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Objective To investigate the effects of groove topography on morphology and migration speed of cervical cancer HeLa cells. Methods HeLa cells were cultured on PDMS substrates with four different surface features, namely, flat substrate, 10 μm width parallel groove, 20 μm width parallel groove, bifurcate groove. Immunofluorescence technique was used to transfect F-actin in HeLa cells, and specific probes Mito-Tracker Green were used to label mitochondria. The location, morphology of cells and distribution of mitochondrial at different moments were obtained through the living cell system. Results Compared with 20 μm width parallel groove and flat substrate, HeLa cells in 10 μm width parallel groove were more orderly arranged and more elongated, but their migration speed was much slower. HeLa cells at the bifurcation protruded into branches and mitochondria were mainly distributed at the protrusion and around the nucleus. The bifurcation reduced the average migration speed of HeLa cells in 10 μm width parallel groove. Conclusions Groove topography has a significant effect on morphology and migration speed of HeLa cells. The research findings help to understand the role of topography in in vivo microenvironment during migration of HeLa cells, and provide references for the subsequent research on invasion and metastasis of cervical cancers.
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Objective:To explore the effect of miR-21 on cell proliferation, apoptosis, invasion and radiosensitivity of cervical cancer HeLa cells and unravel the underlying mechanism.Methods:RT-qPCR assay was used to detect the expression levels of miR-21 in cervical cancer tissues and adjacent non-tumor tissues, normal cervical epithelial cells (H8) and cervical cancer cell lines (HeLa, SiHa, ME180). HeLa cell line with inhibition of miR-21 or knockdown of RECK were constructed. CCK-8, Caspase3/7 live cell apoptosis detection, wound healing test, Transwell invasion, clone formation assay, Western blot and immunofluorescence were performed to detect cell viability, apoptosis, migration, invasion, radiosensitivity and related proteins. The dual luciferase assay verified whether miR-21 targeted RECK.Results:MiR-21 level in the cervical cancer tissues was significantly higher than that in its corresponding adjacent non-tumor tissues ( P<0.05). The expression levels of miR-21 in cervical cancer cell lines HeLa, SiHa and ME180 were significantly up-regulated compared with those in normal cervical epithelial cells H8(all P<0.05). MiR-21 knockdown significantly inhibited HeLa cell viability, promoted cell apoptosis, reduced radiation tolerance, down-regulated the expression of Cyclin D 1,Bcl-2, MMP-2 and MMP-9, and up-regulated the expression P21 and Bax proteins (all P<0.05). miR-21 targeted the 3’-UTR of RECK mRNA and negatively regulated the expression of RECK. Silencing RECK reversed the effects of miR-21 knockdown on HeLa cell apoptosis, migration, invasion and radiosensitivity. Conclusions:Inhibiting the expression of miR-21 significantly decreases cell viability, induces cell apoptosis, weakens cell migration and invasion capabilities, and enhances the radiosensitivity of HeLa cells. The potential mechanism is closely related to the targeted up-regulation of RECK.
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Objective:To evaluate the effect of resveratrol combined with γ-ray irradiation on the biological behavior of cervical cancer cells, and to explore its possible mechanism.Methods:The proliferation of cell populations after different concentrations of resveratrol solution±γ-ray irradiation was detected by CCK-8 assay. Scratch test and Transwell chamber test were used to detect cell migration and invasion. Flow cytometry and Annexin V-FITC/PI double staining were employed to assess cell apoptosis. Western blot was performed to measure the expression levels of PI3K, Akt, p-Akt, mTOR and p-mTOR proteins.Results:Compared with the normal control (NC) group, the resveratrol group±γ-ray irradiation could inhibit the proliferation, migration, and invasion and promote cell apoptosis of human cervical cancer Hela cells, and the combined effect was more obvious. Compared with the NC group, resveratrol and γ-ray irradiation could significantly down-regulate the expression levels of Bcl-2, PI3K, p-Akt and p-mTOR proteins, up-regulate the expression level of Bax protein, but did not significantly alter the expression levels of Akt and mTOR proteins in human cervic1 255al cancer Hela cells.Conclusions:Resveratrol combined with γ-ray irradiation can dramatically inhibit the proliferation, migration, invasion, and promote the apoptosis of cervical cancer Hela cells. The mechanism may be related to the inhibition of the PI3K/Akt/mTOR signaling pathway and down-regulating the expression levels of downstream related proteins.
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The study aimed to achieve enhanced targeted cytotoxicity and cell-internalization of cisplatin-loaded deoxyribonucleic acid-nanothread (CPT-DNA-NT),mediated by scavenger receptors into HeLa cells.DNA-NT was developed with stiff-topology utilizing circular-scaffold to encapsulate CPT.Atomic force microscopy (AFM) characterization of the DNA-NT showed uniformity in the structure with a diameter of 50-150 nm and length of 300-600 nm.The successful fabrication of the DNA-NT was confirmed through native-polyacrylamide gel electrophoresis analysis,as large the molecular-weight (polymeric) DNA-NT did not split into constituting strands under applied current and voltage.The results of cell viability confirmed that blank DNA-NT had the least cytotoxicity at the highest concentration (512 nM) with a viability of 92％ as evidence of its biocompatibility for drug delivery.MTT assay showed superior cyto-toxicity of CPT-DNA-NT than that of the free CPT due to the depot release of CPT after DNA-NT inter-nalization.The DNA-NT exhibited targeted cell internalizations with the controlled intracellular release of CPT (from DNA-NT),as illustrated in confocal images.Therefore,in vitro cytotoxicity assessment through flow cytometry showed enhanced apoptosis (72.7％) with CPT-DNA-NT (compared to free CPT;64.4％).CPT-DNA-NT,being poly-anionic,showed enhanced endocytosis via scavenger receptors.
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AIM: To study the effects of forsythiaside B on HeLa cells proliferation by inhibiting NF-κB through p21/Cyclin E/CDK2 signaling pathway. METHODS: The cell activity of HeLa cells was observed by MTT assay after 24 h, 48 h and 72 h treatment with different concentration (1, 2, 4 μmol/L) of forsythiaside B. The clone formation of HeLa cells was evaluated with the condition of different concentrations of forsythiaside B. The expression of transcription factor NF-κB (p-p65 and p65) was determined, and the expression of p21, Cyclin E and CDK2 were detected as well.RESULTS:Forsythioside B inhibited the clone formation of HeLa cells in concentration dependent manner, and the inhibitory effect on HeLa cells' proliferation by forsythioside B was in both time and concentration dependent manner. Forsythioside B up-regulated p21 protein expression and down-regulated p-p65, Cyclin E and CDK2 protein expression. CONCLUSION:Forsythioside B inhibits the transcription factor NF-κB and affects the p21/Cyclin E/CDK2 signaling pathway, thus inhibiting the proliferation of HeLa cells.
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@#[Abstract] Objective: To explore the regulatory effect of lncRNA maternal imprinting gene 3 (MEG3) on proliferation, migration, invasion and epithelial-mesenchymal transition (EMT) of cervical cancer cells via miR-9-5p/SOCS5 axis. Methods: A total of 20 pairs of cancer and para-cancerous tissue specimens resected from cervical cancer patients in Chongqing Hospital of Traditional Chinese Medicine from January 2017 to June 2019 were collected for this study. Using liposome transfection technology, pcDNA3.1-MEG3,si-MEG3, miR-9-5p mimics, miR-9-5p inhibitor and their control plasmids were transfected into cervical cancer HeLa and SiHa cells respectively to construct overexpression and silence cell model. qPCR was used to detect the expression levels of MEG3, miR-9-5p and SOCS5 in cervical cancer tissues and cell lines. CCK-8 method and Transwell chamber method were used to detect cell proliferation, migration and invasion ability. The expression levels of E-cadherin and vimentin in cells were detected by cellular immunofluorescence experiments. Target genes were predicted through the Online Bioinformatics TargetScan database. Dual luciferase reporter gene assay was used to verify the targeting relationship between miR-9-5p and MEG3, SOCS5, respectively. Results: Compared with para-cancerous tissues and cervical epithelial HcerEpic cells, the expressions of MEG3 and SOCS5 were significantly down-regulated and the expression of miR-9-5p was significantly up-regulated in cervical cancer tissues and cell lines (all P<0.01). TargetScan database analysis and Dual luciferase reporter gene assay confirmed the targeting relationship between miR-9-5p and MEG3 or SOCS5. MEG3 and SOCS5 significantly inhibited while miR-9-5p significantly promoted cell proliferation, migration and invasion ability (all P<0.01). MEG3 and SOCS5 promoted E-cadherin expression and inhibited vimentin expression, while miR-9-5p inhibited E-cadherin expression and promoted vimentin expression (P<0.05 or P<0.01). Conclusion: lncRNA MEG3 regulates proliferation, migration, invasion and EMT of cervical cancer cells via miR-9-5p/SOCS5 axis.
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@# Objective： ：To investigate the expression of miR-137 in cervical cancer tissues and cells, and to explore its effect on proliferation, migration and invasion of cervical cancer cells as well as the mechanisms. Methods： ：Thirty-two pairs of cervical cancer tissues and corresponding para-cancerous tissues that surgically resected at the Department of Gynecology and Obstetrics of Dongguan People's Hospital from January 2017 to March 2018 were collected for this study. In addition, cervical cancer cell lines C33A, HeLa, SiHa and cervical epithelial immortalized cell line H8 were also collected. The expression of miR-137 in cervical cancer tissues and cell lines was detected by RT-PCR. miR-137 mimics and miR-137 NC were respectively transfected into C33Aand HeLa cells, and the effects of miR-137 over-expression on proliferation, migration and invasion of cervical cancer cell lines were observed by CCK-8 and Transwell assay. Luciferase reporter gene assay and WB were used to determine the relationship between miR-137 and Wnt5a in cervical cancer. Wnt5a over-expression vector was constructed, and the effects of simultaneous over-expression of Wnt5a and miR-137 on proliferation, migration and invasion of C33Aand HeLa cells were observed. Results： ：The expression level of miR-137 was significantly down-regulated in cervical cancer tissues and cell lines, as compared to para-cancerous tissues and H8 cells (all P<0.05). The over-expression of miR-137 significantly inhibited cell proliferation, migration and invasion of C33A and HeLa cells (all P<0.05). Moreover, Wnt5a was identified as a target of miR-137 by luciferase reporter gene assay. Furthermore, Wnt5a over-expression, to a certain degree, attenuated the suppressive effects of miR-137 on the proliferation, migration and invasion of C33A and HeLa cells. Conclusion： ：miR-137 can inhibit the proliferation, migration and invasion of cervical cancer cells via targeting Wnt5a, which may be an effective target for the treatment of cervical cancer.
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@#Objective: To construct indocyanine green-loaded silica nanoparticles (ICG@MSNs) and evaluate their killing effect on cervical cancer HeLa cells. Methods: Mesoporous silica nanoparticles (MSNs) were synthesized by template method, and indocyanine green (ICG) containing photothermal agent was loaded to prepare ICG@MSNs with photothermal effect, which were applied in the research of HeLa cells in vitro. Results: The particle of ICG@MSNs was uniform and in regular spherical shape with the size about 200 nm. ICG@MSNs was similar photothermal effect with pure ICG. Cell endocytosis experiments showed that ICG encapsulated in silica nanoparticles is more likely to be endocytosed by tumor cells, and then played a photothermal role in killing cervical cancer HeLa cells. On the other hand, cytotoxicity experiments showed that under the irradiation of 808 nm laser, ICG@MSNs significantly increased cytotoxicity, which could significantly kill cervical cancer HeLa cells. Conclusion: ICG@MSNs has good stability and biocompatibility, as well as good thermogenesis. It’s photothermal treatment effect on tumor is obvious, which has a good prospect for the treatment of cervical cancer.
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@# Objective: To investigate the effects and mechanisms of miR-129-5p on invasion, migration and epithelial-mesenchymal transition (EMT) of cervical cancer HeLa cells. Methods:Cervical cancer HeLa cells were selected. The target gene of miR-129-5p was screened by bioinformatics prediction software, and the targeting relationship between miR-129-5p and MAPK1 was verified by dual luciferase reporter gene assay. HeLa cells were transfected with miR-129-5p mimic, miR-129-5p inhibitor and pcDNA-MAPK1 alone or in combination.The expressions of miR-129-5p and MAPK1 in HeLa cells were detected by qPCR; the invasion and migration ability of HeLa cells were detected by Transwell and scratch-healing experiments, respectively; and the expressions of E-cadherin, N-cadherin, MAPK1, STAT3 and Bcl-xL were detected by WB. The subcutaneous xenograft model of HeLa cells in nude mice was constructed to observe the effect of miR-129-5p over-expression on the growth of transplanted tumors. The expressions of EMT and MAPK1 pathwayrelated proteins in transplanted tumor tissues were detected by WB. Results: miR-129-5p could bind with the 3'UTR region of MAPK1, and over-expression of miR-129-5p targetedly inhibited the expression of MAPK1 (P<0.01). Compared with the control group, the number of invasive cells in the miR-129-5p mimic group decreased (P<0.01), the scratch healing rate decreased (all P<0.01); The expression of E-cadherin was up-regulated, and the expressions of N-cadherin, MAPK1, STAT3 and Bcl-xL were down-regulated (all P< 0.01), while co-transfection of MAPK1 reversed the above phenomenon.The nude mice HeLa cell xenograft model was successfully established. Compared with the control group, the tumor mass of the miR-128-3p mimic group was reduced; the expression of E-cadherin was up-regulated in tumor tissues, while the expressions of N-cadherin, MAPK1, STAT3 and Bcl-xL were down-regulated (all P<0.01). Conclusion: Over-expression of miR-129-5p inhibits invasion, migration and epithelial-mesenchymal transition of cervical cancer HeLa cells by targeting MAPK1.
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@# Objective: To investigate the effect of lncRNA SBF2-AS1 on epithelial-mesenchymal transition (EMT) of cervical cancer HeLa cell via regulating miR-140-5p/VEGFA (vascular endothelial growth factor A) axis. Methods: After cell culture and transfection, the cells were divided into 5 groups: NC group, miR-140-5p mimic group, miR-140-5p mimic+pcDNA-VEGFA group, si-lncRNA SBF2-AS1+pcDNA-VEGFA group and si-lncRNA SBF2-AS1+miR-140-5p mimic group. The expression level of lncRNA SBF2-AS1 in cervical cancer tissues and cell lines was detected by qPCR. The targeted relationship between lncRNA SBF2-AS1, miR-140-5p and VEGFA was confirmed by Dual luciferase reporter gene assay. The expression levels of VEGFA and EMT-related proteins N-cadherin, Vimentin and E-cadherin in HeLa cells were detected by WB. The invasion and migration of HeLa cells were detected by Transwell. Results: lncRNA SBF2-AS1 was highly expressed in cervical cancer tissues and cell lines (P<0.05 or P<0.01). Dual luciferase reporter gene assay confirmed that lncRNASBF2-AS1 targetedly combined with miR-140-5p and VEGFAwas a target gene of miR-140-5p (P< 0.05). Knockdown of lncRNA SBF2-AS1 inhibited invasion and migration as well as EMT of HeLa cells. Further experiment confirmed that lncRNA SBF2-AS1 up-regulated the expression level of VEGFA via miR-140-5p, thereby promoting invasion, migration and EMT of HeLa cells. Conclusion: lncRNASBF2-AS1 promotes EMT of HeLa cells via miR-140-5p/VEGFAaxis.
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@# Objective: To investigate whether miR-140 could increase the sensitivity of cervical cancer (CC) to oxaliplatin by downregulating the expression of programmed death-1 (PD-L1). Methods: qPCR was used to analyze miR-140 expression in normal human cervical cells, CC cells and oxaliplatin-resistant CC cells. Cells were transfected with miR-140 mimic, and then, the proliferation of CC cells and oxaliplatin-resistant CC cells was detected by using CCK-8 assay, and the colony formation rate of CC cells was obtained by using colony formation assay. Starbase and TargetScan were used to predict the targeted binding site of miR-140 and PD-L1, and the influence of miR-140 on the expression of PD-L1 was validated by dual luciferase reporter gene assay.Annexin V FITC/PI double staining and Wb assays were used to detect the effect of over-expression of miR-140 or both over-expression of PD-L1 and miR140 on the apoptosis, migration and expression of apoptosis-related proteins in CC cells after treatment with oxaliplatin. Moreover, transplantation tumor of CC cell lines was established in nude mice to assess the effects of miR-140 on enhancing the sensitivity of tumors to oxaliplatin. Results: The expression of miR-140 was significantly decreased in oxaliplatin-resistant CC cells (P<0.01). Over-expression of miR140 could significantly increase the sensitivity of oxaliplatin-resistant CC cells to oxaliplatin (P<0.05), and inhibit the CC cells proliferation and colony formation (P<0.01). miR-140 showed targeted binding to PD-L1 3'-UTR and inhibited its expression. Over-expression of miR-140 significantly promoted CC cell migration and apoptosis (P<0.01). However, co-transfection of PD-L1 counteracts the effects of miR-140 on cell metastasis and apoptosis (all P<0.05). In addition, xenograft tumor model in mice also verified that miR-140 could promote the sensitivity of tumors to oxaliplatin. Conclusion: miR-140 increases the sensitivity of CC to oxaliplatin through inhibition of PD-L1 expression. Therefore, up-regulation of miR-140 or down-regulation of PD-L1 in combination with oxaliplatin may be a novel strategy for the treatment of Oxaliplatin-resistant CC.
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OBJECTIVES: Pseudolaric acid B (PAB) has been shown to inhibit the growth of various tumor cells, but the molecular details of its function are still unknown. This study investigated the molecular mechanisms by which PAB induces apoptosis in HeLa cells. METHODS: The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assays were performed to investigate the effect of PAB treatment in various cervical cancer cell lines. Annexin V/propidium iodide staining combined with flow cytometry and Hoechst 33258 staining were used to assess PAB-induced apoptosis. Additionally, we performed bioinformatics analyses and identified a paired box 2 (PAX2) binding site on the BAX promoter. We then validated the binding using luciferase and chromatin immunoprecipitation assays. Finally, western blotting assays were used to investigate PAB effect on the Wnt signaling and the involved signaling molecules. RESULTS: PAB promotes apoptosis and downregulates PAX2 expression in HeLa cells in a time- and concentration-dependent manner. PAX2 binds to the promoter of BAX and inhibits its expression; therefore, PAX2 inhibition is associated with increased levels of BAX, which induces apoptosis of HeLa cells via the mitochondrial pathway. Additionally, PAB inhibits classical Wnt signaling. CONCLUSION: PAB effectively inhibits Wnt signaling and PAX2 expression, and increases BAX levels, which induce apoptosis in HeLa cells. Therefore, PAB is a promising natural molecule for the treatment of cervical cancer.
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Humans , Apoptosis , Binding Sites , Bisbenzimidazole , Blotting, Western , Cell Line , Chromatin Immunoprecipitation , Computational Biology , Flow Cytometry , HeLa Cells , Luciferases , Mitochondria , Uterine Cervical Neoplasms , Wnt Signaling PathwayABSTRACT
OBJECTIVE: To investigate the effect of genistein on the anticancer effects of chemotherapeutic agents, we examined the effect of a genistein and cisplatin combination on CaSki human cervical cancer cells. METHODS: After the cervical cancer cells (HeLa cells, CaSki cells) had been cultured, cisplatin and genistein were added to the culture medium, and the cell activity was measured using MTT assay. The CaSki cells were cultured in a medium containing cisplatin and genistein, and then, the cells were collected in order to measure p53, Bcl2, ERK, and caspase 3 levels by western blotting. RESULTS: Both the HeLa and CaSki cells had decreased cell viabilities when the cisplatin concentration was 10 μM or higher. When combined with genistein, the cell viabilities of the HeLa and CaSki cells decreased at cisplatin concentrations of 8 μM and 6 μM, respectively. The administration of genistein increased the toxicity of cisplatin in the HeLa and CaSki cells. In the CaSki cells, the p-ERK1/2 level decreased by 37%, the p53 expression level increased by 304%, and the cleaved caspase 3 level increased by 115% in the cisplatin+genistein group compared to that in the cisplatin group. Bcl2 expression was reduced by 69% in the cisplatin+genistein group compared to that in the cisplatin group. CONCLUSION: Genistein enhances the anticancer effect of cisplatin in CaSki cells, and can be used as a chemotherapeutic adjuvant to increase the activity of a chemotherapeutic agent.
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Humans , Blotting, Western , Caspase 3 , Cell Line , Cell Survival , Cisplatin , Genistein , HeLa Cells , Uterine Cervical NeoplasmsABSTRACT
Objective To observe the effect of different fixative solutions on cancer cell morphology and membrane permeability. Methods Human pancreatic acinar epithelial carcinona(HPAC) cells of human pancreatic cancer and HeLa cells of human cervical cancer were fixed with 4 fixation solutions: Freshly prepared 0.25% paraformaldehyde solution; Freshly prepared 4% paraformaldehyde solution; 75% ethanol solution; 90% ethanol solution. The fixation lime is 30 minutes. PBS solution and complete medium were used as the controls. Cell morphology of each group was observed under optical microscope. Changes in cell jncmbrane permeability were observed by fluorescence staining with 7-aminoactinomycin (7-AAD) , which is not cell membrane permeable in intact cells but permeable in damaged cells. Hoechst33342 was used for staining both intact and damaged cells. Results The cells in the complete medium group were similar to unfixed cells in morphology, and the fluorescence staining of 7-AAD was the weakest. The cells in the complete medium group have typical eel! morphology and low 7-AAD permeability. The 0.25% paraformaldehyde solution group had similar cell morphology to the complete medium group, and the 7-AAD fluorescence staining was weak. The morphology of cells in the 4% paraformaldehyde solution group was typical, but the fluorescence staining of 7-AAD was strong. The cells in the 90% ethanol solution group showed swelling, with a larger volume than the unfixed cells and a stronger fluorescence staining of 7-AAD. The cell swelling in 75% ethanol solution group was not as obvious as that in 90% ethanol solution group, and the fluorescence staining of 7-AAD was strong. The cells in PBS group were round, and the fluorescence staining of 7-AAD was strong. Conclusion 0. 25% paraformaldehyde solution can not only fix tumor cells, but also maintain the integrity of cell membrane.
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Objective: To investigate the effect of carmofur on the proliferation and invasion of cervical cancer cell line HeLa and to study its mechanism. Methods: Carmofur was prepared into different concentrations (0.4 mg/L, 0.8 mg/L, and 1.2 mg/L) and the control group was set. Cell counting kit-8(CCK-8) method was used to detect the viability of the cells. Transwell technique was used to detect cell invasion. The expression levels of matrix metalloproteinase(MMP) -7, β-catenin and P120 catenin(P120 ctn) and mRNA were detected by Western blotting and Real-time PCR. Results: Compared with the control group, the cell proliferation ability decreased with the increase of the concentration of carmofur in the drug treatment group, and the cell proliferation was inhibited after 24 hours of carmofur exposure, and the cell proliferation rate decreased significantly with the prolongation of the action time (all P<0.05). Compared with the control group, carmofur group inhibited the invasion of HeLa cells (P<0.05). Compared with the control group, the carmofur group decreased MMP-7, β-catenin and PI20 catenin levels and mRNA levels (P<0.05). Conclusion: HCFU can inhibit the proliferation of HeLa cells, and its mechanism might be related to the down-regulation of MMP-7, β-catenin and P120 catenin expression levels.
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This study aimed to investigate the possible sensitivity of Astragalus polysaccharides, in order to improve the chemosensitivity of cervical cancer HeLa cells to cisplatin by regulating the cell autophagy, and explore its possible mechanism. In this study, HeLa cells were divided into control group, cisplatin group, Astragalus polysaccharide group, and Astragalus polysaccharide combined with cisplatin group. MTT assay was used to detect the proliferation of cervical cancer HeLa cells. Flow cytometry was used to detect the apoptosis and cycle of HeLa cells in each experimental group. RT-PCR was used to detect the mRNA expression of autophagy-related proteins beclin1, LC3Ⅱ and p62. The expression levels of autophagy-related proteins beclin1, LC3Ⅱ, LC3Ⅰ and p62 were detected by WB method. MTT results showed that compared with the control group, the proliferation of HeLa cells was significantly inhibited in each administration group(<0.05), and the inhibitory effect of the combination group was more significant(<0.01). The apoptotic rate of HeLa cells was significantly increased(<0.05), and the apoptotic rate of the combination group was significantly increased(<0.01) compared with the control group(<0.05).In conclusion, G₀/G₁ phase showed the most significant differences between the two groups. RT-PCR and WB results showed that the gene and protein expressions of beclin1 and LC3Ⅱ were up-regulated, while the gene and protein expressions of p62 were down-regulated compared with the control group. The above-mentioned changes in the combination group were more significant. Through the analysis of the above experimental results, it is speculated that Astragalus polysaccharides may increase the sensitivity of cervical cancer HeLa cells to cisplatin by regulating the cell autophagy. Its possible mechanism of action is correlated with the up-regulation of autophagy-related proteins beclin1, the promote the conversion from LC3Ⅰ to LC3Ⅱ, the down-regulation of labeled protein p62, and the enhancement of HeLa cell autophagic activity, thereby increasing the sensitivity of HeLa cells to cisplatin chemotherapy.
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Humans , Apoptosis , Astragalus Plant , Chemistry , Autophagy , Cell Cycle , Cisplatin , Pharmacology , Drug Resistance, Neoplasm , HeLa Cells , Microtubule-Associated Proteins , Metabolism , Polysaccharides , PharmacologyABSTRACT
Objective To observe the effect of apatinib (APA) combined with radiotherapy on cell cycle and apoptosis of cervical cancer HeLa cells in vitro. Methods HeLa cells in logarithmic growth phase were divided into control group, drug group, radiotherapy group and joint group. Cell cycle and apoptosis were detected by flow cytometry, and the changes of HeLa cell cycle and apoptosis after radiotherapy were analyzed. Results Retardant of G0/G1 phase for joint group was obviously higher than control and radiotherapy group (P ＜ 0. 05). S phase percentage of joint group was minimum when compared with control group (P ＜0. 05), while there was no significant difference when compared with radiotherapy group. Apoptosis rate of joint group was higher than control group and drug group (P ＜ 0. 05 ). Conclusion APA combined with radiotherapy shows significant G0/G1 phase cell cycle arrest, but no significant induction of apoptosis.
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Objective:To establish a paclitaxel (PTX)-resistance cell line of human cervical carcinoma Hela/PTX and investigate its biological features. Methods:Using Hela as the negative cells,drug-resistant cervical cancer Hela/PTX cells were applied as the target cells. The morphology changes were observed under an optical microscope;the drug resistance of Hela/PTX to different chemotherapeutic drugs was detected by CCK-8 method. The accumulation of drugs in the cells was determined by HPLC. The expression of P-glycoprotein (P-gp), Survivin protein and mRNA was detected by Western blot and RT-PCR. Results:Hela/PTX multidrug resistant cell line was successfully established. The resistance index to PTX was 58.40 and the resistance index of cross-resistance to docetaxel and 5-fluorouracil was 8.11 and 4.62,respectively. The mRNA expression level of P-gp and Survivin protein in Hela/PTX cells respectively was(9.93 +0.90) times(P < 0.05) and(3.02 +0.57) times(P <0.05) of that in Hela cells. The expression of P-gp and Survivin protein in drug-resistant cells was significantly higher than that in parental cells.Conclusion:Human cervical cancer resistant cell line Hela/PTX with the basic biological characteristics of multidrug resistance is successfully established, which can be used as an in vitro model for the studies on paclitaxel resistance mechanism and provide experimental basis for the studies of multidrug resistance mechanism of cervical cancer.
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OBJECTIVE:To study induction effect of euphornin on the apoptosis of cervical cancer Hela cells and its mechanism. METHODS:The cervical cancer Hela cells were divided into blank control group,cisplatin group(positive control, 10 mg/L) and euphornin low-dose,medium-dose and high-dose groups (50,100,200 mg/L). They were treated with relevant medicine. The inhibitory effect of Hela cells proliferation was tested by MTT assay after 24,48,72 h of medicine treatment. The apoptotic rate of Hela cells was measured by flow cytometry after 48 h of medicine treatment. Morphology of nucleus was detected by Hoechst 33258 staining. The protein expression of Cyt-C,Bcl-2,Bax,Caspase-3,Caspase-8,Caspase-9 and Caspase-10 were detected by Western blot assay. RESULTS:Compared with blank control group,inhibitory rate of cell proliferation and cell apoptosis rate were increased significantly in cisplatin group and euphornin groups(P＜0.05 or P＜0.01),and obvious staining, deformation,shrinking,fragmentation or apoptotic bodies was found in nucleus. Compared with blank control group,the protein expression levels of Cyt-C,Caspase-8 and Caspase-9 in euphornin low-dose,medium-dose and high-dose groups were increased significantly,while the protein expression level of Bcl-2 and Bcl-2/Bax ratio were decreased significantly(P＜0.05 or P＜0.01);the protein expression levels of Bax,Caspase-3 and Caspase-10 in euphornin medium-dose and high-dose groups were increased significantly(P＜0.05 or P＜0.01). CONCLUSIONS:Euphornin can significantly inhibit the proliferation of Hela cell and promote cell apoptosis,the effect of which will be achieved by activating the Caspase-dependent mitochondrion apoptosis pathway.
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@# Objective: To investigate the expression of HOPX gene in cervical cancer tissues and blood serum as well as its effect on cervical cancer HeLa cells, and to analyze its correlation to tumor maker CEAand CA125. Methods: 50 pairs of cervical cancer tissues and para-cancerous tissues as well as the peripheral blood samples from patients with cervical cancer, who were treated at Tianjin Binhai People’s Hospital and Tianjin Wuqing People’s Hospital from June 2015 to December 2017, were collected for this study; in addition, 50 samples of blood serum from healthy people were used as control. Real-time quantitative PCR (qRT-PCR) and immumohistochemical staining (IHC) were used to detect mRNA and protein expressions of HOPX in tissue and serum samples, NCBI-GEO data base and TCGA data base were used to collect the information on HOPX gene and patients’prognosis, and the correlation between HOPX expression and patients’prognosis was analyzed. Vectors over-expressing HOPX or control vectors were transfected into HeLa cells; MTT assay and colony formation assay were used to examine the proliferation ability of HeLa cells, Tranwell assay was used to detect the migration and invasion of HeLa cells, and Western blotting was used to detect the expression of EMT-related proteins. Results: Both sample examination and data base information showed that the expression level of HOPX was down-regulated in tissue and serum samples of cervical cancer patients and was positively related with the survival of patients (r=0.736， P<0.05); while it’s expression was negatively related to the level of CEAand CA125 in cervical cancer tissues and serum (r=-0.678, P<0.05). HOPX over-expression inhibited cell proliferation, migration and invasion, promoted the expression of E-cadherin but inhibited the expression of Vimentin and ICAM1 (all P<0.05 or P<0.001). Conclusion: HOPX is low expressed in cervical cancer tissues and blood samples, and negatively correlated with CEA and CA125, but positively correlated with the survival of patients. Thus, combination of HOPX and CEA/CA125 may improve the early diagnosis rate of cervical cancer and provide a new strategy for precision treatment of cervical cancer in future.