目的:研究重楼皂苷Ⅰ(PPⅠ)对去势抵抗性人前列腺癌PC3细胞生长的抑制作用及其分子机制。方法:体外培养PC3细胞,予不同浓度(0.4、0.8、1.2、1.6、2.0、2.4μmol/L)的PPⅠ处理24、48、72 h,另设对照组,MTT法观察PPⅠ对PC3细胞增殖的影响...目的:研究重楼皂苷Ⅰ(PPⅠ)对去势抵抗性人前列腺癌PC3细胞生长的抑制作用及其分子机制。方法:体外培养PC3细胞,予不同浓度(0.4、0.8、1.2、1.6、2.0、2.4μmol/L)的PPⅠ处理24、48、72 h,另设对照组,MTT法观察PPⅠ对PC3细胞增殖的影响;流式细胞术检测细胞凋亡率;Western印迹检测PPⅠ对磷酸化细胞外调节蛋白激酶1/2(p-ERK1/2)、细胞外调节蛋白激酶1/2(ERK1/2)、核转录因子kappa B(NF-κB)/p65以及DNA甲基转移酶1(DNMT1)蛋白表达的影响,并加入ERK1/2抑制剂(PD98059)后检测PPⅠ对NF-κB/p65表达的影响。结果:MTT结果显示,PPⅠ能抑制PC3细胞的体外增殖,与对照组相比,PC3细胞从给药浓度0.4μmol/L(0.85±0.05 vs 1.00±0.00,P<0.01)开始明显下降,且呈时间和剂量依赖性。流式细胞术检测显示,PPⅠ能明显诱导PC3细胞早期凋亡,与对照组相比,PC3细胞早期凋亡率从给药浓度0.8μmol/L(13.83±2.97 vs 4.83±0.95)开始明显增加(P均<0.01),且呈剂量依赖性;Western印迹显示,PPⅠ以时间依赖性上调p-ERK1/2和ERK1/2蛋白的激活与表达,与对照组相比,给药时间从2 h(1.73±0.17 vs 1.00±0.00,P<0.01)开始,p-ERK1/2明显被激活表达,PPⅠ以剂量依赖性下调NF-κB/p65、DNMT1蛋白的表达,与对照组相比,给药浓度分别从1.6μmol/L(0.67±0.11 vs 1.00,P<0.01)与1.2μmol/L(0.63±0.06 vs 1.00±0.00,P<0.01)开始,NF-κB/p65、DNMT1表达明显下调;抑制ERK1/2磷酸化能逆转重楼皂苷Ⅰ对NF-κB/p65蛋白表达的下调,与PPⅠ组相比,PD98059+PPⅠ组NF-κB/p65蛋白表达(0.86±0.18 vs 0.43±0.09,P<0.05)明显上调。结论:PPⅠ可能通过介导ERK1/2通路抑制NF-κB/p65和DNMT1蛋白表达,诱导PC3细胞早期凋亡,进而抑制细胞增殖。展开更多
AIM： To explore the relationship between DNA methyltransferase 1 （DNMT1） and hepatitis B virus （HBV）-related hepatocellular carcinoma （HCC） and its biological significance in primary HCC. METHODS： We carried o...AIM： To explore the relationship between DNA methyltransferase 1 （DNMT1） and hepatitis B virus （HBV）-related hepatocellular carcinoma （HCC） and its biological significance in primary HCC. METHODS： We carried out an immunohistochemical examination of DNMT1 in both HCC and paired nonneoplastic liver tissues from Chinese subjects. DNMT1 mRNA was further examined in HCC cell lines by real-time PCR. We inhibited DNMT1 using siRNA and detected the effect of depletion of DNMT1 on cell proliferation ability and cell apoptosis in the HCC celt line SMMC-7721. RESULTS： DNMT1 protein expression was increased in HCCs compared to histologically normal nonneoplastic liver tissues and the incidence of DNMT1 immunoreactivity in HCCs correlated significantly with poor tumor differentiation （P = 0.014）. There were more cases with DNMT1 overexpression in HCC with HBV （42.85%） than in HCC without HBV （28.57%）. However, no significant difference in DNMT1 expression was found in HBV-positive and HBV-negative cases in the Chinese HCC group. There was a trend that DNMT1 RNA expression increased more in HCC cell lines than in pericarcinoma cell lines and normal liver cell lines. In addition, we inhibited DNMT1 using siRNA in the SMMC-7721 HCC cell line and found depletion of DNMT1 suppressed cells growth independent of expression of proliferating cell nuclear antigen （PCNA）, even in HCC cell lines where DNMT1 was stably decreased. CONCLUSION： The findings implied that DNMT1 plays a key role in HBV-retated hepatocellular tumorigenesis. Depletion of DNMT1 mediates growth suppression in SMMC-7721 cells.展开更多
BACKGROUND In recent years,the incidence of gastrointestinal(GI)cancer in China has increased annually.Early detection and appropriate therapy are considered to be the key to treat GI cancer.DNMT1 takes an active part...BACKGROUND In recent years,the incidence of gastrointestinal(GI)cancer in China has increased annually.Early detection and appropriate therapy are considered to be the key to treat GI cancer.DNMT1 takes an active part in the advancement of GI cancer,which will change as the disease progresses.But its expression characteristics in the dynamic variations of GI carcinogenesis are still unclear.AIM To investigate the expression characteristics of DNMT1 in different GI diseases.METHODS We detected the expression of DNMT1 in 650 cases of different GI diseases by immunohistochemistry,including 90 cases of chronic superficial gastritis(CSG),72 cases of atrophic gastritis with intestinal metaplasia(AG/GIM),54 cases of low-grade intraepithelial neoplasia(GLIN),66 cases of high-grade intraepithelial neoplasia(GHIN),71 cases of early gastric cancer(EGC),90 cases of normal intestinal mucosa(NIM),54 cases of intestinal low-grade intraepithelial neoplasia(ILIN),71 cases of intestinal high-grade intraepithelial neoplasia(IHIN),and 82 cases of early colorectal cancer(ECRC).RESULTS In the CSG group,all cases showed weakly positive or negative expression of DNMT1.However,in other four groups(AG/GIM,GLIN,GHIN,and EGC),the positive expression rate gradually increased with the severity of the diseases;the negative or weakly positive cases accounted for 55.56%(40/72),38.89%(21/54),1.52%(1/66),and 1.41%(1/71),respectively.Besides,the moderately positive cases were 44.44%(32/72),57.41%(31/54),80.30%(53/66),and 43.66%(31/71),respectively.The strongly positive cases only existed in the GLIN(3.70%,2/54),GHIN(18.18%,12/66),and EGC(54.93%,39/71)groups.The differences between any two groups were statistically significant(P<0.05).Similarly,in the NIM group,cases with weakly positive expression of DNMT1 were predominant(91.11%,82/90),and the rest were moderately positive cases(8.89%,8/90).In the ILIN,IHIN,and ECRC groups,the rates of cases with weak or negative expression of DNMT1 were 46.30%(25/54),12.68%(9/71),and 4.88%(4/82),respectively;with展开更多
文摘目的:研究重楼皂苷Ⅰ(PPⅠ)对去势抵抗性人前列腺癌PC3细胞生长的抑制作用及其分子机制。方法:体外培养PC3细胞,予不同浓度(0.4、0.8、1.2、1.6、2.0、2.4μmol/L)的PPⅠ处理24、48、72 h,另设对照组,MTT法观察PPⅠ对PC3细胞增殖的影响;流式细胞术检测细胞凋亡率;Western印迹检测PPⅠ对磷酸化细胞外调节蛋白激酶1/2(p-ERK1/2)、细胞外调节蛋白激酶1/2(ERK1/2)、核转录因子kappa B(NF-κB)/p65以及DNA甲基转移酶1(DNMT1)蛋白表达的影响,并加入ERK1/2抑制剂(PD98059)后检测PPⅠ对NF-κB/p65表达的影响。结果:MTT结果显示,PPⅠ能抑制PC3细胞的体外增殖,与对照组相比,PC3细胞从给药浓度0.4μmol/L(0.85±0.05 vs 1.00±0.00,P<0.01)开始明显下降,且呈时间和剂量依赖性。流式细胞术检测显示,PPⅠ能明显诱导PC3细胞早期凋亡,与对照组相比,PC3细胞早期凋亡率从给药浓度0.8μmol/L(13.83±2.97 vs 4.83±0.95)开始明显增加(P均<0.01),且呈剂量依赖性;Western印迹显示,PPⅠ以时间依赖性上调p-ERK1/2和ERK1/2蛋白的激活与表达,与对照组相比,给药时间从2 h(1.73±0.17 vs 1.00±0.00,P<0.01)开始,p-ERK1/2明显被激活表达,PPⅠ以剂量依赖性下调NF-κB/p65、DNMT1蛋白的表达,与对照组相比,给药浓度分别从1.6μmol/L(0.67±0.11 vs 1.00,P<0.01)与1.2μmol/L(0.63±0.06 vs 1.00±0.00,P<0.01)开始,NF-κB/p65、DNMT1表达明显下调;抑制ERK1/2磷酸化能逆转重楼皂苷Ⅰ对NF-κB/p65蛋白表达的下调,与PPⅠ组相比,PD98059+PPⅠ组NF-κB/p65蛋白表达(0.86±0.18 vs 0.43±0.09,P<0.05)明显上调。结论:PPⅠ可能通过介导ERK1/2通路抑制NF-κB/p65和DNMT1蛋白表达,诱导PC3细胞早期凋亡,进而抑制细胞增殖。
基金Supported by National Natural Science Foundation of China,No.30470950
文摘AIM： To explore the relationship between DNA methyltransferase 1 （DNMT1） and hepatitis B virus （HBV）-related hepatocellular carcinoma （HCC） and its biological significance in primary HCC. METHODS： We carried out an immunohistochemical examination of DNMT1 in both HCC and paired nonneoplastic liver tissues from Chinese subjects. DNMT1 mRNA was further examined in HCC cell lines by real-time PCR. We inhibited DNMT1 using siRNA and detected the effect of depletion of DNMT1 on cell proliferation ability and cell apoptosis in the HCC celt line SMMC-7721. RESULTS： DNMT1 protein expression was increased in HCCs compared to histologically normal nonneoplastic liver tissues and the incidence of DNMT1 immunoreactivity in HCCs correlated significantly with poor tumor differentiation （P = 0.014）. There were more cases with DNMT1 overexpression in HCC with HBV （42.85%） than in HCC without HBV （28.57%）. However, no significant difference in DNMT1 expression was found in HBV-positive and HBV-negative cases in the Chinese HCC group. There was a trend that DNMT1 RNA expression increased more in HCC cell lines than in pericarcinoma cell lines and normal liver cell lines. In addition, we inhibited DNMT1 using siRNA in the SMMC-7721 HCC cell line and found depletion of DNMT1 suppressed cells growth independent of expression of proliferating cell nuclear antigen （PCNA）, even in HCC cell lines where DNMT1 was stably decreased. CONCLUSION： The findings implied that DNMT1 plays a key role in HBV-retated hepatocellular tumorigenesis. Depletion of DNMT1 mediates growth suppression in SMMC-7721 cells.
文摘BACKGROUND In recent years,the incidence of gastrointestinal(GI)cancer in China has increased annually.Early detection and appropriate therapy are considered to be the key to treat GI cancer.DNMT1 takes an active part in the advancement of GI cancer,which will change as the disease progresses.But its expression characteristics in the dynamic variations of GI carcinogenesis are still unclear.AIM To investigate the expression characteristics of DNMT1 in different GI diseases.METHODS We detected the expression of DNMT1 in 650 cases of different GI diseases by immunohistochemistry,including 90 cases of chronic superficial gastritis(CSG),72 cases of atrophic gastritis with intestinal metaplasia(AG/GIM),54 cases of low-grade intraepithelial neoplasia(GLIN),66 cases of high-grade intraepithelial neoplasia(GHIN),71 cases of early gastric cancer(EGC),90 cases of normal intestinal mucosa(NIM),54 cases of intestinal low-grade intraepithelial neoplasia(ILIN),71 cases of intestinal high-grade intraepithelial neoplasia(IHIN),and 82 cases of early colorectal cancer(ECRC).RESULTS In the CSG group,all cases showed weakly positive or negative expression of DNMT1.However,in other four groups(AG/GIM,GLIN,GHIN,and EGC),the positive expression rate gradually increased with the severity of the diseases;the negative or weakly positive cases accounted for 55.56%(40/72),38.89%(21/54),1.52%(1/66),and 1.41%(1/71),respectively.Besides,the moderately positive cases were 44.44%(32/72),57.41%(31/54),80.30%(53/66),and 43.66%(31/71),respectively.The strongly positive cases only existed in the GLIN(3.70%,2/54),GHIN(18.18%,12/66),and EGC(54.93%,39/71)groups.The differences between any two groups were statistically significant(P<0.05).Similarly,in the NIM group,cases with weakly positive expression of DNMT1 were predominant(91.11%,82/90),and the rest were moderately positive cases(8.89%,8/90).In the ILIN,IHIN,and ECRC groups,the rates of cases with weak or negative expression of DNMT1 were 46.30%(25/54),12.68%(9/71),and 4.88%(4/82),respectively;with
