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Volume 20,Issue 6,2013 Table of Contents
2013, 20(6):631-636.
DOI: 10.3872/j.issn.1007-385X.2013.06.001
Abstract:
Alternative splicing of pre-mRNA is an important mechanism of the proteomic diversity, and is closely related to the cancer development. Serine/arginine-rich splicing factor 1 (SRSF1), a typical member of alternative splicing family, is involved in diverse events including alternative splicing and processing, intracellur location and transportation of precursor mRNA. SRSF1 has been identified as an oncoprotein and is up-regualted in many cancers, including those of the lung, breast, as well as in leukemia. SRSF1 contributes to tumorigenesis and development by interacting with cancer-related genes, regulating cell cycle and apoptosis and other cell processes. Additionally, SRSF1 plays an oncogenic role through regulating the splicing of genes involved in several signaling pathways including PI3K-Akt-mTOR, Ras-MNK-MAPK and Wnt-β-catenin. For splicing disorders, the current treatment is to correct the missplicing of genes with antisense oligonucleotides, which has been widely applied in some solid tumors such as lung cancer and breast cancer. On this basis, further exploration in site selection and mechanism of alternative splcing will elucidate tumorigenesis and improve the diagnosis and treatment of cancer.
Alternative splicing of pre-mRNA is an important mechanism of the proteomic diversity, and is closely related to the cancer development. Serine/arginine-rich splicing factor 1 (SRSF1), a typical member of alternative splicing family, is involved in diverse events including alternative splicing and processing, intracellur location and transportation of precursor mRNA. SRSF1 has been identified as an oncoprotein and is up-regualted in many cancers, including those of the lung, breast, as well as in leukemia. SRSF1 contributes to tumorigenesis and development by interacting with cancer-related genes, regulating cell cycle and apoptosis and other cell processes. Additionally, SRSF1 plays an oncogenic role through regulating the splicing of genes involved in several signaling pathways including PI3K-Akt-mTOR, Ras-MNK-MAPK and Wnt-β-catenin. For splicing disorders, the current treatment is to correct the missplicing of genes with antisense oligonucleotides, which has been widely applied in some solid tumors such as lung cancer and breast cancer. On this basis, further exploration in site selection and mechanism of alternative splcing will elucidate tumorigenesis and improve the diagnosis and treatment of cancer.
2013, 20(6):637-644.
DOI: 10.3872/j.issn.1007-385X.2013.06.002
Abstract:
Objective:To construct a recombinant adenovirus Ad5/F35 containing human melanoma-associated antigen 3 (MAGE-A3), and to observe the effect of adenovirus-mediated MAGE-A3 overexpression on the maturation and apoptosis of dendritic cells (DCs) in patients with melanoma. Methods: To choose the adenoviral vectors with the highest transfection efficiency, and then to construct recombinant adenovirus vectors and packaging adenovirus particles Ad5/F35-MAGE-A3. Immunohistochemistry and Western blotting were performed to detect the influence of Ad5/F35-MAGE-A3 infection on the expression of MAGE-A3 of DCs in healthy people and patients with kidney cancer or melanoma. Flow cytometry was used to detect the effect of Ad5/F35-MAGE-A3 infection on the maturation and apoptosis of DCs in patients with melanoma. Results: The recombinant adenovirus vector containing human MAGE-A3 was successfully constructed and adenovirus particles Ad5/F35-MAGE-A3 were packaged with the infective titer of 7.94×108 IU/ml. Ad5/F35-MAGE-A3 infection improved the MAGE-A3 expression of DCs in healthy people and kidney cancer patients (P<0.05), and it did not affect MAGE-A3 expression of DCs in melanoma patients (\[0.3352±0.1272\] vs \[0.4672±0.0704\], P＞0.05). After Ad5/F35-MAGE-A3 infection, the co-stimulatory molecule CD80 (\[20.42±0.58\]% vs \[10.22±104\]%, \[895±0.2\]%), CD86 (\[85.3±3.98\]% vs \[39.85±2.86\]%, \[34.1±4.32\]%) and HLA-DR (\[8687±436\]% vs \[63.68±3.15\]%, \[60.69±4.81\]%) which expressed on the surface of DCs was significantly higher than that of the negative control group and the blank control group (all P<0.05), but no significant difference existed in apoptosis rate (\[1.18±0.09\]% vs \[1.09±0.11\]%, P>0.05).Conclusion: Recombinant adenovirus vector can efficiently affect DCs. Ad5/F35-MAGE-A3 infection may not affect the expression of MAGE-A3 of DCs in melanoma patients, and promote the maturation of DCs without obvious cytotoxicity.
Objective:To construct a recombinant adenovirus Ad5/F35 containing human melanoma-associated antigen 3 (MAGE-A3), and to observe the effect of adenovirus-mediated MAGE-A3 overexpression on the maturation and apoptosis of dendritic cells (DCs) in patients with melanoma. Methods: To choose the adenoviral vectors with the highest transfection efficiency, and then to construct recombinant adenovirus vectors and packaging adenovirus particles Ad5/F35-MAGE-A3. Immunohistochemistry and Western blotting were performed to detect the influence of Ad5/F35-MAGE-A3 infection on the expression of MAGE-A3 of DCs in healthy people and patients with kidney cancer or melanoma. Flow cytometry was used to detect the effect of Ad5/F35-MAGE-A3 infection on the maturation and apoptosis of DCs in patients with melanoma. Results: The recombinant adenovirus vector containing human MAGE-A3 was successfully constructed and adenovirus particles Ad5/F35-MAGE-A3 were packaged with the infective titer of 7.94×108 IU/ml. Ad5/F35-MAGE-A3 infection improved the MAGE-A3 expression of DCs in healthy people and kidney cancer patients (P<0.05), and it did not affect MAGE-A3 expression of DCs in melanoma patients (\[0.3352±0.1272\] vs \[0.4672±0.0704\], P＞0.05). After Ad5/F35-MAGE-A3 infection, the co-stimulatory molecule CD80 (\[20.42±0.58\]% vs \[10.22±104\]%, \[895±0.2\]%), CD86 (\[85.3±3.98\]% vs \[39.85±2.86\]%, \[34.1±4.32\]%) and HLA-DR (\[8687±436\]% vs \[63.68±3.15\]%, \[60.69±4.81\]%) which expressed on the surface of DCs was significantly higher than that of the negative control group and the blank control group (all P<0.05), but no significant difference existed in apoptosis rate (\[1.18±0.09\]% vs \[1.09±0.11\]%, P>0.05).Conclusion: Recombinant adenovirus vector can efficiently affect DCs. Ad5/F35-MAGE-A3 infection may not affect the expression of MAGE-A3 of DCs in melanoma patients, and promote the maturation of DCs without obvious cytotoxicity.
Wu Quanrui
,
Zhao Yongqiang
,
Chu Datong
,
Xu Binghe
,
Liao Meilin
,
Jiang Liyan
,
Xu Jianmin
,
Wang Huaying
,
Li Jin
,
Hou Mei
,
Zhou Qinghua
,
Zhang Lijian
,
Zhang Shucai
,
Xia Zhongjun
,
Jiang Wenqi
,
Lv Yue
,
Zhai Ming
,
Meng Fanyi
,
Wang Dongxing
,
Wang Jianmin
,
Chen Zhengtang
,
Guan Huajun
,
Wang Qingyu
,
Chen Xiequn
,
Liu Jiwei
,
Zhang Yang
,
Song Shanjun
,
Liu Wenli
,
Yu Shiying
,
Xu Jianming
,
Song Shuping
,
Xu Jian
,
Li Liqing
,
Zhang Mei
,
Sun Hong
,
Jiang Bin
2013, 20(6):645-653.
DOI: 10.3872/j.issn.1007-385X.2013.06.003
Abstract:
Objective:To assess the effectiveness and safety of recombinant human thrombopoietin(rh-TP0) on chemotherapy-induced thrombocytopenia(CIT) in patients with solid tumor. Methods: A total of 276 cases were enrolled in 3 studies, 63 cases in Phase Ⅱ study, 154 cases in Phase Ⅲ study and 59 cases in complement study. A total of 230 cases were enrolled pre-protocol (PP) group, except of 5 protocol deviations, 41 lost-follow-ups. All of the patients had solid tumor which was confirmed by histology or cytology. Phase Ⅱ and complement trials were randomized crossover self-control trials. It was divided into randomized-crossover-self-control part and non-randomized-crossover-self-control part in Phase Ⅲ trial depending on if the subject was joined by randomized-crossover. The period when rh-TPO subject was administered was defined as an administration period, and the other period was defined as a control period. The chemical therapeutic regimen and dosage did not change during the trials. All of the trials’ data were pooled together to compare the efficacy and safety profile.Results:They showed the utilized changes both in intention-to-treat (ITT) group and PP group when compared with the treated cycle and control cycle (following example with ITT group data). Compared with the control period, rh-TPO could significantly decrease the degree of CIT (minimal mean platelet count: (\[63.02±46.48\]×109 vs \[49.47±31.41\]×109/L, P=0002); shorten the thrombocytopenia duration induced by chemotherapy(days of platelet count recovered to 75×109/L：\[11.18±9.71\] vs \[17.8±10.46\] d, P=0.000); and highly increase recover level of platelet count (platelet count of last visit: \[211.21±119.20\]×109 vs \[138.13±71.54\]×109/L, P=0.000; maximal mean platelet count: \[262.78±162.60\]×109 vs \[149.36±73.26\]×109/L, P=0.000; platelet count’s difference between the last visit and baseline: \[79.64±118.06\] vs \[-8.92±102.50\], P=0.000). On the other hand, rh-TPO could decrease the ratio of patients who had to be transfused with platelet (12.21% vs 19.85%,P=0017); and decrease the time (\[022±0.72\] vs \[0.37±0.90\] time, P=0.010) and the quantity (\[1.66±6.09\] vs \[2.77±7.08\] U, P=0.009) of platelet transfusion. It was much more significantly in the complement trial (1379% vs 33.93%, P=00082). There were no significant differences before and after treatment on Hb, WBC, liver and renal function and blood coagulation (P＞0.05).There were only 11 cases who met side effects during the studies, most of whom presented fever (6 cases) and chill (2 cases). Conclusion: Administration of rh-TPO after chemotherapycan may significantly reduce the degree and duration of thrombocytopenia induced by chemotherapy and promote platelet recovery of patients with solid tumors. It also decreased the capability of platelet transfusion, and severe side effects were not found．
Objective:To assess the effectiveness and safety of recombinant human thrombopoietin(rh-TP0) on chemotherapy-induced thrombocytopenia(CIT) in patients with solid tumor. Methods: A total of 276 cases were enrolled in 3 studies, 63 cases in Phase Ⅱ study, 154 cases in Phase Ⅲ study and 59 cases in complement study. A total of 230 cases were enrolled pre-protocol (PP) group, except of 5 protocol deviations, 41 lost-follow-ups. All of the patients had solid tumor which was confirmed by histology or cytology. Phase Ⅱ and complement trials were randomized crossover self-control trials. It was divided into randomized-crossover-self-control part and non-randomized-crossover-self-control part in Phase Ⅲ trial depending on if the subject was joined by randomized-crossover. The period when rh-TPO subject was administered was defined as an administration period, and the other period was defined as a control period. The chemical therapeutic regimen and dosage did not change during the trials. All of the trials’ data were pooled together to compare the efficacy and safety profile.Results:They showed the utilized changes both in intention-to-treat (ITT) group and PP group when compared with the treated cycle and control cycle (following example with ITT group data). Compared with the control period, rh-TPO could significantly decrease the degree of CIT (minimal mean platelet count: (\[63.02±46.48\]×109 vs \[49.47±31.41\]×109/L, P=0002); shorten the thrombocytopenia duration induced by chemotherapy(days of platelet count recovered to 75×109/L：\[11.18±9.71\] vs \[17.8±10.46\] d, P=0.000); and highly increase recover level of platelet count (platelet count of last visit: \[211.21±119.20\]×109 vs \[138.13±71.54\]×109/L, P=0.000; maximal mean platelet count: \[262.78±162.60\]×109 vs \[149.36±73.26\]×109/L, P=0.000; platelet count’s difference between the last visit and baseline: \[79.64±118.06\] vs \[-8.92±102.50\], P=0.000). On the other hand, rh-TPO could decrease the ratio of patients who had to be transfused with platelet (12.21% vs 19.85%,P=0017); and decrease the time (\[022±0.72\] vs \[0.37±0.90\] time, P=0.010) and the quantity (\[1.66±6.09\] vs \[2.77±7.08\] U, P=0.009) of platelet transfusion. It was much more significantly in the complement trial (1379% vs 33.93%, P=00082). There were no significant differences before and after treatment on Hb, WBC, liver and renal function and blood coagulation (P＞0.05).There were only 11 cases who met side effects during the studies, most of whom presented fever (6 cases) and chill (2 cases). Conclusion: Administration of rh-TPO after chemotherapycan may significantly reduce the degree and duration of thrombocytopenia induced by chemotherapy and promote platelet recovery of patients with solid tumors. It also decreased the capability of platelet transfusion, and severe side effects were not found．
2013, 20(6):654-660.
DOI: 10.3872/j.issn.1007-385X.2013.06.004
Abstract:
Objective:To observe the effect of culture with IL-2 and IL-15 in vitro on the proportion, the cell phenotype, the cytotoxic activity against tumor cells and the adhesion activity of NK cells, NKT cells and T cells subsets, and to discuss the possible mechanism. Methods:Five patients with breast cancer were obtained in the Department of Biotherapy, Tianjin Medical University Cancer Institute and Hospital from May 2012 to July 2012, and PBMCs of those patients were isolated by and cultured with the NK cells proliferation scheme contained IL-2+IL-5. The cell expansion fold and the proportion of different lymphocyte subsets were observed after cultured by IL-2 and IL-15 for 15 d. The changes of the cell immunophenotype and cell surface receptors were detected by flow cytometry. The anti-tumor cytotoxicity against HLA-match or HLA-mismatch cancer cells among three lymphocyte subsets were measured using LDH cytotoxicity assay and CD107a release assay. Adhesion between the total expansion products and HLA-match or HLA-mismatch tumor cells was detected by Live Cell Station. Results: After expansion by IL-2+IL-15 for 15 days, compared with preamplification, the proportions of NK cells (\[36.74±17.10\]% vs \[16.34±3.12\]%, P<0.05) and NKT cells (\[24.88±12.34\]% vs \[4.06±2.05\]%, P<0.05\] were significantly increased; the proportions of CD4+T cells and Treg cells were significantly decreased (P<0.05), the proportions of CD8+ T cells were significantly increased (P<0.05); the expressions of surface activation receptors NKp30 (\[92.38±7.19\]% vs (32.14±17.64)%, P<0.05\], NKp44 \[(74.26±1628\]% vs \[1.94±1.60\]%, P<0.05\] and NKG2D (\[98.58±1.28)% vs (66.50±24.84)%, P<0.05\] expressed on NK cells were increased significantly, while CD16 was obviously decreased (\[85.12±7.66\]% vs \[95.60±253\]%, P<0.05\]; the activation receptors DNAM-1 and NKG2D expressed on both NKT cells and T cells were significantly increased (P<0.05). The killing rates of total expansion cells, NK cells and NKT cells against HLA-mismatched tumor cells were significantly higher than those against HLA-matched tumor cells (P<0.05). In the 84th minute of co-incubation, the adherent number of total expansion cells combined to HLA-mismatched tumor cells were significantly higher than that of HLA-matched tumor cells (\[4.80±0.53\] vs \[2.25±0.35\], P<0.05\]. Conclusion: Not only NK cells but also NKT cells can be amplificated by cultured with IL-2 and IL-15, indicating that it can be used to expand CD56+ cells. Expansion products kill the tumor cells mainly by the NK cells killing activity without HLA-restricted.
Objective:To observe the effect of culture with IL-2 and IL-15 in vitro on the proportion, the cell phenotype, the cytotoxic activity against tumor cells and the adhesion activity of NK cells, NKT cells and T cells subsets, and to discuss the possible mechanism. Methods:Five patients with breast cancer were obtained in the Department of Biotherapy, Tianjin Medical University Cancer Institute and Hospital from May 2012 to July 2012, and PBMCs of those patients were isolated by and cultured with the NK cells proliferation scheme contained IL-2+IL-5. The cell expansion fold and the proportion of different lymphocyte subsets were observed after cultured by IL-2 and IL-15 for 15 d. The changes of the cell immunophenotype and cell surface receptors were detected by flow cytometry. The anti-tumor cytotoxicity against HLA-match or HLA-mismatch cancer cells among three lymphocyte subsets were measured using LDH cytotoxicity assay and CD107a release assay. Adhesion between the total expansion products and HLA-match or HLA-mismatch tumor cells was detected by Live Cell Station. Results: After expansion by IL-2+IL-15 for 15 days, compared with preamplification, the proportions of NK cells (\[36.74±17.10\]% vs \[16.34±3.12\]%, P<0.05) and NKT cells (\[24.88±12.34\]% vs \[4.06±2.05\]%, P<0.05\] were significantly increased; the proportions of CD4+T cells and Treg cells were significantly decreased (P<0.05), the proportions of CD8+ T cells were significantly increased (P<0.05); the expressions of surface activation receptors NKp30 (\[92.38±7.19\]% vs (32.14±17.64)%, P<0.05\], NKp44 \[(74.26±1628\]% vs \[1.94±1.60\]%, P<0.05\] and NKG2D (\[98.58±1.28)% vs (66.50±24.84)%, P<0.05\] expressed on NK cells were increased significantly, while CD16 was obviously decreased (\[85.12±7.66\]% vs \[95.60±253\]%, P<0.05\]; the activation receptors DNAM-1 and NKG2D expressed on both NKT cells and T cells were significantly increased (P<0.05). The killing rates of total expansion cells, NK cells and NKT cells against HLA-mismatched tumor cells were significantly higher than those against HLA-matched tumor cells (P<0.05). In the 84th minute of co-incubation, the adherent number of total expansion cells combined to HLA-mismatched tumor cells were significantly higher than that of HLA-matched tumor cells (\[4.80±0.53\] vs \[2.25±0.35\], P<0.05\]. Conclusion: Not only NK cells but also NKT cells can be amplificated by cultured with IL-2 and IL-15, indicating that it can be used to expand CD56+ cells. Expansion products kill the tumor cells mainly by the NK cells killing activity without HLA-restricted.
2013, 20(6):661-666.
DOI: 10.3872/j.issn.1007-385X.2013.06.005
Abstract:
Objective:To study the effects of overexpression of BCSC-1 gene on the proliferation, invasion, adherence and migration abilities of hepatoma carcinoma Bel-7402 cells, and to investigate the possible mechanism. Methods: Bel-7402 cells were transfected by pcDNA3.1/v5-HisB-BCSC-1 and pcDNA3.1/v5-HisB respectively as BCSC-1 group and empty vector group, and the wild-type Bel-7402 cells as a wild group. Proliferation, invasion, adherence and migration abilities of the Bel-7402 cells were detected by MTT assay, Transwell assay, adhesion experiment in vitro and scratch test, respectively. Real-time PCR was performed to detect the expression of BCSC-1 and osteopontin (OPN), ICAM-1, PTTG mRNA, which were correlated with the cell growth and adhesion in Bel-7402 cells. Results: After transfected by pcDNA3.1/v5-HisB-BCSC-1, the expression of BCSC-1 mRNA in Bel-7402 cells was significantly raised compared with that of empty vector group and the wild-type group (\[10.58±0.56\] vs \[1.10±0.22\], \[1.00±0.01\]; all P＜0.01). The Bel-7402 cell line stably overexpressing BCSC-1 was successfully established. Compared with the empty vector group and wild group, the growth rate of Bel-7402 cells in the BCSC-1 group was obviously slower (72 h: \[0.29±0.003\] vs \[0.34±0.014\], \[0.35±0.013\]; all P<0.05), the invasion rate (\[76.20±1.85\]% vs \[93.42±3.24\]%, \[10000±105\]%; all P<0.01) and the adhesion rate (\[58.57±0.84\]% vs \[97.14±0.84\]%, \[100.00±130\]%; all P<0.01) were obviously decreased, the migratory distance was significantly reduced (\[116.60±10.58\] vs \[231.33±1026\], \[237.96±11.58\] μm; all P<0.01\]. The expression of OPN mRNA in the Bel-7402 cells which the BCSC-1 was up-regulated was significantly decreased (\[0.12±0.06\] vs \[0.95±0.14\], \[1.00±0.08\]; all P<001). No significant changes were observed in the expression of ICAM-1 and PTTG mRNA. Conclusion: Overexpression of BCSC-1 can inhibit the proliferation, invasion, adherence and migration abilities of Bel-7402 cells, which may be related to the decreased expression of OPN.
Objective:To study the effects of overexpression of BCSC-1 gene on the proliferation, invasion, adherence and migration abilities of hepatoma carcinoma Bel-7402 cells, and to investigate the possible mechanism. Methods: Bel-7402 cells were transfected by pcDNA3.1/v5-HisB-BCSC-1 and pcDNA3.1/v5-HisB respectively as BCSC-1 group and empty vector group, and the wild-type Bel-7402 cells as a wild group. Proliferation, invasion, adherence and migration abilities of the Bel-7402 cells were detected by MTT assay, Transwell assay, adhesion experiment in vitro and scratch test, respectively. Real-time PCR was performed to detect the expression of BCSC-1 and osteopontin (OPN), ICAM-1, PTTG mRNA, which were correlated with the cell growth and adhesion in Bel-7402 cells. Results: After transfected by pcDNA3.1/v5-HisB-BCSC-1, the expression of BCSC-1 mRNA in Bel-7402 cells was significantly raised compared with that of empty vector group and the wild-type group (\[10.58±0.56\] vs \[1.10±0.22\], \[1.00±0.01\]; all P＜0.01). The Bel-7402 cell line stably overexpressing BCSC-1 was successfully established. Compared with the empty vector group and wild group, the growth rate of Bel-7402 cells in the BCSC-1 group was obviously slower (72 h: \[0.29±0.003\] vs \[0.34±0.014\], \[0.35±0.013\]; all P<0.05), the invasion rate (\[76.20±1.85\]% vs \[93.42±3.24\]%, \[10000±105\]%; all P<0.01) and the adhesion rate (\[58.57±0.84\]% vs \[97.14±0.84\]%, \[100.00±130\]%; all P<0.01) were obviously decreased, the migratory distance was significantly reduced (\[116.60±10.58\] vs \[231.33±1026\], \[237.96±11.58\] μm; all P<0.01\]. The expression of OPN mRNA in the Bel-7402 cells which the BCSC-1 was up-regulated was significantly decreased (\[0.12±0.06\] vs \[0.95±0.14\], \[1.00±0.08\]; all P<001). No significant changes were observed in the expression of ICAM-1 and PTTG mRNA. Conclusion: Overexpression of BCSC-1 can inhibit the proliferation, invasion, adherence and migration abilities of Bel-7402 cells, which may be related to the decreased expression of OPN.
2013, 20(6):667-672.
DOI: 10.3872/j.issn.1007-385X.2013.06.006
Abstract:
Objective:To investigate the effect of epidermal growth factor-like domain 7 ( EGFL7 ) on the proliferation, apoptosis, invasion and angiogenesis of lung carcinoma A549 cells through silence EGFL7 by lentivirus infection. Methods: Lentivirus mediated small hairpin RNA target EGFL7 (LV-RNAi) were transfected into A549 cells. Three groups were designed in this study: a LV-RNAi group (A549 cells transfected with LV-RNAi), a LV-NC group (A549 cells transfected with the control GFP-lentivirus) and a control group (A549 cells). Real-time PCR and Western blotting were performed to detect the effect of LV-RNAi infection on the expression of EGFL7 mRNA and protein in A549 cells. MTT assay and flow cytometry were used to analyz the influence of silence EGFL7 on the growth and apoptosis of A549 cells, respectively. Transwell invasion assay and chick embryo chorioallantoic membrane (CAM) assay were used to detect the influence of silence EGFL7 on the angiogenesis and invasion, respectively. Results: Lentivirus infected A549 cells stably and efficiently. Expression of EGFL7 mRNA （\[0.18±0.03\] vs \[0.98±0.05\], \[1.03±0.09\]; P<0.05） and protein of the LV-RNAi group were significantly reduced compared with that of the LV-NC group and control group. Compared with the LV-NC group and control group, silencing EGFL7 showed no obvious influence on the proliferation capacity (120 h after infection: \[0.73±0.22\] vs \[0.79±0.08\] vs \[0.81±0.11\], P>0.05) and apoptosis rate (120 h after infection: \[1.92%±0.06\] vs \[2.11%±0.07\] vs \[1.85%±0.13\], P>0.05) of A549 cells in the LV-RNAi group; meanwhile, the invasion cell number (\[61.7±14.5\] vs \[272.8±21.5\], \[286.6±40.0\]/mm2; P<0.05) and the angiogenesis index (\[31.7±4.1\] vs \[96.3±4.4\], \[103.3±6.5\]; P<0.05) were significantly decreased in the LV-RNAi group. Conclusion: Silencing EGFL7gene may inhibit the invasion ability and the angiogenesis ability of A549 cells without affecting the proliferation and apoptosis.
Objective:To investigate the effect of epidermal growth factor-like domain 7 ( EGFL7 ) on the proliferation, apoptosis, invasion and angiogenesis of lung carcinoma A549 cells through silence EGFL7 by lentivirus infection. Methods: Lentivirus mediated small hairpin RNA target EGFL7 (LV-RNAi) were transfected into A549 cells. Three groups were designed in this study: a LV-RNAi group (A549 cells transfected with LV-RNAi), a LV-NC group (A549 cells transfected with the control GFP-lentivirus) and a control group (A549 cells). Real-time PCR and Western blotting were performed to detect the effect of LV-RNAi infection on the expression of EGFL7 mRNA and protein in A549 cells. MTT assay and flow cytometry were used to analyz the influence of silence EGFL7 on the growth and apoptosis of A549 cells, respectively. Transwell invasion assay and chick embryo chorioallantoic membrane (CAM) assay were used to detect the influence of silence EGFL7 on the angiogenesis and invasion, respectively. Results: Lentivirus infected A549 cells stably and efficiently. Expression of EGFL7 mRNA （\[0.18±0.03\] vs \[0.98±0.05\], \[1.03±0.09\]; P<0.05） and protein of the LV-RNAi group were significantly reduced compared with that of the LV-NC group and control group. Compared with the LV-NC group and control group, silencing EGFL7 showed no obvious influence on the proliferation capacity (120 h after infection: \[0.73±0.22\] vs \[0.79±0.08\] vs \[0.81±0.11\], P>0.05) and apoptosis rate (120 h after infection: \[1.92%±0.06\] vs \[2.11%±0.07\] vs \[1.85%±0.13\], P>0.05) of A549 cells in the LV-RNAi group; meanwhile, the invasion cell number (\[61.7±14.5\] vs \[272.8±21.5\], \[286.6±40.0\]/mm2; P<0.05) and the angiogenesis index (\[31.7±4.1\] vs \[96.3±4.4\], \[103.3±6.5\]; P<0.05) were significantly decreased in the LV-RNAi group. Conclusion: Silencing EGFL7gene may inhibit the invasion ability and the angiogenesis ability of A549 cells without affecting the proliferation and apoptosis.
2013, 20(6):673-678.
DOI: 10.3872/j.issn.1007-385X.2013.06.007
Abstract:
Objective:To investigate the effect of down-regulation of lentivirus-mediated N-acetylglucosaminyltransferase Ⅳa (GnTⅣa) on the proliferation and invasion of mouse hepatoma Hca-F cells in vitro. Methods: Expression plasmid pll3.7/Mgat4a-shRNA with interference shRNA sequences gene target Mgat4agene, were encoding GnTⅣa was constructed. The RNAi lentiviral particles were packaged to infect Hca-F cells. The effect of Mgat4a-shRNA infection on the Mgat4a mRNA and GnTⅣa expression of Hca-F cells was detected by RT-PCR and flowcytometry, respectively. The effect of recombinant lentiviral infection on the proliferation and invasion capability of Hca-F cells was determined by CCK-8 and transwell assay in vitro, respectively. Results: The mouse hepotoma Hca-F cells could be infected by the lentivirus with a high efficiency, and the infection efficiency of Mgat4a-shRNA group reached 72%. Compared with the Hca-F cells and Mock-shRNA group, the Mgat4a mRNA (\[0.53±0.2\] vs \[2.97±0.2\], \[2.90±0.1\]; P<0.05) and GnTⅣa (\[47.80±2.25\] vs \[39461±5.27), \[378.61±4.38\]) expression of the Mgat4a-shRNA group was significantly decreased, and the cell proliferation (inhibitory rate \[46.9±0.02\]% vs \[0.82±0.03\]%, 0; all P<0.01) and invasion (invasive cell numbers: \[18±3\] vs \[42±4\], \[38±3\], all P<0.05) capability were inhibited significantly. Conclusion: Down-regulation of lentivirus-mediated GnTⅣa expression inhibits the proliferation and invasion of mouse hepatoma Hca-F cells in vitro.
Objective:To investigate the effect of down-regulation of lentivirus-mediated N-acetylglucosaminyltransferase Ⅳa (GnTⅣa) on the proliferation and invasion of mouse hepatoma Hca-F cells in vitro. Methods: Expression plasmid pll3.7/Mgat4a-shRNA with interference shRNA sequences gene target Mgat4agene, were encoding GnTⅣa was constructed. The RNAi lentiviral particles were packaged to infect Hca-F cells. The effect of Mgat4a-shRNA infection on the Mgat4a mRNA and GnTⅣa expression of Hca-F cells was detected by RT-PCR and flowcytometry, respectively. The effect of recombinant lentiviral infection on the proliferation and invasion capability of Hca-F cells was determined by CCK-8 and transwell assay in vitro, respectively. Results: The mouse hepotoma Hca-F cells could be infected by the lentivirus with a high efficiency, and the infection efficiency of Mgat4a-shRNA group reached 72%. Compared with the Hca-F cells and Mock-shRNA group, the Mgat4a mRNA (\[0.53±0.2\] vs \[2.97±0.2\], \[2.90±0.1\]; P<0.05) and GnTⅣa (\[47.80±2.25\] vs \[39461±5.27), \[378.61±4.38\]) expression of the Mgat4a-shRNA group was significantly decreased, and the cell proliferation (inhibitory rate \[46.9±0.02\]% vs \[0.82±0.03\]%, 0; all P<0.01) and invasion (invasive cell numbers: \[18±3\] vs \[42±4\], \[38±3\], all P<0.05) capability were inhibited significantly. Conclusion: Down-regulation of lentivirus-mediated GnTⅣa expression inhibits the proliferation and invasion of mouse hepatoma Hca-F cells in vitro.
2013, 20(6):679-684.
DOI: 10.3872/j.issn.1007-385X.2013.06.008
Abstract:
Objective:To investigate the effects of parthenolide (PTL) on the proliferation, bcl-2 expression and susceptibility of CD34+CD38- KG-1a leukemia cells to cytotoxicity of allogeneic natural killer cells (allo-NK cells). Methods: CD34+CD38- KG-1a cells were separated from KG-1a cells by magnetic activated cell sorting (MACS). The effect of PTL on the proliferation of CD34+CD38- KG-1a cells was assessed by XTT assay. The expression of Bcl-2 mRNA and protein in CD34+CD38- KG-1a cells regulated by parthenolide were evaluated by real-time reverse transcriptase-polymerase chain reaction analysis (RT-PCR) and Western blotting, respectively. LDH-releasing assay was performed to observe the effect of PTL on the cytotoxicity of allo-NK cells against CD34+CD38- KG-1a cells. Results: PTL inhibited the proliferation of CD34+CD38- KG-1a cells with a concentration-dependent manner. As the concentration of PTL was 2.5 μmol/L, the cellular proliferation inhibition rate of CD34+CD38- KG-1a cells in PTL group was significantly higher than that of the control group (\[4.89±1.07\]% vs 0, P＜0.01). The IC50 of PTL to inhibit the proliferation of CD34+CD38- KG-1a leukemia cells was 20 μmol/L. By treated with 20 μmol/L PTL, the expression of bcl-2 mRNA (\[0.105±0007\] vs \[0.307±0.013\], P＜0.01) and protein in CD34+CD38- KG-1a cells were significantly lower than those of the control group. At effect-to-target ratio of 10∶1, 20∶1 and 40∶1, the cytotoxicity rate of allo-NK cell against CD34+CD38- KG-1a cells in the PTL group and in the positive control group was increasing and was higher than that of the negative control group (\[19.76±1.01\]%, \[30.14±0.96\]% and \[51.48±3.15\]% vs \[12.50±1.42\]%, \[16.90±093\]% and \[31.70±1.53\]%; all P<0.01). In addition, at effect-to-target ratio of 40∶1, the cytotoxicity rate of allo-NK cell against CD34+CD38- KG-1a cells in the PTL group was significantly higher than that of the positive control group (\[5148±315\]% vs \[43.08±2.81\]%, P<0.05). Conclusion: PTL can suppress the proliferation of CD34+CD38- KG-1a cells and enhance its susceptibility to the cytotoxicity of allo-NK cells, which may be related with the down-regulation of Bcl-2 by PTL．
Objective:To investigate the effects of parthenolide (PTL) on the proliferation, bcl-2 expression and susceptibility of CD34+CD38- KG-1a leukemia cells to cytotoxicity of allogeneic natural killer cells (allo-NK cells). Methods: CD34+CD38- KG-1a cells were separated from KG-1a cells by magnetic activated cell sorting (MACS). The effect of PTL on the proliferation of CD34+CD38- KG-1a cells was assessed by XTT assay. The expression of Bcl-2 mRNA and protein in CD34+CD38- KG-1a cells regulated by parthenolide were evaluated by real-time reverse transcriptase-polymerase chain reaction analysis (RT-PCR) and Western blotting, respectively. LDH-releasing assay was performed to observe the effect of PTL on the cytotoxicity of allo-NK cells against CD34+CD38- KG-1a cells. Results: PTL inhibited the proliferation of CD34+CD38- KG-1a cells with a concentration-dependent manner. As the concentration of PTL was 2.5 μmol/L, the cellular proliferation inhibition rate of CD34+CD38- KG-1a cells in PTL group was significantly higher than that of the control group (\[4.89±1.07\]% vs 0, P＜0.01). The IC50 of PTL to inhibit the proliferation of CD34+CD38- KG-1a leukemia cells was 20 μmol/L. By treated with 20 μmol/L PTL, the expression of bcl-2 mRNA (\[0.105±0007\] vs \[0.307±0.013\], P＜0.01) and protein in CD34+CD38- KG-1a cells were significantly lower than those of the control group. At effect-to-target ratio of 10∶1, 20∶1 and 40∶1, the cytotoxicity rate of allo-NK cell against CD34+CD38- KG-1a cells in the PTL group and in the positive control group was increasing and was higher than that of the negative control group (\[19.76±1.01\]%, \[30.14±0.96\]% and \[51.48±3.15\]% vs \[12.50±1.42\]%, \[16.90±093\]% and \[31.70±1.53\]%; all P<0.01). In addition, at effect-to-target ratio of 40∶1, the cytotoxicity rate of allo-NK cell against CD34+CD38- KG-1a cells in the PTL group was significantly higher than that of the positive control group (\[5148±315\]% vs \[43.08±2.81\]%, P<0.05). Conclusion: PTL can suppress the proliferation of CD34+CD38- KG-1a cells and enhance its susceptibility to the cytotoxicity of allo-NK cells, which may be related with the down-regulation of Bcl-2 by PTL．
2013, 20(6):685-689.
DOI: 10.3872/j.issn.1007-385X.2013.06.009
Abstract:
Objective:To study the effect of silencing Slug gene on proliferation, cell cycle and invasion ability of lung cancer A549 cells by RNA interference technique. Methods: The recombinant plasmid expressing short hairpin RNA (shRNA) targeting Slug gene was constructed and named as pGPU6-GFP-Neo-Slug. A549 cells were transfected with pGPU6-GFP-Neo-Slug and the negative control plasmid pNeg-shRNA with liposome method. Real-time PCR and Western blotting analysis was performed to determine the expression of Slug mRNA and protein after transfection, respectively. The proliferation, cell cycle distribution and cell invasion of A549 cells were detected by CCK-8 assay, flow cytometry and transwell assay, respectively. Results: pGPU6-GFP-Neo-Slug vector was successfully constructed and transfected into A549 cells with transfection rate of nearly 90%. Compared to the control and pNeg-shRNA group, Slug mRNA (\[0.23±0.01\] vs \[0.97±0.08\], \[1.0±0.09\]; all P＜0.05) and protein (\[0.20±0.09\] vs \[1.0±0.32\], \[1.13±0.26\]; all P＜0.05) expression level was significantly reduced in pSlug-shRNA group. Compared to the control and pNeg-shRNA group, the inhibition rate of proliferation in Slug-silencing A549 cells was significantly increased (\[35.3±5.4\]% vs \[15±02%\], \[3.3±0.7%\]; all P< 0.05); the cell multiplication index was significantly decreased(\[32.92±069\] vs \[48.19±0.71\], \[42.88±0.75\]; all P<0.05); the cell number in G1 phase was significantly increased(\[67.08±092\] vs \[52.81±0.78\], \[56.12±0.73\]; all P<0.05); and the cell invasion ability of A549 was significantly reduced (number of alive A549 cells through the matrigel chamber：\[55±9\] vs \[169±12\], \[173±15\]; all P<0.01). Conclusion: pGPU6-GFP-Neo-Slug vector transfected into lung cancer A549 cell can effectively silence Slug gene expression, inhibit cell proliferation, influence cell cycle and inhibit cell invasion of A549 cells.
Objective:To study the effect of silencing Slug gene on proliferation, cell cycle and invasion ability of lung cancer A549 cells by RNA interference technique. Methods: The recombinant plasmid expressing short hairpin RNA (shRNA) targeting Slug gene was constructed and named as pGPU6-GFP-Neo-Slug. A549 cells were transfected with pGPU6-GFP-Neo-Slug and the negative control plasmid pNeg-shRNA with liposome method. Real-time PCR and Western blotting analysis was performed to determine the expression of Slug mRNA and protein after transfection, respectively. The proliferation, cell cycle distribution and cell invasion of A549 cells were detected by CCK-8 assay, flow cytometry and transwell assay, respectively. Results: pGPU6-GFP-Neo-Slug vector was successfully constructed and transfected into A549 cells with transfection rate of nearly 90%. Compared to the control and pNeg-shRNA group, Slug mRNA (\[0.23±0.01\] vs \[0.97±0.08\], \[1.0±0.09\]; all P＜0.05) and protein (\[0.20±0.09\] vs \[1.0±0.32\], \[1.13±0.26\]; all P＜0.05) expression level was significantly reduced in pSlug-shRNA group. Compared to the control and pNeg-shRNA group, the inhibition rate of proliferation in Slug-silencing A549 cells was significantly increased (\[35.3±5.4\]% vs \[15±02%\], \[3.3±0.7%\]; all P< 0.05); the cell multiplication index was significantly decreased(\[32.92±069\] vs \[48.19±0.71\], \[42.88±0.75\]; all P<0.05); the cell number in G1 phase was significantly increased(\[67.08±092\] vs \[52.81±0.78\], \[56.12±0.73\]; all P<0.05); and the cell invasion ability of A549 was significantly reduced (number of alive A549 cells through the matrigel chamber：\[55±9\] vs \[169±12\], \[173±15\]; all P<0.01). Conclusion: pGPU6-GFP-Neo-Slug vector transfected into lung cancer A549 cell can effectively silence Slug gene expression, inhibit cell proliferation, influence cell cycle and inhibit cell invasion of A549 cells.
Zhang Tao
,
Li Dong
,
Qi Zhongchun
,
Peng Jingjing
,
Chen Tao
,
Tan Yong
,
Zhou Jinjun
,
Xu Tao
,
Fu Zengqiang
,
Liu Yu
,
Li Hua
2013, 20(6):690-695.
DOI: 10.3872/j.issn.1007-385X.2013.06.010
Abstract:
Objective:To build a method for enrichment, culture and identification of a cell subset which has the characteristics of liver cancer stem cells (CSC) in vivo. Methods: Huh7 liver cancer cells were subjected to enrichment culture in spheric culture condition with cancer stem cell enrichment medium to obtain liver cancer stem cell-like cells. The stem cell-like cells were continuously cultured and proliferated to form liver cancer stem cell sphere in vitro. Flow cytometry assay was performed to detect the expression of the bio-marker moleculars of CSC such as EpCAM (also named CD326), CD90 and CD133. Colony formation assay and tumorigenicity experiments in nude mice were used to detect the colony formation abilities and tumorigenicity of Huh7 cells and Huh7 stem cell-like cells, respectively. Results: After enrichment culture for 3-7 days, Huh7 cells formed cell spheres. The liver cancer stem cell-like sphere-forming cells exhibited self-renewal ability and multiplication capacity. The proportion of EpCAM positive cells was significantly increased compared to Huh7 cells (\[99.6%±0.31\]% vs \[0.12%±0.05\]%，P<0.01). However, there is no significant difference in the expression of CD90 (\[0.11%±0.06\]% vs \[0.09%±0.07\]，P>0.05) and CD 133 (\[0.17%±0.08\]% vs \[0.15%±0.05\]，P>0.05) between the Huh7 cells and sphere-forming cells. The number of colonies formed by enriched sphere-forming cells was significantly more than that formed by Huh7 cells (\[188.67±12.5\] vs \[79±16.7\]，P＜0.01). Compared with the nude mice inoculation of Huh7 cells in the control group，tumor-forming time of nude mice noculation of Huh7 stem cell-like cells in the experimental group was obviously shorter (11 vs 30 d), and showed a higher ratio of tumor formation(100% vs 16.67%) when all the mice of two groups were seeded with 5×104 cells. Both tumor volume (\[171.90±10.94\] vs \[86.39±11.21\] mm3, P＜0.01) and weight (\[2.98±0.82\] vs \[0.32±0.17\]g, P＜001) of nude mice in the experimental group was respectively bigger and heavier than that of the control group when the seeding cell number was 5×105 cells. Conclusion: Liver cancer stem cell-like cells could be enriched and transformed from Huh7 cells with sphere-forming culture method in vitro, and these cells have more powerful tumorigenicity than Huh7 cells do.
Objective:To build a method for enrichment, culture and identification of a cell subset which has the characteristics of liver cancer stem cells (CSC) in vivo. Methods: Huh7 liver cancer cells were subjected to enrichment culture in spheric culture condition with cancer stem cell enrichment medium to obtain liver cancer stem cell-like cells. The stem cell-like cells were continuously cultured and proliferated to form liver cancer stem cell sphere in vitro. Flow cytometry assay was performed to detect the expression of the bio-marker moleculars of CSC such as EpCAM (also named CD326), CD90 and CD133. Colony formation assay and tumorigenicity experiments in nude mice were used to detect the colony formation abilities and tumorigenicity of Huh7 cells and Huh7 stem cell-like cells, respectively. Results: After enrichment culture for 3-7 days, Huh7 cells formed cell spheres. The liver cancer stem cell-like sphere-forming cells exhibited self-renewal ability and multiplication capacity. The proportion of EpCAM positive cells was significantly increased compared to Huh7 cells (\[99.6%±0.31\]% vs \[0.12%±0.05\]%，P<0.01). However, there is no significant difference in the expression of CD90 (\[0.11%±0.06\]% vs \[0.09%±0.07\]，P>0.05) and CD 133 (\[0.17%±0.08\]% vs \[0.15%±0.05\]，P>0.05) between the Huh7 cells and sphere-forming cells. The number of colonies formed by enriched sphere-forming cells was significantly more than that formed by Huh7 cells (\[188.67±12.5\] vs \[79±16.7\]，P＜0.01). Compared with the nude mice inoculation of Huh7 cells in the control group，tumor-forming time of nude mice noculation of Huh7 stem cell-like cells in the experimental group was obviously shorter (11 vs 30 d), and showed a higher ratio of tumor formation(100% vs 16.67%) when all the mice of two groups were seeded with 5×104 cells. Both tumor volume (\[171.90±10.94\] vs \[86.39±11.21\] mm3, P＜0.01) and weight (\[2.98±0.82\] vs \[0.32±0.17\]g, P＜001) of nude mice in the experimental group was respectively bigger and heavier than that of the control group when the seeding cell number was 5×105 cells. Conclusion: Liver cancer stem cell-like cells could be enriched and transformed from Huh7 cells with sphere-forming culture method in vitro, and these cells have more powerful tumorigenicity than Huh7 cells do.
Zhang Zongqin
,
Chen Lei
,
Li Pengpeng
,
Zhang Xiaofeng
,
Sun Bin
,
Qian Haihua
,
Shi Lehua
,
Yin Zhengfeng
2013, 20(6):696-699.
DOI: 10.3872/j.issn.1007-385X.2013.06.011
Abstract:
Objective : To develop a novel method for isolation and culture of the oligoclonal tumor-infiltrating lymphocytes (TILs) with a stronger cytotoxic activity to autologous hepatoma carcinoma cells. Methods: Fresh human hepatocellular carcinoma (HCC) tissues were dissected to prepare conventional TILs and oligoclonal TILs by the way of enzymatic digestion with physical disaggregation and the new way of oligoclonal TILs culture from a single tumor fragment in microcultures. After 2-week culture, the oligoclonal TILs with a stronger cytotoxic activity were mergered for further expending. Different TILs-mediated lysis of HCC cells in vitro was determined by colorimetric tetrazolium (MTT) assay to compare the specific cytotoxicity between different oligoclonal TILs and between bulk TILs and oligoclonal TILs. Results: Lymphocytes could gradually migrate out of each tumor fragment and expand in the culture with high levels of IL-2. Oligoclonal TILs generated from different fragments showed different cytotoxic activities against autologous HCC cells, and the difference in cytotoxic activity between different oligoclonal TILs was significant (P<0.01). Moreover, the cytotoxic activity of oligoclonal TILs was significantly stronger than that of bulk TILs (\[72.56±6.69\]% vs \[46.24±403\]%, P<0.01). Conclusion: The cytotoxic activity against autologous HCC cells of oligoclonal TILs generated from HCC tissues is much stronger than that of conventional TILs.
Objective : To develop a novel method for isolation and culture of the oligoclonal tumor-infiltrating lymphocytes (TILs) with a stronger cytotoxic activity to autologous hepatoma carcinoma cells. Methods: Fresh human hepatocellular carcinoma (HCC) tissues were dissected to prepare conventional TILs and oligoclonal TILs by the way of enzymatic digestion with physical disaggregation and the new way of oligoclonal TILs culture from a single tumor fragment in microcultures. After 2-week culture, the oligoclonal TILs with a stronger cytotoxic activity were mergered for further expending. Different TILs-mediated lysis of HCC cells in vitro was determined by colorimetric tetrazolium (MTT) assay to compare the specific cytotoxicity between different oligoclonal TILs and between bulk TILs and oligoclonal TILs. Results: Lymphocytes could gradually migrate out of each tumor fragment and expand in the culture with high levels of IL-2. Oligoclonal TILs generated from different fragments showed different cytotoxic activities against autologous HCC cells, and the difference in cytotoxic activity between different oligoclonal TILs was significant (P<0.01). Moreover, the cytotoxic activity of oligoclonal TILs was significantly stronger than that of bulk TILs (\[72.56±6.69\]% vs \[46.24±403\]%, P<0.01). Conclusion: The cytotoxic activity against autologous HCC cells of oligoclonal TILs generated from HCC tissues is much stronger than that of conventional TILs.
2013, 20(6):700-705.
DOI: 10.3872/j.issn.1007-385X.2013.06.012
Abstract:
Objective:To observe the effect of recombinant super-compound interferon (rSIFN-co) on the proliferation, apoptosis and resistance to epirubicin of human breast cancer MCF-7/ADR cells (a multi-drug resistance \[MDR\] strain), and to investigate the possible mechanism. Methods: MCF-7/ADR cells were treated with rSIFN-co, epirubicin alone or combaination, and the MCF-7 cells were used as control. MTT assay and flow cytometry were performed to detect the effect of rSIFN-co on the proliferation and apoptosis of MCF-7/ADR cells, respectively. Immunohistochemical staining was used to detect the influence of rSIFN-co on the P-gp expression level in MCF-7/ADR cells. Results: After treated by different drugs for 24 h, the growth inhibition rate of MCF-7/ADR cells treated by 0.078 μg/ml rsIFN-co or 0.02 μg/ml rsIFN-co combined with 15.00 μg/ml epirubicin was significantly higher than that treated by 100.00 μg/ml epirubicin (\[29.7±1.4\]%, \[23.0±2.1\]% vs \[17.1±1.5\]%; all P<0.01). The inhibition effect of each drug had a dose and time dependence. Synergistic effect of rSIFN-co with epirubicin was also observed after being treated for 72 h. Epirubicin showed no significant effect on MCF-7/ADR cells′ apoptosis after treated for 24 h (P>0.05); however, use of rSIFN-co alone or combined with epirubicin significantly enhanced the apoptosis rate than did epirubicin alone after 24 h (\[3537±1.40\]%, \[61.37±1.76\]% vs \[9.80±1.66\]%; all P<0.01), and the effects on cell apoptosis had a time dependence (P<0.01); and the synergistic effect of rSIFN-co with epirubicin was also observed. Compared with the control group (3.94±0.0088), the P-gp expression was increased in the epirubicin group (4.17±0.0252, P<0.01), but decreased in rSIFN-co group (2.59±0.0260, P<0.01) and the combined group (2.62±0.0100, P<0.01).There was no significant difference between the combined group and rsIFN-co group in P-gp expression (P=0.948). Conclusion: rSIFN-co can inhibit cell growth, induce cell apoptosis of human breast cancer MCF-7/ADR cells, and reverse the multi-drug resistance by decreasing the expression of P-gp.
Objective:To observe the effect of recombinant super-compound interferon (rSIFN-co) on the proliferation, apoptosis and resistance to epirubicin of human breast cancer MCF-7/ADR cells (a multi-drug resistance \[MDR\] strain), and to investigate the possible mechanism. Methods: MCF-7/ADR cells were treated with rSIFN-co, epirubicin alone or combaination, and the MCF-7 cells were used as control. MTT assay and flow cytometry were performed to detect the effect of rSIFN-co on the proliferation and apoptosis of MCF-7/ADR cells, respectively. Immunohistochemical staining was used to detect the influence of rSIFN-co on the P-gp expression level in MCF-7/ADR cells. Results: After treated by different drugs for 24 h, the growth inhibition rate of MCF-7/ADR cells treated by 0.078 μg/ml rsIFN-co or 0.02 μg/ml rsIFN-co combined with 15.00 μg/ml epirubicin was significantly higher than that treated by 100.00 μg/ml epirubicin (\[29.7±1.4\]%, \[23.0±2.1\]% vs \[17.1±1.5\]%; all P<0.01). The inhibition effect of each drug had a dose and time dependence. Synergistic effect of rSIFN-co with epirubicin was also observed after being treated for 72 h. Epirubicin showed no significant effect on MCF-7/ADR cells′ apoptosis after treated for 24 h (P>0.05); however, use of rSIFN-co alone or combined with epirubicin significantly enhanced the apoptosis rate than did epirubicin alone after 24 h (\[3537±1.40\]%, \[61.37±1.76\]% vs \[9.80±1.66\]%; all P<0.01), and the effects on cell apoptosis had a time dependence (P<0.01); and the synergistic effect of rSIFN-co with epirubicin was also observed. Compared with the control group (3.94±0.0088), the P-gp expression was increased in the epirubicin group (4.17±0.0252, P<0.01), but decreased in rSIFN-co group (2.59±0.0260, P<0.01) and the combined group (2.62±0.0100, P<0.01).There was no significant difference between the combined group and rsIFN-co group in P-gp expression (P=0.948). Conclusion: rSIFN-co can inhibit cell growth, induce cell apoptosis of human breast cancer MCF-7/ADR cells, and reverse the multi-drug resistance by decreasing the expression of P-gp.
2013, 20(6):706-710.
DOI: 10.3872/j.issn.1007-385X.2013.06.013
Abstract:
Objective : To detect down regulation of the expression of BAG-1 gene in lung cancer A549 by transfected a vector contained shRNA, and to investigate its effects on the cisplatin (DDP) resistance of A549 cells. Methods: Interference vector pGCsi-BAG-1 target BAG-1 was constructed and stable transferred into A549 cells. Cells stable transferred by pGCsi-BAG-1 were used as an experimental group (BAG-1-shRNA), cells transferred by non-sense vector were used as a negative control group (SC-shRNA), and the parent A549 cells were used as a control group. Western blotting was performed to detect the effect of pGCsi-BAG-1 transfection on the BCL-2 and BAG-1 expression of A549 cells. MTT method and flow cytometry was used to detect the influence on proliferation and apoptosis of A549 cells after DDP treatment. Results: An A549 cell line where BAG-1 was stably interfered was successfully constructed. And the expression of BCL-2 protein in cells of the BAG-1- shRNA group was significantly lower than that of SC-shRNA group and the control group (all P<005). With increasing of the concentration of DDP, the proliferation inhibition rate of each cell was increased. When DDP concentration was 2.5 μg/ml, the cell proliferation inhibition rate of the BAG-1-shRNA group was significantly higher than that of SC-shRNA group and the control group (\[8.12±4.09\] % vs \[10.07± 3.82\] %, \[22.26±489\]%; all P<0.05). Compared with the SC-shRNA group and the control group, the apoptosis rate of the BAG-1-shRNA group was significantly increased after 24 h treatment with DDP (\[37.84±3.62\]% vs \[16.80±2.81\]%, \[1710±311\]%, P< 0.05). Conclusion: Down-regulation of BAG-1 expression can inhibit the proliferation of A549 cells after DDP treatment and promote its apoptosis.
Objective : To detect down regulation of the expression of BAG-1 gene in lung cancer A549 by transfected a vector contained shRNA, and to investigate its effects on the cisplatin (DDP) resistance of A549 cells. Methods: Interference vector pGCsi-BAG-1 target BAG-1 was constructed and stable transferred into A549 cells. Cells stable transferred by pGCsi-BAG-1 were used as an experimental group (BAG-1-shRNA), cells transferred by non-sense vector were used as a negative control group (SC-shRNA), and the parent A549 cells were used as a control group. Western blotting was performed to detect the effect of pGCsi-BAG-1 transfection on the BCL-2 and BAG-1 expression of A549 cells. MTT method and flow cytometry was used to detect the influence on proliferation and apoptosis of A549 cells after DDP treatment. Results: An A549 cell line where BAG-1 was stably interfered was successfully constructed. And the expression of BCL-2 protein in cells of the BAG-1- shRNA group was significantly lower than that of SC-shRNA group and the control group (all P<005). With increasing of the concentration of DDP, the proliferation inhibition rate of each cell was increased. When DDP concentration was 2.5 μg/ml, the cell proliferation inhibition rate of the BAG-1-shRNA group was significantly higher than that of SC-shRNA group and the control group (\[8.12±4.09\] % vs \[10.07± 3.82\] %, \[22.26±489\]%; all P<0.05). Compared with the SC-shRNA group and the control group, the apoptosis rate of the BAG-1-shRNA group was significantly increased after 24 h treatment with DDP (\[37.84±3.62\]% vs \[16.80±2.81\]%, \[1710±311\]%, P< 0.05). Conclusion: Down-regulation of BAG-1 expression can inhibit the proliferation of A549 cells after DDP treatment and promote its apoptosis.
2013, 20(6):711-716.
DOI: 10.3872/j.issn.1007-385X.2013.06.014
Abstract:
Objective : To investigate the clinical effect and safety of autologous dendritic cells (DCs) stimulated by cytokine-induced killer (CIK) cells in patients with advanced renal cell carcinoma (RCC). Methods: During July 2011 to June 2012, peripheral blood mononuclear cells (PBMCs) were collected from 22 patients (12 males and 10 females, median age 60.8 years \[21-79 years\]) with advanced renal carcinoma in the Department of Medical Oncology of Suzhou Hospital affiliated to Nanjing Medical University, and DC-CIK cells were individually prepared from these PBMCs. Flow cytometry was performed to analyz the proportion of CD3+T, CD8+T, CD4+T, NK（CD3-CD56+）and NKT（CD3+CD56+）cells in the DC-CIK cells. The cytotoxicity of DC-CIK cells on leukemia K562 cells (sensitive to NK cells) and renal carcinoma 786-0 cells (insensitive to NK cells) was detected by MTT assay. DC-CIK cells therapy was started when the conventional therapy (surgery+chemotherapy+radiotherapy/ cytokine therapy) finished and continued for 4 weeks. Each time, about (5.0±0.5)×108 DC-CIK cells were returned through intravenous transfusion, and each patient received 5 times (one cycle) of DC-CIK cells intravenous infusion. The immunological index such as lymphocyte subsets and cytokine concentration in the peripheral blood were detected before and after DC-CIK infusion. The adverse reactions during the treatment were closely observed and recorded. Results: In the total number of DC-CIK cells, the proportion of CD3 positive (CD3+) lymphocytes was (86.92±5.32)%, while the proportions of CD3+CD56+NKT cells and CD3-CD56+ NK cells were (52.04±7.33)% and(7.85±3.15)% respectively. DC-CIK cells showed similar in vitro killing activities on 786-0 cells and K562 cells in vitro, with a killing rate of (16.5±1.7) % and (18.4±1.9) % respectively (P=0.014), when the ratio of effect cells and target cells was 3 ∶1. After patients received DC-CIK cell therapy, the proportion of peripheral lymphocyte sub-group (CD3+T, CD4+T, CD8+T and CD3-CD56+NK cells) showed no significant change (P>0.05); compared to the cytokine levels prior to DC-CIK cells infusion, the levels of interleukine 2 (IL-2), interleukin 12 (IL-12) and interferon gamma (IFN-r) increased significantly (P<0.05), while the levels of tumor necrosis factor-α (TNF-α) and interleukine 10 (IL-10) changed unremarkably (P>0.05). Two patients experienced a transient fever lasting 4-6 hours and one felt a short time of fatigue after DC-CIK cells infusion. Conclusion: DC-CIK cells can kill 786-0 cells and K562 cells effectively. The DC-CIK cells infusion may improve immune response of some patients with a safe level, and it can be one of the assistant treatment methods for advanced RCC patients.
Objective : To investigate the clinical effect and safety of autologous dendritic cells (DCs) stimulated by cytokine-induced killer (CIK) cells in patients with advanced renal cell carcinoma (RCC). Methods: During July 2011 to June 2012, peripheral blood mononuclear cells (PBMCs) were collected from 22 patients (12 males and 10 females, median age 60.8 years \[21-79 years\]) with advanced renal carcinoma in the Department of Medical Oncology of Suzhou Hospital affiliated to Nanjing Medical University, and DC-CIK cells were individually prepared from these PBMCs. Flow cytometry was performed to analyz the proportion of CD3+T, CD8+T, CD4+T, NK（CD3-CD56+）and NKT（CD3+CD56+）cells in the DC-CIK cells. The cytotoxicity of DC-CIK cells on leukemia K562 cells (sensitive to NK cells) and renal carcinoma 786-0 cells (insensitive to NK cells) was detected by MTT assay. DC-CIK cells therapy was started when the conventional therapy (surgery+chemotherapy+radiotherapy/ cytokine therapy) finished and continued for 4 weeks. Each time, about (5.0±0.5)×108 DC-CIK cells were returned through intravenous transfusion, and each patient received 5 times (one cycle) of DC-CIK cells intravenous infusion. The immunological index such as lymphocyte subsets and cytokine concentration in the peripheral blood were detected before and after DC-CIK infusion. The adverse reactions during the treatment were closely observed and recorded. Results: In the total number of DC-CIK cells, the proportion of CD3 positive (CD3+) lymphocytes was (86.92±5.32)%, while the proportions of CD3+CD56+NKT cells and CD3-CD56+ NK cells were (52.04±7.33)% and(7.85±3.15)% respectively. DC-CIK cells showed similar in vitro killing activities on 786-0 cells and K562 cells in vitro, with a killing rate of (16.5±1.7) % and (18.4±1.9) % respectively (P=0.014), when the ratio of effect cells and target cells was 3 ∶1. After patients received DC-CIK cell therapy, the proportion of peripheral lymphocyte sub-group (CD3+T, CD4+T, CD8+T and CD3-CD56+NK cells) showed no significant change (P>0.05); compared to the cytokine levels prior to DC-CIK cells infusion, the levels of interleukine 2 (IL-2), interleukin 12 (IL-12) and interferon gamma (IFN-r) increased significantly (P<0.05), while the levels of tumor necrosis factor-α (TNF-α) and interleukine 10 (IL-10) changed unremarkably (P>0.05). Two patients experienced a transient fever lasting 4-6 hours and one felt a short time of fatigue after DC-CIK cells infusion. Conclusion: DC-CIK cells can kill 786-0 cells and K562 cells effectively. The DC-CIK cells infusion may improve immune response of some patients with a safe level, and it can be one of the assistant treatment methods for advanced RCC patients.
2013, 20(6):717-724.
DOI: 10.3872/j.issn.1007-385X.2013.06.015
Abstract:
Objective : To investigate the expression of growth arrest and DNA-damage-inducible 45 (Gadd45) gene family members in esophageal squamous cell carcinoma (ESCC) and discuss its clinical significance. Methods: ESCC tissues and paracancerous tissues of one hundred and twenty-eight patients with ESCC from the Fourth Hospital Affiliated to Hebei Medical University during 2004-2008 were included in this study. The expressions of Gadd45A, Gadd45B and Gadd45G mRNA in the ESCC tissues and paracancerous tissues were detected by RT-PCR. The protein expressions of Gadd45A, Gadd45B and Gadd45G in ESCC tissues and paracancerous tissues were detected by using immunohistochemistry methods. The relation was analyzed between the expressions of Gadd45A, Gadd45B and Gadd45G and the clinical pathologic characteristics and prognosis of ESCC. Results: Compared to the paracancerous tissues, the expression of Gadd45A mRNA in the tumor tissues was significantly increased (\[0.8924±0.3457\] vs \[0.6123±0.2132\], P<005\], Gadd45B mRNA showed no significantly changes (\[0.8256±0.3110\] vs \[0.8173±0.3069), P>0.05\], Gadd45G mRNA was significantly decreased (\[0.3855±0.1210\] vs \[0.8214±0.3069\], P<0.05, and the expression of Gadd45G mRNA was closely associated with TNM stage and histological differentiation. The expression of Gadd45A in the paracancerous tissues was mainly stained in nucleolus, while part of the tumor tissues demonstrated a cytoplasm expression mode. Compared with the paracancerous tissues, the expression of Gadd45A in the tumor tissues was significantly increased (χ2=55.603, P=0.000), Gadd45B showed no significant difference (χ2=0.456, P=0.500), and Gadd45G was significantly decreased (χ2= 70.134, P=0.000). The expression of Gadd45G in patients in phase Ⅲ and Ⅳwas significantly lower than that in patients in phaseⅠ and Ⅱ (χ2=5.768, P=0.016), and was significantly lower in the low differentiation group than in the high and moderate differentiation groups (χ2=4.483, P=0.034). The 5-year survival rate of patients with Gadd45G-positive was significantly higher than that of patients with Gadd45G-negative (44% vs 21%, P<0.05); Cox model analysis shows that Gadd45G expression was an independent prognostic factor for ESCC. Conclusion: Aberrant expression of Gadd45A and Gadd45G may participate in the development and progression of ESCC.
Objective : To investigate the expression of growth arrest and DNA-damage-inducible 45 (Gadd45) gene family members in esophageal squamous cell carcinoma (ESCC) and discuss its clinical significance. Methods: ESCC tissues and paracancerous tissues of one hundred and twenty-eight patients with ESCC from the Fourth Hospital Affiliated to Hebei Medical University during 2004-2008 were included in this study. The expressions of Gadd45A, Gadd45B and Gadd45G mRNA in the ESCC tissues and paracancerous tissues were detected by RT-PCR. The protein expressions of Gadd45A, Gadd45B and Gadd45G in ESCC tissues and paracancerous tissues were detected by using immunohistochemistry methods. The relation was analyzed between the expressions of Gadd45A, Gadd45B and Gadd45G and the clinical pathologic characteristics and prognosis of ESCC. Results: Compared to the paracancerous tissues, the expression of Gadd45A mRNA in the tumor tissues was significantly increased (\[0.8924±0.3457\] vs \[0.6123±0.2132\], P<005\], Gadd45B mRNA showed no significantly changes (\[0.8256±0.3110\] vs \[0.8173±0.3069), P>0.05\], Gadd45G mRNA was significantly decreased (\[0.3855±0.1210\] vs \[0.8214±0.3069\], P<0.05, and the expression of Gadd45G mRNA was closely associated with TNM stage and histological differentiation. The expression of Gadd45A in the paracancerous tissues was mainly stained in nucleolus, while part of the tumor tissues demonstrated a cytoplasm expression mode. Compared with the paracancerous tissues, the expression of Gadd45A in the tumor tissues was significantly increased (χ2=55.603, P=0.000), Gadd45B showed no significant difference (χ2=0.456, P=0.500), and Gadd45G was significantly decreased (χ2= 70.134, P=0.000). The expression of Gadd45G in patients in phase Ⅲ and Ⅳwas significantly lower than that in patients in phaseⅠ and Ⅱ (χ2=5.768, P=0.016), and was significantly lower in the low differentiation group than in the high and moderate differentiation groups (χ2=4.483, P=0.034). The 5-year survival rate of patients with Gadd45G-positive was significantly higher than that of patients with Gadd45G-negative (44% vs 21%, P<0.05); Cox model analysis shows that Gadd45G expression was an independent prognostic factor for ESCC. Conclusion: Aberrant expression of Gadd45A and Gadd45G may participate in the development and progression of ESCC.
2013, 20(6):725-728.
DOI: 10.3872/j.issn.1007-385X.2013.06.016
Abstract:
目的：检测胃癌患者外周血各类干扰素（Interferon，IFN）水平，探讨其与临床病理特征的关系。方法：收集天津医科大学附属肿瘤医院胃部肿瘤科2012年3月至2012年5月收治的47例初治胃腺癌患者外周血，以同期收集的25例健康人外周血为对照，ELISA法检测血清中IFN-α、IFN-β、IFN-γ、IFN-λ1以及IFN-λ2/3含量，分析各类IFN含量与患者性别、年龄、肿瘤分化程度、原发部位及各项生化指标间的相关性。结果：与健康对照相比，IFN-β在Ⅰ、Ⅱ期胃癌中表达升高（P=0009），IFN-λ1在Ⅲ、Ⅳ期胃癌中表达下降（P<0.001），而IFN-γ和λ2/3在Ⅲ、Ⅳ期胃癌中表达升高（P=0.029；P=0.004）。IFN-α、β含量与肿瘤发生部位有密切关系（P=0.003；P=0.004）；IFN-γ与LDH水平密切相关（P=0.013）。结论：胃癌患者外周血中不同IFN亚型呈现不同的表达变化，其变化与胃癌发生部位、分期及LDH水平相关，提示IFN家族成员可能对肿瘤的发生发展发挥各自的作用。
目的：检测胃癌患者外周血各类干扰素（Interferon，IFN）水平，探讨其与临床病理特征的关系。方法：收集天津医科大学附属肿瘤医院胃部肿瘤科2012年3月至2012年5月收治的47例初治胃腺癌患者外周血，以同期收集的25例健康人外周血为对照，ELISA法检测血清中IFN-α、IFN-β、IFN-γ、IFN-λ1以及IFN-λ2/3含量，分析各类IFN含量与患者性别、年龄、肿瘤分化程度、原发部位及各项生化指标间的相关性。结果：与健康对照相比，IFN-β在Ⅰ、Ⅱ期胃癌中表达升高（P=0009），IFN-λ1在Ⅲ、Ⅳ期胃癌中表达下降（P<0.001），而IFN-γ和λ2/3在Ⅲ、Ⅳ期胃癌中表达升高（P=0.029；P=0.004）。IFN-α、β含量与肿瘤发生部位有密切关系（P=0.003；P=0.004）；IFN-γ与LDH水平密切相关（P=0.013）。结论：胃癌患者外周血中不同IFN亚型呈现不同的表达变化，其变化与胃癌发生部位、分期及LDH水平相关，提示IFN家族成员可能对肿瘤的发生发展发挥各自的作用。
2013, 20(6):729-734.
DOI: 10.3872/j.issn.1007-385X.2013.06.017
Abstract:
表观遗传学是研究基因序列不发生改变的情况下，基因表达发生了可以遗传的改变的一门学科。DNA甲基化是表观遗传学中进化上比较保守、相对比较稳定的一种表观修饰。DNA甲基化，特别是CpG岛的高甲基化，通常能够介导基因表达的稳定沉默。研究表明，DNA甲基化参与调控免疫系统的分化发育，DNA甲基化的异常也是包括肿瘤在内的众多免疫相关疾病发生、发展的关键致病因素。表观遗传学与免疫学的碰撞也为理解免疫学现象的分子调控机制提供了崭新的视角。本文将对近年来DNA甲基化调控在免疫学中的进展作一综述。
表观遗传学是研究基因序列不发生改变的情况下，基因表达发生了可以遗传的改变的一门学科。DNA甲基化是表观遗传学中进化上比较保守、相对比较稳定的一种表观修饰。DNA甲基化，特别是CpG岛的高甲基化，通常能够介导基因表达的稳定沉默。研究表明，DNA甲基化参与调控免疫系统的分化发育，DNA甲基化的异常也是包括肿瘤在内的众多免疫相关疾病发生、发展的关键致病因素。表观遗传学与免疫学的碰撞也为理解免疫学现象的分子调控机制提供了崭新的视角。本文将对近年来DNA甲基化调控在免疫学中的进展作一综述。
2013, 20(6):735-738.
DOI: 10.3872/j.issn.1007-385X.2013.06.018
Abstract:
线粒体融合素基因-2（mitofusin-2 gene， Mfn-2 ）是作用于线粒体外膜的一种增殖抑制基因，在维持线粒体的形态、功能等方面起着重要作用。随着研究的不断深入， Mfn-2 在细胞信号转导、能量代谢、增殖及凋亡等生命过程中的作用日益显现。而肿瘤发生与细胞过度增殖及凋亡不足等密切相关，因此，如何抑制肿瘤细胞的过度增殖和促进细胞凋亡已成为目前的研究热点。 Mfn-2 通过多条通路参与多种肿瘤传代细胞系的增殖和凋亡，其表达异常或功能缺失可能是肿瘤发生、发展的重要原因。 Mfn-2 在许多肿瘤组织中低表达，且表达情况与肿瘤病理类型及生物学行为密切相关，提示其有可能是新的抑癌基因；同时， Mfn-2 过表达与喜树碱、放线菌酮（cycloheximide，CHX）等化疗药物联用具有协同作用，提示具有成为化疗增敏靶点的潜力。本文就 Mfn-2 的功能与肿瘤发生、发展的关系及其治疗学意义的相关研究进展进行综述。
线粒体融合素基因-2（mitofusin-2 gene， Mfn-2 ）是作用于线粒体外膜的一种增殖抑制基因，在维持线粒体的形态、功能等方面起着重要作用。随着研究的不断深入， Mfn-2 在细胞信号转导、能量代谢、增殖及凋亡等生命过程中的作用日益显现。而肿瘤发生与细胞过度增殖及凋亡不足等密切相关，因此，如何抑制肿瘤细胞的过度增殖和促进细胞凋亡已成为目前的研究热点。 Mfn-2 通过多条通路参与多种肿瘤传代细胞系的增殖和凋亡，其表达异常或功能缺失可能是肿瘤发生、发展的重要原因。 Mfn-2 在许多肿瘤组织中低表达，且表达情况与肿瘤病理类型及生物学行为密切相关，提示其有可能是新的抑癌基因；同时， Mfn-2 过表达与喜树碱、放线菌酮（cycloheximide，CHX）等化疗药物联用具有协同作用，提示具有成为化疗增敏靶点的潜力。本文就 Mfn-2 的功能与肿瘤发生、发展的关系及其治疗学意义的相关研究进展进行综述。
2013, 20(6):739-743.
DOI: 10.3872/j.issn.1007-385X.2013.06.019
Abstract:
自噬是机体内的一种代谢方式，参与调控多种生理病理过程，受不同信号级联反应调节，在维持细胞生长与代谢等方面起关键作用。哺乳动物雷帕霉素靶蛋白（mammalian target of rapamycin，mTOR）是一种蛋白激酶，也是调节细胞生长和增殖的重要信号分子，其通路的异常激活与肿瘤的发生发展密切相关且可通过调节细胞自噬治疗肿瘤。 p53 为抑癌基因，具有抑制癌症形成的作用。近来研究显示，胞质和胞核中均存在P53蛋白，在核内P53可通过活化腺苷酸活化的蛋白激酶（adenosine monophosphate activated protein kinase，AMPK）激活抑癌基因与张力蛋白同源10q丢失的磷酸酶基因（phosphatase and tensin homolog deleted on chromosome ten，PTEN）的转录等机制来调节mTOR信号通路相关分子，抑制mTOR活性，促使UNC-51激酶1（unc-51-like kinase1，ULK1）去磷酸化而增强自噬，亦可介导某些凋亡蛋白（Bcl-2、Bax等）及调控因子（DRAM、PUMA等）的信号通路参与自噬调节，但与mTOR通路的关系尚待研究；细胞质中的P53蛋白以非依赖转录的方式如降解P53、抑制RB1CC1/FIP200活性等，参与mTOR途径调控，抑制自噬。本文就P53蛋白介导mTOR信号途径影响肿瘤细胞自噬的研究进展作一综述。
自噬是机体内的一种代谢方式，参与调控多种生理病理过程，受不同信号级联反应调节，在维持细胞生长与代谢等方面起关键作用。哺乳动物雷帕霉素靶蛋白（mammalian target of rapamycin，mTOR）是一种蛋白激酶，也是调节细胞生长和增殖的重要信号分子，其通路的异常激活与肿瘤的发生发展密切相关且可通过调节细胞自噬治疗肿瘤。 p53 为抑癌基因，具有抑制癌症形成的作用。近来研究显示，胞质和胞核中均存在P53蛋白，在核内P53可通过活化腺苷酸活化的蛋白激酶（adenosine monophosphate activated protein kinase，AMPK）激活抑癌基因与张力蛋白同源10q丢失的磷酸酶基因（phosphatase and tensin homolog deleted on chromosome ten，PTEN）的转录等机制来调节mTOR信号通路相关分子，抑制mTOR活性，促使UNC-51激酶1（unc-51-like kinase1，ULK1）去磷酸化而增强自噬，亦可介导某些凋亡蛋白（Bcl-2、Bax等）及调控因子（DRAM、PUMA等）的信号通路参与自噬调节，但与mTOR通路的关系尚待研究；细胞质中的P53蛋白以非依赖转录的方式如降解P53、抑制RB1CC1/FIP200活性等，参与mTOR途径调控，抑制自噬。本文就P53蛋白介导mTOR信号途径影响肿瘤细胞自噬的研究进展作一综述。
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Supported by:Beijing E-Tiller Technology Development Co., Ltd.