent prostate cancer PC-3 cells were detected by WST-8 assay. The morphology of PC-3 cells was observed under the fluorescence microscope, and the cell cycle and apoptosis were analyzed by flow cytometry. The mRNA and protein levels of PTEN and P27kip1 were measured by RT-PCR and Western blot.

RESULTSLAMS inhibited the proliferation of androgen-independent prostate cancer PC-3 cells in a dose- and time-dependent manner, and the cell cycle analysis showed that PC-3 cells were arrested in the S phase after treated with different concentrations of LAMS. The rate of apoptosis was increased and many typical apoptotic morphological features were observed under the fluorescence microscope. The PTEN and P27kip1 expressions at mRNA and protein levels were increased in a dose-dependent manner.

CONCLUSIONLAMS can inhibit the proliferation, arrest the cell cycle in the S phase and induce apoptosis of prostate cancer PC-3 cells. The significantly increased expressions of PTEN and P27kip1 may be one of the mechanisms for LAMS inhibiting prostate cancer PC-3 cells.

Apoptosis ; drug effects ; Cell Cycle ; drug effects ; Cell Line, Tumor ; drug effects ; Cell Proliferation ; drug effects ; Cyclin-Dependent Kinase Inhibitor p27 ; genetics ; Humans ; Male ; PTEN Phosphohydrolase ; genetics ; Polysaccharides ; pharmacology ; Prostatic Neoplasms ; blood ; genetics ; RNA, Messenger ; genetics

OBJECTIVETo investigate the role of Mcl-1 gene in resistance of all-trans retinoic acid (ATRA) of leukemia cells.

METHODSLong-term, intermittent and repetitive exposure of HL-60 cells to ATRA was used to establish a multidrug-resistance cell line (HL-60/ATRA). HL-60/ATRA cells were transfected with Mcl-1 small interference RNA (siRNA) by Lipofectamine 2000. Western blot was used to detect the expression of Mcl-1. The proliferation, apoptosis and differentiation were evaluated by MTT assay, in situ nick end-labeling (TUNEL) and NBT assay, respectively.

RESULTSThe HL-60/ATRA could keep its undifferentiated and proliferative status to a high concentration of ATRA (100 nmol/L) with highly expressed Mcl-1 protein (relative grey scale 0.624 +/- 0.127). Mcl-1 gene knockdown by siRNA (relative grey scale 0.267 +/- 0.086) could reverse the resistance of ATRA of HL-60/ATRA by inhibiting proliferation, and inducing differentiation and apoptosis [apoptosis rate (18.5 +/- 4.5)%].

CONCLUSIONMcl-1 gene might be involved in ATRA resistance in HL-60 cells and inhibiting its expression could be a new approach to ATRA resistance reversion.

Apoptosis ; drug effects ; genetics ; Cell Differentiation ; drug effects ; genetics ; Cell Proliferation ; drug effects ; Drug Resistance, Neoplasm ; genetics ; HL-60 Cells ; drug effects ; metabolism ; Humans ; Proto-Oncogene Proteins c-bcl-2 ; genetics ; metabolism ; RNA, Small Interfering ; Tretinoin ; pharmacology

The study was aimed to investigate the synergistically effect of interferon-α (IFN-α) and homoharringtonine (HHT) on the proliferation, apoptosis, cell cycle of K562 cells and the expression of β-catenin. The proliferation, apoptosis, cell cycle and β-catenin mRNA expression of K562 cells treated with IFN-α and/or HHT were assayed with MTT, flow cytometry or RT-PCR respectively. The results showed that HHT alone, but not IFN-α alone, displayed a proliferation inhibition, apoptosis induction, G(0)/G(1) phase block and down-regulation of β-catenin expression in K562 cells with concentration- and time-dependent manners. The expression level of β-catenin mRNA after being treated with HHT was 0.5576 ± 0.0373, which were lower than that in control group (0.9369 ± 0.0142). The down-regulation of β-catenin expression in group of IFN-α combined with HHT was higher significantly than that in HHT group (0.3737 ± 0.0529 vs 0.5576 ± 0.0373, P < 0.05). Otherwise, HHT combined with IFN-α did not demonstrate obvious toxicologic effect on bone marrow mononuclear cells. It is concluded that IFN-α combined with HHT can enhance the cytotoxic effect of HHT on K562 cells, which may be associated with down-regulation of β-catenin expression.
Cell Proliferation ; drug effects ; Harringtonines ; pharmacology ; Humans ; Interferon-alpha ; pharmacology ; K562 Cells ; beta Catenin ; genetics ; metabolism