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Objective:To investigate the impact of connexin 43(CX43) on the connection of S24 glioblastoma multiforme (S24-GBM) cellular network and to explore its role on radio-resistance.Methods:Specific lentiviral vectors were used to knockout CX43 in S24-GBM stem cells (S24-GBMSCs). Alternatively, carbenoxolone (CBX) was used to block transmission of CX43. Subsequently, the animal subjects grafted with S24-GBMSCs were monitored under a multiphoton laser scanning microscope (MPLSM). Dynamic changes of tumor microtubes (TMs) and transmission of Ca 2+ and SR101 in the cellular network were recorded. To study the radiosenstivity of S24-GBM before and after CX43 inhibition, MRI scanning of the brains was taken before and after radiation to assess the tumor sizes. Survival time of each subject was also recorded. Results:In comparison with control group, knockout of CX43 in S24-GBMSCs led to shorter TMs, less TM connected cells, lower Ca 2+ synchronicity and SR101 fluorescence, as well as decreased tumor sizes and prolonged survival time (all P<0.01), which were independent from radiation. However, CBX only demonstrated inhibition on the growth of tumors and the diffussion of Ca 2+ and SR101, without affecting TMs formation. These above-mentioned alterations could be enhanced by the combination of gap43 knockout in S24-GBMSCs with blockage of CX43 by CBX (all P<0.05). Conclusion:CX43 plays a critical role in the radioresistance of S24-GBM by influencing the formation of S24-GBM cellular network and the transmission of important signaling molecules including Ca 2+ and SR101.
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Objective To exPlore the effect of mammalian target of raPamycin ( mTOR ) inhibitor eVerolimus on radiosensitiVity of human non_small cell lung cancer cell line in vitro by using eVerolimus to inhibit mTOR signaling Pathway of A549. Methods Human non_small cell lung cancer cell line A549 was subjected to radiation alone or in combination with eVerolimus treatment. The 50%inhibition concentration ( IC50 ) of eVerolimus in A549 cells was detected by methylthiazol tetrazolium ( MTT) assay in vitro. EVerolimus at the 20%inhibition concentration ( IC20 ) was used to Pretreat A549 cells for 24 h. Cells were then irradiated by X_ray with 2,4,6,8 Gy. The cell surViVal fraction was comPuted by clone formation. Cell surViVal curVe was fitted by multitarget one_hit model, and mean lethal dose ( D0 ), dose quasithreshold ( Dq ), surViVal fraction at 2 Gy ( SF2 ), and sensitization enhancement ratio (SER) were calculated. The exPression ofγ_H2AX was determined by Western blotting and then the relatiVe gray Values were analyzed. Results EVerolimus significantly imProVed the sensitiVity of A549 cells to radiation. The D0 , Dq and SF2 of eVerolimus+irradiation grouP were significantly lower than those of irradiation grouP. The SER was 1. 36. The residual amount of γ_H2AX Protein in the eVerolimus + irradiation grouP was significantly higher than that of the irradiation grouP. Conclusion EVerolimus inhibiting mTOR signaling Pathway can increase the radiosensitiVity of A549 cells.
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Background and purpose:Hypoxia induced the decreased radiosensitivity of tumor cells, which was the cause of tumor radioresistance and relapse and metastasis. During the course, HIF-1a played the most important role in the regulation of hypoxia. However, it’s still unknown about the effect of HIF-1a on the radiosensitivity of hypoxia tumor cells and the relationship with autophagy. This study was to inhibit HIF-1αexpression in hypoxic lung adenocarcinoma cell line A549 with RNA interference (RNAi), and explore its effect on hypoxic cell radiosensitivity and autophagy. Methods: Plasmids pHIF-1α-shRNA and Neg-shRNA (negative control) were constructed and transfected into hypoxic A549 cells, this positive clone was named A549/HIF-1α-shRNA. Clone formation array was applied to calculate the value of D0, SF2, SER. The expression of HIF-1α, LC3, c-parp was detected by Western blot. Results:The SF2 of hypoxic A549 cell was 0.62, which was higher than that of normoxic A549 cell, SER was 1.45. The level of LC3Ⅱincreased significantly and the level of c-parp decreased after the radiation of hypoxic A549 cell. The level of HIF-1a increased in hypoxic A549 cells. The expression of HIF-1αin hypoxic A549 cells was suppressed markedly after transfection of HIF-1α-shRNA;this clone was named A549/HIF-1α-shRNA. The SF2 and SER were significantly lower in A549/HIF-1α-shRNA cells, 0.45 and 0.72 respectively. Under the hypoxic condition and after the inhibition of HIF-1α, the expression of LC3Ⅱ decreased significantly and the expression of c-parp increased. Conclusion:We successfully established a cell model that HIF-1αexpression was suppressed almost completely by RNAi. The inhibition of HIF-1αby shRNA may raise the radiosensitivity and decrease the autophagy of hypoxic A549 cells in vitro.
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This study examined the radiation-induced ERBB2 nuclear transport in the BT474 breast cancer cell line and the relationship between caveolin-1 and radiation-induced ERBB2 nuclear transport. The BT474 cells were treated with herceptin (200 nmol/L), PP2 (a caveolin-1 inhibitor, 100 nmol/L) and irradiation combined or alone. Confocal microscopy was used to observe the nuclear import of ERBB2 and caveolin-1 after irradiation. Western blotting was employed to detect the expression of ERBB2, caveolin-1 and DNA-PKcs after irradiation, and immunoprecipitation to identify the ERBB2 and caveolin-1 complex before perinuclear ERBB2 localization. Confocal microscopy showed the transport of ERBB2 and caveolin-1 from the cell membrane to the nucleus 15 min after irradiation and the proteins accumulated at the perinuclear region within 45 min. Western blotting revealed that the expression levels of ERBB2, caveolin-1 and DNA-PKcs were increased after irradiation and reached a peak 45 min later. Both herceptin and PP2 treatments were found to decrease ERBB2 expression. An immune complex composed of ERBB2 and caveolin-1 was found in the herceptin group after irradiation. It was concluded that after irradiation, ERBB2 may be transported from the cell membrane to the nucleus and activate DNA-PKcs to trigger DNA double-strand break (DSB) repair; caveolin-1 may participate in this process. Treatments involving the downregulation of caveolin-1 may increase the radiosensitization of breast cancer cells.
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Chemotherapy-induced neuropathy is a serious clinical problem for patients receiving cancer treatment. The aim of this study was to investigate the potential efficacy of pregabalin in chemotherapy-induced neuropathy in rats. A total of 35 male Sprague-Dawley rats were randomly divided into 5 groups: group 1, naive control; group 2, treated with pregabalin (30 mg/kg p.o., for 8 days); group 3, docetaxel was given by single intravenous infusion at 10 mg/kg; groups 4 and 5, pregabalin at 10 mg/kg and 30 mg/kg respectively was orally administered for 8 days after the docetaxel treatment. On day 8, behavioral test was performed, and substance P and CGRP release in dorsal root ganglion (DRG) and sciatic nerve were analyzed by electron microscope. Our results showed that docetaxel induced mechanical allodynia, mechanical hyperalgesia, heat hypoalgesia, cold allodynia, and sciatic nerve impairment and substance P and CGRP release in DRG. However, oral administration of pregabalin (10 mg/kg and 30 mg/kg) for 8 consecutive days significantly attenuated docetaxel-induced neuropathy by ameliorating heat hypoalgesia, cold allodynia, impairment of sciatic nerve and reducing the release of substance P and CGRP. The findings in the present study reveal that pregabalin may be a potential treatment agent against chemotherapy-induced neuropathy.
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This study examined the radiation-induced ERBB2 nuclear transport in the BT474 breast cancer cell line and the relationship between caveolin-1 and radiation-induced ERBB2 nuclear transport. The BT474 cells were treated with herceptin (200 nmol/L), PP2 (a caveolin-1 inhibitor, 100 nmol/L) and irradiation combined or alone. Confocal microscopy was used to observe the nuclear import of ERBB2 and caveolin-1 after irradiation. Western blotting was employed to detect the expression of ERBB2, caveolin-1 and DNA-PKcs after irradiation, and immunoprecipitation to identify the ERBB2 and caveolin-1 complex before perinuclear ERBB2 localization. Confocal microscopy showed the transport of ERBB2 and caveolin-1 from the cell membrane to the nucleus 15 min after irradiation and the proteins accumulated at the perinuclear region within 45 min. Western blotting revealed that the expression levels of ERBB2, caveolin-1 and DNA-PKcs were increased after irradiation and reached a peak 45 min later. Both herceptin and PP2 treatments were found to decrease ERBB2 expression. An immune complex composed of ERBB2 and caveolin-1 was found in the herceptin group after irradiation. It was concluded that after irradiation, ERBB2 may be transported from the cell membrane to the nucleus and activate DNA-PKcs to trigger DNA double-strand break (DSB) repair; caveolin-1 may participate in this process. Treatments involving the downregulation of caveolin-1 may increase the radiosensitization of breast cancer cells.
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Female , Humans , Active Transport, Cell Nucleus , Physiology , Breast Neoplasms , Metabolism , Caveolin 1 , Metabolism , Cell Line, Tumor , Protein Transport , Physiology , Radiation , Receptor, ErbB-2 , MetabolismABSTRACT
Objective To investigate factors influencing the quality of life (QOL) of cancer patients with bone metastases. Methods Eighty-two cancer patients with hone metastasis were investigated.A questionnaire designed according to European Organization for Treatment of Cancer quality of life questionnaire ( EORTC QLQ-C30),Monroe Dunaway Anderson symptom inventory-Chinese edition (MADSI-C) and hospital anxiety and depression scale (HADS) standards was used to collect the information. Results The average total standardized QOL score of these cancer patients was 53.28±19.20.Among the function subscales,social function got the lowest average score (47.54),while among the symptom subscales fatigue got the highest average score (56.65).According to the MDASI-C,the most serious symptom burdens were fatigue,distress and pain; working and walking experienced the most interference.The symptom burdens correlated significantly with the QOL results.Twenty-four of the patients (29.3％) had been diagnosed with anxiety,and 17 (20.7％) were diagnosed with depression.Anxiety and depression continued to be significantly associated with overall QOL and its various dimensions. Conclusions The results show that the burden of fatigue and pain,as well as of anxiety and depression are significantly associated with impaired QOL among cancer patients with bone metastasis.Work (housework) and walking were the most severely affected activities.Psychological rehabilitation should be focused on the comprehensive treatment of patients with bone metastasis along with other appropriate rehabilitation strategies to enhance their overall functioning,relieve their symptoms and improve their QOL.
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Chemotherapy-induced neuropathy is a serious clinical problem for patients receiving cancer treatment. The aim of this study was to investigate the potential efficacy of pregabalin in chemotherapy-induced neuropathy in rats. A total of 35 male Sprague-Dawley rats were randomly divided into 5 groups: group 1, naive control; group 2, treated with pregabalin (30 mg/kg p.o., for 8 days); group 3, docetaxel was given by single intravenous infusion at 10 mg/kg; groups 4 and 5, pregabalin at 10 mg/kg and 30 mg/kg respectively was orally administered for 8 days after the docetaxel treatment. On day 8, behavioral test was performed, and substance P and CGRP release in dorsal root ganglion (DRG) and sciatic nerve were analyzed by electron microscope. Our results showed that docetaxel induced mechanical allodynia, mechanical hyperalgesia, heat hypoalgesia, cold allodynia, and sciatic nerve impairment and substance P and CGRP release in DRG. However, oral administration of pregabalin (10 mg/kg and 30 mg/kg) for 8 consecutive days significantly attenuated docetaxel-induced neuropathy by ameliorating heat hypoalgesia, cold allodynia, impairment of sciatic nerve and reducing the release of substance P and CGRP. The findings in the present study reveal that pregabalin may be a potential treatment agent against chemotherapy-induced neuropathy.
Subject(s)
Animals , Male , Rats , Ganglia, Spinal , Nervous System Diseases , Drug Therapy , Pregabalin , Rats, Sprague-Dawley , Sciatic Nerve , Taxoids , gamma-Aminobutyric Acid , PharmacologyABSTRACT
Objective To observe the influence of trastuzumab on DNA break repair and Her-2 nuclear import after radiation in breast cancer cell line SKBR3,and discuss the radiosensitivity mechanism of trastuzumab.Methods Clone formation assay was used to analyze the difference of survival fractions between radiation group and radiation plus trastuzumab group.Confocal microscopy was applied to observe the influence of trastuzumab in the nuclear import process of Her-2 and the expression of γH2AX after radiation,which is considered as the marker of DNA double strand break.Western blotting was used to detect the expression of Her-2 and DNA-PKcs in nuclei after radiation.Results The result of clone formation assayshowed that the SF2 in radiation group was 0.547±0.046 and 0.321±0.022 in the radiation plus trastuzumab group were significantly decreased,the results of confocal microscopy showed that trastuzumab postponed the nuclear import process of Her-2 (52.80±19.74 in radiation group,21.41±10.55 in the radiation group),and increased expression of γH2AX after radiation (85.40±25.63 in radiation group,18.53±44.32 in the radiation group),and western blotting revealed trastuzumab reduced the expression of Her-2,DNA-PKcs in nuclei.Conclusion Trastuzumab can inhibit the radiation induced nuclear import of Her-2,and decrease Her-2,DNA-PKcs in nuclei to increase the DSB on early stage after radiation.
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Objective To investigate the effect of histone deacetylases-1(HDAC-1)siRNA on the HDAC-1 protein expression,cell growth and apoptosis in HeLa cells.Methods The HDAC-1 protein was knocked down by HDAC-1 siRNA.HDAC-1 protein was detected by Western blot.The cell growth inhibition was assesses by MTT and clone forming assay.Apoptosis was measured by flow cytometry.Results Forty-eight h after transfection of HDAC-1 siRNA,HDAC-1 protein expression in HeLa cells was down-regulated obviously.The down-regulation of HDAC-1 significantly reduced the colony formation rate,inhibited cell proliferation and induced apoptosis in HeLa cells.Conclusion HDAC-1 siRNA may play an anti-proliferation role in HeLa cells by inducing apoptosis.
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This study examined the role of PI3K/Akt pathway in radiosensitization of DNA damage of cervical carcinoma cells. The 50% inhibition concentration (IC(50)) of cisplatin and docetaxel in HeLa cells was detected by Mono-nuclear cell direct cytotoxicity assay (MTT) in vitro. HeLa cells were treated by cisplatin/docetaxel of 10 percent of IC(20) alone or combined with LY294002 for 24 h, and then radiated by different doses of X-ray. The cell survival ratio was obtained by means of clone formation. One-hit multi-target model was fitted to the cell survival curve to calculate dose quasithreshold (Dq), mean lethal dose (D(0)), 2Gy survival fraction (SF(2)) and sensitization enhancement ratio (SER). The pAkt and total Akt expression was detected by Western blotting and DNA damage by neutro-comet electrophoresis. The HeLa cells were randomly divided into 7 groups in terms of different treatments: Control; radiation treatment (RT) group; LY294002+RT group; cisplatin+RT group; docetaxel+RT group; LY294002+cisplatin+RT group; LY294002+docetaxel+RT group. The apoptosis ratio of each group was measured by flow cytometry. The results showed that docetaxel and cisplatin significantly enhanced the phosphorylation of Akt in radiation-treated HeLa cells. The Dq, D(0) and SF2 in LY294002-contained groups were lower than those in docetaxel or cisplatin+RT group. The SER in the LY294002+docetaxel+RT group was 1.35 times that of the docetaxel+RT group, and that in the LY294002+cisplatin+RT group 1.26 times that of the cisplatin+RT group. The Comet electrophoresis showed that tail distance in the LY294002+cisplatin+RT group or LY294002+docetaxel+ RT group was longer than in the cisplatin+RT group or docetaxel+RT group. The apoptosis ratio in the LY294002+cisplatin+RT group or LY294002+docetaxel +RT group was higher than in the cisplatin+RT group or docetaxel+RT group. It was concluded that inhibiting PI3K/Akt pathway can increase the effect of docetaxel and cisplatin on the radiosensitivity of HeLa cells and DNA damage resulted from radiation.
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In order to investigate the inhibitory effects of Endostar (rh-endostatin, YH-16) in combination with radiotherapy on lung adenocarcinoma A549 in mice and the interaction mechanisms of combined therapy, the transplantation tumor models of A549 lung adenocarcinoma were established. When the largest diameter of tumor reached 1.0 cm, all nude mice were randomly divided into 4 groups: Endostar group, radiotherapy group, radiotherapy plus Endostar (combined treatment) group, and control group (n=6 in each group). The largest diameter and the vertical diameter of tumor were measured at different time points. At the 16th day, mice were executed, and the tumors were applied to analysis of rate of tumor cell apoptosis, and the expression levels of basic fibroblast growth factor (bFGF) mRNA were detected by reverse transcription-polymerase chain reaction (RT-PCR) and those of vascular endothelial growth factor (VEGF) by immunohistochemistry. The results demonstrated that the rate of tumor inhibition in combined treatment group was higher than that in other groups. And the rate of tumor cell apoptosis in combined treatment group was also higher than that in other groups. Meanwhile, the levels of bFGF mRNA and VEGF expression in combined treatment group were lower than those in other groups. It was concluded that Endostar obviously enhanced the curative effectiveness of radiotherapy on lung adenocarcinoma A549 in mice. The underlying mechanisms may involve the down-regulation of bFGF mRNA and VEGF expression to inhibit angiogenesis by Endostar and the cooperative effect of Endostar and radiotherapy to synergistically promote tumor cell apoptosis. And Endostar inhibits angiogenesis by down-regulating the expression of bFGF mRNA and VEGF.
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Objective To determine the reliability, validity and feasibility of using International Classifica-tion of Functioning, Disability and Health (ICF) core set for breast cancer in Chinese patients. Methods Quality of life (QOL) of 110 patients with breast cancer were investigated using ICF breast cancer core set and SF-36 scale. Statistical analysis tested their reliability, validity and feasibility. Results It was shown that the ICF core set for breast cancer had good reliability, validity and feasibility. Conclusions The ICF core set for breast cancer is a re-liable and valid instrument for Chinese breast cancer patients.
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Objective To evaluate the efficacy and safety of sorafenib in the treatment of Chi-nese patients with metastatic renal cell carcinoma. Methods This muhicenter phase Ⅱ clinical trial was performed from May 2006 to December 2006. Sixty-two patients with metastatic renal cell carci-noma not suitable for curative treatment were enrolled. All patients received oral sorafenib as single a-gent at the dose of 400 mg twice a day until disease progression or intolerable toxicities occurred. Re-salts Partial responses were recorded as best response in 11 patients, while complete remission was found in 1 patient and stable diseases were found in another 35 patients. According to the intents-to-treatment population, the overall response rate was 19.4% (12/62), and the disease control rate was 77.4%(48/62). The median progression free survival time was 9.6 months with 1-year progression-free survival rate of 41.9%. However, the median survival time had not reached due to the short fol-low-up. The most frequent adverse events included alopecia (66.1%), diarrhea (62.9%), hand-foot syndrome (58.1%), anorexia (40.3%), rash(37.1%), fatigue (37.1%), hypertension (35.5%), hoarseness(32.3%), joint pain (25.8%), hypophosphatemia (21.0%), fever (19.4%), nausea (19.4%), abnormal transeaminase( 11.3% ), elevated total bilirubicin( 16.1% ), leucopenia( 12.9% ), bleeding under nail(16.1%), and gum bleeding(11.3%). Grade 3 adverse events included hand-foot syndrome (16.1%), hypertension (12.9%), diarrhea(6.5%), hypophosphatemia (4.8%), joint pain (3.2%), and leucopenia(3.2%). Conclusions Sorafenib has prominent anti-tumor activity in Chi-nese metastatic renal cell cancer patients with most adverse events being grade 1 or 2. More attention should be paid to hypertension and cardio-cerebral vascular events during the application of sorafenib.
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Objective To evaluate the influence of belly beard device and the distended bladder on the dose distribution of PTV and the dose-volume histograms(DVHs)of organs at risk(OARs)for postoperative radiation tIlerapy of rectal cancer.Methods A total of 23 patients(8 and 15)with distended bladder receiving 3-field postoperative radiation therapy were dealed with or without a special belly beard in the prone position.At the same time,15 cages with belly board were scanned with empty bladder.The volume of irradiated small bowel was calculated for doses between 5-50 Gy at 5 Gy intervals.With prescription dose in plan target volume(PTV)of 50 Gy,we compared the dose distribution,DVH of OARs,conformity index(CIPTV),the volume of irradiated small bowel and the acute toxicity under the condition of thlee different moulds.Results There was no significant difference in PTV's converge,DVHs of femoral head and CI among 3 moulds(P＞0.05).With the belly board,the high-dose volume of irradiated small bowel(V20-V52.5)was significantly decreased(P＜0.05),specially with distended bladder.However,the low dose volume(V5-V15)was slightly increased.The bladder distension significanfly decreases the volumes of the irradiated small howel at dose levels from 15-52.5 Gy(P＜0.05).Furthermore,the mean volume(V5-V30)of irradiated small bowel differed significantly between patients experiencing Grade 0.1 and ≥2 diarrhea(P＜0.05).Conclusions The combination of belly board and distended bladder was more effectively to reduce the irradiated small bowel volume among 3 moulds,so as to minimized acute diarrhea toxicity.
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Objective To observe the EGFR nuclear translocation in cervical carcinoma cell lines after irradiation and its possible role in radiation tolerance.Methods Western blotting was used to detect the nuclear EGFR and cytoplastic EGFR after irradiation.The effect of Cetuximab on expression of nuclear EGFR and survival fractions were investigated.Results After irradiation,compared with control group,the expression of nuclear EGFR protein increased in irradiated cervical carcinoma cell.Cetuximab inhibited the radiation-induced nuclear EGFR expression with decreased survival fractions.Conclusion Radiation could induce EGFR nuclear translocation in cervical carcinoma cell lines and nuclear EGFR might be correlated with radiation tolerance in Cervical carcinoma cell.
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Objective To evaluate the role of postoperative electron beam radiotherapy in the management of ke-loids. Methods Between January 2004 and December 2006,116 cases of keloids were treated with 6 MeV electron beam radiotherapy within 24 h after surgical resection of the keloids. All patients had received a total of 21 Gy in seven daily 3-Gy fractions. Treatment was started at the 24th h after surgery. Patients were followed up and the information regarding treatment results, early and late radiation toxicity,and the satisfaction level by self-assessments was recorded. Results Recurrence occurred in 17 cases of keloids(14. 7%),and was correlated with site of the lesions(χ~2 =29. 91,P<0. 01). Most recurrences were observed at site of sternum (10/43) and shoulder (5/13 ). Keloid associated symptoms, e. g. itching and pain, were improved in 88. %%. For early toxicity outcomes, 100% had grade 1～2 skin erythema,7. 8% had wound delayed union,and 4. 3% had infection. For late toxicity outcomes, 30. 2% reported grade 1～2 hyperpigmentation, 11. 2% grade 1～2 hypopigmentation, and 5. 2% grade 1 telangiectasia. No severe complications or secondary malignancies were observed. 72. 4% patients described the results of treatment as excellent or good,and 15. 5% patients were not satisfied with the treatment results. Conclusion Postoperative electron radiotherapy is well tolerated and effective in preventing keloid recurrence.
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In order to investigate the role of antiapoptosis gene, survivin in the resistance to palcitaxel, the expression of survivin mRNA and protein in the process of paclitaxel treatment in breast cancer cell line MCF-7 was detected MCF-7 cells were incubated with paclitaxel at different concentrations. The growth inhibition rate of MCF-7 was investigated by tetrazolium bromide (MTT) colorimetry. The change of apoptosis was detected by Annexin-V/PI methods. The changes in the expression of survivin mRNA and protein were studied by reverse transcription polymerase chain reaction (RT-PCR) and Western-blot assay respectively. The growth inhibition rate of MCF-7 was increased in a concentration-and time-dependent manner. Paclitaxel of higher concentration could effectively induce apoptosis in MCF-7 cells after 48 h, while the expression of survivin was increased at early time (within 6 h) and decreased after 24 h regardless of treatment concentrations of paclitaxel. It suggested that tumor cells might evade the paclitaxel-induced cell cycle arrest and apoptosis by increasing the level of survivin at early treatment time.
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In order to investigate the role of antiapoptosis gene, survivin in the resistance to palcitaxel, the expression of survivin mRNA and protein in the process of paclitaxel treatment in breast cancer cell line MCF-7 was detected. MCF-7 cells were incubated with paclitaxel at different concentrations. The growth inhibition rate of MCF-7 was investigated by tetrazolium bromide (MTT) colorimetry. The change of apoptosis was detected by Annexin-V/PI methods. The changes in the expression of survivin mRNA and protein were studied by reverse transcription polymerase chain reaction (RT-PCR) and Western-blot assay respectively. The growth inhibition rate of MCF-7 was increased in a concentration- and time-dependent manner. Paclitaxel of higher concentration could effectively induce apoptosis in MCF-7 cells after 48 h, while the expression of survivin was increased at early time (within 6 h) and decreased after 24 h regardless of treatment concentrations of paclitaxel. It suggested that tumor cells might evade the paclitaxel-induced cell cycle arrest and apoptosis by increasing the level of survivin at early treatment time.
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In order to investigate the effects of short hairpin RNA (shRNA) on the expression of Survivin, cell cycle and cell proliferation in MCF-7 cells, using a pEGFP vector which contained a U6 promoter shRNA plasmid targeted against survivin was constructed and transfected into MCF-7 cells. The change of the expression of Survivin and cell proliferation rates were detected by semiquantitative reverse transcription polymerase chain reaction (RT-PCR) and MTT methods respectively. The change of cell cycle after transfection was analyzed by flow cytometry. The results indicated that the recombinant plasmid containing Survivin shRNA was constructed successfully, which could suppress the expression of Survivin at mRNA and protein level. The growth of MCF-7 cells was arrested in G1 phase of the cell cycle and the proliferation activity was suppressed after transfection. It was concluded that Survivin shRNA plasmid could knock down the expression of Survivin in MCF-7 cells specifically. In addition, Survivin shRNA plasmid could lead to G1 arrest and inhibit the proliferation of MCF-7 cells, which suggested that Survivin shRNA might be used as a new therapeutic method for breast cancer.