Objective To investigate the impact of all-trans retinoic acid （ATRA） on MDM2 gene expression in astrocytoma cell line SHG-44, and to provide basic data for further research on the progression mechanism and gene the...Objective To investigate the impact of all-trans retinoic acid （ATRA） on MDM2 gene expression in astrocytoma cell line SHG-44, and to provide basic data for further research on the progression mechanism and gene therapy of human astrocytoma. Methods The differential expressions of MDM2 gene and protein in SHG-44 cells were detected by cDNA microarray and Western blot, respectively, before and after treatment of ATRA. The expressions of MDM2 protein in WHO grade Ⅱ and grade Ⅳ astrocytomas were determined by immunohistochemical streptavidin-peroxidase method. Some differentially expressed genes were selected randomly for Northern blot analysis. Results The intensity ratio of ATRA-treated to untreated SHG-44 cell was 0.37 in the cDNA microarray, suggesting that the expression of MDM2 gene was down-regulated in SHG-44 cells after treatment with ATRA. Some genes differentially expressed in the microarray were confirmed by Northern blot. Western blot demonstrated that the optical density ratios of MDM2 to β-actin in ATRA-treated and untreated SHG-44 were 14.02±0.35 and 21.40±0.58 （t = 24.728, P = 0.000）, respectively, suggesting that the expression of MDM2 protein was inhibited in ATRA-treated SHG-44 cells. Moreover, the percentages of MDM2-positive protein were 24.00% （6/25） and 56.52% （13/23） （x^2 = 5.298, P = 0.021） in WHO grade Ⅱ and grade Ⅳ astrocytomas, respectively, suggesting that the expression of MDM2 protein may increase along with the elevation of astrocytoma malignancy. Conclusion ATRA can inhibit MDM2 gene expression in SHG-44 cells, and MDM2 is related to astrocytoma progression.展开更多
Objective:To explore the suppressing effect ofγ-secretase inhibitor DAPT on proliferation of human glioma cell line SHG-44 in vitro and its mechanism.Methods:The SHG-44 cell was treated by DAPT with different concent...Objective:To explore the suppressing effect ofγ-secretase inhibitor DAPT on proliferation of human glioma cell line SHG-44 in vitro and its mechanism.Methods:The SHG-44 cell was treated by DAPT with different concentration.The proliferation of cells was detected by MTT assay;cell cycle and TSC of CD133^+were determined by flow cytometry analysis technique;the key factor in Notch signaling pathway(Notch-1,Delta-1,Hes-1)was measured by reverse transcrip tase-polymerase chain reaction and western blotting.Results:DAPT inhibited the growth and proliferation of SHG-44 cells significantly(P<0.05).And the inhibiting effect on SHG-44 cells produced by DAPT showed a dose-dependent manner.DAPT increased the rate of cells in G_0/G_1 phase of SHG-44 cells,while it decreased the rate of cells in S phase.TSC of CD133^+was significantly reduced after DAPT treated SHC-44 cells.The expression of protein and mRNA of Notch-1,Delta-1 and Hes-1 were gradually downregulated with the increase of DAPT doses.Conclusions:DAPT can downregulate these key factor in Notch signaling pathway,reduce the TSC of CD133+and inhibit the proliferation of SHC-44 cells.展开更多
AIM: To observe the effects of salvianolic add B (SalB) on in vitro growth inhibition and apoptosis induction of retinoblastoma HXO-RB44 cells. METHODS: The effects of SalB on the HXO-RB44 cells proliferation in vitro...AIM: To observe the effects of salvianolic add B (SalB) on in vitro growth inhibition and apoptosis induction of retinoblastoma HXO-RB44 cells. METHODS: The effects of SalB on the HXO-RB44 cells proliferation in vitro were observed by MTT colorimetric method. The morphological changes of apoptosis before and after the treatment of SalB were observed by Hoechst 33258 fluorescent staining method. Apoptosis rate and cell cycle changes of HXO-RB44 cells were detected by flow cytometer at 48 hours after treated by SalB. The expression changes of Caspase-3 protein in HXO-RB44 cells were detected by Western Blot. RESULTS: SalB significantly inhibited the growth of HXO-RB44 cells, while the inhibition was in a concentration-and time-dependent manner. The results of fluorescent staining method indicated that HXO-RB44 cells showed significant phenomenon of apoptosis including karyorrhexis, fragmentation and the formation of apoptotic bodies, etc. after 24, 48 and 72 hours co-culturing of SalB and HXO-RB44 cells. The results of flow cytometer showed that the apoptosis rate and the proportion of cells in S phase were gradually increased at 48 hours and 72 hours after treated by different concentrations of SalB. Western Blot strip showed that the expression of Caspase-3 protein in HXO-RB44 cells was gradually increased with the increase of the concentration of SalB. CONCLUSION: SalB can significantly affect on HXO-RB44 cells growth inhibition and apoptosis induction which may be achieved through the up-regulation of Caspase-3 expression and the induction of cell cycle arrest.展开更多
BACKGROUND： Progenitors of the immortalized human glioma cell line, SHG-44, are significantly less sensitive to irradiation. Two hypotheses regarding the mechanism of this effect exist： several studies have suggeste...BACKGROUND： Progenitors of the immortalized human glioma cell line, SHG-44, are significantly less sensitive to irradiation. Two hypotheses regarding the mechanism of this effect exist： several studies have suggested that there is a subgroup with different radiosensitivities in identical cell group, and the progenitors of irradiate is a adaptive response subgroup, so its radiosensitivity is descend. A second hypothesis suggests that irradiated glioma progeny have a stronger ability to repair DNA damage. This would suggest that when progeny are continuously irradiated, resistance to irradiation-induced DNA increases, and radiosensitivity decreases. OBJECTIVE： To investigate radiosensitivity and growth features after irradiation to progeny of the human glioma cell line SHG-44. DESIGN, TIME AND SETTING： A randomized, controlled experiment, which was performed at the Department of Radiology Laboratory, the First Hospital Affiliated to Soochow University, between September 2004 and January 2006. MATERIALS： The glioma cell line SHG-44 was provided by the Institute of Neuroscience, First Affiliated Hospital of Suzhou University. Propidium iodide reagent was provided by Coulter Corporation. A linear accelerator, KD-2 type, was provided by Siemens, Germany. The flow cytometer EPICS-XL was provided by Coulter Corporation. METHODS： Brain glioma SHG-44 cells were divided into four groups： SHG-44, SHG-44-2, SHG-44-6, and SHG-44-10 . The SHG-44-2, SHG-44-6, and SHG-44-10 cells were vertically irradiated with varying doses of 2, 6 and 10 Gy by a linear accelerator （6 MVX）. The cells were passaged for 15 generations and cultured in RPMI-1640 culture media. MAIN OUTCOME MEASURES： Community re-double time, mean lethal dose （D0）, extrapolation number （N）, fraction surviving fraction irradiated by 2 Gy dose （SF2）, quasi-threshold dose （Dq）, and cell cycle. RESULTS： The Population doubling time （PDT） of SHG-44-2, SHG-44-6, and SHG-44-10 cell groups was not significant （P = 0.052）. Compared to these three groups, the PDT of the SHG-44 cell group was significantly difference （F = 7.878, P 〈 0.002）. SHG-44 cell clone ratewas 26.5%, and SHG-44-10 cell group was 15.5%. The SHG-44-10 cell group also exhibited radiosensitivity, but was less than the radiosensitivity of the SHG-44 cell group. Compared to the SHG-44 cell group, the ratio of the G2/M phase was decreased in the SHG-44-10 cell group, and the radio of S phase was increased. The SHG-44 and SHG-44-10 cell groups were irradiated with 8 Gy. After 12 hours, the G2/M ratio was compared to pre-irradiation times, indicating a significantly higher ratio in the pre-irradiated groups （P 〈 0.01）. The cells between S HG-44 and SHG-44-10 groups were harvested 12 hours after irradiation： G2 phase of SHG-44-10 cells was arrested and the G2/M ratio was increased, which was intensified with increasing irradiation doses. CONCLUSION： In the present study, the proliferation delay and decreased radiosensitivity were confirmed in progeny of irradiated human glioma cells, and radiosensitivity was dose-dependent.展开更多
Objective： To study the effects of acetaminophen （ACE） combined with radiation on the progeny of the human glioma cell line SHG-44, and to investigate if ACE may be an useful therapeutic radiosensitivity agent in t...Objective： To study the effects of acetaminophen （ACE） combined with radiation on the progeny of the human glioma cell line SHG-44, and to investigate if ACE may be an useful therapeutic radiosensitivity agent in the treatment of recurrent human glioma. Methods： A randomized, controlled experiment, was performed at the Department of Radiology Laboratory, the First Hospital Affiliated to Soochow University, between September 2004 and January 2006. Brain glioma SHG-44 cells were divided into three groups： SHG-44, SHG-44-10, and SHG-44-10 ＋ ACE cells groups. The SHG-44-10 cells group was irradiated with dose of 10 Gy by a linear accelerator （6 MVX）. It was passaged for 15 generations and cultured in RPMI-1640 culture media. Then SHG-44-10 ＋ ACE cells group was treated with ACE. Measures： Community re-double time, mean lethal dose （DO）, extrapolation number （N）, fraction surviving fraction irradiated by 2 Gy dose （SF2）, quasi-threshold dose （Dq）, and cell cycle. Results： The SF2 of the SHG-44, SHG-44-10, and SHG-44-10 ＋ ACE cells groups were 70.8%, 80.6% and 45.2%, respectively, with significance （P = 0.040）. The SHG-44-10 and SHG-44-10 ＋ ACE cells groups were irradiated with 8 Gy. After 12 hours, the G2/M ratio of the SHG-44-10 and SHG-44-10 ＋ ACE cells groups were indicating significantly higher ratio compared to pre-irradiated groups （P 〈 0.01）. After 24 hours, the G2/M ratio of the SHG-44-10 cells group decreased rapidly, while the ratio of the SHG-44-10 ＋ ACE cells group still maintained in high level. Conclusion： In the present study, Subtoxic dose of ACE increased the radiosensitivity of the progeny of irradiated human glioma cell. ACE may be an useful radiosensitivity agent in the treatment of recrudescent human malignant glioma.展开更多
基金a grant from the Bureau of Health, Sichuan Province, China (No. 050209).
文摘Objective To investigate the impact of all-trans retinoic acid （ATRA） on MDM2 gene expression in astrocytoma cell line SHG-44, and to provide basic data for further research on the progression mechanism and gene therapy of human astrocytoma. Methods The differential expressions of MDM2 gene and protein in SHG-44 cells were detected by cDNA microarray and Western blot, respectively, before and after treatment of ATRA. The expressions of MDM2 protein in WHO grade Ⅱ and grade Ⅳ astrocytomas were determined by immunohistochemical streptavidin-peroxidase method. Some differentially expressed genes were selected randomly for Northern blot analysis. Results The intensity ratio of ATRA-treated to untreated SHG-44 cell was 0.37 in the cDNA microarray, suggesting that the expression of MDM2 gene was down-regulated in SHG-44 cells after treatment with ATRA. Some genes differentially expressed in the microarray were confirmed by Northern blot. Western blot demonstrated that the optical density ratios of MDM2 to β-actin in ATRA-treated and untreated SHG-44 were 14.02±0.35 and 21.40±0.58 （t = 24.728, P = 0.000）, respectively, suggesting that the expression of MDM2 protein was inhibited in ATRA-treated SHG-44 cells. Moreover, the percentages of MDM2-positive protein were 24.00% （6/25） and 56.52% （13/23） （x^2 = 5.298, P = 0.021） in WHO grade Ⅱ and grade Ⅳ astrocytomas, respectively, suggesting that the expression of MDM2 protein may increase along with the elevation of astrocytoma malignancy. Conclusion ATRA can inhibit MDM2 gene expression in SHG-44 cells, and MDM2 is related to astrocytoma progression.
基金supported by Shaanxi Province Health Department Key Funds(sx201227273)
文摘Objective:To explore the suppressing effect ofγ-secretase inhibitor DAPT on proliferation of human glioma cell line SHG-44 in vitro and its mechanism.Methods:The SHG-44 cell was treated by DAPT with different concentration.The proliferation of cells was detected by MTT assay;cell cycle and TSC of CD133^+were determined by flow cytometry analysis technique;the key factor in Notch signaling pathway(Notch-1,Delta-1,Hes-1)was measured by reverse transcrip tase-polymerase chain reaction and western blotting.Results:DAPT inhibited the growth and proliferation of SHG-44 cells significantly(P<0.05).And the inhibiting effect on SHG-44 cells produced by DAPT showed a dose-dependent manner.DAPT increased the rate of cells in G_0/G_1 phase of SHG-44 cells,while it decreased the rate of cells in S phase.TSC of CD133^+was significantly reduced after DAPT treated SHC-44 cells.The expression of protein and mRNA of Notch-1,Delta-1 and Hes-1 were gradually downregulated with the increase of DAPT doses.Conclusions:DAPT can downregulate these key factor in Notch signaling pathway,reduce the TSC of CD133+and inhibit the proliferation of SHC-44 cells.
基金National "Eleventh Five-year Plan" Science and Technology Support Project (No. 2006BAI06 A15-3)
文摘AIM: To observe the effects of salvianolic add B (SalB) on in vitro growth inhibition and apoptosis induction of retinoblastoma HXO-RB44 cells. METHODS: The effects of SalB on the HXO-RB44 cells proliferation in vitro were observed by MTT colorimetric method. The morphological changes of apoptosis before and after the treatment of SalB were observed by Hoechst 33258 fluorescent staining method. Apoptosis rate and cell cycle changes of HXO-RB44 cells were detected by flow cytometer at 48 hours after treated by SalB. The expression changes of Caspase-3 protein in HXO-RB44 cells were detected by Western Blot. RESULTS: SalB significantly inhibited the growth of HXO-RB44 cells, while the inhibition was in a concentration-and time-dependent manner. The results of fluorescent staining method indicated that HXO-RB44 cells showed significant phenomenon of apoptosis including karyorrhexis, fragmentation and the formation of apoptotic bodies, etc. after 24, 48 and 72 hours co-culturing of SalB and HXO-RB44 cells. The results of flow cytometer showed that the apoptosis rate and the proportion of cells in S phase were gradually increased at 48 hours and 72 hours after treated by different concentrations of SalB. Western Blot strip showed that the expression of Caspase-3 protein in HXO-RB44 cells was gradually increased with the increase of the concentration of SalB. CONCLUSION: SalB can significantly affect on HXO-RB44 cells growth inhibition and apoptosis induction which may be achieved through the up-regulation of Caspase-3 expression and the induction of cell cycle arrest.
文摘BACKGROUND： Progenitors of the immortalized human glioma cell line, SHG-44, are significantly less sensitive to irradiation. Two hypotheses regarding the mechanism of this effect exist： several studies have suggested that there is a subgroup with different radiosensitivities in identical cell group, and the progenitors of irradiate is a adaptive response subgroup, so its radiosensitivity is descend. A second hypothesis suggests that irradiated glioma progeny have a stronger ability to repair DNA damage. This would suggest that when progeny are continuously irradiated, resistance to irradiation-induced DNA increases, and radiosensitivity decreases. OBJECTIVE： To investigate radiosensitivity and growth features after irradiation to progeny of the human glioma cell line SHG-44. DESIGN, TIME AND SETTING： A randomized, controlled experiment, which was performed at the Department of Radiology Laboratory, the First Hospital Affiliated to Soochow University, between September 2004 and January 2006. MATERIALS： The glioma cell line SHG-44 was provided by the Institute of Neuroscience, First Affiliated Hospital of Suzhou University. Propidium iodide reagent was provided by Coulter Corporation. A linear accelerator, KD-2 type, was provided by Siemens, Germany. The flow cytometer EPICS-XL was provided by Coulter Corporation. METHODS： Brain glioma SHG-44 cells were divided into four groups： SHG-44, SHG-44-2, SHG-44-6, and SHG-44-10 . The SHG-44-2, SHG-44-6, and SHG-44-10 cells were vertically irradiated with varying doses of 2, 6 and 10 Gy by a linear accelerator （6 MVX）. The cells were passaged for 15 generations and cultured in RPMI-1640 culture media. MAIN OUTCOME MEASURES： Community re-double time, mean lethal dose （D0）, extrapolation number （N）, fraction surviving fraction irradiated by 2 Gy dose （SF2）, quasi-threshold dose （Dq）, and cell cycle. RESULTS： The Population doubling time （PDT） of SHG-44-2, SHG-44-6, and SHG-44-10 cell groups was not significant （P = 0.052）. Compared to these three groups, the PDT of the SHG-44 cell group was significantly difference （F = 7.878, P 〈 0.002）. SHG-44 cell clone ratewas 26.5%, and SHG-44-10 cell group was 15.5%. The SHG-44-10 cell group also exhibited radiosensitivity, but was less than the radiosensitivity of the SHG-44 cell group. Compared to the SHG-44 cell group, the ratio of the G2/M phase was decreased in the SHG-44-10 cell group, and the radio of S phase was increased. The SHG-44 and SHG-44-10 cell groups were irradiated with 8 Gy. After 12 hours, the G2/M ratio was compared to pre-irradiation times, indicating a significantly higher ratio in the pre-irradiated groups （P 〈 0.01）. The cells between S HG-44 and SHG-44-10 groups were harvested 12 hours after irradiation： G2 phase of SHG-44-10 cells was arrested and the G2/M ratio was increased, which was intensified with increasing irradiation doses. CONCLUSION： In the present study, the proliferation delay and decreased radiosensitivity were confirmed in progeny of irradiated human glioma cells, and radiosensitivity was dose-dependent.
文摘Objective： To study the effects of acetaminophen （ACE） combined with radiation on the progeny of the human glioma cell line SHG-44, and to investigate if ACE may be an useful therapeutic radiosensitivity agent in the treatment of recurrent human glioma. Methods： A randomized, controlled experiment, was performed at the Department of Radiology Laboratory, the First Hospital Affiliated to Soochow University, between September 2004 and January 2006. Brain glioma SHG-44 cells were divided into three groups： SHG-44, SHG-44-10, and SHG-44-10 ＋ ACE cells groups. The SHG-44-10 cells group was irradiated with dose of 10 Gy by a linear accelerator （6 MVX）. It was passaged for 15 generations and cultured in RPMI-1640 culture media. Then SHG-44-10 ＋ ACE cells group was treated with ACE. Measures： Community re-double time, mean lethal dose （DO）, extrapolation number （N）, fraction surviving fraction irradiated by 2 Gy dose （SF2）, quasi-threshold dose （Dq）, and cell cycle. Results： The SF2 of the SHG-44, SHG-44-10, and SHG-44-10 ＋ ACE cells groups were 70.8%, 80.6% and 45.2%, respectively, with significance （P = 0.040）. The SHG-44-10 and SHG-44-10 ＋ ACE cells groups were irradiated with 8 Gy. After 12 hours, the G2/M ratio of the SHG-44-10 and SHG-44-10 ＋ ACE cells groups were indicating significantly higher ratio compared to pre-irradiated groups （P 〈 0.01）. After 24 hours, the G2/M ratio of the SHG-44-10 cells group decreased rapidly, while the ratio of the SHG-44-10 ＋ ACE cells group still maintained in high level. Conclusion： In the present study, Subtoxic dose of ACE increased the radiosensitivity of the progeny of irradiated human glioma cell. ACE may be an useful radiosensitivity agent in the treatment of recrudescent human malignant glioma.
