We investigated the ability of NK4, an antagonist of human hepatocyte growth factor （HGF）, to inhibit the influence of HGF on proliferation, migration, invasion and apoptosis of human prostate cancer cells. Expressi...We investigated the ability of NK4, an antagonist of human hepatocyte growth factor （HGF）, to inhibit the influence of HGF on proliferation, migration, invasion and apoptosis of human prostate cancer cells. Expression vector pBudCE4.1-EGFP-NK4 containing NK4 cDNA was used to transfect human prostate cancer DU145 cells, and the effects of the autocrine NK4 on tumor cell proliferation, migration, invasion and apoptosis were assessed in vitro. in vivo, we subcutaneously implanted DU145 cells, mock-transfected clone （DU145/empty vector） cells and NK4- transfected clone （DU145/NK4） cells into nude mice, and then evaluated tumor growth, cell proliferation and cell apoptosis in vivo. We found that DU145/NK4 cells expressed NK4 protein. In the in vitro study, autocrine NK4 at- tenuated the HGF-induced tumor cell proliferation, migration and invasion, and stimulated apoptosis. Furthermore, autocrine NK4 effectively inhibited the HGF-induced phosphorylation of c-Met, extracellular signal-regulated kinase-1 （ERK1）. and protein kinase B 1/2 （Aktl/2）. Histological examination revealed that autocrine NK4 inhibited prolifera- tion and accelerated apoptosis of prostate cancer cells. These results show that genetic modification of DU145 cells with NK4 cDNA yields a significant effect on their proliferation, migration, invasion and apoptosis. Molecular targeting of HGF/c-Met by NK4 could be applied as a novel therapeutic approach to prostate cancer.展开更多
The proteasome inhibitor, bortezomib, has been demonstrated to sensitize tumor cells to tumor necrosis factor-related apoptosis-inducing ligand （TRAIL）-mediated apoptosis. Natural killer （NK） cells represent poten...The proteasome inhibitor, bortezomib, has been demonstrated to sensitize tumor cells to tumor necrosis factor-related apoptosis-inducing ligand （TRAIL）-mediated apoptosis. Natural killer （NK） cells represent potent antitumor effector cells. They also express TRAIL. Therefore, we investigated whether bortezomib could sensitize tumor cells to NK cell-mediated killing, and have the same effect in human prostate cancer cell lines （LNCaP and DU145）. We found that bortezomib strongly inhibits proliferation in both cell lines. Furthermore, compared with LNCaP cells, DU145 cells are more sensitive to bortezomib-induced apoptosis. However, bortezomib is unable to sensitize these two cell lines to NK cell-mediated killing in short-term assays. In long-term assays, we found that killing mediated by activated NK cells following bortezomib treatment leads to greater antitumor effects than either treatment alone. In addition, treatment with bortezomib causes these cells to upregulate apoptosis-related mRNA as well as death receptors and downregulate the major histocompatibility class （MHC）-I molecule on the cell surface of DU145 cells. In contrast, LNCaP cells are not sensitized by this treatment. Death receptors and the MHC-I molecule did not change in this cell line. These data suggest that bortezomib can be used to sensitize prostate cancer cells to NK cell-mediated killing and improve current cancer therapies. This theral）eutic stratelzv may be more effective in I）atients with androeen-insensitive orostate cancer.展开更多
目的探讨汉防己甲素(Tet)联合阿霉素(DOX)体外逆转耐药型前列腺癌(DU145/DOX)细胞耐药的可行性及潜在机制。方法采用经典的DOX浓度梯度诱导法制备DU145/DOX细胞,然后采用3-(4,5-二甲基噻唑-2)-2,5-二苯基四氮唑溴盐(MTT)法测定Tet干预对...目的探讨汉防己甲素(Tet)联合阿霉素(DOX)体外逆转耐药型前列腺癌(DU145/DOX)细胞耐药的可行性及潜在机制。方法采用经典的DOX浓度梯度诱导法制备DU145/DOX细胞,然后采用3-(4,5-二甲基噻唑-2)-2,5-二苯基四氮唑溴盐(MTT)法测定Tet干预对DU145/DOX细胞生长抑制的作用,凋亡试剂盒测定联合用药对耐药细胞凋亡诱导的影响,流式细胞仪和荧光分光光度计分别测定Tet对DOX和荧光探针罗丹明123(R123)被DU145/DOX细胞摄取行为,Pgp-Glo TM分析系统测定Tet对P-糖蛋白(P-gp)ATP酶活性的影响,Western blot法测定Tet对耐药细胞膜上P-gp蛋白表达的影响,RT-PCR法测定Tet对DU145/DOX细胞MDR1 m RNA表达的影响。结果 Tet组、DOX组、DOX+Tet(1/5)组对DU145/DOX细胞的半数抑制浓度(IC50,以COX浓度计算)分别为(7.16±0.54)、(1.62±0.09)、(0.37±0.03)μM,Tet能明显增强DOX的抗肿瘤作用。DOX+Tet(1/5)组诱导DU145/DOX细胞凋亡率是DOX组的2.1倍。Tet的参与能明显提高DU145/DOX细胞内DOX和R123的累积量,其机制可能与Tet刺激细胞内P-gp ATP酶活性、降低细胞膜上P-gp蛋白表达有关。Tet可能通过降低耐药基因表达的机制,联合DOX提高对DU145/DOX细胞杀伤作用。结论 Tet与DOX体外联合使用具有逆转DU145/DOX细胞耐药的潜力,作用机制可能涉及刺激耐药泵酶活性、降低耐药蛋白表达量、下调MDR1 m RNA表达等。展开更多
文摘目的:观察白花蛇舌草提取物对人前列腺癌DU145细胞增殖和凋亡的影响,并初步探讨其作用机制。方法:不同浓度(0.25 g·L-1、0.5 g·L-1、1 g·L-1)白花蛇舌草提取物作用人前列腺癌DU145细胞24 h、48 h、72 h后,使用CCK-8法检测白花蛇舌草提取物对DU145细胞增殖的影响;流式细胞术检测细胞凋亡的改变;实时定量PCR检测COX-2 m RNA和PCNA m RNA的表达;Western bloting法检测Bcl-2、Bax、Caspase3、Cyclin D1蛋白表达。结果:随着作用时间的延长和剂量的增加,白花蛇舌草提取物对DU145细胞增殖抑制率明显提高,呈时效和量效依赖效应。白花蛇舌草提取物作用48 h后,可明显提高细胞凋亡率,下调DU145细胞COX-2 m RNA、PCNA m RNA及Bcl-2、Cyclin D1蛋白表达,上调Bax、Caspase3蛋白表达,呈量效依赖效应。结论:白花蛇舌草提取物能降低人前列腺癌DU145细胞增殖能力,并诱导其凋亡,其机制可能与改变细胞周期和影响凋亡相关基因表达有关。
文摘We investigated the ability of NK4, an antagonist of human hepatocyte growth factor （HGF）, to inhibit the influence of HGF on proliferation, migration, invasion and apoptosis of human prostate cancer cells. Expression vector pBudCE4.1-EGFP-NK4 containing NK4 cDNA was used to transfect human prostate cancer DU145 cells, and the effects of the autocrine NK4 on tumor cell proliferation, migration, invasion and apoptosis were assessed in vitro. in vivo, we subcutaneously implanted DU145 cells, mock-transfected clone （DU145/empty vector） cells and NK4- transfected clone （DU145/NK4） cells into nude mice, and then evaluated tumor growth, cell proliferation and cell apoptosis in vivo. We found that DU145/NK4 cells expressed NK4 protein. In the in vitro study, autocrine NK4 at- tenuated the HGF-induced tumor cell proliferation, migration and invasion, and stimulated apoptosis. Furthermore, autocrine NK4 effectively inhibited the HGF-induced phosphorylation of c-Met, extracellular signal-regulated kinase-1 （ERK1）. and protein kinase B 1/2 （Aktl/2）. Histological examination revealed that autocrine NK4 inhibited prolifera- tion and accelerated apoptosis of prostate cancer cells. These results show that genetic modification of DU145 cells with NK4 cDNA yields a significant effect on their proliferation, migration, invasion and apoptosis. Molecular targeting of HGF/c-Met by NK4 could be applied as a novel therapeutic approach to prostate cancer.
文摘The proteasome inhibitor, bortezomib, has been demonstrated to sensitize tumor cells to tumor necrosis factor-related apoptosis-inducing ligand （TRAIL）-mediated apoptosis. Natural killer （NK） cells represent potent antitumor effector cells. They also express TRAIL. Therefore, we investigated whether bortezomib could sensitize tumor cells to NK cell-mediated killing, and have the same effect in human prostate cancer cell lines （LNCaP and DU145）. We found that bortezomib strongly inhibits proliferation in both cell lines. Furthermore, compared with LNCaP cells, DU145 cells are more sensitive to bortezomib-induced apoptosis. However, bortezomib is unable to sensitize these two cell lines to NK cell-mediated killing in short-term assays. In long-term assays, we found that killing mediated by activated NK cells following bortezomib treatment leads to greater antitumor effects than either treatment alone. In addition, treatment with bortezomib causes these cells to upregulate apoptosis-related mRNA as well as death receptors and downregulate the major histocompatibility class （MHC）-I molecule on the cell surface of DU145 cells. In contrast, LNCaP cells are not sensitized by this treatment. Death receptors and the MHC-I molecule did not change in this cell line. These data suggest that bortezomib can be used to sensitize prostate cancer cells to NK cell-mediated killing and improve current cancer therapies. This theral）eutic stratelzv may be more effective in I）atients with androeen-insensitive orostate cancer.
文摘目的探讨汉防己甲素(Tet)联合阿霉素(DOX)体外逆转耐药型前列腺癌(DU145/DOX)细胞耐药的可行性及潜在机制。方法采用经典的DOX浓度梯度诱导法制备DU145/DOX细胞,然后采用3-(4,5-二甲基噻唑-2)-2,5-二苯基四氮唑溴盐(MTT)法测定Tet干预对DU145/DOX细胞生长抑制的作用,凋亡试剂盒测定联合用药对耐药细胞凋亡诱导的影响,流式细胞仪和荧光分光光度计分别测定Tet对DOX和荧光探针罗丹明123(R123)被DU145/DOX细胞摄取行为,Pgp-Glo TM分析系统测定Tet对P-糖蛋白(P-gp)ATP酶活性的影响,Western blot法测定Tet对耐药细胞膜上P-gp蛋白表达的影响,RT-PCR法测定Tet对DU145/DOX细胞MDR1 m RNA表达的影响。结果 Tet组、DOX组、DOX+Tet(1/5)组对DU145/DOX细胞的半数抑制浓度(IC50,以COX浓度计算)分别为(7.16±0.54)、(1.62±0.09)、(0.37±0.03)μM,Tet能明显增强DOX的抗肿瘤作用。DOX+Tet(1/5)组诱导DU145/DOX细胞凋亡率是DOX组的2.1倍。Tet的参与能明显提高DU145/DOX细胞内DOX和R123的累积量,其机制可能与Tet刺激细胞内P-gp ATP酶活性、降低细胞膜上P-gp蛋白表达有关。Tet可能通过降低耐药基因表达的机制,联合DOX提高对DU145/DOX细胞杀伤作用。结论 Tet与DOX体外联合使用具有逆转DU145/DOX细胞耐药的潜力,作用机制可能涉及刺激耐药泵酶活性、降低耐药蛋白表达量、下调MDR1 m RNA表达等。
