It is well-known that risk for endometrial adenocarcinoma increases in patients with high level ofestrogen that is unopposed by progestin. And activation of extracellular signal-regulated kinase （ERK） and phosphatid...It is well-known that risk for endometrial adenocarcinoma increases in patients with high level ofestrogen that is unopposed by progestin. And activation of extracellular signal-regulated kinase （ERK） and phosphatidylinositol 3 kinase/protein kinase B （PI3K/PKB） pathway are responsible for hormone-dependent cell growth in endometrial carcinoma. PI3K produces phosphatidylinositol- 3-phosphates by phosphory-lating the D3 hydroxyl of phosphoinositides, leading to membrane translocation of PKB,展开更多
Background Endometrial carcinoma is one of the most common female tract genital malignant tumors. Nifedipine, an L-type calcium channel antagonist can inhibit cell proliferation of carcinomas. Recent studies indicated...Background Endometrial carcinoma is one of the most common female tract genital malignant tumors. Nifedipine, an L-type calcium channel antagonist can inhibit cell proliferation of carcinomas. Recent studies indicated that a rise in the free cytosolic calcium （[Ca2±]c） was a potent inducer of autophagy. Here, we investigated the relationship between nifedipine and autophagy in Hec-IA cells. Methods Cells were cultured with nifedipine （10 μmol/L） and harvested at different times for counting cell number. MTT assay was applied to evaluate the cell viability and transwell assay to reveal cell migration. Apoptotic cells were detected with annexin V/PI assay. Then cells were treated with 3-methyladenine （3-MA） （2.5 mmol/L） for 0, 5, 15, 30, 60, and 120 minutes and the expression of the L-type calcium channel alphalD （Cavl.3） protein was detected. At last, cells were cultured and assigned to four groups with different treatment： untreated （control group）, 10 μmol/L nifedipine （N group）, 2.5 mmol/L 3-MA （3-MA group）, and 10 μmol/L nifedipine plus 2.5 mmol/L 3-MA （N±3MA group）. Autophagy was detected with GFP-LC3 modulation by fluorescent microscopy, and expression of the autophagy-associated proteins （LC3, Beclinl and P70s6K） by Western blotting and monodansylcadaverine （MDC） labeled visualization. Results Proliferation of Hec-lA cells was obviously suppressed by nifedipine compared with that of the untreated cells for 24, 48, and 96 hours （P=0.000 for each day）. The suppression of migration ability of the nifedipine-treated cells （94.0±8.2） was significantly different from that of the untreated cells （160.00±9.50, P=0.021 ）. The level of early period cell apoptosis induced by nifedipine was （2.21_±0.19）%, which was （2.90±0.13）% in control group （P=-0.052）, whereas the late period apoptosis level reached （10.38_±0.96）% and （4.40_±0.60）% （P=0.020）, respectively. The 3-MA group induced a slight increase in the Cavl.3 levels within 15 minutes, but significantly attenuated the Cavl.3 levels after 30 minutes. There were more autophagic vacuoles labeled by MDC in the N group （20.63_±3.36） than the control group （6.29_±0.16, P=-0.015）. GFP-LC3 localization revealed that the LC3 levels of cells in 3-MA group, N±3MA group, 3-MA group were 2.80_±0.29, 2.30_±0.17, and 1.80±0.21, respectively. Cells in the N group showed significant augmentation of autophagy （P 〈0.05）. Western blotting analysis confirmed the down-regulation of LC3 levels in 3-MA group （0.85±0.21） and N±3MA group （1.21±0.12） compared with nifedipine treatment （2.64±0.15, P 〈0.05）. The annexin-V-FITC/PI assay showed that the level of early period cell apoptosis induced in the N＋3-MA group （（11.22±0.91）%） differed significantly from that of the control group （（2.51±0.70）%） and N group （（3.47±0.39）%）. Similarly, the late period level of the N＋3-MA group （（55.19±2.51）%） differed significantly from that of the control group（（15.81±1.36）%） and the N group （（22.09±2.48）%, P 〈0.05）. The down-regulated expression of P70s6k and up-regulated expression of the Beclinl revealed significant differences between the N＋3-MA group and control group （P=0.025; Beclinl： P=-0.015）. Conclusions Proliferation and migration in vitro of endometrial carcinoma Hec-lA cells are significantly suppressed by nifedipine. The nifedipine leads autophagy to oppose Hec-lA cells apoptosis. Autophagy inhibition by 3-MA leads down-regulation of Cavl.3 and enhances nifedipine-induced cell death. The nifedipine-induced autophagy is linked to Beclinl and mTOR pathways.展开更多
Background Histone deacetylase （HDAC） inhibitors are a group of small chemical molecules that inhibit histone deacetylase. At cell level, HDAC inhibitors have multiple biological effects such as cell cycle arrest, a...Background Histone deacetylase （HDAC） inhibitors are a group of small chemical molecules that inhibit histone deacetylase. At cell level, HDAC inhibitors have multiple biological effects such as cell cycle arrest, apoptosis, cell differentiation and auotophagy. At molecular level, HDAC inhibitors cause histone and nonhistone acetylation and induce gene expression. HDAC inhibitors are widely used in cancer therapy because of its function of inducing apoptosis. However, the mechanisms of apoptosis effect are not fully understood. TSA is a classical HDAC inhibitor and widely used in epigenetic and anti-cancer research. In this study, we selected Trichostatin A （TSA） to investigate the mechanisms of HDAC inhibitors apoptotic effect on cancer cells. Methods Cervical cancer cell lines such as Hela, Caski and normal human keratinocyte line HaCaT were treated with various concentrations of TSA. Crystal violent assay and 3-（4,5-Dimethylthiazol-2-yl）-2,5-diphenyltetrazolium bromide （MTT） assay were performed to determine cell number. PARP cleavage and FITC-AnexinV were performed to determine apoptosis. DNA-methyltransferase （DNMT）I, DNMT3A and DNMT3B were determined by regular PCR, qPCR and Western Blotting. Small interfering RNA （SiRNAi） was used to knock down DNMT3B. Results HDAC inhibitors only induce cervical cancer cell apoptosis. At 1 Iumol/L of TSA, 86% of Hela cell and 76% of Caski went apoptosis. For normal cells, HDAC inhibitors have no cytotoxic effect at therapeutic dosage, （90.0＋8.4）% of normal cell survive after treated with 1 IJmol/L of TSA. We compared 1 pmol/L group with untreated control with t-test. There was no significance between 1 pmol/L group and untreated control for normal cell （P 〉0.05）. HDAC inhibitors decreased DNMT3B in cancer cell but not in normal cell. Manually knock-down of DNMT3B induced Hela and Caski cell apoptosis. More than 99% of Hela and Caski cell went apoptosis after deprived of DNMT3B. Conclusions DNMT3B was essential to cervical cancer cell survival. Down-regulated DNMT3B by HDAC inhibitors may play an important role in the toxicity of HDAC inhibitors on cervical cancer cells.展开更多
Background Phenotypic and genotypic heterogeneity is a known feature of many cancers.Whether serum tumor marker kinds vary and change following chemotherapy is still unclear.The aim of this study was to investigate wh...Background Phenotypic and genotypic heterogeneity is a known feature of many cancers.Whether serum tumor marker kinds vary and change following chemotherapy is still unclear.The aim of this study was to investigate whether there is a change in the expression of serum tumor markers following chemotherapy,and the potential clinical significance in patients with epithelial ovarian carcinoma （EOC） or primary serous peritoneal carcinoma （PSPC）.Methods Samples were collected before surgery,during chemotherapy and during follow-up for enzyme-linked immunosorbent assay （ELISA）-based evaluation of serum CA-125,CA19-9 and CP2 levels in patients with EOC or PSPC who had received primary debulking surgery followed by adjuvant chemotherapy.In total,72 patients were examined,including 37 patients with recurrent lesions and 35 patients receiving first-line chemotherapy.Results In 35 de novo patients,20% （7/35） demonstrated a significant changed serum tumor marker kinds among whom the patients with mucinous carcinoma （57.1%,4/7） showed resistance to chemotherapy.In the 37 recurrent patients,51.4% （19/37） had changed serum tumor markers,of whom 57.9% （11/19） presented with serous carcinoma.There was no significant difference in median progression-free survival or overall survival in patients with drug-sensitive or drug-resistant recurrence in patients with changed tumor marker kinds relative to those with unchanged markers.However,for patients with changed serum tumor markers there was a trend towards prolonged survival compared with the unchanged serum tumor marker group.In the 17 patients with secondary recurrence,37.5% （6/17） had changed tumor marker levels.The ratios of CA-125/CP2 and CA-125/CA19-9 were significantly different after either chemotherapy or recurrence.Conclusions Serum tumor marker expression in patients with EOC or PSPC may change after chemotherapy or recurrence,indicating that in addition to the markers that are abnormal before surgery,those markers that are normalshould also be monitored during chemotherapy and follow-up.展开更多
基金This study was partially supported by a grant from the Scientific Research Fund for Capital Medicine Development (No.ZD 199911).
文摘It is well-known that risk for endometrial adenocarcinoma increases in patients with high level ofestrogen that is unopposed by progestin. And activation of extracellular signal-regulated kinase （ERK） and phosphatidylinositol 3 kinase/protein kinase B （PI3K/PKB） pathway are responsible for hormone-dependent cell growth in endometrial carcinoma. PI3K produces phosphatidylinositol- 3-phosphates by phosphory-lating the D3 hydroxyl of phosphoinositides, leading to membrane translocation of PKB,
基金This study was supported by a grant from the National Natural Science Foundation of China （No. 30973182）.
文摘Background Endometrial carcinoma is one of the most common female tract genital malignant tumors. Nifedipine, an L-type calcium channel antagonist can inhibit cell proliferation of carcinomas. Recent studies indicated that a rise in the free cytosolic calcium （[Ca2±]c） was a potent inducer of autophagy. Here, we investigated the relationship between nifedipine and autophagy in Hec-IA cells. Methods Cells were cultured with nifedipine （10 μmol/L） and harvested at different times for counting cell number. MTT assay was applied to evaluate the cell viability and transwell assay to reveal cell migration. Apoptotic cells were detected with annexin V/PI assay. Then cells were treated with 3-methyladenine （3-MA） （2.5 mmol/L） for 0, 5, 15, 30, 60, and 120 minutes and the expression of the L-type calcium channel alphalD （Cavl.3） protein was detected. At last, cells were cultured and assigned to four groups with different treatment： untreated （control group）, 10 μmol/L nifedipine （N group）, 2.5 mmol/L 3-MA （3-MA group）, and 10 μmol/L nifedipine plus 2.5 mmol/L 3-MA （N±3MA group）. Autophagy was detected with GFP-LC3 modulation by fluorescent microscopy, and expression of the autophagy-associated proteins （LC3, Beclinl and P70s6K） by Western blotting and monodansylcadaverine （MDC） labeled visualization. Results Proliferation of Hec-lA cells was obviously suppressed by nifedipine compared with that of the untreated cells for 24, 48, and 96 hours （P=0.000 for each day）. The suppression of migration ability of the nifedipine-treated cells （94.0±8.2） was significantly different from that of the untreated cells （160.00±9.50, P=0.021 ）. The level of early period cell apoptosis induced by nifedipine was （2.21_±0.19）%, which was （2.90±0.13）% in control group （P=-0.052）, whereas the late period apoptosis level reached （10.38_±0.96）% and （4.40_±0.60）% （P=0.020）, respectively. The 3-MA group induced a slight increase in the Cavl.3 levels within 15 minutes, but significantly attenuated the Cavl.3 levels after 30 minutes. There were more autophagic vacuoles labeled by MDC in the N group （20.63_±3.36） than the control group （6.29_±0.16, P=-0.015）. GFP-LC3 localization revealed that the LC3 levels of cells in 3-MA group, N±3MA group, 3-MA group were 2.80_±0.29, 2.30_±0.17, and 1.80±0.21, respectively. Cells in the N group showed significant augmentation of autophagy （P 〈0.05）. Western blotting analysis confirmed the down-regulation of LC3 levels in 3-MA group （0.85±0.21） and N±3MA group （1.21±0.12） compared with nifedipine treatment （2.64±0.15, P 〈0.05）. The annexin-V-FITC/PI assay showed that the level of early period cell apoptosis induced in the N＋3-MA group （（11.22±0.91）%） differed significantly from that of the control group （（2.51±0.70）%） and N group （（3.47±0.39）%）. Similarly, the late period level of the N＋3-MA group （（55.19±2.51）%） differed significantly from that of the control group（（15.81±1.36）%） and the N group （（22.09±2.48）%, P 〈0.05）. The down-regulated expression of P70s6k and up-regulated expression of the Beclinl revealed significant differences between the N＋3-MA group and control group （P=0.025; Beclinl： P=-0.015）. Conclusions Proliferation and migration in vitro of endometrial carcinoma Hec-lA cells are significantly suppressed by nifedipine. The nifedipine leads autophagy to oppose Hec-lA cells apoptosis. Autophagy inhibition by 3-MA leads down-regulation of Cavl.3 and enhances nifedipine-induced cell death. The nifedipine-induced autophagy is linked to Beclinl and mTOR pathways.
基金This article was supported by the grants from the National Natural Science Foundation of China，the “985” Project of the Peking University Health Science Center and the Scientific Research Foundation for the Returned Overseas Chinese Scholars,State Education Ministry
文摘Background Histone deacetylase （HDAC） inhibitors are a group of small chemical molecules that inhibit histone deacetylase. At cell level, HDAC inhibitors have multiple biological effects such as cell cycle arrest, apoptosis, cell differentiation and auotophagy. At molecular level, HDAC inhibitors cause histone and nonhistone acetylation and induce gene expression. HDAC inhibitors are widely used in cancer therapy because of its function of inducing apoptosis. However, the mechanisms of apoptosis effect are not fully understood. TSA is a classical HDAC inhibitor and widely used in epigenetic and anti-cancer research. In this study, we selected Trichostatin A （TSA） to investigate the mechanisms of HDAC inhibitors apoptotic effect on cancer cells. Methods Cervical cancer cell lines such as Hela, Caski and normal human keratinocyte line HaCaT were treated with various concentrations of TSA. Crystal violent assay and 3-（4,5-Dimethylthiazol-2-yl）-2,5-diphenyltetrazolium bromide （MTT） assay were performed to determine cell number. PARP cleavage and FITC-AnexinV were performed to determine apoptosis. DNA-methyltransferase （DNMT）I, DNMT3A and DNMT3B were determined by regular PCR, qPCR and Western Blotting. Small interfering RNA （SiRNAi） was used to knock down DNMT3B. Results HDAC inhibitors only induce cervical cancer cell apoptosis. At 1 Iumol/L of TSA, 86% of Hela cell and 76% of Caski went apoptosis. For normal cells, HDAC inhibitors have no cytotoxic effect at therapeutic dosage, （90.0＋8.4）% of normal cell survive after treated with 1 IJmol/L of TSA. We compared 1 pmol/L group with untreated control with t-test. There was no significance between 1 pmol/L group and untreated control for normal cell （P 〉0.05）. HDAC inhibitors decreased DNMT3B in cancer cell but not in normal cell. Manually knock-down of DNMT3B induced Hela and Caski cell apoptosis. More than 99% of Hela and Caski cell went apoptosis after deprived of DNMT3B. Conclusions DNMT3B was essential to cervical cancer cell survival. Down-regulated DNMT3B by HDAC inhibitors may play an important role in the toxicity of HDAC inhibitors on cervical cancer cells.
文摘Background Phenotypic and genotypic heterogeneity is a known feature of many cancers.Whether serum tumor marker kinds vary and change following chemotherapy is still unclear.The aim of this study was to investigate whether there is a change in the expression of serum tumor markers following chemotherapy,and the potential clinical significance in patients with epithelial ovarian carcinoma （EOC） or primary serous peritoneal carcinoma （PSPC）.Methods Samples were collected before surgery,during chemotherapy and during follow-up for enzyme-linked immunosorbent assay （ELISA）-based evaluation of serum CA-125,CA19-9 and CP2 levels in patients with EOC or PSPC who had received primary debulking surgery followed by adjuvant chemotherapy.In total,72 patients were examined,including 37 patients with recurrent lesions and 35 patients receiving first-line chemotherapy.Results In 35 de novo patients,20% （7/35） demonstrated a significant changed serum tumor marker kinds among whom the patients with mucinous carcinoma （57.1%,4/7） showed resistance to chemotherapy.In the 37 recurrent patients,51.4% （19/37） had changed serum tumor markers,of whom 57.9% （11/19） presented with serous carcinoma.There was no significant difference in median progression-free survival or overall survival in patients with drug-sensitive or drug-resistant recurrence in patients with changed tumor marker kinds relative to those with unchanged markers.However,for patients with changed serum tumor markers there was a trend towards prolonged survival compared with the unchanged serum tumor marker group.In the 17 patients with secondary recurrence,37.5% （6/17） had changed tumor marker levels.The ratios of CA-125/CP2 and CA-125/CA19-9 were significantly different after either chemotherapy or recurrence.Conclusions Serum tumor marker expression in patients with EOC or PSPC may change after chemotherapy or recurrence,indicating that in addition to the markers that are abnormal before surgery,those markers that are normalshould also be monitored during chemotherapy and follow-up.
