【摘要】 目的 研究肿瘤坏死因子相关凋亡诱导配体（TRAIL）蛋白对SKOV3移植瘤细胞半胱天冬氨酸蛋白酶-3（Caspase-3）表达的影响及其与肿瘤细胞凋亡的关系。 方法 建立雌性裸小鼠SKOV3移植瘤24只，随机分为4组，每组6只。TRAIL组单用重组人TRAIL蛋白（10 μg/kg），顺铂（DDP）组单用DDP（3 mg/kg），TRAIL+DDP组联合使用TRAIL蛋白（10 μg/kg）和DDP（3 mg/kg），空白对照组给予0.5 mL生理盐水。经处理后，各组的组织切片用免疫组织化学染色检测Caspase-3的表达和末端脱氧核苷酸转移酶介导核苷酸缺口标记技术（TUNEL）检测肿瘤细胞凋亡指数。 结果 Caspase-3的表达水平在TRAIL组（171.67±14.38）、DDP组（172.50±14.75）、联合组（230.00±40.99）中均明显高于对照组（135.83±16.25）（Plt;0.05）。SKOV3移植瘤细胞凋亡指数在空白对照组、TRAIL组、DDP组和联合组分别为16.67±5.43、33.17±8.42、24.33±4.59和40.50±6.16，TRAIL组和联合组细胞凋亡指数较空白对照组和DDP组明显增高（Plt;0.05）。 结论 TRAIL蛋白使卵巢癌移植瘤细胞的Caspase-3表达增强，TRAIL蛋白促进肿瘤细胞凋亡发生。【Abstract】 Objective To investigate the effects of Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) on the expression of Cysteine/aspartic acid specific protease- 3 (Caspase-3) in SKOV3 ovarian tumor cells and its relationship with the apoptosis of the ovarian tumor xenografts in nude mice. Methods Twenty-four nude mice with SKOV3 cell line ovarian tumor were randomly divided into four groups with 6 in each group. TRAIL (10 μg/kg) was given to the mice in the TRAIL group; DDP (3 μg/kg) was given to the mice in the DDP group; TRAIL (10 μg/kg) and DDP (3 μg/kg) were given to the mice in the TRAIL+DDP group; and 0.5 mL of saline solution was give to the mice in the control group. The expression of Caspase-3 was detected with immunohistochemistry. The apoptosis index (AI) of cells was determined by Terminal deoxynucleotidyl transferase mediated-dUTP nick end labeling (TUNEL). Results The expression of Caspase-3 in the TRAIL group (171.67±14.38), DDP group (172.50±14.75), and TRAIL+DDP group (230.00±40.99) was significantly higher than that in the control group (135.83±16.25) (Plt;0.05). The apoptosis index for the control group, TRAIL group, DDP group and TRAIL+DDP group was 16.67±5.43, 33.17±8.42, 24.33±4.59, and 40.50±6.16, respectively. The apoptosis index for the TRAIL group and the TRAIL+DDP group was significantly higher than that in the control group and the DDP group (Plt;0.05). Conclusion Soluble TRAIL has an effect on enhancing the expression of Caspase-3 in implanted tumor in nude mice. TRAIL protein can inhibit the growth of SKOV3 cells in nude mice by inducing cell apoptosis.
Objective To investigate the expressions of tumor necrosis factor related apoptosis inducing ligand （TRAIL） and its receptors (DR4, DcR1) in human rectal cancer tissues and normal rectal tissues. MethodsThe expressions of TRAIL and its receptors (DR4, DcR1) in 31 cases of human rectal cancer tissues and 20 cases of normal rectal tissues were detected by immunohistochemical staining. ResultsThe positive expression rates of TRAIL, DR4 and DcR1 （32.26%， 29.03%， 0） were lower than those of normal rectal tissues （55.00%， 70.00%， 65.00%）, the difference was statistically significant(P=0.015， P=0.000， P=0.000). There were no relation between the expressions of TRAIL, DR4 and DcR1 and clinicopathologic characteristics (Pgt;0.05). ConclusionThe expressions of TRAIL and its receptors (DR4, DcR1) in human rectal cancer tissues were lower than those of normal rectal tissues, which may suggest that the apoptotic effect induced by the interaction between TRAIL and its receptors has attenuated in human rectal cancer.
Objective To review the current researches about tumor necrosis factor-related apoptosis ligand (TRAIL) and its receptors in gastric carcinoma. Methods Relevant articles of researches on TRAIL and its receptors in gastric carcinoma were searched in electronic databases of PUB-MEDLINE and Chinese Journal Fulltext Database. Results The reported TRAIL expression level of gastric carcinoma was diverse, which was highly correlated to the histological differentiation degree, serosa invasion and lymph node metastasis. Its receptors DR4 and DR5 were both positive in gastric carcinoma tissue, while some researches reported DcR1 and DcR2 were also positive expressed. caspase-3, -8 and survivin were the important factors for regulation of TRAIL signal pathway. 5-Aza-CdR, doxorubicin, 5-fluorouracil, α-TOS and X-ray irradiation might enhance the TRAIL-induced apoptosis of gastric carcinoma cells. Conclusion Gastric carcinoma may be potentially sensitive to TRAIL targeting therapy, but the mechanism of TRAIL-induced apoptosis is quite complex and is regulated by multi-factors. Up to now, there are still many issues to research further, such as how to efficiently enhance and regulate the TRAIL-induced apoptosis of gastric carcinoma, whether any potential toxicities existing, etc.
【Abstract】 Objective To investigate the expression and significance of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) receptors in pancreatic cancer. Methods Thirty-two samples of pancreatic cancer tissue were collected from year 2002 to 2004. All of them were verified by histopathology and there were 9 cases of well-differentiated, 12 of moderately differentiated, and 11 of poorly differentiated, in which 12 cases were in the stage of Ⅰor Ⅱand 20 in the stage of Ⅲ or Ⅳ according to the TNM staging method. Eighteen normal pancreatic tissues were used as control group. The expressions of TRAIL receptors (death receptor 4, death receptor 5, decoy receptor 4 and decoy receptor 5) mRNA were assayed by semi-quantitive reverse transcription polymerase chain reaction (RT-PCR) in the pancreatic cancer tissues and the normal pancreatic tissues. Results The expressions of death receptor 4 (DR4) and death receptor 5 (DR5) were detected in all the pancreatic cancer tissues and the normal pancreatic tissues and the levels of DR4 and DR5 were significantly higher than those of the normal pancreatic tissues (P＜0.01). Decoy receptor 1 (DcR1) and decoy receptor 2 (DcR2) were also expressed in normal pancreatic tissues, whereas DcR1 and DcR2 were only expressed in 18 and 20 pancreatic cancer tissues, respectively. However, there were no significant difference of the expression of DcR1 and DcR2 between the pancreatic cancer tissues and the normal pancreatic tissues （Pgt;0.05）. The expression level of DR5 in pancreatic cancer tissue was correlated with tumor differentiation and clinical stage, and the levels in stage Ⅲ and stage Ⅳwere significantly lower than those of stageⅠand stageⅡ(P＜0.05). The expressions of DR4, DcR1 and DcR2 were not correlated with tumor differentiation and clinical stage (Pgt;0.05). Conclusion ①The expression of TRAIL receptors in pancreatic cancer tissues is prevalent, but the types of receptors expressed in different tissues were also different. High expression of death receptors may play an important role in TRAIL recptors regulated pancreatic cancer apoptosis. ②The expression of DR5 is correlated with the differentiation degree of pancreatic cancer cell and clinical stage of tumor. The expressions of DR4, DcR1 and DcR2 should not be considered as related indexes of differentiation degree or clinical stage of pancreatic cancer.
【Abstract】ObjectiveTo construct the eukaryotic expressing plasmid of tumor necrosis factor-related apoptosisinducing ligand（TRAIL），and study its inhibitory effect on hepatic tumor which implanted subcutaneously in nude BALB/c mice.MethodsTotal RNA of U937 cell was extracted, and its extracellular domain (114-281aa) was amplified by RTPCR, then signal peptide was ligated. The recombinant secreting plasmid for TRAIL was constructed successfully which was confirmed by enzyme cleavage identification and sequencing identification. Liver cancer cell (strain No.7402) was implanted subcutaneously in 32 nude BALB/c mice. These mice were randomly divided into two groups: study group and control group. The mice in study group received muscular injection of plasmids for transfection, and the mice in control group received the injection of normal saline at the same time. The size of implanted tumors were measured continuously till the day of sacrificing, tumor cell apoptosis effect was examined by TUNEL method. ResultsIn study group，tumor volume was smaller than that in control group and the bluepurple apoptosis cells were observed under microscope. ConclusionTRAIL plasmid can induce apoptosis of liver cancer cell and can inhibit the growth of liver tumor.
Human tumor necrosis factor-related apoptosis-inducing ligand (hTRAIL) might be developed as a novel anti-tumor drug due to its selective cytotoxicity in tumor cells. The predicted Macaca mulatta TRAIL (mmTRAIL) is highly homologous to hTRAIL in nucleotide acid as well as amino acid sequence, suggesting that mmTRAIL might induce apoptosis of human cancer cells. However, the cytotoxicity of mmTRAIL in human cancer cells has not been investigated. In this paper, it is reported that the gene encoding mmTRAIL has been cloned by using reverse-transcriptase polymerase chain reaction (RT-PCR) from monkey peripheral blood mononuclear cells (PBMCs) in our laboratory. Subsequently, an expression plasmid was constructed by inserting mmTRAIL gene into pQE30 plasmid. After induction by addition of Isopropylβ-D-1-Thiogalactopyranoside (IPTG), mmTRAIL was expressed. MmTRAIL was recovered from supernatant of sonicated bacteria by Ni-NTA agarose affinity chromatography. SDS-PAGE and gel filtration chromatography demonstrated that mmTRAIL forms trimer in solution. In vitro assays indicated that mmTRAIL was cytotoxic to human COLO205 tumor cells but not to normal cells at low concentration of nanomole. In addition, antitumor effect of mmTRAIL was evaluated in mice bearing human COLO205 tumor xenografts. Intratumorally injected mmTRAIL significantly inhibited growth of tumor grafts. These results suggested that mmTRAIL was valuable as candidate drug for cancer-targeted therapy.