Physicochemical Characterization, Cytotoxic Activity, and Caspase-9 Expression of Nanogold-Parijoto (Medinilla Speciosa Reinw .Ex, Bl ) in Hela Cell Lines

To explore the effects of paeoniflorin on the proliferation and apoptosis of HeLa cells. HeLa cells treated with paeoniflorin at different concentrations for different hours were assessed by the method of methyl thiazolyl tetrazolium (MTT). Cell apoptosis rate and cycle change were detected by flow cytometry with annexin V-fluorescein isothiocyanate (FITC)/propidium iodide (PI). The morphological change of HeLa cells was observed by transmission electron microscope (TEM). The expressions of Bcl-2, Bax and Caspase-3 in HeLa cells induced by paeoniflorin were detected by immunocytochemistry. After treatment with different concentrations of paeoniflorin for different hours, the proliferation of HeLa cells was inhibited in a dose and time-dependent manner with an IC50 value of 5054, 2965, 2459 µg/ml at 24, 48 and 72 hours respectively (P < 0.05). After treated by different concentrations of paeoniflorin (control group, 1000 and 2000 µg/ml), apoptosis was induced in HeLa cells at a rate of 0.94%, 10.94% and 13.95% respectively. Such effects were dose-dependent (P < 0.05) and the proportion of HeLa cells in Sphase was under a rising trend. Typical apoptotic changes of HeLa cells under the exposure to paeoniflorin were observed under TEM. There was a lowered expression of Bcl-2 and an elevated expression of Bax and Caspase-3 genes versus the control group after a 48-hour paeoniflorin treatment (P < 0.05). Paeoniflorin can significantly induce the apoptosis of HeLa cells through a down-regulation of anti-apoptotic gene Bcl-2 and an up-regulation of pro-apoptotic genes Bax and Caspase-3.

Caspases, members of the cysteine protease family, execute UVB-induced apoptosis in several cell lines and keratinocytes. Several researchers investigating UVB-induced apoptosis have demonstrated a dose-dependent protective effect of the synthetic peptide caspase inhibitor zVAD-fmk. However, zVAD-fmk displays a dose-dependent protective effect against UVB-induced apoptosis, even at doses higher than those required to block all known proapoptotic caspases. In addition, it is known that zVAD-fmk also inhibits other cysteine proteases including cathepsins and calpains, and these proteases have recently been demonstrated to play a role in the execution of programmed cell death induced by other stimuli, e.g. TNF-alpha. The purpose of the present study was therefore to investigate whether inhibitors of cysteine cathepsins and calpains could prevent UVB-induced apoptosis in HeLa cells and keratinocytes. This was done by investigating the effect of the irreversible cysteine protease inhibitor zFA-fmk, the cathepsin B inhibitor CA-074-Me and the calpain inhibitor ALLN on the viability of UVB-irradiated human keratinocytes and HeLa cells. At concentrations of 10 microM and above zVAD-fmk conferred partial dose-dependent protection against UVB-induced apoptosis in HeLa cells and keratinocytes. Moreover, caspase-3 activity was completely blocked at zVAD-fmk concentrations of 1 microM in HeLa cells. This indicates that caspase-independent mechanisms could be involved in UVB-induced apoptosis. However, the protease inhibitors zFA-fmk, CA-074-Me and ALLN all failed to prevent UVB-induced apoptosis in HeLa cells and keratinocytes. In conclusion, the protective effect of zVAD-fmk at high concentrations indicates that other proteases than caspases are active in the execution of UVB-induced apoptosis but further studies are needed to identify these proteases.

Cassia fistula L. bark fraction modulated GSK3β/ p53 expression for mitochondrial mediated apoptosis in HeLa cells

Antiproliferative and apoptosis inducing activity of Markhamia tomentosa leaf extract on HeLa cells

Cervical cancer (CC) is considered a public health problem. Circular RNAs (circRNAs) serve important roles in different types of cancer, including CC. However, the mechanisms used by circRNAs to facilitate CC progression are currently unclear. The present study analyzed the effects of hsa_circ_0009910 knockdown on microRNA (miRNA/miR)-198 and mesenchymal-epithelial transition factor (c-Met) expression levels and its impact on apoptosis and the viability of HeLa cells. Differentially expressed circRNAs in CC were identified using analysis of circRNA microarray data. Bioinformatics analysis was performed to predict circRNA-microRNA (miRNA) and miRNA-mRNA interactions. The knockdown of hsa_circ_0009910 in HeLa cells was performed using small interfering RNA and the expression levels of hsa_circ_0009910, miR-198 and c-Met were assessed using reverse transcription-quantitative PCR. The viability and apoptosis of HeLa cells were evaluated using MTT, neutral red uptake and ApoLive-Glo™ multiplex assays. Hsa_circ_0009910 was significantly upregulated in HeLa cells and the knockdown of hsa_circ_0009910 increased miRNA-198 expression levels, reduced c-Met expression levels and decreased cellular viability, but not apoptosis, in HeLa cells. Overall, these results indicated that hsa_circ_0009910 could act as a molecular sponge of miRNA-198 and contribute to the upregulation of c-Met expression levels. The hsa_circ_0009910/miRNA-198/c-Met interaction network affects the viability, but not apoptosis, of HeLa cells. Based on this mechanism, the present study suggests that hsa_circ_0009910 may be a promising biomarker for CC.

Chrysin and its phosphate ester inhibit cell proliferation and induce apoptosis in Hela cells

BackgroundHOTAIR was known to enhance radioresistance in several cancers. However, the function of HOTAIR on radioresistance involving the regulation of HIF-1α in cervical cancer has not been reported.MethodsBALB/c nude mice were injected subcutaneously with HeLa cells and irradiated by X-ray. The tumor volume was measured and the expression of HOTAIR in tumors was detected by quantitative real-time PCR. Western blot was performed to detect the protein level of HIF-1α. MTT (3-(4,5-Dimethylthiazol-2-yl) 22,5-diphenyltetrazolium bromide) assay and the terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay was used to examine the cell viability and cell apoptosis of HeLa cells and C33A cells exposed to radiation.ResultsRadiotherapy inhibited the tumor growth in mice bearing HeLa cells. Radiotherapy reduced the expression of HOTAIR and HIF-1α in tumor tissues and HeLa cells or C33A cells. HOTAIR overexpression abrogated the effect of radiation on the cell viability and cell apoptosis of HeLa and C33A cells. HOTAIR also upregulated the expression of HIF-1α in HeLa and C33A cell exposed to radiation. HIF-1α knockdown reversed increasing cell viability and reducing apoptosis of HeLa and C33A cell induced by HOTAIR overexpression. HOTAIR overexpression promoted tumor growth in mice bearing HeLa and exposed to radiation.ConclusionRadiotherapy might inhibit cervical cancer cell growth through HOTAIR/HIF-1α pathway.

Cervical cancer is the second common cause of cancer deaths in women worldwide. Radioresistancy of cancer is a principal cause of treatment impairing. Inhibitor of apoptosis proteins (IAPs) widely block apoptosis against apoptotic stimuli, including current chemo- and radiation therapies. Apollon, a membrane of IAP, can support cells against apoptosis and is over expressed in some treatment-resistant cancer cells. The aim of this study was to evaluate the effects of apollon knockdown on induction of apoptosis and also its potential for enhancement of radiosensitvity on hela cells. plasmid encoding shRNA which has been confirmed its effect against apollon, transfected into hela cells. Consequent effects on the level of P53 , Bax and BAK analyzed by real time PCR. Apoptotic phenotype of transfected cells was monitored by Tunnel assay. Viability of hela cells after radiotherapy was analyzed by MTT assay. shRNA1 effectively increased transcription of p53, Bax and BAK and induced apoptosis phenotype of treated hela cells. Radiosensitivity of transfected cell was increased after knock-down of apollon obviously. Apollon knockdown induces apoptosis in hela cell . Also it can be as new molecular target for radio-sensitizing strategies in these cells. So, apollon can be a potentially considerable therapeutic object for cervical cancer.

BackgroundCervical cancer threatens women’s health worldwide. Verteporfin (VP), a small-molecule YAP1 inhibitor, inhibits cancer cell growth. This study investigated whether VP could inhibit the proliferation and promote the apoptosis of cervical cancer cells by decreasing SULT2B1 expression.Material/MethodsNormal and cancerous cervical cell proliferation after VP treatment was detected by CCK-8 assay. HeLa cell migration, invasion, and apoptosis after VP treatment and transfection were analyzed by wound healing assay, transwell assay, and TUNEL assay, respectively. The expression of related proteins was determined by western blot analysis. Western blot and RT-qPCR analysis detected mRNA and protein expression of SULT2B1.ResultsDifferent VP concentrations (0.5, 1, 2, and 5 μM) inhibited the viability of HeLa cells and had no obvious effect on H8 cells. Therefore, 5 μM VP was selected for subsequent experiments. VP inhibited the proliferation, migration, and invasion of HeLa cells and promoted their apoptosis. Bcl-2 expression decreased, and expression of Bax, caspase-3, and caspase-9 in VP-treated HeLa cells increased. SULT2B1 expression increased in cervical cancer cells compared with normal cervical cells. Furthermore, SULT2B1 expression increased in HeLa cells and VP suppressed SULT2B1 expression. SULT2B1 overexpression reduced the inhibiting effect of VP on the proliferation, migration, and apoptosis of HeLa cells, and reduced VP effect on apoptosis of HeLa cells. SULT2B1 overexpression upregulated the Bcl-2 expression and downregulated the expression of Bax, caspase-3, and caspase-9 in VP-treated HeLa cells.ConclusionsVP inhibited the proliferation, migration, and invasion and promoted apoptosis of cervical cancer cells by decreasing SULT2B1 expression.

This paper investigated the effect of total saponins from Rhizoma Panacis Majoris on the proliferation, apoptosis, and autophagy of human cervical carcinoma HeLa cells. The saponin content was detected by ultraviolet-visible spectrophotometry. Cell coun-ting kit-8(CCK-8) assay, 4,6-diamidino-2-phenylindole(DAPI) staining, and flow cytometry were used to detect the effects of total saponins of Panacis Majoris Rhizoma on cell viability, morphology, cell cycle and apoptosis of HeLa cells. Western blot was used to detect the expression of apoptosis-related proteins B cell lymphoma-2(Bcl-2), Bcl-2-associated X protein(Bax), cleaved caspase-9, and cleaved caspase-3, autophagy-related proteins Beclin-1 and SQSTM1(p62), and the proteins related to the phosphatidylinositol 3-kinase/protein kinase B/mammalian target of rapamycin(PI3K/Akt/mTOR) and mitogen-activated protein kinase(MAPK) signaling pathways. It was found that the yield and saponin content of total saponins from Rhizoma Panacis Majoris were 6.3% and 78.3%, respectively. Total saponins from Rhizoma Panacis Majoris could significantly inhibit the proliferation(P&lt;0.001), effect the nuclear morphology, block the G_0/G_1 cycle, and induce cell apoptosis in HeLa cells with a concentration-dependent manner. In addition, total saponins from Rhizoma Panacis Majoris up-regulated the expression of pro-apoptotic proteins Bax, cleaved caspase-9, and cleaved caspase-3, and autophagy-related protein p62(P&lt;0.05), while down-regulated the expression of anti-apoptotic protein Bcl-2 and autophagy-related protein Beclin-1(P&lt;0.01). Total saponins from Rhizoma Panacis Majoris could promote the expression of p-p38/p38, p-Jun N-terminal kinase(JNK)/JNK, p-PI3K/PI3K, p-Akt/Akt, p-mTOR/mTOR proteins in PI3K/Akt/mTOR and MAPK signaling pathways(P&lt;0.05). In contrast, the effect on p-ERK/ERK expression was not obvious. Therefore, total saponins from Rhizoma Panacis Majoris may inhibit autophagy and promote apoptosis of HeLa cells through the activation of the PI3K/Akt/mTOR, c-JNK, and p38 MAPK signaling pathways, which indicates that total saponins from Rhizoma Panacis Majoris may have a potential role in cervical cancer treatment.

To investigate the pro-apoptotic effects of alpha1-adrenergic inhibitor doxazosin in HeLa cells and the potential involvement of transcription factor activator protein-2alpha (AP-2alpha) in doxazosin-induced apoptosis. The HeLa cells were exposed to various concentrations of doxazosin for 16 h. Apoptosis was detected using a DNA fragmentation assay, Hoechst 33258 staining, and flow cytometric analysis. The expression of AP-2alpha and caspase-3 was detected by relative quantitative RT-PCR and Western blot assays, respectively. After the respective transfections of the HeLa cells with AP-2alpha overexpressing constructs and an antisense oligonucleotide against AP-2alpha, apoptosis was assessed by flow cytometric analysis, and the expression of AP-2alpha and caspase-3 was detected by relative quantitative RT-PCR and Western blot assays. The colorimetric assay was performed to detect the caspase-3 activity. Treatment with various concentrations of doxazosin for 16 h increased the apoptotic rate and total cell death rate of the HeLa cells in a dose-dependent manner and upregulated the expression of AP-2alpha and caspase-3 in a dose-dependent manner. A dose-dependent increase was observed in the caspase-3 activity. Overexpressing AP-2alpha led to the increased rate of doxazosin-induced apoptosis and the total cell death, whereas doxazosin-induced apoptosis and the total cell death in HeLa cells decreased by antisense AP-2alpha. Furthermore, overexpressing AP-2alpha increased the expression and activity of caspase-3, whereas antisense AP-2alpha in part abolished the increased effects of doxazosin on caspase-3 expression and activity. Doxazosin induces apoptosis in HeLa cells in a dose-dependent manner, and transcription factor AP-2alpha is functionally involved in doxazosin-induced HeLa cell apoptosis.

Pseudolaric acid B (PAB) is a diterpene acid isolated from the bark of the root and trunk of Pseudolarix kaempferi Gordon (Pinaceae), which has demonstrated cytotoxic effects against various types of cancer. However, the mechanisms underlying the anticancer effects of PAB have remained to be elucidated. In the present study, the effects of PAB on the viability and apoptosis of HeLa cells were investigated by MTT assay, flow cytometric analysis of Annexin V-fluorescein isothiocyanate/propidium iodide staining, Rhodamine 123 staining and western blot analysis. The results demonstrated that PAB had antiproliferative and apoptosis-inducing effects on HeLa cells. PAB markedly inhibited HeLa cell viability in a time- and concentration-dependent manner. Flow cytometric analysis indicated that PAB induced apoptosis in HeLa cells in a dose-dependent manner. Treatment with PAB suppressed the expression of anti-apoptotic factor B cell lymphoma-2, and promoted the expression of pro-apoptotic factor Bcl-2-associated X protein. In addition, PAB induced an increase in Caspase-3 activity and loss of mitochondrial membrane potential, suggesting that this apoptosis may be mediated by mitochondrial pathways. Furthermore, the results of western blot analysis indicated that PAB was able to reduce Akt phosphorylation, thereby inhibiting the Akt pathway. These results suggested that PAB inhibited cell proliferation and induced apoptosis in HeLa cells, and that the anti-tumor effects of PAB were associated with inhibition of the Akt pathway. In conclusion, the results of the present study suggested that PAB may represent a novel therapeutic strategy for the treatment of human cervical cancer. However, additional studies are required to investigate the underlying apoptotic mechanisms.

Tubeimoside I (TBMS1), a triterpenoid saponin, isolated from the tubers of Bolbostemma paniculatum, showed potent antitumor and antitumor-promoting effects. The objective of this study is to investigate the role of mitochondria and mitochondria cytochrome c in TBMS1-mediated apoptosis of human cervical carcinoma HeLa cell line. Viability of HeLa cells was measured by MTT assay. Apoptotic induction by TBMS1 was determined by fluorescence microscopy, flow cytometry and gel electrophoresis of fragmented DNA. Mitochondrial transmembrane potential (Deltapsim) was assayed by flow cytometry. Cytochrome c (Cyt c) was detected by Western blotting. The results showed that Cyclosporin A (CsA) partly protected HeLa cells from growth inhibitory effect of TBMS1, and partly countered the ability of TBMS1 to rapidly induce apoptosis in HeLa cells, and that TBMS1 decreased Deltapsim and induced Cyt c release by a mechanism inhibited by CsA, and that TBMS1 induced apoptosis of HeLa cells dose-dependently in accordance with increase of cytosolic Cyt c. TBMS1 opens the permeability transition (PT) pore, thereby decreasing Deltapsim, releasing Cyt c from mitochondria, and further causing a series of events consistent with established mechanistic models of apoptosis.

Growth inhibition and induction of apoptosis in human cancerous HeLa cells by Maytenus procumbens

HPV+ cervical carcinomas and cell lines display altered expression of caspases