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Volume 21,Issue 6,2014 Table of Contents
2014, 21(6):603-609.
DOI: 10.3872/j.issn.1007-385X.2014.6.001
Abstract:
MicoRNAs (miRNAs) are small non-coding RNA molecules of approximately 22 nucleotides in length, endogenously present in humans, animals, plants, and some viruses, which function as post-transcriptional regulators in numerous biological processes. MiRNAs can be either oncogenic or anti-oncogenic. Aberrant miRNA expression is the hallmarks of cancer and thus miRNA profiling may help monitor tumor growth and progression. Accumulating evidence suggest that circulating miRNAs, stably expressed in human peripheral blood, can accurately reflect the status of cancer, thus having great potential to serve as novel diagnostic and/or prognostic biomarkers for cancer. Conventionally, miRNA detection mainly involves Northern blotting, microarray, qRT-PCR, and lately next-generation sequencing. In recent years, significant improvement has been made in miRNA-based therapeutic strategies including miRNA mimicking and lipidosome-mediated miRNA delivery. However, some technological difficulties remain, which has restricted the wider application of these strategies in clinics. It is highly anticipated that miRNAs will play more significant roles in cancer diagnose and treatment in clinics when these difficulties are well managed.
MicoRNAs (miRNAs) are small non-coding RNA molecules of approximately 22 nucleotides in length, endogenously present in humans, animals, plants, and some viruses, which function as post-transcriptional regulators in numerous biological processes. MiRNAs can be either oncogenic or anti-oncogenic. Aberrant miRNA expression is the hallmarks of cancer and thus miRNA profiling may help monitor tumor growth and progression. Accumulating evidence suggest that circulating miRNAs, stably expressed in human peripheral blood, can accurately reflect the status of cancer, thus having great potential to serve as novel diagnostic and/or prognostic biomarkers for cancer. Conventionally, miRNA detection mainly involves Northern blotting, microarray, qRT-PCR, and lately next-generation sequencing. In recent years, significant improvement has been made in miRNA-based therapeutic strategies including miRNA mimicking and lipidosome-mediated miRNA delivery. However, some technological difficulties remain, which has restricted the wider application of these strategies in clinics. It is highly anticipated that miRNAs will play more significant roles in cancer diagnose and treatment in clinics when these difficulties are well managed.
2014, 21(6):610-616.
DOI: 10.3872/j.issn.1007-385X.2014.6.002
Abstract:
Objective: To investigate the effect of high-mobility group box 1 ( HMGB1 ) gene silencing on chemotherapeutic agent Doxorubicin (DOX)-induced autophagy and apoptosis in human gastric cancer BGC-823 cells. Methods: Cell viability, microtubule light chain 3 (LC3-Ⅰ, LC3-Ⅱ) protein content and autophagic vesicle formation in DOX-induced BGC-823 cells were assessed by MTT assay, Western blotting and confocal microscopy, respectively. BGC-823 cells were transfected with pSuper-shHMGBl (a shRNA targeting the HMGB1 gene) and pSuper-shNC (a control vector) respectively and subjected to G418 selection. BGC-823 cells stably expressing pSuper-shHMGBl or pSuper-shNC were treated with DOX. After treatment, autophagic vesicle formation was assessed by confocal microscopy. Cell viability and cell apoptosis were determined by the MTT assay and flow cytometry respectively. HMGB1, LC3, Bcl-2, Bcl-XL, Mcl-1 and cleaved caspase-3 proteins were assessed by Western blotting. Results: Treatment with DOX increased the protein content of LC3-Ⅱ and the formation of autophagic vesicles in BGC-823 cells ( P <0.01). As compared to the control, HMGB1 silencing inhibited DOX-induced autophagy ( P <0.01), promoted apoptosis (\[46.12±3.15\]% vs \[12.37±2.84\]%, P <0.05) and up-regulated the expression of cleaved caspase-3 in BGC-823 cells. Furthermore, antiapoptotic Mcl-1 protein degradation was increased in BGC-823 cells after induction of autophagy, however, Bcl-2 and Bcl-XL protein levels did not change significantly. HMGB1 gene silencing significantly attenuated the degradation of Mcl-1 protein. Conclusion: Autophagy can be induced by Doxorubicin in BGC-823 cells, which may contribute to DOX resistance. Knockdown of HMGB1 attenuates the DOX-induced autophagy and promotes apoptosis in gastric cancer cells.
Objective: To investigate the effect of high-mobility group box 1 ( HMGB1 ) gene silencing on chemotherapeutic agent Doxorubicin (DOX)-induced autophagy and apoptosis in human gastric cancer BGC-823 cells. Methods: Cell viability, microtubule light chain 3 (LC3-Ⅰ, LC3-Ⅱ) protein content and autophagic vesicle formation in DOX-induced BGC-823 cells were assessed by MTT assay, Western blotting and confocal microscopy, respectively. BGC-823 cells were transfected with pSuper-shHMGBl (a shRNA targeting the HMGB1 gene) and pSuper-shNC (a control vector) respectively and subjected to G418 selection. BGC-823 cells stably expressing pSuper-shHMGBl or pSuper-shNC were treated with DOX. After treatment, autophagic vesicle formation was assessed by confocal microscopy. Cell viability and cell apoptosis were determined by the MTT assay and flow cytometry respectively. HMGB1, LC3, Bcl-2, Bcl-XL, Mcl-1 and cleaved caspase-3 proteins were assessed by Western blotting. Results: Treatment with DOX increased the protein content of LC3-Ⅱ and the formation of autophagic vesicles in BGC-823 cells ( P <0.01). As compared to the control, HMGB1 silencing inhibited DOX-induced autophagy ( P <0.01), promoted apoptosis (\[46.12±3.15\]% vs \[12.37±2.84\]%, P <0.05) and up-regulated the expression of cleaved caspase-3 in BGC-823 cells. Furthermore, antiapoptotic Mcl-1 protein degradation was increased in BGC-823 cells after induction of autophagy, however, Bcl-2 and Bcl-XL protein levels did not change significantly. HMGB1 gene silencing significantly attenuated the degradation of Mcl-1 protein. Conclusion: Autophagy can be induced by Doxorubicin in BGC-823 cells, which may contribute to DOX resistance. Knockdown of HMGB1 attenuates the DOX-induced autophagy and promotes apoptosis in gastric cancer cells.
Fan Fangfang
,
Xu Hui
,
Sun Huiying
,
Liu Jiawei
,
Zhang Haiyuan
,
Li Yilan
,
Ning Xuelian
,
Bai Jing
,
Fu Songbin
,
Zhou Chunshui
2014, 21(6):617-623.
DOI: 10.3872/j.issn.1007-385X.2014.6.003
Abstract:
Objective:To evaluate the antitumor activity of a novel synthetic cationic peptide designated as AIK, and to elucidate the underlying mechanism(s) in vitro and in vivo . Methods：In experiments in vitro , the optimal dosage and treatment duration for AIK to exert its maximal cell proliferation inhibitory activity were determined by MTS cell proliferation assays in human leukemia HL-60 cells. And then, the antitumor activity of AIK at the optimal dosage and for the optimal duration of treatment was assessed in ten tumor cell lines (i.e., 95C, 95D, HL-60, HeLa, B95-8, HO-8910PM, HO-8910, SMMC-7721, U2OS, A549) and one human normal liver cell line HL-7702. The effects of AIK on HeLa cell apoptosis and morphology were examined by flow cytometry and microscopy respectively. In experiments in vivo, mouse liver cancer H22 cells were transplanted into the armpits of male Kunming mice. Thirty-six mice that developed tumors of a similar size as confirmed 5 days after H22 cell transplantation were randomized to receive subcutaneously PBS (vehicle control), low dose AIK (8 mg/\[kg?d\]), high dose AIK (15 mg/\[kg?d\]) and methotrexate (1 mg/\[kg?d\]) of as a positive control daily for 10 days, during which body weight was measured every day. Animals were sacrificed 24 h after the last treatment. Subcutaneous tumors were counted in each animal. Data on the weight and volume of the individual tumors were obtained and analyzed. Results： In vitro, AIK exhibited a potent cytotoxic activity against all types of cancer cells tested, in particular, the pulmonary giant cell carcinoma 95C cells, the leukemia HL-60 cells, and the cervical cancer HeLa cells. AIK treatment, as compared with vehicle control, resulted in significant increases in apoptosis (\[ 4.88± 0.57\]% vs \[0.51±0.19\]%, P <0.05) and necrosis (\[2.96±0.50\]% vs \[1.87±0.27\]%, P <0.05) in HeLa cells. Microscopic assessment showed necrotic phenomena including dilated, fragmented cell membrane and cell lysis in more than 50% of AIK treated H22 cells. In vivo, tumor volume and weight were significantly smaller in both AIK-and methotrexate-treated animals than in PBS-treated animals ( P <0.01). Body weight declined in methotrexate-treated animals but tended to increase in other 3 groups of animals 5 days after treatment. Nevertheless, the overall differences in body weight changes between the 4 groups of animals failed to reach a statistical significance ( P >0.05). Conclusion: The novel cationic peptide AIK possesses a potent antitumor activity both in vitro and in vivo , thus having a great therapeutic potential for various types of cancer.
Objective:To evaluate the antitumor activity of a novel synthetic cationic peptide designated as AIK, and to elucidate the underlying mechanism(s) in vitro and in vivo . Methods：In experiments in vitro , the optimal dosage and treatment duration for AIK to exert its maximal cell proliferation inhibitory activity were determined by MTS cell proliferation assays in human leukemia HL-60 cells. And then, the antitumor activity of AIK at the optimal dosage and for the optimal duration of treatment was assessed in ten tumor cell lines (i.e., 95C, 95D, HL-60, HeLa, B95-8, HO-8910PM, HO-8910, SMMC-7721, U2OS, A549) and one human normal liver cell line HL-7702. The effects of AIK on HeLa cell apoptosis and morphology were examined by flow cytometry and microscopy respectively. In experiments in vivo, mouse liver cancer H22 cells were transplanted into the armpits of male Kunming mice. Thirty-six mice that developed tumors of a similar size as confirmed 5 days after H22 cell transplantation were randomized to receive subcutaneously PBS (vehicle control), low dose AIK (8 mg/\[kg?d\]), high dose AIK (15 mg/\[kg?d\]) and methotrexate (1 mg/\[kg?d\]) of as a positive control daily for 10 days, during which body weight was measured every day. Animals were sacrificed 24 h after the last treatment. Subcutaneous tumors were counted in each animal. Data on the weight and volume of the individual tumors were obtained and analyzed. Results： In vitro, AIK exhibited a potent cytotoxic activity against all types of cancer cells tested, in particular, the pulmonary giant cell carcinoma 95C cells, the leukemia HL-60 cells, and the cervical cancer HeLa cells. AIK treatment, as compared with vehicle control, resulted in significant increases in apoptosis (\[ 4.88± 0.57\]% vs \[0.51±0.19\]%, P <0.05) and necrosis (\[2.96±0.50\]% vs \[1.87±0.27\]%, P <0.05) in HeLa cells. Microscopic assessment showed necrotic phenomena including dilated, fragmented cell membrane and cell lysis in more than 50% of AIK treated H22 cells. In vivo, tumor volume and weight were significantly smaller in both AIK-and methotrexate-treated animals than in PBS-treated animals ( P <0.01). Body weight declined in methotrexate-treated animals but tended to increase in other 3 groups of animals 5 days after treatment. Nevertheless, the overall differences in body weight changes between the 4 groups of animals failed to reach a statistical significance ( P >0.05). Conclusion: The novel cationic peptide AIK possesses a potent antitumor activity both in vitro and in vivo , thus having a great therapeutic potential for various types of cancer.
2014, 21(6):624-629.
DOI: 10.3872/j.issn.1007-385X.2014.6.004
Abstract:
Objective:To evaluate the cytotoxicity of dendritic cells-cytokine-induced killer cells (DC-CIK cells) derived from patients with remission leukemia or a non-remission disease against leukemia cells. Methods: Blood was sampled from 7 seven patients with refractory non-remission acute leukemia and 7 patients with remission leukemia who were admitted to the Department of Hematology, Hebei Medical University-Affiliated Second Hospital between April 25, 2013 and June 17, 2014. Peripheral blood mononuclear cells (PBMCs) were isolated and induced into DC-CIK cells by conventional methods. The proportions of CD3+CD8+ and CD3+CD56+ cells in the induced DC-CIK cell population were assessed by flow cytometry. Their proliferation rate was calculated and their killing activity against K562 cells and mononuclear cells were determined by CCK-8 assay. Results: DC-CIK cells derived from both patients with a remission disease and patients with a non-remission disease proliferated at similarly rapid rates ( P >0.05). Proportions of CD3+CD8+ and CD3+CD56+ cells increased dramatically with time in both groups of DC-CIK cells ( P <0.05) but there was no significant difference between the two groups ( P >0.05). After 15 days of induction, no leukemia cells were found and the proportion of CD34+ cells was significantly reduced in DC-CIK cell preparations as compared with untreated PBMCs (\[0.1±0.05\]% vs \[8.3±3.1\]%, P <0.05). At effector to target cell ratios from 5 ∶1 to 20 ∶1, DC-CIKs derived from both groups of patients displayed a similar cytotoxic activity against K562 cells ( P >0.05), but DC-CIKs derived from patients with a non-remission disease had a significantly higher cytotoxic activity against mononuclear cells ( P <0.05). Conclusion: DC-CIK cells derived from PBMCs of both patients with non-remission leukemia and patients with remission leukemia have similar proliferative activities, similar profiles of CD3+CD8+ and CD3+CD56+ expression and similar cytotoxic activities against K562 cells. However, DC-CIK cells from patients with non-remission leukemia display a higher sensitivity against autologous mononuclear cells.
Objective:To evaluate the cytotoxicity of dendritic cells-cytokine-induced killer cells (DC-CIK cells) derived from patients with remission leukemia or a non-remission disease against leukemia cells. Methods: Blood was sampled from 7 seven patients with refractory non-remission acute leukemia and 7 patients with remission leukemia who were admitted to the Department of Hematology, Hebei Medical University-Affiliated Second Hospital between April 25, 2013 and June 17, 2014. Peripheral blood mononuclear cells (PBMCs) were isolated and induced into DC-CIK cells by conventional methods. The proportions of CD3+CD8+ and CD3+CD56+ cells in the induced DC-CIK cell population were assessed by flow cytometry. Their proliferation rate was calculated and their killing activity against K562 cells and mononuclear cells were determined by CCK-8 assay. Results: DC-CIK cells derived from both patients with a remission disease and patients with a non-remission disease proliferated at similarly rapid rates ( P >0.05). Proportions of CD3+CD8+ and CD3+CD56+ cells increased dramatically with time in both groups of DC-CIK cells ( P <0.05) but there was no significant difference between the two groups ( P >0.05). After 15 days of induction, no leukemia cells were found and the proportion of CD34+ cells was significantly reduced in DC-CIK cell preparations as compared with untreated PBMCs (\[0.1±0.05\]% vs \[8.3±3.1\]%, P <0.05). At effector to target cell ratios from 5 ∶1 to 20 ∶1, DC-CIKs derived from both groups of patients displayed a similar cytotoxic activity against K562 cells ( P >0.05), but DC-CIKs derived from patients with a non-remission disease had a significantly higher cytotoxic activity against mononuclear cells ( P <0.05). Conclusion: DC-CIK cells derived from PBMCs of both patients with non-remission leukemia and patients with remission leukemia have similar proliferative activities, similar profiles of CD3+CD8+ and CD3+CD56+ expression and similar cytotoxic activities against K562 cells. However, DC-CIK cells from patients with non-remission leukemia display a higher sensitivity against autologous mononuclear cells.
Liang Ying
,
Zhou Danni
,
Fan Xiaohui
,
Song Dezhi
,
Gao Lingxi
,
Xiao Qing
,
Huang Panliu
,
Sun Pan
,
Zhang Bin
,
Lai Zhenping
2014, 21(6):630-634.
DOI: 10.3872/j.issn.1007-385X.2014.6.005
Abstract:
Objective: To evaluate the liver metastasis-inhibiting and immune-stimulating effects of Newcastle disease virus (NDV) strain 7793 in a mouse model of colon cancer. Methods: CT26 colon cancer cells (1×106 cells in 0.1 ml PBS) were injected into the subcapsules of the spleen of BALB/c mice aged 4-6 weeks through intra-peritoneal injection. Surviving mice bearing CT26 colon cancer cells were randomized to receive PBS (0.1 ml/d), NDV7793 (512 HU/kg) and fluorouracil (5-FU, 20 mg/kg), respectively, for 5 days. Body weight and general health status were recorded before colon cancer cell transplantation, before the designated treatments and on post-treatment days 1, 5, 10 and 15, respectively. On post-treatment day 16, animals were sacrificed. Liver metastasis was assessed gross pathology. Thymus and liver were collected. Histologic assessment was performed on paraffin sections of the liver after H-E staining. Thymus index and metastasis inhibition rate were calculated. IFN-γ levels in the liver were measured by ELISA. Results: The metastatic foci in colon cancer cell-bearing mice were significantly less ( P <0.05) after treatment with NDV (20.40±5.20) and 5-FU (205.50±19.21) respectively than with PBS (265.30±35.73). Liver metastasis inhibition rate was significantly higher for NDV7793 than for 5-FU group (75.4% vs 48.0%, P ＜0.05). The mean survival time of colon cancer cell-bearing mice was 30 days after NDV7793 treatment, 22 days after 5-FU treatment but only 17 days after PBS treatment ( P< 0.05). Compared with PBS control group and 5-FU group, the increased Thymus index and liver concentrations of IFN-γ were significantly higher after treatment with NDV 7793 than with 5-FU and PBS respectively ( P ＜0.05). Conclusion: NDV 7793 appears effective to inhibit liver metastasis of colon cancer cells. This effect may be mediated by elevated IFN-γ in liver microenvironment.
Objective: To evaluate the liver metastasis-inhibiting and immune-stimulating effects of Newcastle disease virus (NDV) strain 7793 in a mouse model of colon cancer. Methods: CT26 colon cancer cells (1×106 cells in 0.1 ml PBS) were injected into the subcapsules of the spleen of BALB/c mice aged 4-6 weeks through intra-peritoneal injection. Surviving mice bearing CT26 colon cancer cells were randomized to receive PBS (0.1 ml/d), NDV7793 (512 HU/kg) and fluorouracil (5-FU, 20 mg/kg), respectively, for 5 days. Body weight and general health status were recorded before colon cancer cell transplantation, before the designated treatments and on post-treatment days 1, 5, 10 and 15, respectively. On post-treatment day 16, animals were sacrificed. Liver metastasis was assessed gross pathology. Thymus and liver were collected. Histologic assessment was performed on paraffin sections of the liver after H-E staining. Thymus index and metastasis inhibition rate were calculated. IFN-γ levels in the liver were measured by ELISA. Results: The metastatic foci in colon cancer cell-bearing mice were significantly less ( P <0.05) after treatment with NDV (20.40±5.20) and 5-FU (205.50±19.21) respectively than with PBS (265.30±35.73). Liver metastasis inhibition rate was significantly higher for NDV7793 than for 5-FU group (75.4% vs 48.0%, P ＜0.05). The mean survival time of colon cancer cell-bearing mice was 30 days after NDV7793 treatment, 22 days after 5-FU treatment but only 17 days after PBS treatment ( P< 0.05). Compared with PBS control group and 5-FU group, the increased Thymus index and liver concentrations of IFN-γ were significantly higher after treatment with NDV 7793 than with 5-FU and PBS respectively ( P ＜0.05). Conclusion: NDV 7793 appears effective to inhibit liver metastasis of colon cancer cells. This effect may be mediated by elevated IFN-γ in liver microenvironment.
2014, 21(6):635-639.
DOI: 10.3872/j.issn.1007-385X.2014.6.006
Abstract:
Objective:To investigate the effects of small interference RNA (siRNA)-mediated TLR4 silencing on proliferation and invasion of human lung adenocarcinoma A549 cells. Methods: Three TLR4-specific siRNAs (siRNA722, siRNA1673 and siRNA2560) and one negative control siRNA were designed and chemically synthesized. One siRNA that could most effectively silence the TLR4 gene in A549 cells was chosen from the three test siRNAs based on their capacity of lowering TLR4 mRNA and protein levels revealed by real-time PCR and Western blotting analysis respectively. A549 cells were transfected with this most effective siRNA and the control siRNA respectively. Proliferation and invasion capacity of the transfectants were assessed by CCK-8 assay and Transwell assay, respectively. Results: TLR4-siRNA1673 was the most effective siRNA which could significantly inhibit the expression of TLR4 at both mRNA (0.45±0.03 vs 0.83±0.02, P <0.01) and protein (0.23±0.06 vs 0.98±0.09) levels at 48 h after transfection into A549 cells as compared with negative control siRNA. The proliferation of the TLR4-siRNA1673-transfected A549 cells was significantly inhibited compared with the NS-siRNA-transfected A549 cells ( P <0.05). Similarly, the invasion capacity was significantly lower in the TLR4-siRNA1673-transfected A549 cells than in the NS-siRNA-transfected cells (23.60±2.88 vs 59.80±5.54, P <0.01). Conclusion: Sequence-specific siRNA may effectively silence TLR4 expression and inhibit the proliferation and invasion ability of human lung carcinoma cells in vitro . This preliminary observation suggests that TLR4 may serve as a putative therapeutic target for lung cancer.
Objective:To investigate the effects of small interference RNA (siRNA)-mediated TLR4 silencing on proliferation and invasion of human lung adenocarcinoma A549 cells. Methods: Three TLR4-specific siRNAs (siRNA722, siRNA1673 and siRNA2560) and one negative control siRNA were designed and chemically synthesized. One siRNA that could most effectively silence the TLR4 gene in A549 cells was chosen from the three test siRNAs based on their capacity of lowering TLR4 mRNA and protein levels revealed by real-time PCR and Western blotting analysis respectively. A549 cells were transfected with this most effective siRNA and the control siRNA respectively. Proliferation and invasion capacity of the transfectants were assessed by CCK-8 assay and Transwell assay, respectively. Results: TLR4-siRNA1673 was the most effective siRNA which could significantly inhibit the expression of TLR4 at both mRNA (0.45±0.03 vs 0.83±0.02, P <0.01) and protein (0.23±0.06 vs 0.98±0.09) levels at 48 h after transfection into A549 cells as compared with negative control siRNA. The proliferation of the TLR4-siRNA1673-transfected A549 cells was significantly inhibited compared with the NS-siRNA-transfected A549 cells ( P <0.05). Similarly, the invasion capacity was significantly lower in the TLR4-siRNA1673-transfected A549 cells than in the NS-siRNA-transfected cells (23.60±2.88 vs 59.80±5.54, P <0.01). Conclusion: Sequence-specific siRNA may effectively silence TLR4 expression and inhibit the proliferation and invasion ability of human lung carcinoma cells in vitro . This preliminary observation suggests that TLR4 may serve as a putative therapeutic target for lung cancer.
2014, 21(6):640-646.
DOI: 10.3872/j.issn.1007-385X.2014.6.007
Abstract:
Objective:To investigate the effect of an injectable Chinese herbal medicine Tanreqing on chemotherapy- induced immunosuppression in a mouse model of lung carcinoma. Methods: A total of 40 C57 BL/6 mice were randomized to five groups: 1) normal control, 2) experimentally induced lung carcinoma, 3) Tanreqing treatment, 4) chemotherapy, and 5) chemotherapy combined with Tanreqing treatment. The designated treatments commenced 10 days after lung carcinoma induction and lasted for 5 days. At the end of the treatments, animals were sacrificed after periphery blood sampling, spleen and thymus were collected, and macrophages were isolated from peritoneal fluid. Thymus index and lymphocytes in peripheral blood were quantified. Apoptosis rate of thymus cells and proportion of CD3+, CD3-NK1.1+ and CD3+NK1.1+ cells in peripheral blood lymphocytes were determined by flow cytometry. The viability of T and B lymphocytes were assessed by MTT assay. Bcl-2 and Fas mRNA levels in thymus were measured by RT-PCR. The cytotoxicity of peritoneal macrophages and their secretion of IL-12 were determined with LDH and ELISA kits. Serum levels of IgG and C3 were measured by immune turbidimetry. Results: As compared with the chemotherapy group, the chemotherapy and Tanreqing combination group had significantly higher thymus index, lymphocyte count and T and B lymphocyte viabilities in the spleen ( P <0.01), significantly higher percentages of CD3+, CD3-NK1.1+ and CD3+NK1.1+ cells in peripheral-blood lymphocytes ( P <0.01), and significantly higher Bcl-2 mRNA abundance ( P <0.01) but significantly lower Fas mRNA abundance ( P <0.01).Peritoneal macrophages isolated from animals in the combination treatment group produced significantly higher levels of IL-2 and displayed significantly higher cytotoxicity than those isolated from animals in the chemotherapy group ( P <0.01). Moreover, serum levels of IgG and C3 were significantly higher in the combination group than in the chemotherapy group ( P <0.01). Conclusion: The injectable Chinese herbal medicine Tanreqing appears to have an immune protective and anti-tumor immunity-enhancing activities in lung carcinoma patients undergoing chemotherapy.
Objective:To investigate the effect of an injectable Chinese herbal medicine Tanreqing on chemotherapy- induced immunosuppression in a mouse model of lung carcinoma. Methods: A total of 40 C57 BL/6 mice were randomized to five groups: 1) normal control, 2) experimentally induced lung carcinoma, 3) Tanreqing treatment, 4) chemotherapy, and 5) chemotherapy combined with Tanreqing treatment. The designated treatments commenced 10 days after lung carcinoma induction and lasted for 5 days. At the end of the treatments, animals were sacrificed after periphery blood sampling, spleen and thymus were collected, and macrophages were isolated from peritoneal fluid. Thymus index and lymphocytes in peripheral blood were quantified. Apoptosis rate of thymus cells and proportion of CD3+, CD3-NK1.1+ and CD3+NK1.1+ cells in peripheral blood lymphocytes were determined by flow cytometry. The viability of T and B lymphocytes were assessed by MTT assay. Bcl-2 and Fas mRNA levels in thymus were measured by RT-PCR. The cytotoxicity of peritoneal macrophages and their secretion of IL-12 were determined with LDH and ELISA kits. Serum levels of IgG and C3 were measured by immune turbidimetry. Results: As compared with the chemotherapy group, the chemotherapy and Tanreqing combination group had significantly higher thymus index, lymphocyte count and T and B lymphocyte viabilities in the spleen ( P <0.01), significantly higher percentages of CD3+, CD3-NK1.1+ and CD3+NK1.1+ cells in peripheral-blood lymphocytes ( P <0.01), and significantly higher Bcl-2 mRNA abundance ( P <0.01) but significantly lower Fas mRNA abundance ( P <0.01).Peritoneal macrophages isolated from animals in the combination treatment group produced significantly higher levels of IL-2 and displayed significantly higher cytotoxicity than those isolated from animals in the chemotherapy group ( P <0.01). Moreover, serum levels of IgG and C3 were significantly higher in the combination group than in the chemotherapy group ( P <0.01). Conclusion: The injectable Chinese herbal medicine Tanreqing appears to have an immune protective and anti-tumor immunity-enhancing activities in lung carcinoma patients undergoing chemotherapy.
2014, 21(6):647-651.
DOI: 10.3872/j.issn.1007-385X.2014.6.008
Abstract:
Objective: To evaluate the safety and clinical efficacy of dendritic cells (DCs) combined with cytokine-induced killer (CIK) cells in the treatment of esophagus cancer and its effect on the lymphocyte subsets of peripheral blood mononuclear cells (PBMCs). Methods: PBMCs were collected from 20 patients with stage Ⅱ-Ⅳ esophagus cancer who were admitted to the Academy of Military Medical Sciences-Affiliated Hospital in Beijing between May 2011 and May 2013. Non-adherent PBMCs were induced to develop into CIK cells by IFN-γ, IL-2 500 U/ml and Anti-CD3 and adherent PBMCs were induced to develop into DCs by TNF-α, followed by transfection with surviving and muc-1 . On days 7, 9, 11, and 13, respectively, after PBMCs collection, DCs (3-10)×107 in 1 ml PBS) were given subcutaneously and days 11 and 13 CIK cells (2-15)×109 in 100 ml PBS. This treatment regimen was repeated at an interval of 3 months until substantial remission was achieved. Clinical outcomes and adverse effects were recorded during the treatment period. One week before treatment and one month after each treatment cycle, peripheral blood was collected and lymphocyte subsets were analyzed by flow cytometry. Results: Complete remission occurred in one case and partial remission in 6 cases while the disease remained at a stable state in 6 cases and at a progressive state in 7 cases after treatment. The overall response rate was 35% and the disease control rate was 65%. Treatment with CIKs and DCs significantly increased the percentage of both Th1 (\[14.5±13.3\]% vs \[3.6±5.9\]% and Th2 (\[1.0±0.7\]% vs \[0.6±0.5\]% as compared with the vehicle control ( P <0.05). No evident adverse events were observed. Conclusion: Infusion of DCs and CIK cells may improve the immunosuppression status and enhance the anti-tumor immunity in patients with stage Ⅱ-Ⅳ esophagus cancer, thereby having potential clinical implications.
Objective: To evaluate the safety and clinical efficacy of dendritic cells (DCs) combined with cytokine-induced killer (CIK) cells in the treatment of esophagus cancer and its effect on the lymphocyte subsets of peripheral blood mononuclear cells (PBMCs). Methods: PBMCs were collected from 20 patients with stage Ⅱ-Ⅳ esophagus cancer who were admitted to the Academy of Military Medical Sciences-Affiliated Hospital in Beijing between May 2011 and May 2013. Non-adherent PBMCs were induced to develop into CIK cells by IFN-γ, IL-2 500 U/ml and Anti-CD3 and adherent PBMCs were induced to develop into DCs by TNF-α, followed by transfection with surviving and muc-1 . On days 7, 9, 11, and 13, respectively, after PBMCs collection, DCs (3-10)×107 in 1 ml PBS) were given subcutaneously and days 11 and 13 CIK cells (2-15)×109 in 100 ml PBS. This treatment regimen was repeated at an interval of 3 months until substantial remission was achieved. Clinical outcomes and adverse effects were recorded during the treatment period. One week before treatment and one month after each treatment cycle, peripheral blood was collected and lymphocyte subsets were analyzed by flow cytometry. Results: Complete remission occurred in one case and partial remission in 6 cases while the disease remained at a stable state in 6 cases and at a progressive state in 7 cases after treatment. The overall response rate was 35% and the disease control rate was 65%. Treatment with CIKs and DCs significantly increased the percentage of both Th1 (\[14.5±13.3\]% vs \[3.6±5.9\]% and Th2 (\[1.0±0.7\]% vs \[0.6±0.5\]% as compared with the vehicle control ( P <0.05). No evident adverse events were observed. Conclusion: Infusion of DCs and CIK cells may improve the immunosuppression status and enhance the anti-tumor immunity in patients with stage Ⅱ-Ⅳ esophagus cancer, thereby having potential clinical implications.
2014, 21(6):652-657.
DOI: 10.3872/j.issn.1007-385X.2014.6.009
Abstract:
Objective:To determine the methylation status and expression of SOX7 gene in association with Wnt signaling and clinicopathological characteristics in gastric cardia adenocarcinoma (GCA). Methods: Paired specimens of primary tumor and corresponding adjacent tissues were collected from 130 patients who were diagnosed with GCA and underwent surgical ablation in the Thoracic Surgery, Fourth Hospital Affiliated to Hebei Medical University in Shijiazhuang between 2006 and 2012. Methylation of CpG sites within the SOX7 gene promoter was assessed by methylation specific PCR (MSP), SOX7 mRNA abundance by RT-PCR, and β-catenin protein by immunohistochemistry, in these speciments. Associations of the methylation status of the SOX7 gene promoter with SOX7 mRNA abundance, β-catenin protein contents, clinicopathologic variables and the history of the upper gastrointestinal cancers (UGIC), respectively, were assessed. Results: The frequency of SOX7 gene methylation was significantly higher in GCA tissue (57.7%) than that in the adjacent non-cancerous tissues ( P <0.01). The hypermethylation of this gene was correlated with lymph node metastasis ( P <0.05) but neither with pathological grade nor clinical stage ( P >0.05). SOX7 mRNA abundance was significantly lower in GCA tissue (0.414±0.054) than that in the corresponding adjacent tissues (0.695±0.034, P <0.01). The ectopic expression frequency of β-catenin was significantly different between GCA and adjacent tissue specimens (85.4% vs 43.1%, P <0.05). SOX7 mRNA and β-catenin protein levels were significantly correlated with the frequency of SOX7 gene methylation in GCA tissue ( P <0.05). In addition，the methylation status of the SOX7 gene was closely related with the history of UGIC ( P <0.01). Conclusion: Hypermethylation of CpG sites within the SOX7 gene promoter and the resultant aberration of canonical Wnt/β-catenin signaling may play an important role in the pathogenesis of GCA.
Objective:To determine the methylation status and expression of SOX7 gene in association with Wnt signaling and clinicopathological characteristics in gastric cardia adenocarcinoma (GCA). Methods: Paired specimens of primary tumor and corresponding adjacent tissues were collected from 130 patients who were diagnosed with GCA and underwent surgical ablation in the Thoracic Surgery, Fourth Hospital Affiliated to Hebei Medical University in Shijiazhuang between 2006 and 2012. Methylation of CpG sites within the SOX7 gene promoter was assessed by methylation specific PCR (MSP), SOX7 mRNA abundance by RT-PCR, and β-catenin protein by immunohistochemistry, in these speciments. Associations of the methylation status of the SOX7 gene promoter with SOX7 mRNA abundance, β-catenin protein contents, clinicopathologic variables and the history of the upper gastrointestinal cancers (UGIC), respectively, were assessed. Results: The frequency of SOX7 gene methylation was significantly higher in GCA tissue (57.7%) than that in the adjacent non-cancerous tissues ( P <0.01). The hypermethylation of this gene was correlated with lymph node metastasis ( P <0.05) but neither with pathological grade nor clinical stage ( P >0.05). SOX7 mRNA abundance was significantly lower in GCA tissue (0.414±0.054) than that in the corresponding adjacent tissues (0.695±0.034, P <0.01). The ectopic expression frequency of β-catenin was significantly different between GCA and adjacent tissue specimens (85.4% vs 43.1%, P <0.05). SOX7 mRNA and β-catenin protein levels were significantly correlated with the frequency of SOX7 gene methylation in GCA tissue ( P <0.05). In addition，the methylation status of the SOX7 gene was closely related with the history of UGIC ( P <0.01). Conclusion: Hypermethylation of CpG sites within the SOX7 gene promoter and the resultant aberration of canonical Wnt/β-catenin signaling may play an important role in the pathogenesis of GCA.
2014, 21(6):658-664.
DOI: 10.3872/j.issn.1007-385X.2014.6.010
Abstract:
Objective: To investigate the promoter region CpG island methylation status and protein levels of NDRG1 and NDRG2 in laryngeal carcinoma. Methods: Larynx tumor tissue ( n =45) and normal lung tissue ( n =18) specimens were collected from 45 patients with laryngeal squamous cell carcinoma (LSCC) who underwent surgery in the Department of Head and Neck Surgery, the Fourth Hospital of Hebei Medical University between January, 2010 and December, 2011. The promoter CpG island methylation status and protein levels of NDRG1 and NDRG2 in the collected specimens were quantified by methylation specific polymerase chain reaction and immunohistochemistry, respectively. Results: The methylation level of promoter CpG island was significantly higher in LSCC tissue and in normal larynx tissue for both NDRG1 (66.7% vs 33.3%, P <0.05) and NDRG2 (53.3% vs 22.2%, P <0.05) genes. The proportion of NDRG1 and NDRG2 promoter methylation in LSCC tissue specimens was associated with lymph node metastasis and clinical stage ( P <0.05), but neither with pathological grade and clinical classification of the lesion nor with smoking history, age and sex ( P >0.05). NDRG1 protein was detected in 37.8% (17/45) of LSCC specimens and in 88.9% (16/18) of normal tissue specimens ( P <0.01) and NDRG2 protein in 33.3% (15/45) of LSCC specimens and in 83.3% (15/18) of normal larynx tissue specimens ( P <0.01). Like promoter methylation status, NDRG1 and NDRG2 protein levels in LSCC tissue were associated with lymph node metastasis and clinical stage ( P <0.05), but not with pathological grading and clinical classification of the lesions, smoking history, age and sex ( P >0.05). CpG island methylation in the promoter region was negatively correlated with protein expression for both NDRG1 ( r 1＝-0.713, P <0.01) and NDRG2 ( r 2＝-0.472 , P <0.01) genes. Conclusion: Both NDRG1 and NDRG2 promoters are hypermethylated, resulting in decreased NDRG1 and NDRG2 protein expression, in LSCC tissue. This observation suggests that anti-methylation of CpG island in the promoter region of the NDRG1 and NDRG2 genes may offer a novel therapeutic strategy for LSCC.
Objective: To investigate the promoter region CpG island methylation status and protein levels of NDRG1 and NDRG2 in laryngeal carcinoma. Methods: Larynx tumor tissue ( n =45) and normal lung tissue ( n =18) specimens were collected from 45 patients with laryngeal squamous cell carcinoma (LSCC) who underwent surgery in the Department of Head and Neck Surgery, the Fourth Hospital of Hebei Medical University between January, 2010 and December, 2011. The promoter CpG island methylation status and protein levels of NDRG1 and NDRG2 in the collected specimens were quantified by methylation specific polymerase chain reaction and immunohistochemistry, respectively. Results: The methylation level of promoter CpG island was significantly higher in LSCC tissue and in normal larynx tissue for both NDRG1 (66.7% vs 33.3%, P <0.05) and NDRG2 (53.3% vs 22.2%, P <0.05) genes. The proportion of NDRG1 and NDRG2 promoter methylation in LSCC tissue specimens was associated with lymph node metastasis and clinical stage ( P <0.05), but neither with pathological grade and clinical classification of the lesion nor with smoking history, age and sex ( P >0.05). NDRG1 protein was detected in 37.8% (17/45) of LSCC specimens and in 88.9% (16/18) of normal tissue specimens ( P <0.01) and NDRG2 protein in 33.3% (15/45) of LSCC specimens and in 83.3% (15/18) of normal larynx tissue specimens ( P <0.01). Like promoter methylation status, NDRG1 and NDRG2 protein levels in LSCC tissue were associated with lymph node metastasis and clinical stage ( P <0.05), but not with pathological grading and clinical classification of the lesions, smoking history, age and sex ( P >0.05). CpG island methylation in the promoter region was negatively correlated with protein expression for both NDRG1 ( r 1＝-0.713, P <0.01) and NDRG2 ( r 2＝-0.472 , P <0.01) genes. Conclusion: Both NDRG1 and NDRG2 promoters are hypermethylated, resulting in decreased NDRG1 and NDRG2 protein expression, in LSCC tissue. This observation suggests that anti-methylation of CpG island in the promoter region of the NDRG1 and NDRG2 genes may offer a novel therapeutic strategy for LSCC.
2014, 21(6):665-668.
DOI: 10.3872/j.issn.1007-385X.2014.6.011
Abstract:
Objective:The aim of this study was to analyze the expression at the protein level and clinic significance of ezrin and CD44 in renal cell carcinoma (RCC). Methods: Paraffin sections were prepared from renal cell carcinoma ( n =56 ) and adjacent tissue ( n =17) specimens collected from 56 RCC patients who were admitted to the Urology Department, Hunan Cancer Hospital between February 2012 and November 2012. Ezrin and CD44 proteins in these sections were assessed immunohistochemical staining. The correlation of ezrin and CD44 protein levels with the clinicopathological features of carcinoma lesions were analyzed. Results: Ezrin was detected in 60.7% (34/56) of RCC tissue specimens but in 29.4% (5/17) of the corresponding normal tissue specimens ( P <0.05). CD44 was detected in 64.3% (36/56) of RCC tissue specimens but in 0 out of 17 normal tissue specimens ( P <0.001). Both ezrin and CD44 proteins were correlated with the histological grade and clinical stage of RCC ( P ＜0.05). In addition, ezrin in tumors >7.0 cm was significantly higher than that in tumors ≤7.0 cm ( P ＜0.05) whereas CD44 was not significantly different between tumors >7.0 cm and tumors ≤7.0 cm (P＞0.05). The rates of both ezrin- and CD44-positive staining were not significantly different between specimens collected from male and female patients ( P >0.05). Ezrin was positively correlated with CD44 in RCC ( r =0.427， P <0.01). Conclusion: Both ezrin and CD44 are positively correlated with the severity of renal cell carcinoma, indicating these two proteins may play an important part in the development and progression of RCC.
Objective:The aim of this study was to analyze the expression at the protein level and clinic significance of ezrin and CD44 in renal cell carcinoma (RCC). Methods: Paraffin sections were prepared from renal cell carcinoma ( n =56 ) and adjacent tissue ( n =17) specimens collected from 56 RCC patients who were admitted to the Urology Department, Hunan Cancer Hospital between February 2012 and November 2012. Ezrin and CD44 proteins in these sections were assessed immunohistochemical staining. The correlation of ezrin and CD44 protein levels with the clinicopathological features of carcinoma lesions were analyzed. Results: Ezrin was detected in 60.7% (34/56) of RCC tissue specimens but in 29.4% (5/17) of the corresponding normal tissue specimens ( P <0.05). CD44 was detected in 64.3% (36/56) of RCC tissue specimens but in 0 out of 17 normal tissue specimens ( P <0.001). Both ezrin and CD44 proteins were correlated with the histological grade and clinical stage of RCC ( P ＜0.05). In addition, ezrin in tumors >7.0 cm was significantly higher than that in tumors ≤7.0 cm ( P ＜0.05) whereas CD44 was not significantly different between tumors >7.0 cm and tumors ≤7.0 cm (P＞0.05). The rates of both ezrin- and CD44-positive staining were not significantly different between specimens collected from male and female patients ( P >0.05). Ezrin was positively correlated with CD44 in RCC ( r =0.427， P <0.01). Conclusion: Both ezrin and CD44 are positively correlated with the severity of renal cell carcinoma, indicating these two proteins may play an important part in the development and progression of RCC.
2014, 21(6):669-674.
DOI: 10.3872/j.issn.1007-385X.2014.6.012
Abstract:
Objective:To analyze the expression of cyclin G2 (CCNG2) in thyroid carcinoma tissue specimens and evaluate the effect of CCNG2 on the proliferation and apoptosis of thyroid carcinoma SW579 cells in vitro . Methods: Sixty-three patients diagnosed with thyroid carcinoma in the Department of Endocrinology of Tangshan Workers Hospital between 2003 and 2008 were recruited. Biopsy specimens were collected from primary tumors and the corresponding adjacent tissues. CCNG2 protein presence and content in these specimens were determined by immunohistochemistry and Western blotting. The relationships of CCNG2 expression with clinicopathologic variables and patient survival were assessed. To evaluate the effect of CCNG2 on thyroid carcinoma cell proliferation and apoptosis in vitro , SW579 cells were transfected with a lentiviral vector expressing CCNG2 and the transfectants were then subjected to reverse transcription-polymerase chain reaction (RT-PCR) determination of CCNG2 mRNA abundance, Western blotting analysis of CCNG2 protein content, MTT assay of cell viability and flow cytometric assessment of apoptosis, respectively. Results: CCNG2 protein was detected at significantly lower levels in thyroid cancer tissue than in the paraneoplastic tissues (0.343±0.033 vs 0.713±0.062, P <0.05). While the level of CCNG2 protein was not correlated with gender, age, and tumor size ( P >0.05), it was significantly correlated with lymph node metastasis, clinic stage, histological grade and 5-year overall survival ( P <0.05). Overexpression of CCNG2 significantly lowered SW579 cell viability and enhanced SW579 cell apoptosis (15.1±2.3)% as compared with the control (5.6±1.1 in vitro , P <0.05). Conclusion: CCNG2 protein content is significantly lower in thyroid carcinoma tissue than in peritumor tissue. Overexpression of CCNG2 in thyroid carcinoma cells in vitro results in a significant inhibition of proliferation and a significant enhancement of apoptosis. These observations suggest that CCNG2 may serve as a potential therapeutic target for thyroid carcinoma.
Objective:To analyze the expression of cyclin G2 (CCNG2) in thyroid carcinoma tissue specimens and evaluate the effect of CCNG2 on the proliferation and apoptosis of thyroid carcinoma SW579 cells in vitro . Methods: Sixty-three patients diagnosed with thyroid carcinoma in the Department of Endocrinology of Tangshan Workers Hospital between 2003 and 2008 were recruited. Biopsy specimens were collected from primary tumors and the corresponding adjacent tissues. CCNG2 protein presence and content in these specimens were determined by immunohistochemistry and Western blotting. The relationships of CCNG2 expression with clinicopathologic variables and patient survival were assessed. To evaluate the effect of CCNG2 on thyroid carcinoma cell proliferation and apoptosis in vitro , SW579 cells were transfected with a lentiviral vector expressing CCNG2 and the transfectants were then subjected to reverse transcription-polymerase chain reaction (RT-PCR) determination of CCNG2 mRNA abundance, Western blotting analysis of CCNG2 protein content, MTT assay of cell viability and flow cytometric assessment of apoptosis, respectively. Results: CCNG2 protein was detected at significantly lower levels in thyroid cancer tissue than in the paraneoplastic tissues (0.343±0.033 vs 0.713±0.062, P <0.05). While the level of CCNG2 protein was not correlated with gender, age, and tumor size ( P >0.05), it was significantly correlated with lymph node metastasis, clinic stage, histological grade and 5-year overall survival ( P <0.05). Overexpression of CCNG2 significantly lowered SW579 cell viability and enhanced SW579 cell apoptosis (15.1±2.3)% as compared with the control (5.6±1.1 in vitro , P <0.05). Conclusion: CCNG2 protein content is significantly lower in thyroid carcinoma tissue than in peritumor tissue. Overexpression of CCNG2 in thyroid carcinoma cells in vitro results in a significant inhibition of proliferation and a significant enhancement of apoptosis. These observations suggest that CCNG2 may serve as a potential therapeutic target for thyroid carcinoma.
2014, 21(6):675-679.
DOI: 10.3872/j.issn.1007-385X.2014.6.013
Abstract:
Objective: To evaluate the clinicopathological, diagnostic and prognostic signficance of kallikrein (KLK) 10 and vascular endothelial growth factor (VEGF) expression in ovarian carcinoma. Methods: Paraffin-embedded sections were prepared from 45 malignant, 10 benign and 12 borderline ovarian tumor tissue specimens collected from patients seeking care in the Department of Gynecology and Obstetrics of Nantong University-Affiliated Hospital between January 2004 and January 2009. Immunohistochemical staining was performed to detect KLK10 and VEGF proteins in these tissue sections. The correlation of KLK10 and VEGF protein levels with the clinicopathological features and prognosis of ovarian carcinoma were analysed. Results: Malignant ovarian tumor specimens, as compared with benign and borderline tumor specimens, had significantly higher percentages of KLK10-positive staining (86.7% vs 10.0%, 58.3%; P <0.05) and VEGF-positive staining (82.2% vs 20.0%, 41.7%, P <0.05). The proportion of KLK10-and VEGF-positive cells in the xxx tissue was significantly correlated with clinical stage, pathologic grade, lymphatic metastasis and 5-year survival rate ( P <0.05), but was independent of age, histological type, serum level of CA125, ascetic fluid or remnant tumor diameter ( P >0.05). The expression of KLK10 was positively correlated with the expression of VEGF ( r =0.5279, P =0.043). Conclusion: Both KLK10 and VEGF are highly expressed and positively correlated in ovarian carcinoma and thereby may serve as putative prognostic markers of the disease.
Objective: To evaluate the clinicopathological, diagnostic and prognostic signficance of kallikrein (KLK) 10 and vascular endothelial growth factor (VEGF) expression in ovarian carcinoma. Methods: Paraffin-embedded sections were prepared from 45 malignant, 10 benign and 12 borderline ovarian tumor tissue specimens collected from patients seeking care in the Department of Gynecology and Obstetrics of Nantong University-Affiliated Hospital between January 2004 and January 2009. Immunohistochemical staining was performed to detect KLK10 and VEGF proteins in these tissue sections. The correlation of KLK10 and VEGF protein levels with the clinicopathological features and prognosis of ovarian carcinoma were analysed. Results: Malignant ovarian tumor specimens, as compared with benign and borderline tumor specimens, had significantly higher percentages of KLK10-positive staining (86.7% vs 10.0%, 58.3%; P <0.05) and VEGF-positive staining (82.2% vs 20.0%, 41.7%, P <0.05). The proportion of KLK10-and VEGF-positive cells in the xxx tissue was significantly correlated with clinical stage, pathologic grade, lymphatic metastasis and 5-year survival rate ( P <0.05), but was independent of age, histological type, serum level of CA125, ascetic fluid or remnant tumor diameter ( P >0.05). The expression of KLK10 was positively correlated with the expression of VEGF ( r =0.5279, P =0.043). Conclusion: Both KLK10 and VEGF are highly expressed and positively correlated in ovarian carcinoma and thereby may serve as putative prognostic markers of the disease.
2014, 21(6):680-686.
DOI: 10.3872/j.issn.1007-385X.2014.6.014
Abstract:
Objective:To investigate the effects of different culture media and different sources of serum on proliferative and functional activities of cytokine-induced killer (CIK) cells. Methods: Mononuclear cells were isolated from peripheral blood of 6 lung cancer patients and were induced to differentiate into CIK cells in 8 different culture conditions: GT-T551+autologous serum, GT-T551 + serum from a healthy individual, GT-T551 + FBS, GT-T551, RPMI 1640 + autologous, RPMI 1640 + serum from a healthy individual, RPMI 1640 + FBS, and RPMI 1640. After induction for 6 days, cell proliferation was assessed carboxyfluorescein diacetate succinimidyl sster (CFSE) labeling assay, and CD3+CD8+ cell proportion, CD3+CD4+ T cell expression of Granzyme B and IFN-γ and perforin and the expression of CD107a on CD3+CD56+, CD3+CD4+ and CD3+CD8+ T cells in the presence of leukemia NB4 and K562 cells as targets were assessed by flow cytometry. Results: The proliferation index of CIK cells cultured in GT-T551 medium supplemented with autologous serum was significantly higher than that in other culture conditions ( P <0.05). The expression of Granzyme B on CD4+ T cells was significantly higher ( P <0.01) in the GT-T551+autologous serum group \[22.85±3.50\]% than in the GT-T551+healthy serum group \[13.28±1.75\]% and significantly higher ( P <0.01) in the RPMI 1640+autologous serum group \[22.57±3.45\]% than in the RPMI 1640+healthy serum group\[15.37±4.08\]%. Furthermore, the amount of IFN-γ secreted from CD8+ T cells was significantly higher in the GT-T551 + autologous serum groups compared to GT-T551+healthy serum group. In the presence of killing NB4 and K562 cells, the expression of CD107a on the CD3+CD56+NKT cells was significantly higher in the GT-T551+autologous serum group (\[7.10±1.94\]%, \[8.00±1.82\]%) than in the GT-T551+healthy serum group (\[2.73±0.79\]%, \[3.03±0.78)%; P <0.01) and in the RPMI 1640+autologous serum group (\[4.45±1.96\]%, \[3.30±1.47\]%; all P <0.01). Conclusion: GT-T551 medium supplemented with autologous serum appears the optimal in vitro system to stimulate proliferation, cytokine secretion and enhance the cytotoxicity of CIK cells. This finding may have some clinical significance in CIK therapy.
Objective:To investigate the effects of different culture media and different sources of serum on proliferative and functional activities of cytokine-induced killer (CIK) cells. Methods: Mononuclear cells were isolated from peripheral blood of 6 lung cancer patients and were induced to differentiate into CIK cells in 8 different culture conditions: GT-T551+autologous serum, GT-T551 + serum from a healthy individual, GT-T551 + FBS, GT-T551, RPMI 1640 + autologous, RPMI 1640 + serum from a healthy individual, RPMI 1640 + FBS, and RPMI 1640. After induction for 6 days, cell proliferation was assessed carboxyfluorescein diacetate succinimidyl sster (CFSE) labeling assay, and CD3+CD8+ cell proportion, CD3+CD4+ T cell expression of Granzyme B and IFN-γ and perforin and the expression of CD107a on CD3+CD56+, CD3+CD4+ and CD3+CD8+ T cells in the presence of leukemia NB4 and K562 cells as targets were assessed by flow cytometry. Results: The proliferation index of CIK cells cultured in GT-T551 medium supplemented with autologous serum was significantly higher than that in other culture conditions ( P <0.05). The expression of Granzyme B on CD4+ T cells was significantly higher ( P <0.01) in the GT-T551+autologous serum group \[22.85±3.50\]% than in the GT-T551+healthy serum group \[13.28±1.75\]% and significantly higher ( P <0.01) in the RPMI 1640+autologous serum group \[22.57±3.45\]% than in the RPMI 1640+healthy serum group\[15.37±4.08\]%. Furthermore, the amount of IFN-γ secreted from CD8+ T cells was significantly higher in the GT-T551 + autologous serum groups compared to GT-T551+healthy serum group. In the presence of killing NB4 and K562 cells, the expression of CD107a on the CD3+CD56+NKT cells was significantly higher in the GT-T551+autologous serum group (\[7.10±1.94\]%, \[8.00±1.82\]%) than in the GT-T551+healthy serum group (\[2.73±0.79\]%, \[3.03±0.78)%; P <0.01) and in the RPMI 1640+autologous serum group (\[4.45±1.96\]%, \[3.30±1.47\]%; all P <0.01). Conclusion: GT-T551 medium supplemented with autologous serum appears the optimal in vitro system to stimulate proliferation, cytokine secretion and enhance the cytotoxicity of CIK cells. This finding may have some clinical significance in CIK therapy.
2014, 21(6):687-689.
DOI: 10.3872/j.issn.1007-385X.2014.6.015
Abstract:
目的：探讨非小细胞肺癌（non-small cell lung cancer，NSCLC）中表皮生长因子受体 (epidermal growth factor receptor， EGFR ）基因突变与核苷酸切除修复交叉互补基因1（excision repair cross-complementing 1， ERCC1 ） mRNA 表达之间的关系。 方法：收集2012年6月1日至2012年12月31日在济南军区总医院肿瘤科经组织病理确诊为NSCLC患者的肿瘤组织标本41例，应用突变扩增阻滞系统(amplification refractory mutation system, ARMS)检测 EGFR 基因突变，采用RT-PCR方法检测 ERCC1 mRNA 表达，应用Spearman相关性检验对 EGFR 突变状态与 ERCC1 mRNA 的表达进行相关性分析。 结果：在41例患者中， EGFR 突变21例， ERCC1 mRNA 高、中、低表达率分别为19.1%（4/21）、57.1%（12/21）和23.8%（5/21）， EGFR 基因突变与 ERCC1 mRNA 表达呈显著相关（ P <0.01）。结论: NSCLC组织中 EGFR 基因突变与 ERCC1 mRNA 表达具有显著相关性。
目的：探讨非小细胞肺癌（non-small cell lung cancer，NSCLC）中表皮生长因子受体 (epidermal growth factor receptor， EGFR ）基因突变与核苷酸切除修复交叉互补基因1（excision repair cross-complementing 1， ERCC1 ） mRNA 表达之间的关系。 方法：收集2012年6月1日至2012年12月31日在济南军区总医院肿瘤科经组织病理确诊为NSCLC患者的肿瘤组织标本41例，应用突变扩增阻滞系统(amplification refractory mutation system, ARMS)检测 EGFR 基因突变，采用RT-PCR方法检测 ERCC1 mRNA 表达，应用Spearman相关性检验对 EGFR 突变状态与 ERCC1 mRNA 的表达进行相关性分析。 结果：在41例患者中， EGFR 突变21例， ERCC1 mRNA 高、中、低表达率分别为19.1%（4/21）、57.1%（12/21）和23.8%（5/21）， EGFR 基因突变与 ERCC1 mRNA 表达呈显著相关（ P <0.01）。结论: NSCLC组织中 EGFR 基因突变与 ERCC1 mRNA 表达具有显著相关性。
2014, 21(6):690-692.
DOI: 10.3872/j.issn.1007-385X.2014.6.016
Abstract:
目的：观察脐血和乙型肝炎病毒感染肝癌患者外周血诱导的CIK细胞的体外增殖能力及杀伤活性的特点。方法： 分别采集解放军第105医院妇产科住院的健康产妇剖宫产胎儿的脐带血5份（A组）、感染科住院的合并乙型肝炎病毒感染的原发性肝癌患者自体外周血15份（B组）、无乙型肝炎病毒感染的原发性肝癌患者自体外周血5份（C组），采用Ficoll两步分离法分离出单个核细胞，细胞因子诱导培养成细胞因子诱导的杀伤(ClK)细胞，流式细胞仪检测CIK细胞的免疫表型CD3＼CD8＼CD16＼CD56，MIT法测定其对白血病K562细胞的杀伤活性。结果：体外培养15 d时， C组CIK细胞体外增殖倍数与A组相似\[（577.24±202.4）vs（600.93±249.1）倍， P >0.05\]，该两组均显著高于B组\[（385.16±117.3）倍， P <0.05\]；A组CIK细胞杀伤活性显著高于B组和C组\[（77.3±5.1）% vs （44.1±3.4）%、（54.5±3.7）%， P <0.01\]。结论： 脐血来源的CIK细胞体外增殖快、杀伤活性强；合并有乙型肝炎病毒感染的肝癌患者CIK细胞体外增殖能力和杀伤能力明显降低，不宜接受自体CIK细胞移植。
目的：观察脐血和乙型肝炎病毒感染肝癌患者外周血诱导的CIK细胞的体外增殖能力及杀伤活性的特点。方法： 分别采集解放军第105医院妇产科住院的健康产妇剖宫产胎儿的脐带血5份（A组）、感染科住院的合并乙型肝炎病毒感染的原发性肝癌患者自体外周血15份（B组）、无乙型肝炎病毒感染的原发性肝癌患者自体外周血5份（C组），采用Ficoll两步分离法分离出单个核细胞，细胞因子诱导培养成细胞因子诱导的杀伤(ClK)细胞，流式细胞仪检测CIK细胞的免疫表型CD3＼CD8＼CD16＼CD56，MIT法测定其对白血病K562细胞的杀伤活性。结果：体外培养15 d时， C组CIK细胞体外增殖倍数与A组相似\[（577.24±202.4）vs（600.93±249.1）倍， P >0.05\]，该两组均显著高于B组\[（385.16±117.3）倍， P <0.05\]；A组CIK细胞杀伤活性显著高于B组和C组\[（77.3±5.1）% vs （44.1±3.4）%、（54.5±3.7）%， P <0.01\]。结论： 脐血来源的CIK细胞体外增殖快、杀伤活性强；合并有乙型肝炎病毒感染的肝癌患者CIK细胞体外增殖能力和杀伤能力明显降低，不宜接受自体CIK细胞移植。
2014, 21(6):693-697.
DOI: 10.3872/j.issn.1007-385X.2014.6.017
Abstract:
吲哚胺2,3-双加氧酶（indoleamine 2,3-dioxygenase, IDO）是一种免疫调节酶，能够抑制效应T细胞和自然杀伤（natural killer, NK)细胞的增殖与功能，并且与调节性T细胞形成了正反馈调节环路，抑制微环境内的抗肿瘤免疫应答，在肿瘤的免疫逃逸中发挥着重要作用。肿瘤细胞自身可以表达IDO，还能够募集表达IDO的树突状细胞（dendritic cells, DC）进入肿瘤微环境，在肿瘤浸润组织和引流淋巴结中都可以发现高水平表达的IDO。IDO的免疫抑制效应可以被其竞争性抑制剂1-甲 基色氨酸（1-methyl-tryptophan, 1-MT）所阻断，其异构体D-1-MT在前临床试验中可以干扰肿瘤细胞的免疫逃逸，显著增强放疗、化疗和免疫治疗的疗效，有望成为一种新的抗肿瘤药物。
吲哚胺2,3-双加氧酶（indoleamine 2,3-dioxygenase, IDO）是一种免疫调节酶，能够抑制效应T细胞和自然杀伤（natural killer, NK)细胞的增殖与功能，并且与调节性T细胞形成了正反馈调节环路，抑制微环境内的抗肿瘤免疫应答，在肿瘤的免疫逃逸中发挥着重要作用。肿瘤细胞自身可以表达IDO，还能够募集表达IDO的树突状细胞（dendritic cells, DC）进入肿瘤微环境，在肿瘤浸润组织和引流淋巴结中都可以发现高水平表达的IDO。IDO的免疫抑制效应可以被其竞争性抑制剂1-甲 基色氨酸（1-methyl-tryptophan, 1-MT）所阻断，其异构体D-1-MT在前临床试验中可以干扰肿瘤细胞的免疫逃逸，显著增强放疗、化疗和免疫治疗的疗效，有望成为一种新的抗肿瘤药物。
2014, 21(6):698-702.
DOI: 10.3872/j.issn.1007-385X.2014.6.018
Abstract:
肿瘤转移与多种细胞信号通路作用有关，其中与JAK2/STAT3/SOCS3通路关系尤为密切，JAK2/STAT3信号通路的激活参与了肿瘤发生、发展、侵袭和转移等多个环节。细胞因子信号转导抑制蛋白3（suppressors of cytokine signaling 3，SOCS3）负性调控JAK2/STAT3通路，进而抑制肿瘤的增殖和生长；信号转导与转录激活因子3（signal transducer and activator of transcription 3，STAT3）信号通路的激活促成了肿瘤炎性微环境的形成，参与了肿瘤血管生成、上皮间质转化和细胞外基质降解等多个环节，在肿瘤的侵袭和转移过程中发挥重要作用。本文着重对JAK2/STAT3/SOCS3信号通路与肿瘤转移的关系进行综述，针对JAK2/STAT3/SOCS3细胞信号动态网络在肿瘤中作用机制研究和药物设计为肿瘤治疗提供了新方向。
肿瘤转移与多种细胞信号通路作用有关，其中与JAK2/STAT3/SOCS3通路关系尤为密切，JAK2/STAT3信号通路的激活参与了肿瘤发生、发展、侵袭和转移等多个环节。细胞因子信号转导抑制蛋白3（suppressors of cytokine signaling 3，SOCS3）负性调控JAK2/STAT3通路，进而抑制肿瘤的增殖和生长；信号转导与转录激活因子3（signal transducer and activator of transcription 3，STAT3）信号通路的激活促成了肿瘤炎性微环境的形成，参与了肿瘤血管生成、上皮间质转化和细胞外基质降解等多个环节，在肿瘤的侵袭和转移过程中发挥重要作用。本文着重对JAK2/STAT3/SOCS3信号通路与肿瘤转移的关系进行综述，针对JAK2/STAT3/SOCS3细胞信号动态网络在肿瘤中作用机制研究和药物设计为肿瘤治疗提供了新方向。
2014, 21(6):703-706.
DOI: 10.3872/j.issn.1007-385X.2014.6.019
Abstract:
溶瘤病毒（oncolyticviru，OV）能够特异性地在肿瘤细胞内复制、扩增并杀死肿瘤细胞，而其正常体细胞中不能复制，以此为基础的OV抗肿瘤治疗方法近年受到广泛关注。肿瘤微环境是肿瘤生长的生态位，在肿瘤的生长发展中具有极其重要的作用，利用溶瘤病毒靶向肿瘤微环境可以从多方面抑制肿瘤的发展。肿瘤微环境中含有的大量生长因子、细胞因子、免疫细胞、肿瘤浸润细胞及其胞外基质等均会抑制溶瘤病毒在肿瘤细胞中的复制增殖，通过靶向这些影响因素，有可能开发出多种改造肿瘤微环境的手段，进而提高OV的溶瘤效率。
溶瘤病毒（oncolyticviru，OV）能够特异性地在肿瘤细胞内复制、扩增并杀死肿瘤细胞，而其正常体细胞中不能复制，以此为基础的OV抗肿瘤治疗方法近年受到广泛关注。肿瘤微环境是肿瘤生长的生态位，在肿瘤的生长发展中具有极其重要的作用，利用溶瘤病毒靶向肿瘤微环境可以从多方面抑制肿瘤的发展。肿瘤微环境中含有的大量生长因子、细胞因子、免疫细胞、肿瘤浸润细胞及其胞外基质等均会抑制溶瘤病毒在肿瘤细胞中的复制增殖，通过靶向这些影响因素，有可能开发出多种改造肿瘤微环境的手段，进而提高OV的溶瘤效率。
2014, 21(6):707-711.
DOI: 10.3872/j.issn.1007-385X.2014.6.020
Abstract:
人抗原R（human antigen R，HuR）属于果蝇胚胎致死异常视觉（embryonic lethal abnormal vision，ELAV）家族的RNA结合蛋白（RNA-binding protein，RBP），1996年克隆成功，随后发现其在神经发育和细胞分化中具有重要作用。近年发现，HuR通过增强肿瘤相关RNA的稳定性而参与了肿瘤发生、发展、转移和血管生成，具有类似原癌基因的功能；但也有研究显示HuR具有双重作用，既能促进肿瘤细胞对化疗药物多柔比星、吉西他滨的敏感性，又与肿瘤细胞对顺铂、紫杉醇类药物耐药相关。本文结合笔者所在课题组前期对HuR的研究基础，就HuR在肿瘤多药耐药及药物敏感性中的作用简要作一综述。
人抗原R（human antigen R，HuR）属于果蝇胚胎致死异常视觉（embryonic lethal abnormal vision，ELAV）家族的RNA结合蛋白（RNA-binding protein，RBP），1996年克隆成功，随后发现其在神经发育和细胞分化中具有重要作用。近年发现，HuR通过增强肿瘤相关RNA的稳定性而参与了肿瘤发生、发展、转移和血管生成，具有类似原癌基因的功能；但也有研究显示HuR具有双重作用，既能促进肿瘤细胞对化疗药物多柔比星、吉西他滨的敏感性，又与肿瘤细胞对顺铂、紫杉醇类药物耐药相关。本文结合笔者所在课题组前期对HuR的研究基础，就HuR在肿瘤多药耐药及药物敏感性中的作用简要作一综述。
2014, 21(6):712-720.
DOI: 10.3872/j.issn.1007-385X.2014.6.021
Abstract:
与放、化疗相比，肿瘤的靶向治疗具有较高的特异性和较低的不良反应，因而受到了广泛的关注。传统的靶向药物或干扰肿瘤的分裂增殖，或抑制癌细胞的侵袭转移，或克服肿瘤耐药。近年来，囊泡运输机制、DNA损伤修复机制和免疫系统靶向药物等相关研究在不断升温。其中，可通过囊泡运输进入成胶质瘤胞内的化合物Vacquinol-1已在小鼠实验中验证了其对肿瘤的杀伤作用。上述领域的细胞和分子生物学新突破，揭示着肿瘤靶向治疗理念与技术渐臻成熟，靶点类型也由传统的单一蛋白分子向信号转导通路的相互作用以及免疫微环境等整体协同效应的转变。
与放、化疗相比，肿瘤的靶向治疗具有较高的特异性和较低的不良反应，因而受到了广泛的关注。传统的靶向药物或干扰肿瘤的分裂增殖，或抑制癌细胞的侵袭转移，或克服肿瘤耐药。近年来，囊泡运输机制、DNA损伤修复机制和免疫系统靶向药物等相关研究在不断升温。其中，可通过囊泡运输进入成胶质瘤胞内的化合物Vacquinol-1已在小鼠实验中验证了其对肿瘤的杀伤作用。上述领域的细胞和分子生物学新突破，揭示着肿瘤靶向治疗理念与技术渐臻成熟，靶点类型也由传统的单一蛋白分子向信号转导通路的相互作用以及免疫微环境等整体协同效应的转变。
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Supported by:Beijing E-Tiller Technology Development Co., Ltd.