Objective Isoliquiritigenin (ISL), a licorice chalconoid, is considered to be a bioactive agent with chemopreventive potential. This study investigates the mechanisms involved in ISL-induced apoptosis in human cervi...Objective Isoliquiritigenin (ISL), a licorice chalconoid, is considered to be a bioactive agent with chemopreventive potential. This study investigates the mechanisms involved in ISL-induced apoptosis in human cervical carcinoma HeLa cells. Methods Cell viability was evaluated using a 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyl-tetrazolium bromide (MTT) assay. Apoptosis was determined by flow cytometry using an Annexin V-FITC Apoptosis Detection Kit. The intracetlular ROS levels were assessed using a 2, 7-dichlorofluorescein probe assay. The mitochondrial membrane potential was measured with the dual-emission potential-sensitive probe 5, 5', 6, 6'-tetra-chloro-1, 1', 3, 3'-tetraethyl-imidacarbocyanine iodide (JC-1). The degradation of poly-ADP-ribose polymerase (PARP) protein, the phosphorylation of PKR-like ER kinase (PERK), the phosphorylation of the a-subunit of eukaryotic initiation factor 2 (elF2a), the expression of the 78 kD glucose-regulated protein (GRP 78), and the activation of caspase-12 were analyzed via western blot analysis. Results ISL significantly inhibited the proliferation, the increase in ROS levels and apoptotic rates of HeLa cells in a concentration-dependent manner. Moreover, ISL induced mitochondrial dysfunction, caspase activation, and PARP cleavage, which displayed features of mitochondria dependent on apoptotic signals. Besides, exposure of HeLa cells to ISL triggered endoplasmic reticulum (ER) stress, as indicated by the increase in p-elF2a and GRP78 expression, ER stress-dependent apoptosis is caused by the activation of ER-specific caspase-12. Conclusion The findings from our study suggest that ISL-induced oxidative stress causes HeLa cel apoptosis via the mitochondrion-dependent and the ER stress-triggered signaling pathways.展开更多
The recombinant plasmids pGenesil-1-BIRC71 and pGenesil-1-BIRC72 were transfected into Hela cells and cisplatin was added with different concentrations in order to study the inhibitory effects of Livin gene, increase ...The recombinant plasmids pGenesil-1-BIRC71 and pGenesil-1-BIRC72 were transfected into Hela cells and cisplatin was added with different concentrations in order to study the inhibitory effects of Livin gene, increase the apoptosis induced by cisplatin, and detect the expression ofBcl-2, Bax, caspase-3, and survivin genes. The pGenesil-1-BIRC71 and pGenesil-1-BIRC72 were transfected into Hela cells, and the expression levels of Livin, Bcl-2, Bax, caspase-3, and survivin genes were detected by using fluorescence quantitative real-time PCR. Then cisplatin at different concentrations (3.0, 6.0 and 9.9 μg/mL) was added into the transfected Hela cells, and 24, and 48 h later, the apoptosis rate was measured by flow cytometry. After transfection of pGenesil-l-BIRC71 and pGenesil-1-BIRC72 into Hela cells, the expression level of Livin gene was obviously reduced, and the apoptosis rate was significantly increased in transfection group as compared with control group (P〈0.05). Cisplatin could increase the apoptosis rate in a dose- and time-dependent manner. After cisplatin was added, the expression levels of Bcl-2 mRNA were reduced, and those of Bax, caspase-3, and survivin mRNA were increased in transfection group as compared with those in control group (P〈0.05). It was concluded that shRNA expression vector targeting Livin gene could inhibit the expression of Livin gene in Hela cells and enhance the apoptosis induced by cisplatin, which was related to the decreased expression of Bcl-2 and activation of Bax and caspase-3. Survivin might play an important role as an antagonist in the process of apoptosis induction.展开更多
Hypocrellin A( HA), a photosensitive perylenequinone compound of Hypocrella bambusae, inhibited the proliferation of several tumor cell lines. Human cervical cancer cells, HeLa ceils, were used as a model to elucida...Hypocrellin A( HA), a photosensitive perylenequinone compound of Hypocrella bambusae, inhibited the proliferation of several tumor cell lines. Human cervical cancer cells, HeLa ceils, were used as a model to elucidate the molecular mechanisms of HA-induced tumor cell death. The results show that HA can induce the oligonucleosomal fragmentation of DNA in HeLa cells and also can increase the expression of apoptosis inducer Bax mRNA and that it decreases the expression of apoptosis suppressor, Bcl-2 mRNA, in mitochondria. It can be concluded from the data that HA-induced apoptosis is related to the balance between Bcl-2 and Bax gene expressions.展开更多
To explore the effect of quercetin on the proliferation and apoptosis of HeLa cells, HeLa cells were incubated with quercetin at different concentrations. Cell viability was evaluated by MTT assay, cell apoptosis was ...To explore the effect of quercetin on the proliferation and apoptosis of HeLa cells, HeLa cells were incubated with quercetin at different concentrations. Cell viability was evaluated by MTT assay, cell apoptosis was detected by Annexin- V/PI double labeled cytometry and DNA ladder assay. Cell cycle was flow cytometrically determined and the morphological changes of the cells were ob- served under a fluorescence microscope after Hoechst 33258 staining and the apoptosis-related pro- teins in the HeLa cells were assessed by Western blotting. The results showed that quercetin signifi- cantly inhibited the growth of HeLa cells and induced obvious apoptosis in vitro in a time- and dose-dependent manner. Moreover, quercetin induced apoptosis of HeLa cells in cell cycle-dependent manner because quercetin could induce arrest of HeLa cells at G0/G1 phase. Quercetin treatment down-regulated the expression of the PI3K and p-Akt. In addition, quercetin could down-regulate ex- pression of bcl-2, up-regulate Bax, but exerted no effect on the overall expression of Akt. We are led to conclude that quercetin induces apoptosis via PI3k/Akt pathways, and quercetin has potential to be used as an anti-tumor agent against human cervix oancer.展开更多
This paper deals with the inhibitory mechanisms of ginsenoside \{G Rh 2\} on the growth of tumor cells. \{G Rh 2\} significantly inhibited the proliferation of human cervical adenocarcinoma HeLa cells in a time ...This paper deals with the inhibitory mechanisms of ginsenoside \{G Rh 2\} on the growth of tumor cells. \{G Rh 2\} significantly inhibited the proliferation of human cervical adenocarcinoma HeLa cells in a time and dose dependent manner. G Rh 2 induced apoptotic manifestations in HeLa cells as evidenced by the changes in the cell morphology, the DNA fragmentation and the activation of caspases. Caspase inhibitors, caspase family inhibitor, z Val Ala Asp fmk(z VAD fmk); caspase 1 inhibitor, Ac Tyr Val Ala Asp chloromethyl ketone(Ac YVAD cmk); caspase 3 inhibitor, z Asp Glu Val Asp fmk(z DEVE fmk) and caspase 8 inhibitor, \{z Ile \}Glu Asp fmk(z IETD fmk) effectively attenuated G Rh 2 induced cell death. The activities of caspase 1 and caspase 3 were increased in the G Rh 2 induced apoptotic process. However, caspase inhibitors can not inhibit G Rh - 2 induced cell death completely. These results suggest that G Rh 2 induced cell death is mediated by the activation of caspase cascade, but there might be some other pathways for induction of this apoptosis.展开更多
Effect of quercetin on HeLa cell system of cervical cancer was studied by methods of MTT and Annexin V-FITC/PI. The results show that quercetin has functions of inhibiting breeding of HeLa cells and inducing apoptosis...Effect of quercetin on HeLa cell system of cervical cancer was studied by methods of MTT and Annexin V-FITC/PI. The results show that quercetin has functions of inhibiting breeding of HeLa cells and inducing apoptosis of the cells. The total apoptosis rate is positively proportional to reaction duration and concentration of quercetin used. The maximum apoptosis rate being (88.76±2.35)% was obtained when the concentration was 50.0 μmol/L and the cells were treated with quercetin for 72 hours. Based on establishing a model of tumor of cervical cancer transplanted into nude mice, quercetin of different concentrations was injected into abdominal cavity of nude mice and situation of tumor growth was reviewed. The result showed that with quercetin concent'ration increasing from 0 to 100.0 μmol/L, the transplantation volume and weight of the tumors decreased from (279.59±70.58) mm^3 and (0.145±0.019) g to (128.72±36.12) mm^3 and (0.089± 0.019) g respectively, while apoptosis rate of the transplanted tumor increased from (9.63±1.85)% to (34,98±0.47)%, which proved that quercetin inhibited increment of volume and weight of transplanted tumor in nude mice bodies.展开更多
The killing effects of lymphocytes on Hela cells expressing interleukin-12 (IL-12) in vitro were explored. By using gene transfection technique, full length IL-12 gene was transfected into Hela cells. The expression...The killing effects of lymphocytes on Hela cells expressing interleukin-12 (IL-12) in vitro were explored. By using gene transfection technique, full length IL-12 gene was transfected into Hela cells. The expression of IL-12 in Hela cells was detected quantitatively by ELISA; Changes in killing effects of lymphocytes on Hela cells expressing IL-12 were observed by MTT. It was found that Hela cells could express IL- 12 between 24 h and 72 h after transfection. Killing activity of lymphocytes on Hela cells expressing IL-12 was significantly enhanced. It was concluded by cell transfection technique, Hela cells could express 1L-12 and were more easily killed by lymphocytes.展开更多
Objective: The histone deacetylase inhibitors (HDACIS) have been shown to inhibit cancer cell proliferation, stimulate apoptosis, an induce cell cycle arrest. Our purpose was to investigate the antiproliferative ef...Objective: The histone deacetylase inhibitors (HDACIS) have been shown to inhibit cancer cell proliferation, stimulate apoptosis, an induce cell cycle arrest. Our purpose was to investigate the antiproliferative effects of a HDACI, trichostatin A (TSA), against human cervical cancer cells (HeLa). Methods: HeLa cells were treated in vitro with various concentrations of TSA. The inhibitory effect of TSA on the growth of HeLa cells was measured by MTT assay. To detect the characteristic of apoptosis chromatin condensation, HeLa cells were stained with Hoechst 33258 in the presence of TSA. Induction of cell cycle arrest was studied by flow cytometry. Changes in gene expression of p53, p21wafl and p27Kipl were studied by semiquantitative RT-PCR. Results: TSA inhibited cell growth in a time- and dose-dependent manner. Hoechst 33258 staining assay showed that TSA induced apoptosis. Cell cycle analysis indicated that treatment with TSA decreased the proportion of cells in S phase and increased the proportion of cells in G0/G1 and/or G2/M phases of the cell cycle. This was concomitant with overexpression of genes related to malignant phenotype, including an increase in p53, p21wall and p27Kipl. Conclusion: These results suggest that TSA is effective in inhibiting growth of HeLa cells in vitro. The findings raise the possibility that TSA may prove particularly effective in treatment of cervical cancers.展开更多
Objective To observe the inhibition and radiation-sensitizing effect of Indoleacetic acid (IAA) combined with horseradish peroxidase(HRP)on Hela cells. Methods Hela cells were cultured in vitro and classified into c...Objective To observe the inhibition and radiation-sensitizing effect of Indoleacetic acid (IAA) combined with horseradish peroxidase(HRP)on Hela cells. Methods Hela cells were cultured in vitro and classified into control group, drug group incubated with different doses of IAA(30, 60, 90μmol·L -1) plus 1.2μg·mL -1 HRP, radiation group (6MV-X, 4Gy ) and group of radiation plus IAA plus HRP(same dose as above). All the above were treated for 24-96 hours.The growth inhibition, radiation-sensitizing effect were observed with methyl thiazolyl tetrazolium (MTT) photocolorimetric assay and trypan blue dye assay. The-effect on cell proliferation cycle was determined by flow cytometry. Results The antiproliferation activities showed a significant time-effect and dose-effect relationship to some extent after Hela cells were treated with IAA combined with HRP. The group of radiation plus 60μmol·L -1 IAA plus 1.2μg·mL -1 HRP and radiation plus 90μmol·L -1 IAA plus 1.2μg·mL -1 HRP showed an obvious radiation sensitizing effect. After treatment with 90μmol·L -1 IAA plus 1.2 μg·mL -1 HRP for 72 hours, the determination of cell cycle showed that the percentages of the cells on stages G 2-M and S were all higher than those of the control group. For the group of radiation plus IAA combined with HRP, the percentages of the cells on stages G 2-M were higher than those of the radiation group. Conclusion The above findings suggest that IAA combined with HRP has an inhibitive and killing effect on Hela cells. The effect was stronger during the cell cycles of G 2-M and S. It also has a radiation sensitizing effect. Its mechanism might be that Hela cells were blocked on stages G 2-M, and it presents a collaborative killing effect during miototic time.展开更多
Objective To investigate the Hela cells growth inhibition and apoptosis possible molecular mechanisms. Methods Hela cells were treated with various concentrations (100μmol/L,200μmol/L, 300 μmol/L, 400 μmol/L) of...Objective To investigate the Hela cells growth inhibition and apoptosis possible molecular mechanisms. Methods Hela cells were treated with various concentrations (100μmol/L,200μmol/L, 300 μmol/L, 400 μmol/L) of NS-398 (selective for COX-2 inhibition). Cell growth was measured by MTT (Thiazolyl blue). Apoptosis was detected by double staining flow cytometry (FCM). Levels of PGE: were measured by radioimmunoassay. The expressions of COX-2 protein were also examined by Western blot analysis. Results After treated with different concentrations of NS-398, the growth of Hela cells was suppressed significantly in a dose-and time-dependent manner (P〈0.01). The NS-398 can induce apoptosis with the apoptosis rates at 8.53%-43.46% by FCM in a dose-dependent manner. The release of PGE2 was reduced in Hela cells with the values of 69.26 ± 2.13, 47.46 ± 2.18, 28.15 ± 1.64 and 17.01 ± 1.12, respectively, there was significant difference compared with control group (83.78 ± 1.11) (P〈0.01). The NS-398 could inhibit the activity and expression of COX-2 in a dose- dependent manner and down-regulated the expression of COX-2 protein greatly. Conclusion NS-398 could inhibit the proliferation and increase apoptosis in human Hela cells. These effects may be depended on the inhibition of the expression of COX-2 and PGE2 by NS-398.展开更多
To construct a single tetracycline-regulatable plasmid vector based on the double tetracycline-regulatable plasmid vector system for regulating HSV-tk gene expression so as to effectively kill HeLa cells. Two tetracyc...To construct a single tetracycline-regulatable plasmid vector based on the double tetracycline-regulatable plasmid vector system for regulating HSV-tk gene expression so as to effectively kill HeLa cells. Two tetracycline operator(TetO2) was cloned into pcDNA3.1 and a cassette was made for a cytomegalovirus-type 2 tetracycline operator(CMV-TetO2) promoter, and the obtained vector was named pcDNA3.1-CMV-TetO2. Herpes simplex virus thymidine kinase(HSV-tk) gene and tetracycline repressor(TR) gene were cloned into pcDNA3.1-CMV-TetO2 and the two genes were linked with internal ribosome entry site(IRES) to gain a vector named pcDNA3.1-CMV-TetO2- HSV-tk-IRES-TR. The HeLa cells were stablly transfected with pcDNA3.1-CMV-TetO2-HSV-tk-IRES-TR plasmid. The expression of HSV-tk and TR were detected by RToPCR, the tumorcidal activity of HSV-tk/GCV was determined by MTT assay. In Hela cells transfected with the above plasmid vector, HSV-tk gene and TR gene can be expressed lowly and the concentration of GCV producing a 50% decrease in cell viability was about 50 lag/mL without adding deoxycycline; in contrast, the expessions of HSV-tk gene and TR gene increased significantly and the concentration of GCV producing a 50% decrease in cell viability was about 5 μg/mL with adding deoxycycline. Therefore tetracycline can regulate the expression and tumorcidal activity of HSV-tk gene in HeLa cells with this single plasmid vector.展开更多
Inhibins play important roles in the reproductive system. To evaluate whether inhibin α (1-32) fusion protein plays a role in cervical cancer growth, the plasmid pVAX-inhα was constructed and its effect on prolifera...Inhibins play important roles in the reproductive system. To evaluate whether inhibin α (1-32) fusion protein plays a role in cervical cancer growth, the plasmid pVAX-inhα was constructed and its effect on proliferation and apoptosis of the human cervical cancer cell line (Hela) was checked by flow cytometry and real-time PCR. The expression and localization of inhibin α protein were detected by RT-PCR and confocal mi- croscopy which showed that inhibin α protein was expressed and localized in the nucleus of Hela cells. Over ex- pression of inhibin α gene significantly induced cell apoptosis and ceased S phase of cell cycle. Furthermore, cell proliferation was significantly suppressed 96 h post-transfection and mRNA level of anti-apoptosis genes (Bcl-2, NFκB) were decreased but pro-apoptosis genes (Bax, wild type p53) and inhibin co receptor (TGFβR3) were increased, indicating that inhibin, through its co-receptor, might activate apoptotic and cell growth cascades which regulate proliferation and apoptosis in Hela cells. These results suggest that inhibin α (1-32) fusion pro- tein, located in the cell nucleus, can regulate Hela cells growth and apoptosis by induction of apoptotic pathways such as NFκB, Bcl-2 and p53 families. These findings may have a significant impact on future research regarding cervical cancer cell lines.展开更多
A method is described which permits transmission electron microscope of single cells treated with HpD plus laser microirradiation. The preselected single cell that was irradiated by laser under light microscope and fo...A method is described which permits transmission electron microscope of single cells treated with HpD plus laser microirradiation. The preselected single cell that was irradiated by laser under light microscope and followed fixation, embedded and sectioning is examined under electron microscope. The results demonstrated that at the light dose of 1.88 ml/μm2 not only the irradiated nucleolus appeared transparent region, but the other parts such as non-irradiated mitochondria in cytoplasm can also be damaged. When partial cytoplasm is irradiated with the light dose of 4.50 ml/μm2, the damages appear in all cytoplasm, but there is little change in the nucleus. The experimental results also demonstrate that cytoplasm is more sensitive than nucleus. It is the mitochondria in cytoplasm that are very sensitive to HpD plus laser.展开更多
This study aimed to investigate the effect of fullerenol combined with cisplatin on the proliferation of cervical cancer HeLa cells in order to provide new ideas and laboratory theoretical basis for the clinical treat...This study aimed to investigate the effect of fullerenol combined with cisplatin on the proliferation of cervical cancer HeLa cells in order to provide new ideas and laboratory theoretical basis for the clinical treatment of cervical cancer. Cervical cancer cell line HeLa cells in vitro were treated with different concentrations of fullerenol, different concentrations of cisplatin, and different concentrations of fullerenol combined with cisplatin, after 24 h, 48 h, 72 h, microscope changes in cell morphology;MTT assay was used to determine the effect of drugs on the proliferation of HeLa cells. Fullerenol and cisplatin alone can inhibit the proliferation of HeLa cells in a dose-dependent and time-dependent manner;compared with cisplatin alone, different concentrations of fullerenol combined with cisplatin significantly increased the apoptosis rate of HeLa cells (P < 0.05). The inhibition of fullerenol combined with cisplatin on HeLa cells in vitro is more significant, resulting in a stronger anti-cancer effect.展开更多
Aging is a decelerating unidirectional process of life. Shortening of telomeric DNA, the (TTAGGG)<sub>n</sub> hexanucleotide repeats, which form the caps at the chromosome ends, is implicated to determine ...Aging is a decelerating unidirectional process of life. Shortening of telomeric DNA, the (TTAGGG)<sub>n</sub> hexanucleotide repeats, which form the caps at the chromosome ends, is implicated to determine the aging process, and more importantly the healthy lifespan itself. Telomerase, a ribonucleoprotein having reverse transcriptase activity, arrests telomere loss through addition of the TTAGGG repeats de novo, to the ends of the chromosome. The telomere/telomerase maintenance is an inevitable necessity to delay aging and for a healthy lifespan. Here, we report the potential of full-spectrum, high concentration Ashwagandha (Withania somnifera), an Ayurvedic medicinal herb, root extract to increase telomerase activity. HeLa cells, when treated with various concentrations of Ashwagandha root extract, showed an increase in telomerase activity measured with the established Telomerase Rapid Amplification Protocol (TRAP) assay. Ashwagandha root extract increased telomerase activity with highest enhancement of ~45% at 10 - 50 μg concentration. Thus, Ashwagandha root extract has the anti-aging inducing potential.展开更多
Objective:To explore the effect of bortezomib on migration and invasion of cervical carcinoma HeLa cell and specific molecular mechanism.Methods:The effect of bortezomib on the viability of HeLa cell was measured by M...Objective:To explore the effect of bortezomib on migration and invasion of cervical carcinoma HeLa cell and specific molecular mechanism.Methods:The effect of bortezomib on the viability of HeLa cell was measured by MTT assay.The effect of bortezomib on cell migration and invasion was measured by Transwell assay and invasion experiment respectively.The activation of Akt/mTOR signaling pathway and expression level of MMP2,MMP9 were assayed by western blot.Results:MTT assay indicated bortezomib(2.5 μM.5 μM,10 μM)could inhibit HeLa cell viability,and the inhibitory rate was highest at 48 h.Transwell assay and invasion experiment results showed that bortezomib inhibited HeLa cell migration and invasion.Western blotting assays presented bortezomib could suppress the phosphorylation of Akt and mTOR.and down-regulate the expression of MMP2 and MMP9.Conclusions:These results suggested bortezomib could inhibit migration and invasion in cervical carcinoma HeLa cell,which might be related to Akt/mTOR signal pathway.展开更多
Objective: To examine the proapoptotic properties of Oroxylum indicum methanol extract on cervical cancer cells. Methods: Methylene blue assay was used to determine the IC50 value of the extract. Western blotting assa...Objective: To examine the proapoptotic properties of Oroxylum indicum methanol extract on cervical cancer cells. Methods: Methylene blue assay was used to determine the IC50 value of the extract. Western blotting assays were done to analyze the expression of HPV oncoproteins (HPV18 E6 and E7) and apoptotic molecules (caspase-3 and caspase-8). Reverse transcriptase PCR assays were performed to determine genetic alteration of tumor suppressors p53 and pRb and apoptosis markers Fas and FasL. Enzyme-linked immunosorbent assay (ELISA) was done to determine the expression of cytokine levels (IL-6 and IL-12). Results: The determination of IC50 value indicated a higher anti-proliferative activity of the extract compared to cisplatin. After 24 hours of treatment, Western blot analysis showed that treated HeLa cells exhibited a significant down-regulation of HPV18 oncoproteins E6 and E7, and a significant induction of caspase-8 and caspase-3 activation level. Meanwhile, the mRNA expressions of p53, pRb, Fas and FasL were significantly upregulated in treated cells. Moreover, ELISA showed an increased IL-12 and decreased IL-6 production after Oroxylum indicum treatment. Conclusions: The methanol extract of Oroxylum indicum has an anti-proliferative activity and proapoptotic potential. It induces localized-immunity improvements by altering cytokine production in HPV-positive cervical cancer cells.展开更多
Recently there has been a wide concern on inorganic nanoparticles as drug delivery carriers. CaCO3 particles have shown promising potential for the development of carriers for drugs, but little research had been perfo...Recently there has been a wide concern on inorganic nanoparticles as drug delivery carriers. CaCO3 particles have shown promising potential for the development of carriers for drugs, but little research had been performed regarding their safe dosage for maximizing the therapeutic activity without harming biosystems. In this study, we assessed the biological safety of porous spherical CaCO3 microparticles on Hela cells. The reactive oxygen species (ROS), glutathione (GSH), carbonyl content in proteins (CCP), DNA-protein crosslinks (DPC) and cell viability were measured. Results showed that with the exposure concentration increase, ROS and CCP in Hela cells presented a significant increase but GSH contents in Hela cells and cell viability showed a significant decrease respectively compared with the control. DPC coefficient ascended, but no statistically significant changes were observed. The results indicated that porous spherical CaCO3 microparticles may induce oxidative damage to Hela cells. But compared with other nanomaterials, porous spherical CaCO3 appeared to have good biocompatibility. The results implied that porous spherical calcium carbonate microparticles could be applied as relatively safe drug vehicles, but with the caveat that the effect of high dosages should not be ignored when attempting to maximize therapeutic activity by increasing the concentration.展开更多
Objective: To explore the impact of s-phase kinase-associated protein 2 (Skp2) on cervical cancer cell proliferation and the relationship between Skp2 and expression of cell regulation factors and transcription factor...Objective: To explore the impact of s-phase kinase-associated protein 2 (Skp2) on cervical cancer cell proliferation and the relationship between Skp2 and expression of cell regulation factors and transcription factors. Methods: RNAi technology was used to silence Skp2 gene in HeLa cells. After interference, RT-PCR was used for detection of Skp-2 mRNA, and Western blotting and flow cytometry were used for protein expression analysis. Results: siRNA significantly inhibited HeLa cell proliferation (P<0.05) and increased HeLa apoptosis, and G1/G0 phase cells were increased significantly (P<0.01). Skp2 siRNA transfected HeLa cells effectively reduced Skp2 protein levels, while p27 and p-p53 protein levels were increased significantly. RT-PCR results showed that after interference Skp2 mRNA, c-myc mRNA and cyclin A mRNA expressions decreased significantly compared with those in control group (P<0.01), and p27mRNA expression level was significantly higher (P<0.01). Conclusion: The change of Skp2 expression affects the expression of the cell cycle protein, thus affecting proliferation and apoptosis of HeLa cells. Skp2 protein plays an important role in the progression of cervical cancer; yet the specific mechanism still needs further study.展开更多
基金supported by the National Natural Science Foundation of China (No. 30960451)Major State Basic Research Development Program (2010CB535003)the Xinjiang production and construction corps funds for distinguished young scientists to ZHENG Qiu Sheng,international cooperation projects (2012BC001)
文摘Objective Isoliquiritigenin (ISL), a licorice chalconoid, is considered to be a bioactive agent with chemopreventive potential. This study investigates the mechanisms involved in ISL-induced apoptosis in human cervical carcinoma HeLa cells. Methods Cell viability was evaluated using a 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyl-tetrazolium bromide (MTT) assay. Apoptosis was determined by flow cytometry using an Annexin V-FITC Apoptosis Detection Kit. The intracetlular ROS levels were assessed using a 2, 7-dichlorofluorescein probe assay. The mitochondrial membrane potential was measured with the dual-emission potential-sensitive probe 5, 5', 6, 6'-tetra-chloro-1, 1', 3, 3'-tetraethyl-imidacarbocyanine iodide (JC-1). The degradation of poly-ADP-ribose polymerase (PARP) protein, the phosphorylation of PKR-like ER kinase (PERK), the phosphorylation of the a-subunit of eukaryotic initiation factor 2 (elF2a), the expression of the 78 kD glucose-regulated protein (GRP 78), and the activation of caspase-12 were analyzed via western blot analysis. Results ISL significantly inhibited the proliferation, the increase in ROS levels and apoptotic rates of HeLa cells in a concentration-dependent manner. Moreover, ISL induced mitochondrial dysfunction, caspase activation, and PARP cleavage, which displayed features of mitochondria dependent on apoptotic signals. Besides, exposure of HeLa cells to ISL triggered endoplasmic reticulum (ER) stress, as indicated by the increase in p-elF2a and GRP78 expression, ER stress-dependent apoptosis is caused by the activation of ER-specific caspase-12. Conclusion The findings from our study suggest that ISL-induced oxidative stress causes HeLa cel apoptosis via the mitochondrion-dependent and the ER stress-triggered signaling pathways.
基金supported by a grant from Natural Sciences Foundation of Hubei Province,China (No.CGZ0285)
文摘The recombinant plasmids pGenesil-1-BIRC71 and pGenesil-1-BIRC72 were transfected into Hela cells and cisplatin was added with different concentrations in order to study the inhibitory effects of Livin gene, increase the apoptosis induced by cisplatin, and detect the expression ofBcl-2, Bax, caspase-3, and survivin genes. The pGenesil-1-BIRC71 and pGenesil-1-BIRC72 were transfected into Hela cells, and the expression levels of Livin, Bcl-2, Bax, caspase-3, and survivin genes were detected by using fluorescence quantitative real-time PCR. Then cisplatin at different concentrations (3.0, 6.0 and 9.9 μg/mL) was added into the transfected Hela cells, and 24, and 48 h later, the apoptosis rate was measured by flow cytometry. After transfection of pGenesil-l-BIRC71 and pGenesil-1-BIRC72 into Hela cells, the expression level of Livin gene was obviously reduced, and the apoptosis rate was significantly increased in transfection group as compared with control group (P〈0.05). Cisplatin could increase the apoptosis rate in a dose- and time-dependent manner. After cisplatin was added, the expression levels of Bcl-2 mRNA were reduced, and those of Bax, caspase-3, and survivin mRNA were increased in transfection group as compared with those in control group (P〈0.05). It was concluded that shRNA expression vector targeting Livin gene could inhibit the expression of Livin gene in Hela cells and enhance the apoptosis induced by cisplatin, which was related to the decreased expression of Bcl-2 and activation of Bax and caspase-3. Survivin might play an important role as an antagonist in the process of apoptosis induction.
基金Supported by the Department of Science and Technology of Jilin Province(No.20020502).
文摘Hypocrellin A( HA), a photosensitive perylenequinone compound of Hypocrella bambusae, inhibited the proliferation of several tumor cell lines. Human cervical cancer cells, HeLa ceils, were used as a model to elucidate the molecular mechanisms of HA-induced tumor cell death. The results show that HA can induce the oligonucleosomal fragmentation of DNA in HeLa cells and also can increase the expression of apoptosis inducer Bax mRNA and that it decreases the expression of apoptosis suppressor, Bcl-2 mRNA, in mitochondria. It can be concluded from the data that HA-induced apoptosis is related to the balance between Bcl-2 and Bax gene expressions.
基金supported by the National Natural Science Foundation of China(No.81071663)
文摘To explore the effect of quercetin on the proliferation and apoptosis of HeLa cells, HeLa cells were incubated with quercetin at different concentrations. Cell viability was evaluated by MTT assay, cell apoptosis was detected by Annexin- V/PI double labeled cytometry and DNA ladder assay. Cell cycle was flow cytometrically determined and the morphological changes of the cells were ob- served under a fluorescence microscope after Hoechst 33258 staining and the apoptosis-related pro- teins in the HeLa cells were assessed by Western blotting. The results showed that quercetin signifi- cantly inhibited the growth of HeLa cells and induced obvious apoptosis in vitro in a time- and dose-dependent manner. Moreover, quercetin induced apoptosis of HeLa cells in cell cycle-dependent manner because quercetin could induce arrest of HeLa cells at G0/G1 phase. Quercetin treatment down-regulated the expression of the PI3K and p-Akt. In addition, quercetin could down-regulate ex- pression of bcl-2, up-regulate Bax, but exerted no effect on the overall expression of Akt. We are led to conclude that quercetin induces apoptosis via PI3k/Akt pathways, and quercetin has potential to be used as an anti-tumor agent against human cervix oancer.
文摘This paper deals with the inhibitory mechanisms of ginsenoside \{G Rh 2\} on the growth of tumor cells. \{G Rh 2\} significantly inhibited the proliferation of human cervical adenocarcinoma HeLa cells in a time and dose dependent manner. G Rh 2 induced apoptotic manifestations in HeLa cells as evidenced by the changes in the cell morphology, the DNA fragmentation and the activation of caspases. Caspase inhibitors, caspase family inhibitor, z Val Ala Asp fmk(z VAD fmk); caspase 1 inhibitor, Ac Tyr Val Ala Asp chloromethyl ketone(Ac YVAD cmk); caspase 3 inhibitor, z Asp Glu Val Asp fmk(z DEVE fmk) and caspase 8 inhibitor, \{z Ile \}Glu Asp fmk(z IETD fmk) effectively attenuated G Rh 2 induced cell death. The activities of caspase 1 and caspase 3 were increased in the G Rh 2 induced apoptotic process. However, caspase inhibitors can not inhibit G Rh - 2 induced cell death completely. These results suggest that G Rh 2 induced cell death is mediated by the activation of caspase cascade, but there might be some other pathways for induction of this apoptosis.
基金Supported by the Natural Science Foundation of Hubei Province (30113075)
文摘Effect of quercetin on HeLa cell system of cervical cancer was studied by methods of MTT and Annexin V-FITC/PI. The results show that quercetin has functions of inhibiting breeding of HeLa cells and inducing apoptosis of the cells. The total apoptosis rate is positively proportional to reaction duration and concentration of quercetin used. The maximum apoptosis rate being (88.76±2.35)% was obtained when the concentration was 50.0 μmol/L and the cells were treated with quercetin for 72 hours. Based on establishing a model of tumor of cervical cancer transplanted into nude mice, quercetin of different concentrations was injected into abdominal cavity of nude mice and situation of tumor growth was reviewed. The result showed that with quercetin concent'ration increasing from 0 to 100.0 μmol/L, the transplantation volume and weight of the tumors decreased from (279.59±70.58) mm^3 and (0.145±0.019) g to (128.72±36.12) mm^3 and (0.089± 0.019) g respectively, while apoptosis rate of the transplanted tumor increased from (9.63±1.85)% to (34,98±0.47)%, which proved that quercetin inhibited increment of volume and weight of transplanted tumor in nude mice bodies.
文摘The killing effects of lymphocytes on Hela cells expressing interleukin-12 (IL-12) in vitro were explored. By using gene transfection technique, full length IL-12 gene was transfected into Hela cells. The expression of IL-12 in Hela cells was detected quantitatively by ELISA; Changes in killing effects of lymphocytes on Hela cells expressing IL-12 were observed by MTT. It was found that Hela cells could express IL- 12 between 24 h and 72 h after transfection. Killing activity of lymphocytes on Hela cells expressing IL-12 was significantly enhanced. It was concluded by cell transfection technique, Hela cells could express 1L-12 and were more easily killed by lymphocytes.
文摘Objective: The histone deacetylase inhibitors (HDACIS) have been shown to inhibit cancer cell proliferation, stimulate apoptosis, an induce cell cycle arrest. Our purpose was to investigate the antiproliferative effects of a HDACI, trichostatin A (TSA), against human cervical cancer cells (HeLa). Methods: HeLa cells were treated in vitro with various concentrations of TSA. The inhibitory effect of TSA on the growth of HeLa cells was measured by MTT assay. To detect the characteristic of apoptosis chromatin condensation, HeLa cells were stained with Hoechst 33258 in the presence of TSA. Induction of cell cycle arrest was studied by flow cytometry. Changes in gene expression of p53, p21wafl and p27Kipl were studied by semiquantitative RT-PCR. Results: TSA inhibited cell growth in a time- and dose-dependent manner. Hoechst 33258 staining assay showed that TSA induced apoptosis. Cell cycle analysis indicated that treatment with TSA decreased the proportion of cells in S phase and increased the proportion of cells in G0/G1 and/or G2/M phases of the cell cycle. This was concomitant with overexpression of genes related to malignant phenotype, including an increase in p53, p21wall and p27Kipl. Conclusion: These results suggest that TSA is effective in inhibiting growth of HeLa cells in vitro. The findings raise the possibility that TSA may prove particularly effective in treatment of cervical cancers.
文摘Objective To observe the inhibition and radiation-sensitizing effect of Indoleacetic acid (IAA) combined with horseradish peroxidase(HRP)on Hela cells. Methods Hela cells were cultured in vitro and classified into control group, drug group incubated with different doses of IAA(30, 60, 90μmol·L -1) plus 1.2μg·mL -1 HRP, radiation group (6MV-X, 4Gy ) and group of radiation plus IAA plus HRP(same dose as above). All the above were treated for 24-96 hours.The growth inhibition, radiation-sensitizing effect were observed with methyl thiazolyl tetrazolium (MTT) photocolorimetric assay and trypan blue dye assay. The-effect on cell proliferation cycle was determined by flow cytometry. Results The antiproliferation activities showed a significant time-effect and dose-effect relationship to some extent after Hela cells were treated with IAA combined with HRP. The group of radiation plus 60μmol·L -1 IAA plus 1.2μg·mL -1 HRP and radiation plus 90μmol·L -1 IAA plus 1.2μg·mL -1 HRP showed an obvious radiation sensitizing effect. After treatment with 90μmol·L -1 IAA plus 1.2 μg·mL -1 HRP for 72 hours, the determination of cell cycle showed that the percentages of the cells on stages G 2-M and S were all higher than those of the control group. For the group of radiation plus IAA combined with HRP, the percentages of the cells on stages G 2-M were higher than those of the radiation group. Conclusion The above findings suggest that IAA combined with HRP has an inhibitive and killing effect on Hela cells. The effect was stronger during the cell cycles of G 2-M and S. It also has a radiation sensitizing effect. Its mechanism might be that Hela cells were blocked on stages G 2-M, and it presents a collaborative killing effect during miototic time.
基金Hubei Provincial Department of Education Funds (D200511008)
文摘Objective To investigate the Hela cells growth inhibition and apoptosis possible molecular mechanisms. Methods Hela cells were treated with various concentrations (100μmol/L,200μmol/L, 300 μmol/L, 400 μmol/L) of NS-398 (selective for COX-2 inhibition). Cell growth was measured by MTT (Thiazolyl blue). Apoptosis was detected by double staining flow cytometry (FCM). Levels of PGE: were measured by radioimmunoassay. The expressions of COX-2 protein were also examined by Western blot analysis. Results After treated with different concentrations of NS-398, the growth of Hela cells was suppressed significantly in a dose-and time-dependent manner (P〈0.01). The NS-398 can induce apoptosis with the apoptosis rates at 8.53%-43.46% by FCM in a dose-dependent manner. The release of PGE2 was reduced in Hela cells with the values of 69.26 ± 2.13, 47.46 ± 2.18, 28.15 ± 1.64 and 17.01 ± 1.12, respectively, there was significant difference compared with control group (83.78 ± 1.11) (P〈0.01). The NS-398 could inhibit the activity and expression of COX-2 in a dose- dependent manner and down-regulated the expression of COX-2 protein greatly. Conclusion NS-398 could inhibit the proliferation and increase apoptosis in human Hela cells. These effects may be depended on the inhibition of the expression of COX-2 and PGE2 by NS-398.
基金Supported Partly by the Program of Jilin Provincial Science & Technology Department,China(Nos.200705335 and 200504057)the Program of Development and Reform Commission of Jilin City,China(No.2004987)
文摘To construct a single tetracycline-regulatable plasmid vector based on the double tetracycline-regulatable plasmid vector system for regulating HSV-tk gene expression so as to effectively kill HeLa cells. Two tetracycline operator(TetO2) was cloned into pcDNA3.1 and a cassette was made for a cytomegalovirus-type 2 tetracycline operator(CMV-TetO2) promoter, and the obtained vector was named pcDNA3.1-CMV-TetO2. Herpes simplex virus thymidine kinase(HSV-tk) gene and tetracycline repressor(TR) gene were cloned into pcDNA3.1-CMV-TetO2 and the two genes were linked with internal ribosome entry site(IRES) to gain a vector named pcDNA3.1-CMV-TetO2- HSV-tk-IRES-TR. The HeLa cells were stablly transfected with pcDNA3.1-CMV-TetO2-HSV-tk-IRES-TR plasmid. The expression of HSV-tk and TR were detected by RToPCR, the tumorcidal activity of HSV-tk/GCV was determined by MTT assay. In Hela cells transfected with the above plasmid vector, HSV-tk gene and TR gene can be expressed lowly and the concentration of GCV producing a 50% decrease in cell viability was about 50 lag/mL without adding deoxycycline; in contrast, the expessions of HSV-tk gene and TR gene increased significantly and the concentration of GCV producing a 50% decrease in cell viability was about 5 μg/mL with adding deoxycycline. Therefore tetracycline can regulate the expression and tumorcidal activity of HSV-tk gene in HeLa cells with this single plasmid vector.
基金National natural Science Foundation of China(31272446)Fundamental Research Funds for the Central Universities(No.2011PY020 and 52902-0900206126).
文摘Inhibins play important roles in the reproductive system. To evaluate whether inhibin α (1-32) fusion protein plays a role in cervical cancer growth, the plasmid pVAX-inhα was constructed and its effect on proliferation and apoptosis of the human cervical cancer cell line (Hela) was checked by flow cytometry and real-time PCR. The expression and localization of inhibin α protein were detected by RT-PCR and confocal mi- croscopy which showed that inhibin α protein was expressed and localized in the nucleus of Hela cells. Over ex- pression of inhibin α gene significantly induced cell apoptosis and ceased S phase of cell cycle. Furthermore, cell proliferation was significantly suppressed 96 h post-transfection and mRNA level of anti-apoptosis genes (Bcl-2, NFκB) were decreased but pro-apoptosis genes (Bax, wild type p53) and inhibin co receptor (TGFβR3) were increased, indicating that inhibin, through its co-receptor, might activate apoptotic and cell growth cascades which regulate proliferation and apoptosis in Hela cells. These results suggest that inhibin α (1-32) fusion pro- tein, located in the cell nucleus, can regulate Hela cells growth and apoptosis by induction of apoptotic pathways such as NFκB, Bcl-2 and p53 families. These findings may have a significant impact on future research regarding cervical cancer cell lines.
文摘A method is described which permits transmission electron microscope of single cells treated with HpD plus laser microirradiation. The preselected single cell that was irradiated by laser under light microscope and followed fixation, embedded and sectioning is examined under electron microscope. The results demonstrated that at the light dose of 1.88 ml/μm2 not only the irradiated nucleolus appeared transparent region, but the other parts such as non-irradiated mitochondria in cytoplasm can also be damaged. When partial cytoplasm is irradiated with the light dose of 4.50 ml/μm2, the damages appear in all cytoplasm, but there is little change in the nucleus. The experimental results also demonstrate that cytoplasm is more sensitive than nucleus. It is the mitochondria in cytoplasm that are very sensitive to HpD plus laser.
文摘This study aimed to investigate the effect of fullerenol combined with cisplatin on the proliferation of cervical cancer HeLa cells in order to provide new ideas and laboratory theoretical basis for the clinical treatment of cervical cancer. Cervical cancer cell line HeLa cells in vitro were treated with different concentrations of fullerenol, different concentrations of cisplatin, and different concentrations of fullerenol combined with cisplatin, after 24 h, 48 h, 72 h, microscope changes in cell morphology;MTT assay was used to determine the effect of drugs on the proliferation of HeLa cells. Fullerenol and cisplatin alone can inhibit the proliferation of HeLa cells in a dose-dependent and time-dependent manner;compared with cisplatin alone, different concentrations of fullerenol combined with cisplatin significantly increased the apoptosis rate of HeLa cells (P < 0.05). The inhibition of fullerenol combined with cisplatin on HeLa cells in vitro is more significant, resulting in a stronger anti-cancer effect.
文摘Aging is a decelerating unidirectional process of life. Shortening of telomeric DNA, the (TTAGGG)<sub>n</sub> hexanucleotide repeats, which form the caps at the chromosome ends, is implicated to determine the aging process, and more importantly the healthy lifespan itself. Telomerase, a ribonucleoprotein having reverse transcriptase activity, arrests telomere loss through addition of the TTAGGG repeats de novo, to the ends of the chromosome. The telomere/telomerase maintenance is an inevitable necessity to delay aging and for a healthy lifespan. Here, we report the potential of full-spectrum, high concentration Ashwagandha (Withania somnifera), an Ayurvedic medicinal herb, root extract to increase telomerase activity. HeLa cells, when treated with various concentrations of Ashwagandha root extract, showed an increase in telomerase activity measured with the established Telomerase Rapid Amplification Protocol (TRAP) assay. Ashwagandha root extract increased telomerase activity with highest enhancement of ~45% at 10 - 50 μg concentration. Thus, Ashwagandha root extract has the anti-aging inducing potential.
基金supported by Tangshan City Science And Technology Research And Development Project(14130246a)
文摘Objective:To explore the effect of bortezomib on migration and invasion of cervical carcinoma HeLa cell and specific molecular mechanism.Methods:The effect of bortezomib on the viability of HeLa cell was measured by MTT assay.The effect of bortezomib on cell migration and invasion was measured by Transwell assay and invasion experiment respectively.The activation of Akt/mTOR signaling pathway and expression level of MMP2,MMP9 were assayed by western blot.Results:MTT assay indicated bortezomib(2.5 μM.5 μM,10 μM)could inhibit HeLa cell viability,and the inhibitory rate was highest at 48 h.Transwell assay and invasion experiment results showed that bortezomib inhibited HeLa cell migration and invasion.Western blotting assays presented bortezomib could suppress the phosphorylation of Akt and mTOR.and down-regulate the expression of MMP2 and MMP9.Conclusions:These results suggested bortezomib could inhibit migration and invasion in cervical carcinoma HeLa cell,which might be related to Akt/mTOR signal pathway.
基金supported by Fundamental Research Grant Scheme Grant 203/PPSK/6171191
文摘Objective: To examine the proapoptotic properties of Oroxylum indicum methanol extract on cervical cancer cells. Methods: Methylene blue assay was used to determine the IC50 value of the extract. Western blotting assays were done to analyze the expression of HPV oncoproteins (HPV18 E6 and E7) and apoptotic molecules (caspase-3 and caspase-8). Reverse transcriptase PCR assays were performed to determine genetic alteration of tumor suppressors p53 and pRb and apoptosis markers Fas and FasL. Enzyme-linked immunosorbent assay (ELISA) was done to determine the expression of cytokine levels (IL-6 and IL-12). Results: The determination of IC50 value indicated a higher anti-proliferative activity of the extract compared to cisplatin. After 24 hours of treatment, Western blot analysis showed that treated HeLa cells exhibited a significant down-regulation of HPV18 oncoproteins E6 and E7, and a significant induction of caspase-8 and caspase-3 activation level. Meanwhile, the mRNA expressions of p53, pRb, Fas and FasL were significantly upregulated in treated cells. Moreover, ELISA showed an increased IL-12 and decreased IL-6 production after Oroxylum indicum treatment. Conclusions: The methanol extract of Oroxylum indicum has an anti-proliferative activity and proapoptotic potential. It induces localized-immunity improvements by altering cytokine production in HPV-positive cervical cancer cells.
文摘Recently there has been a wide concern on inorganic nanoparticles as drug delivery carriers. CaCO3 particles have shown promising potential for the development of carriers for drugs, but little research had been performed regarding their safe dosage for maximizing the therapeutic activity without harming biosystems. In this study, we assessed the biological safety of porous spherical CaCO3 microparticles on Hela cells. The reactive oxygen species (ROS), glutathione (GSH), carbonyl content in proteins (CCP), DNA-protein crosslinks (DPC) and cell viability were measured. Results showed that with the exposure concentration increase, ROS and CCP in Hela cells presented a significant increase but GSH contents in Hela cells and cell viability showed a significant decrease respectively compared with the control. DPC coefficient ascended, but no statistically significant changes were observed. The results indicated that porous spherical CaCO3 microparticles may induce oxidative damage to Hela cells. But compared with other nanomaterials, porous spherical CaCO3 appeared to have good biocompatibility. The results implied that porous spherical calcium carbonate microparticles could be applied as relatively safe drug vehicles, but with the caveat that the effect of high dosages should not be ignored when attempting to maximize therapeutic activity by increasing the concentration.
文摘Objective: To explore the impact of s-phase kinase-associated protein 2 (Skp2) on cervical cancer cell proliferation and the relationship between Skp2 and expression of cell regulation factors and transcription factors. Methods: RNAi technology was used to silence Skp2 gene in HeLa cells. After interference, RT-PCR was used for detection of Skp-2 mRNA, and Western blotting and flow cytometry were used for protein expression analysis. Results: siRNA significantly inhibited HeLa cell proliferation (P<0.05) and increased HeLa apoptosis, and G1/G0 phase cells were increased significantly (P<0.01). Skp2 siRNA transfected HeLa cells effectively reduced Skp2 protein levels, while p27 and p-p53 protein levels were increased significantly. RT-PCR results showed that after interference Skp2 mRNA, c-myc mRNA and cyclin A mRNA expressions decreased significantly compared with those in control group (P<0.01), and p27mRNA expression level was significantly higher (P<0.01). Conclusion: The change of Skp2 expression affects the expression of the cell cycle protein, thus affecting proliferation and apoptosis of HeLa cells. Skp2 protein plays an important role in the progression of cervical cancer; yet the specific mechanism still needs further study.