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Objective:To determine mechanisms for the role of high mobility group box-1 (HMGB1) to platinum based chemo-sensitivity in cervical cancer.Methods:Using Western blot,we examined the effect of cisplatin on expressions of LC3,Beclin1 and P62 in cervical cancer.After treated with autophagy inhibitor and/or cisplatin in HeLa and CaSki cells,cell counting kit-8 (CCK-8) assay was used to evaluate the chemo-sensitivity.The expression of LC3,Beclin1 and P62 were detected by Western blot.We established HeLashHMGB1,CaSki-shHMGB1 and HeLa-CTR,CaSki-CTR stable cell lines.CCK-8 assay was used to determine the chemosensitivity in these cell lines.Using Western blot,we examined the correlation among the expressions of HMGB1,LC3,Beclin1,and P62.Results:With the increase of cisplatin,the expression of LC3 and Beclin1 was increased,while the expression of P62 was decreased in HeLa and CaSki cell lines.Combination of cisplatin with autophagy inhibitor could increase the cellular sensitivity to cisplatin (P＜0.05).The expression of HMGB1 was related to chemo-sensitivity and autophagy related protein in cervical cancer cells.HMGB1 was positively correlated with LC3 and Beclin1 while negatively correlated with P62 (P＜0.05).Conclusion:HMGB 1 might play an important role in chemo-sensitivity via regulating autophagic activity.The combination of cisplatin with autophagy inhibitor might become a possible strategy for cervical cancer.HMGB1 might serve as a potential biomarker for predicting chemo-therapeutic responses.
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Objective: To investigate the chemo-sensitivity enhancing effect and its major active mechanism of Shengmai Injection (SMI). Methods: MTT assay was used to determine the chemo-sensitivity enhancing activity of SMI combined with various current- used chemotherapeutic drugs. Flow cytometry was conducted to determine the effect of Epirubicin combined with SMI on the cell cycle distribution and apoptotic cell death, and the mRNA expression level of MDR-1 gene was determined by RT-PCR. Results: SMI (30 μL/mL) combined with gemcitabine, cisplatin, paclitaxel, and Epirubicin exihibited the potent antiproliferative effects on human lung carcinoma A549, gastric carcinoma SGC-7901, breast carcinoma MCF-7, and hepatocellular carcinoma HepG-2 cell lines, respectively. SMI showed the most significant enhancing effects on HepG-2 cells in combination with Epirubicin, and the sensitivity of HepG-2 to Epirubicin was increased by 16 fold. Whereas SMI (30 μL/mL) alone did not affect the cell cycle distribution and apoptosis of HepG-2 cells, it enhanced the cellular responses when combined with Epirubincin together with a down-regulated mRNA level of MDR-1 gene. Conclusion: SMI combined with the different chemotherapeutic drugs could enhance the sensitivity of cancer cells significantly via down-regulating the mRNA expression level of MDR-1 and the expression of P-glucoprotein with the excretion of drugs.
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The effects of synthetic Smac peptide (SmacN7) on chemotherapeutic sensitivity of bladder cancer cells were investigated. SmacN7 penetratin peptide was synthesized and delivered into T24 cells. MTT assay was used to evaluate the viability of T24 cells induced by low-dosage of MMC. Flow cytometry was used to analyze the proportions of apoptosis. Western blot was used to detect the expression of XIAP and Caspase-3. The activity of Caspase-3 was measured and the effect of SmacN7 combined with MMC on T24 cell lines was also determined. The results showed that SmacN7 penetratin peptide could successfully interact with endogenous XIAP, increase the proportions of apoptosis of T24 cell lines induced by low-dosage of MMC in a dose-and time-dependent manner. An obvious down-regulation of XIAP expression and up-regulation of Caspase-3 was identified by Western blot. The activity of Caspase-3 in experimental group was significantly increased as compared with that in the control group. As compared with MMC group, the viability of T24 cells in SmacN7 penetratin peptide + MMC group was markedly decreased to 2.22 and 3.61 folds at 24h and 48h respectively. It was concluded that SmacN7 penetratin peptide could act as a cell-permeable IAP inhibitor, inhibit the proliferation, induce apoptosis and enhance the chemo-sensitivity of bladder cancer cells to MMC. These findings indicate that SmacN7 penetratin peptide may be a very promising ageut for bladder cancer treatment when used in combination with chemotherapy.
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Background and purpose:Smac/DIABLO was the only apoptosis-related protein that could inhibit IAPs directly and simultaneously.The four amino-residual AVPI(Ala-Val-Pro-lie)in its N-terminal was the very important domain that could stimulate apoptosis.This study investigated the effect of synthetic Smac peptide (SmacN7) on chemotherapy sensitivity of bladder cancer cells.Methods:SmacN7 penetratin peptide was synthesized and delivered into T24 cells.MTT assay was adopted to evaluate the viability of T24 cells induced by low-dosage of MMC. Flow cytometry was applied to analyze the proportion of apoptosis and Western blot was used to detect the expression of XIAP and caspase-3;The activity of caspase-3 was measured and the effect of SmacN7 combined with MMC on T24 cell lines was also determined.Results:SmacN7 penetratin peptide could successfully interact with endogenous XIAP and increase the proportions of apoptosis of T24 cell lines induced by low-dosage of MMC in a dose-and time- dependent manner.An obvious down-regulation of XIAP expression and up-regulation of caspase-3 was identified by Western blot.The activity of caspase-3 in experimental group was significantly increased as compared with that in the control group;Combining the treatment with SmacN7 penetratin peptide,the viability of T24 cells decreased to 55% and 72.7% in 24 hrs and 48 hrs respectively.Conclusion:SmacN7 penetratin peptide could act as a cell-permeable IAP inhibitor,inhibit the proliferation,induce apoptosis and enhance the chemo-sensitivity of bladder cancer cells to MMC. When combined with chemotherapy,it may be a very promising strategy for bladder cancer therapy.
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Objective:To study the enhancing effect of bFGF-targeted antisense phosphorothioate oligonucleotide (APO)on the chemosensitivity of human laryngeal squamous carcinoma cell line Hep2 to Doxorubicin,5-Fluorouracil, and Cisplatin.Methods:bFGF-specific APO was designed,constructed and transfected into Hep2 cells with jetPEI (polyethyleneimine).Expression of bFGF mRNA was evaluated by semi-quantitative RT-PCR after transfection;immuno- cytochemical method was used to examine the expression of bEGF expression before and after transfection of Hep2;the in- duction of cell apoptosis was analyzed by flow cytometry;cell proliferation was then analYzed by MTT assay after treatment with bFGF-specific APO or chemotherapeutic drugs,or a combination of both.Results:bFGF-specific APO inhibited the growth of Hep2 cells in a dose-and time-dependent manner,with the peak inhibitory rate being 25.5%.The expression of bFGF mRNA and protein decreased by 52.0% and 41.1%,respectively.The apoptosis rate of Hep2 cells was 20.5% after transfection,bFGF-specifie APO reduced the 50% inhibitory concentration of Doxorubicin,5-Fluorouracil,and Cis- platin in Hep2 cells by 75.5%,83.5% and 65.4%,respectively.Conclusion:bFGF-specific APO can enhance the chemosensitivity of Hep2 cells,which paves a new way for potential biologic chemotherapy of laryngeal squamous carcino- ma.
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Objective:To observe the sensitivities of breast cancer cells and its implanted tumors to chemotherapeutic drug epirubicin after down-regulation of HER-2 expression by RNA interference(RNAi).Methods:HER-2siRNApU6 vector containing HER-2 RNAi was constructed and was used to transfect HER-2 positive breast cancer cell line SKBR-3.The expression of HER-2 mRNA and protein were analyzed by RT-PCR and Western blotting,respectively.Transfected SKBR-3 cells were treated with different concentrations of epirubicin;the growth of SKBR-3 cells and IC50 of epirubicin were observed by MTT.SKBR-3 cells were injected into nude mouse to establish breast cancer model;the sensitivity of mouse model to epirubicin was observed after HER-2shRNApU6 treatment.Results:Expression of HER-2 mRNA and HER-2 protein were down-regulated in SKBR-3 cells after transfection with HER-2shRNApU6.Furthermore,the proliferation of SKBR-3 cells transfected with HER-2shRNApU6 was significantly decreased compared with mock tansfected group(P